Germline Stem Cells: Origin and Destiny

PubMed Central

Lehmann, Ruth

2012-01-01

Germline stem cells are key to genome transmission to future generations. Over recent years, there have been numerous insights into the regulatory mechanisms that govern both germ cell specification and the maintenance of the germline in adults. Complex regulatory interactions with both the niche and the environment modulate germline stem cell function. This perspective highlights some examples of this regulation to illustrate the diversity and complexity of the mechanisms involved. PMID:22704513

The microRNA-200 family coordinately regulates cell adhesion and proliferation in hair morphogenesis.

PubMed

Hoefert, Jaimee E; Bjerke, Glen A; Wang, Dongmei; Yi, Rui

2018-06-04

The microRNA (miRNA)-200 (miR-200) family is highly expressed in epithelial cells and frequently lost in metastatic cancer. Despite intensive studies into their roles in cancer, their targets and functions in normal epithelial tissues remain unclear. Importantly, it remains unclear how the two subfamilies of the five-miRNA family, distinguished by a single nucleotide within the seed region, regulate their targets. By directly ligating miRNAs to their targeted mRNA regions, we identify numerous miR-200 targets involved in the regulation of focal adhesion, actin cytoskeleton, cell cycle, and Hippo/Yap signaling. The two subfamilies bind to largely distinct target sites, but many genes are coordinately regulated by both subfamilies. Using inducible and knockout mouse models, we show that the miR-200 family regulates cell adhesion and orientation in the hair germ, contributing to precise cell fate specification and hair morphogenesis. Our findings demonstrate that combinatorial targeting of many genes is critical for miRNA function and provide new insights into miR-200's functions. © 2018 Hoefert et al.

Myb proteins: angels and demons in normal and transformed cells

PubMed Central

Zhou, Ye; Ness, Scott A.

2013-01-01

A key regulator of proliferation, differentiation and cell fate, the c-Myb transcription factor regulates the expression of hundreds of genes and is in turn regulated by numerous pathways and protein interactions. However, the most unique feature of c-Myb is that it can be converted into an oncogenic transforming protein through a few mutations that completely change its activity and specificity. The c-Myb protein is a myriad of interactions and activities rolled up in a protein that controls proliferation and differentiation in many different cell types. Here we discuss the background and recent progress that have led to a better understanding of this complex protein, and outline the questions that have yet to be answered. PMID:21196221

Collecting Duct Intercalated Cell Function and Regulation

PubMed Central

Roy, Ankita; Al-bataineh, Mohammad M.

2015-01-01

Intercalated cells are kidney tubule epithelial cells with important roles in the regulation of acid-base homeostasis. However, in recent years the understanding of the function of the intercalated cell has become greatly enhanced and has shaped a new model for how the distal segments of the kidney tubule integrate salt and water reabsorption, potassium homeostasis, and acid-base status. These cells appear in the late distal convoluted tubule or in the connecting segment, depending on the species. They are most abundant in the collecting duct, where they can be detected all the way from the cortex to the initial part of the inner medulla. Intercalated cells are interspersed among the more numerous segment-specific principal cells. There are three types of intercalated cells, each having distinct structures and expressing different ensembles of transport proteins that translate into very different functions in the processing of the urine. This review includes recent findings on how intercalated cells regulate their intracellular milieu and contribute to acid-base regulation and sodium, chloride, and potassium homeostasis, thus highlighting their potential role as targets for the treatment of hypertension. Their novel regulation by paracrine signals in the collecting duct is also discussed. Finally, this article addresses their role as part of the innate immune system of the kidney tubule. PMID:25632105

Mammalian Krüppel-Like Factors in Health and Diseases

PubMed Central

McConnell, Beth B.; Yang, Vincent W.

2010-01-01

The Krüppel-like factor (KLF) family of transcription factors regulates diverse biological processes that include proliferation, differentiation, growth, development, survival, and responses to external stress. Seventeen mammalian KLFs have been identified, and numerous studies have been published that describe their basic biology and contribution to human diseases. KLF proteins have received much attention because of their involvement in the development and homeostasis of numerous organ systems. KLFs are critical regulators of physiological systems that include the cardiovascular, digestive, respiratory, hematological, and immune systems and are involved in disorders such as obesity, cardiovascular disease, cancer, and inflammatory conditions. Furthermore, KLFs play an important role in reprogramming somatic cells into induced pluripotent stem (iPS) cells and maintaining the pluripotent state of embryonic stem cells. As research on KLF proteins progresses, additional KLF functions and associations with disease are likely to be discovered. Here, we review the current knowledge of KLF proteins and describe common attributes of their biochemical and physiological functions and their pathophysiological roles. PMID:20959618

NFκB signaling regulates embryonic and adult neurogenesis

PubMed Central

ZHANG, Yonggang; HU, Wenhui

2013-01-01

Both embryonic and adult neurogenesis involves the self-renewal/proliferation, survival, migration and lineage differentiation of neural stem/progenitor cells. Such dynamic process is tightly regulated by intrinsic and extrinsic factors and complex signaling pathways. Misregulated neurogenesis contributes much to a large range of neurodevelopmental defects and neurodegenerative diseases. The signaling of NFκB regulates many genes important in inflammation, immunity, cell survival and neural plasticity. During neurogenesis, NFκB signaling mediates the effect of numerous niche factors such as cytokines, chemokines, growth factors, extracellular matrix molecules, but also crosstalks with other signaling pathways such as Notch, Shh, Wnt/β-catenin. This review summarizes current progress on the NFκB signaling in all aspects of neurogenesis, focusing on the novel role of NFκB signaling in initiating early neural differentiation of neural stem cells and embryonic stem cells. PMID:24324484

Regulation of Memory T Cells by Interleukin-23.

PubMed

Li, Yanchun; Wang, Hongbo; Lu, Honghua; Hua, Shucheng

2016-01-01

Interleukin-23 (IL-23), a member of the IL-12 family of cytokines, is a heterodimeric cytokine. It is composed of subunits p40 (shared with IL-12) and p19 (an IL-12 p35-related subunit) and is secreted by several types of immune cells, such as natural killer cells and dendritic cells. The IL-23 receptor is composed of the subunit IL-12Rβ1 and the IL-23-specific subunit IL-23R. The binding of IL-23 to its specific cell surface receptor regulates a number of functions, including proliferation and differentiation of cells and secretion of cell factors. Memory T cells are a subset of T cells that secrete numerous important cell factors, and they function in the immune response to infection and diseases like cancer, autoimmune disease and bronchial asthma. IL-23R is expressed on the surface of memory T cells, which suggests that it can specifically regulate memory T cell function. IL-23 has been widely used as a clinical indicator in immune-related diseases and shows potential for use in disease treatment. Here we review the current progress in the study of the role of IL-23 in the regulation of memory T cells. © 2016 S. Karger AG, Basel.

Modulation of cell function by electric field: a high-resolution analysis

PubMed Central

Taghian, T.; Narmoneva, D. A.; Kogan, A. B.

2015-01-01

Regulation of cell function by a non-thermal, physiological-level electromagnetic field has potential for vascular tissue healing therapies and advancing hybrid bioelectronic technology. We have recently demonstrated that a physiological electric field (EF) applied wirelessly can regulate intracellular signalling and cell function in a frequency-dependent manner. However, the mechanism for such regulation is not well understood. Here, we present a systematic numerical study of a cell-field interaction following cell exposure to the external EF. We use a realistic experimental environment that also recapitulates the absence of a direct electric contact between the field-sourcing electrodes and the cells or the culture medium. We identify characteristic regimes and present their classification with respect to frequency, location, and the electrical properties of the model components. The results show a striking difference in the frequency dependence of EF penetration and cell response between cells suspended in an electrolyte and cells attached to a substrate. The EF structure in the cell is strongly inhomogeneous and is sensitive to the physical properties of the cell and its environment. These findings provide insight into the mechanisms for frequency-dependent cell responses to EF that regulate cell function, which may have important implications for EF-based therapies and biotechnology development. PMID:25994294

Anticancer effect of triterpenes from Ganoderma lucidum in human prostate cancer cells.

PubMed

Qu, Lijun; Li, Sumei; Zhuo, Yumin; Chen, Jianfan; Qin, Xiaoping; Guo, Guoqing

2017-12-01

Ganoderma lucidum , within the Polyporaceae family of Basidiomycota, is a popular traditional remedy medicine used in Asia to promote health and longevity. Compounds extracted from G. lucidum have revealed anticancer, antioxidant and liver protective effects. G. lucidum has been associated with prostate cancer cells. G. lucidum extracts contain numerous bioactive components; however, the exact functional monomer is unknown and the role of triterpenes from G. lucidum (GLT) in prostate cancer remain obscure. The present study investigated the effects of GLT on cell viability, migration, invasion and apoptosis in DU-145 human prostate cancer cells. The results demonstrated that a high dose (2 mg/ml) of GLT inhibits cell viability in a dose- and time-dependent manner by the regulation of matrix metalloproteases. Furthermore, GLT induced apoptosis of DU-145 cells. In general, GLT exerts its effect on cancer cells via numerous mechanisms and may have potential therapeutic use for the prevention and treatment of cancer.

Specificity of arrestin subtypes in regulating airway smooth muscle G protein-coupled receptor signaling and function.

PubMed

Pera, Tonio; Hegde, Akhil; Deshpande, Deepak A; Morgan, Sarah J; Tiegs, Brian C; Theriot, Barbara S; Choi, Yeon H; Walker, Julia K L; Penn, Raymond B

2015-10-01

Arrestins have been shown to regulate numerous G protein-coupled receptors (GPCRs) in studies employing receptor/arrestin overexpression in artificial cell systems. Which arrestin isoforms regulate which GPCRs in primary cell types is poorly understood. We sought to determine the effect of β-arrestin-1 or β-arrestin-2 inhibition or gene ablation on signaling and function of multiple GPCRs endogenously expressed in airway smooth muscle (ASM). In vitro [second messenger (calcium, cAMP generation)], ex vivo (ASM tension generation in suspended airway), and in vivo (invasive airway resistance) analyses were performed on human ASM cells and murine airways/whole animal subject to β-arrestin-1 or -2 knockdown or knockout (KO). In both human and murine model systems, knockdown or KO of β-arrestin-2 relative to control missense small interfering RNA or wild-type mice selectively increased (40-60%) β2-adrenoceptor signaling and function. β-arrestin-1 knockdown or KO had no effect on signaling and function of β2-adrenoceptor or numerous procontractile GPCRs, but selectively inhibited M3 muscarinic acetylcholine receptor signaling (∼50%) and function (∼25% ex vivo, >50% in vivo) without affecting EC50 values. Arrestin subtypes differentially regulate ASM GPCRs and β-arrestin-1 inhibition represents a novel approach to managing bronchospasm in obstructive lung diseases. © FASEB.

Regulation of PCP by the Fat signaling pathway

PubMed Central

Matis, Maja; Axelrod, Jeffrey D.

2013-01-01

Planar cell polarity (PCP) in epithelia, orthogonal to the apical–basal axis, is essential for numerous developmental events and physiological functions. Drosophila model systems have been at the forefront of studies revealing insights into mechanisms regulating PCP and have revealed distinct signaling modules. One of these, involving the atypical cadherins Fat and Dachsous and the ectokinase Four-jointed, appears to link the direction of cell polarization to the tissue axes. We discuss models for the function of this signaling module as well as several unanswered questions that may guide future investigations. PMID:24142873

AMP-activated protein kinase, stress responses and cardiovascular diseases

PubMed Central

WANG, Shaobin; SONG, Ping; ZOU, Ming-Hui

2012-01-01

AMPK (AMP-activated protein kinase) is one of the key players in maintaining intracellular homoeostasis. AMPK is well known as an energy sensor and can be activated by increased intracellular AMP levels. Generally, the activation of AMPK turns on catabolic pathways that generate ATP, while inhibiting cell proliferation and biosynthetic processes that consume ATP. In recent years, intensive investigations on the regulation and the function of AMPK indicates that AMPK not only functions as an intracellular energy sensor and regulator, but is also a general stress sensor that is important in maintaining intracellular homoeostasis during many kinds of stress challenges. In the present paper, we will review recent literature showing that AMPK functions far beyond its proposed energy sensor and regulator function. AMPK regulates ROS (reactive oxygen species)/redox balance, autophagy, cell proliferation, cell apoptosis, cellular polarity, mitochondrial function and genotoxic response, either directly or indirectly via numerous downstream pathways under physiological and pathological conditions. PMID:22390198

Managing the complexity of communication: regulation of gap junctions by post-translational modification

PubMed Central

Axelsen, Lene N.; Calloe, Kirstine; Holstein-Rathlou, Niels-Henrik; Nielsen, Morten S.

2013-01-01

Gap junctions are comprised of connexins that form cell-to-cell channels which couple neighboring cells to accommodate the exchange of information. The need for communication does, however, change over time and therefore must be tightly controlled. Although the regulation of connexin protein expression by transcription and translation is of great importance, the trafficking, channel activity and degradation are also under tight control. The function of connexins can be regulated by several post translational modifications, which affect numerous parameters; including number of channels, open probability, single channel conductance or selectivity. The most extensively investigated post translational modifications are phosphorylations, which have been documented in all mammalian connexins. Besides phosphorylations, some connexins are known to be ubiquitinated, SUMOylated, nitrosylated, hydroxylated, acetylated, methylated, and γ-carboxyglutamated. The aim of the present review is to summarize our current knowledge of post translational regulation of the connexin family of proteins. PMID:24155720

Study on the regulation of focal adesions and cortical actin by matrix nanotopography in 3D environment

NASA Astrophysics Data System (ADS)

Han, Jingjing; Lin, Keng-Hui; Chew, Lock Yue

2017-11-01

Matrix nanotopography plays an important role in regulating cell behaviors by providing spatial as well as mechanical cues for cells to sense. It has been proposed that nanoscale topography is possible to modulate the tensions which direct the formation of cytoskeleton and the organization of the membrane receptor within the cell, which in turn regulate intracellular mechanical and biochemical signaling. With current studies on this topic being performed mainly in 2D platforms, the question on how nanotopography can influence cell bahaviors in 3D environments has yet to be addressed. In this paper, we explored this question by placing cells in 3D hollow spherical polydimethylsiloxane scaffolds. After culturing rat embryonic fibroblast cells in two kinds of scaffold, one with smooth surface and the other with numerous nano-spikes, we observed that cells in the smooth scaffold have more anchoring sites and more focal adhesions than in the etched scaffold. Moreover, we found the presence of correlation between cortical actin, the important component for supporting cell attachment, and local cell geometry.

Biophysical Regulation of Cell Behavior—Cross Talk between Substrate Stiffness and Nanotopography

PubMed Central

Yang, Yong; Wang, Kai; Gu, Xiaosong; Leong, Kam W.

2017-01-01

The stiffness and nanotopographical characteristics of the extracellular matrix (ECM) influence numerous developmental, physiological, and pathological processes in vivo. These biophysical cues have therefore been applied to modulate almost all aspects of cell behavior, from cell adhesion and spreading to proliferation and differentiation. Delineation of the biophysical modulation of cell behavior is critical to the rational design of new biomaterials, implants, and medical devices. The effects of stiffness and topographical cues on cell behavior have previously been reviewed, respectively; however, the interwoven effects of stiffness and nanotopographical cues on cell behavior have not been well described, despite similarities in phenotypic manifestations. Herein, we first review the effects of substrate stiffness and nanotopography on cell behavior, and then focus on intracellular transmission of the biophysical signals from integrins to nucleus. Attempts are made to connect extracellular regulation of cell behavior with the biophysical cues. We then discuss the challenges in dissecting the biophysical regulation of cell behavior and in translating the mechanistic understanding of these cues to tissue engineering and regenerative medicine. PMID:29071164

An RNA-binding protein, Qki5, regulates embryonic neural stem cells through pre-mRNA processing in cell adhesion signaling.

PubMed

Hayakawa-Yano, Yoshika; Suyama, Satoshi; Nogami, Masahiro; Yugami, Masato; Koya, Ikuko; Furukawa, Takako; Zhou, Li; Abe, Manabu; Sakimura, Kenji; Takebayashi, Hirohide; Nakanishi, Atsushi; Okano, Hideyuki; Yano, Masato

2017-09-15

Cell type-specific transcriptomes are enabled by the action of multiple regulators, which are frequently expressed within restricted tissue regions. In the present study, we identify one such regulator, Quaking 5 (Qki5), as an RNA-binding protein (RNABP) that is expressed in early embryonic neural stem cells and subsequently down-regulated during neurogenesis. mRNA sequencing analysis in neural stem cell culture indicates that Qki proteins play supporting roles in the neural stem cell transcriptome and various forms of mRNA processing that may result from regionally restricted expression and subcellular localization. Also, our in utero electroporation gain-of-function study suggests that the nuclear-type Qki isoform Qki5 supports the neural stem cell state. We next performed in vivo transcriptome-wide protein-RNA interaction mapping to search for direct targets of Qki5 and elucidate how Qki5 regulates neural stem cell function. Combined with our transcriptome analysis, this mapping analysis yielded a bona fide map of Qki5-RNA interaction at single-nucleotide resolution, the identification of 892 Qki5 direct target genes, and an accurate Qki5-dependent alternative splicing rule in the developing brain. Last, our target gene list provides the first compelling evidence that Qki5 is associated with specific biological events; namely, cell-cell adhesion. This prediction was confirmed by histological analysis of mice in which Qki proteins were genetically ablated, which revealed disruption of the apical surface of the lateral wall in the developing brain. These data collectively indicate that Qki5 regulates communication between neural stem cells by mediating numerous RNA processing events and suggest new links between splicing regulation and neural stem cell states. © 2017 Hayakawa-Yano et al.; Published by Cold Spring Harbor Laboratory Press.

BARHL2 differentially regulates the development of retinal amacrine and ganglion neurons

PubMed Central

Ding, Qian; Chen, Hui; Xie, Xiaoling; Libby, Richard T.; Tian, Ning; Gan, Lin

2009-01-01

Summary Through transcriptional regulations the BarH family of homeodomain proteins play essential roles in cell fate specification, cell differentiation, migration and survival. Barhl2, a member of the Barh gene family, is expressed in retinal ganglion cells (RGCs), amacrine cells (ACs) and horizontal cells. Here, to investigate the role of Barhl2 in retinal development, Barhl2 deficient mice were generated. Analysis of AC subtypes in Barhl2 deficient retinas suggests that Barhl2 plays a critical role in AC subtype determination. A significant reduction of glycinergic and GABAergic ACs with a substantial increase in the number of cholinergic ACs was observed in Barhl2-null retinas. Barhl2 is also critical for the development of a normal complement of RGCs. Barhl2 deficiency resulted in a 35% increase in RGCs undergoing apoptosis during development. Genetic analysis revealed that Barhl2 functions downstream of the Atoh7-Pou4f3 regulatory pathway and regulates the maturation and/or survival of RGCs. Thus, BARHL2 appears to have numerous roles in retinal development, including regulating neuronal subtype specification, differentiation, and survival. PMID:19339595

Contrasting mechanisms of growth in two model rod-shaped bacteria

PubMed Central

Billaudeau, Cyrille; Chastanet, Arnaud; Yao, Zhizhong; Cornilleau, Charlène; Mirouze, Nicolas; Fromion, Vincent; Carballido-López, Rut

2017-01-01

How cells control their shape and size is a long-standing question in cell biology. Many rod-shaped bacteria elongate their sidewalls by the action of cell wall synthesizing machineries that are associated to actin-like MreB cortical patches. However, little is known about how elongation is regulated to enable varied growth rates and sizes. Here we use total internal reflection fluorescence microscopy and single-particle tracking to visualize MreB isoforms, as a proxy for cell wall synthesis, in Bacillus subtilis and Escherichia coli cells growing in different media and during nutrient upshift. We find that these two model organisms appear to use orthogonal strategies to adapt to growth regime variations: B. subtilis regulates MreB patch speed, while E. coli may mainly regulate the production capacity of MreB-associated cell wall machineries. We present numerical models that link MreB-mediated sidewall synthesis and cell elongation, and argue that the distinct regulatory mechanism employed might reflect the different cell wall integrity constraints in Gram-positive and Gram-negative bacteria. PMID:28589952

Selenium as an essential micronutrient: roles in cell cycle and apoptosis.

PubMed

Zeng, Huawei

2009-03-23

Selenium is an essential trace element for humans and animals, and selenium deficiency is associated with several disease conditions such as immune impairment. In addition, selenium intakes that are greater than the recommended daily allowance (RDA) appear to protect against certain types of cancers. In humans and animals, cell proliferation and death must be regulated to maintain tissue homeostasis, and it has been well documented that numerous human diseases are directly related to the control of cell cycle progression and apoptosis. Thus, the elucidation of the mechanisms by which selenium regulates the cell cycle and apoptosis can lead to a better understanding of the nature of selenium's essentiality and its role in disease prevention. This article reviews the status of knowledge concerning the effect of selenium on cell cycle and apoptosis.

MYBL2 (B-Myb): a central regulator of cell proliferation, cell survival and differentiation involved in tumorigenesis

PubMed Central

Musa, Julian; Aynaud, Marie-Ming; Mirabeau, Olivier; Delattre, Olivier; Grünewald, Thomas GP

2017-01-01

Limitless cell proliferation, evasion from apoptosis, dedifferentiation, metastatic spread and therapy resistance: all these properties of a cancer cell contribute to its malignant phenotype and affect patient outcome. MYBL2 (alias B-Myb) is a transcription factor of the MYB transcription factor family and a physiological regulator of cell cycle progression, cell survival and cell differentiation. When deregulated in cancer cells, MYBL2 mediates the deregulation of these properties. In fact, MYBL2 is overexpressed and associated with poor patient outcome in numerous cancer entities. MYBL2 and players of its downstream transcriptional network can be used as prognostic and/or predictive biomarkers as well as potential therapeutic targets to offer less toxic and more specific anti-cancer therapies in future. In this review, we summarize current knowledge on the physiological roles of MYBL2 and highlight the impact of its deregulation on cancer initiation and progression. PMID:28640249

Transcriptional and post-transcriptional regulation of NK cell development and function

PubMed Central

Leong, Jeffrey W.; Wagner, Julia A.; Ireland, Aaron R.; Fehniger, Todd A.

2016-01-01

Natural killer (NK) cells are specialized innate lymphoid cells that survey against viral infections and malignancy. Numerous advances have improved our understanding of the molecular mechanisms that control NK cell development and function over the past decade. These include both studies on the regulatory effects of transcription factors and translational repression via microRNAs. In this review, we summarize our current knowledge of DNA-binding transcription factors that regulate gene expression and thereby orchestrate NK cell development and activation, with an emphasis on recent discoveries. Additionally, we highlight our understanding of how RNA-bindings microRNAs fine tune the NK cell molecular program. We also underscore the large number of open questions in field that are now being addressed using new technological approaches and genetically engineered model organisms. Ultimately, a deeper understanding of the basic molecular biology of NK cells will facilitate new strategies to manipulate NK cells for the treatment of human disease. PMID:26948928

The cell biology of inflammasomes: Mechanisms of inflammasome activation and regulation

PubMed Central

2016-01-01

Over the past decade, numerous advances have been made in the role and regulation of inflammasomes during pathogenic and sterile insults. An inflammasome complex comprises a sensor, an adaptor, and a zymogen procaspase-1. The functional output of inflammasome activation includes secretion of cytokines, IL-1β and IL-18, and induction of an inflammatory form of cell death called pyroptosis. Recent studies have highlighted the intersection of this inflammatory response with fundamental cellular processes. Novel modulators and functions of inflammasome activation conventionally associated with the maintenance of homeostatic biological functions have been uncovered. In this review, we discuss the biological processes involved in the activation and regulation of the inflammasome. PMID:27325789

Suppression of B-cell development genes is key to glucocorticoid efficacy in treatment of acute lymphoblastic leukemia.

PubMed

Kruth, Karina A; Fang, Mimi; Shelton, Dawne N; Abu-Halawa, Ossama; Mahling, Ryan; Yang, Hongxing; Weissman, Jonathan S; Loh, Mignon L; Müschen, Markus; Tasian, Sarah K; Bassik, Michael C; Kampmann, Martin; Pufall, Miles A

2017-06-01

Glucocorticoids (GCs), including dexamethasone (dex), are a central component of combination chemotherapy for childhood B-cell precursor acute lymphoblastic leukemia (B-ALL). GCs work by activating the GC receptor (GR), a ligand-induced transcription factor, which in turn regulates genes that induce leukemic cell death. Which GR-regulated genes are required for GC cytotoxicity, which pathways affect their regulation, and how resistance arises are not well understood. Here, we systematically integrate the transcriptional response of B-ALL to GCs with a next-generation short hairpin RNA screen to identify GC-regulated "effector" genes that contribute to cell death, as well as genes that affect the sensitivity of B-ALL cells to dex. This analysis reveals a pervasive role for GCs in suppression of B-cell development genes that is linked to therapeutic response. Inhibition of phosphatidylinositol 3-kinase δ (PI3Kδ), a linchpin in the pre-B-cell receptor and interleukin 7 receptor signaling pathways critical to B-cell development (with CAL-101 [idelalisib]), interrupts a double-negative feedback loop, enhancing GC-regulated transcription to synergistically kill even highly resistant B-ALL with diverse genetic backgrounds. This work not only identifies numerous opportunities for enhanced lymphoid-specific combination chemotherapies that have the potential to overcome treatment resistance, but is also a valuable resource for understanding GC biology and the mechanistic details of GR-regulated transcription. © 2017 by The American Society of Hematology.

Characteristics of hepatic stem/progenitor cells in the fetal and adult liver.

PubMed

Koike, Hiroyuki; Taniguchi, Hideki

2012-11-01

The liver is an essential organ that maintains vital activity through its numerous important functions. It has a unique capability of fully regenerating after injury. Regulating a balance between self-renewal and differentiation of hepatic stem cells that are resources for functional mature liver cells is required for maintenance of tissue homeostasis. This review describes the characteristics of hepatic stem/progenitor cells and the regulatory mechanism of their self-renewal and differentiation capacity. In liver organogenesis, undifferentiated hepatic stem/progenitor cells expand their pool by repeated self-renewal in the early stage of liver development and then differentiate into two different types of cell lineage, namely hepatocytes and cholangiocytes. Liver development is regulated by expression of stem cell transcription factors in a complex multistep process. Recent studies suggest that stem cells are maintained by integrative regulation of gene expression patterns related to self-renewal and differentiation by epigenetic mechanisms such as histone modification and DNA methylation. Analysis of the proper regulatory mechanism of hepatic stem/progenitor cells is important for regenerative medicine that utilizes hepatic stem cells and for preventing liver cancer through clarification of the carcinogenetic mechanism involved in stem cell system failure.

Machineries regulating the activity of the small GTPase Arf6 in cancer cells are potential targets for developing innovative anti-cancer drugs.

PubMed

Yamauchi, Yohei; Miura, Yuki; Kanaho, Yasunori

2017-01-01

The Small GTPase ADP-ribosylation factor 6 (Arf6) functions as the molecular switch in cellular signaling pathways by cycling between GDP-bound inactive and GTP-bound active form, which is precisely regulated by two regulators, guanine nucleotide exchange factors (GEFs) and GTPase-activating proteins (GAPs). Numerous studies have shown that these machineries play critical roles in tumor angiogenesis/growth and cancer cell invasion/metastasis through regulating the cycling of Arf6. Here, we summarize accumulating knowledge for involvement of Arf6 GEFs/GAPs and small molecule inhibitors of Arf6 signaling/cycling in cancer progression, and discuss possible strategies for developing innovative anti-cancer drugs targeting Arf6 signaling/cycling. Copyright © 2016 Elsevier Ltd. All rights reserved.

Regulation of Glycan Structures in Animal Tissues

PubMed Central

Nairn, Alison V.; York, William S.; Harris, Kyle; Hall, Erica M.; Pierce, J. Michael; Moremen, Kelley W.

2008-01-01

Glycan structures covalently attached to proteins and lipids play numerous roles in mammalian cells, including protein folding, targeting, recognition, and adhesion at the molecular or cellular level. Regulating the abundance of glycan structures on cellular glycoproteins and glycolipids is a complex process that depends on numerous factors. Most models for glycan regulation hypothesize that transcriptional control of the enzymes involved in glycan synthesis, modification, and catabolism determines glycan abundance and diversity. However, few broad-based studies have examined correlations between glycan structures and transcripts encoding the relevant biosynthetic and catabolic enzymes. Low transcript abundance for many glycan-related genes has hampered broad-based transcript profiling for comparison with glycan structural data. In an effort to facilitate comparison with glycan structural data and to identify the molecular basis of alterations in glycan structures, we have developed a medium-throughput quantitative real time reverse transcriptase-PCR platform for the analysis of transcripts encoding glycan-related enzymes and proteins in mouse tissues and cells. The method employs a comprehensive list of >700 genes, including enzymes involved in sugar-nucleotide biosynthesis, transporters, glycan extension, modification, recognition, catabolism, and numerous glycosylated core proteins. Comparison with parallel microarray analyses indicates a significantly greater sensitivity and dynamic range for our quantitative real time reverse transcriptase-PCR approach, particularly for the numerous low abundance glycan-related enzymes. Mapping of the genes and transcript levels to their respective biosynthetic pathway steps allowed a comparison with glycan structural data and provides support for a model where many, but not all, changes in glycan abundance result from alterations in transcript expression of corresponding biosynthetic enzymes. PMID:18411279

Regulation of Mouse NK Cell Development and Function by Cytokines

PubMed Central

Marçais, Antoine; Viel, Sébastien; Grau, Morgan; Henry, Thomas; Marvel, Jacqueline; Walzer, Thierry

2013-01-01

Natural Killer (NK) cells are innate lymphocytes with an important role in the early defense against intracellular pathogens and against tumors. Like other immune cells, almost every aspects of their biology are regulated by cytokines. Interleukin (IL)-15 is pivotal for their development, homeostasis, and activation. Moreover, numerous other activating or inhibitory cytokines such as IL-2, IL-4, IL-7, IL-10, IL-12, IL-18, IL-21, Transforming growth factor-β (TGFβ) and type I interferons regulate their activation and their effector functions at different stages of the immune response. In this review we summarize the current understanding on the effect of these different cytokines on NK cell development, homeostasis, and functions during steady-state or upon infection by different pathogens. We try to delineate the cellular sources of these cytokines, the intracellular pathways they trigger and the transcription factors they regulate. We describe the known synergies or antagonisms between different cytokines and highlight outstanding questions in this field of investigation. Finally, we discuss how a better knowledge of cytokine action on NK cells could help improve strategies to manipulate NK cells in different clinical situations. PMID:24376448

A mex3 homolog is required for differentiation during planarian stem cell lineage development

PubMed Central

Zhu, Shu Jun; Hallows, Stephanie E; Currie, Ko W; Xu, ChangJiang; Pearson, Bret J

2015-01-01

Neoblasts are adult stem cells (ASCs) in planarians that sustain cell replacement during homeostasis and regeneration of any missing tissue. While numerous studies have examined genes underlying neoblast pluripotency, molecular pathways driving postmitotic fates remain poorly defined. In this study, we used transcriptional profiling of irradiation-sensitive and irradiation-insensitive cell populations and RNA interference (RNAi) functional screening to uncover markers and regulators of postmitotic progeny. We identified 32 new markers distinguishing two main epithelial progenitor populations and a planarian homolog to the MEX3 RNA-binding protein (Smed-mex3-1) as a key regulator of lineage progression. mex3-1 was required for generating differentiated cells of multiple lineages, while restricting the size of the stem cell compartment. We also demonstrated the utility of using mex3-1(RNAi) animals to identify additional progenitor markers. These results identified mex3-1 as a cell fate regulator, broadly required for differentiation, and suggest that mex3-1 helps to mediate the balance between ASC self-renewal and commitment. DOI: http://dx.doi.org/10.7554/eLife.07025.001 PMID:26114597

The cell adhesion molecule nectin-1 is critical for normal enamel formation in mice

PubMed Central

Barron, Martin J.; Brookes, Steven J.; Draper, Clare E.; Garrod, David; Kirkham, Jennifer; Shore, Roger C.; Dixon, Michael J.

2008-01-01

Nectin-1 is a member of a sub-family of immunoglobulin-like adhesion molecules and a component of adherens junctions. In the current study, we have shown that mice lacking nectin-1 exhibit defective enamel formation in their incisor teeth. Although the incisors of nectin-1-null mice were hypomineralized, the protein composition of the enamel matrix was unaltered. While strong immunostaining for nectin-1 was observed at the interface between the maturation-stage ameloblasts and the underlying cells of the stratum intermedium (SI), its absence in nectin-1-null mice correlated with separation of the cell layers at this interface. Numerous, large desmosomes were present at this interface in wild-type mice; however, where adhesion persisted in the mutant mice, the desmosomes were smaller and less numerous. Nectins have been shown to regulate tight junction formation; however, this is the first report showing that they may also participate in the regulation of desmosome assembly. Importantly, our results show that integrity of the SI–ameloblast interface is essential for normal enamel mineralization. PMID:18703497

Modern Perspectives on Numerical Modeling of Cardiac Pacemaker Cell

PubMed Central

Maltsev, Victor A.; Yaniv, Yael; Maltsev, Anna V.; Stern, Michael D.; Lakatta, Edward G.

2015-01-01

Cardiac pacemaking is a complex phenomenon that is still not completely understood. Together with experimental studies, numerical modeling has been traditionally used to acquire mechanistic insights in this research area. This review summarizes the present state of numerical modeling of the cardiac pacemaker, including approaches to resolve present paradoxes and controversies. Specifically we discuss the requirement for realistic modeling to consider symmetrical importance of both intracellular and cell membrane processes (within a recent “coupled-clock” theory). Promising future developments of the complex pacemaker system models include the introduction of local calcium control, mitochondria function, and biochemical regulation of protein phosphorylation and cAMP production. Modern numerical and theoretical methods such as multi-parameter sensitivity analyses within extended populations of models and bifurcation analyses are also important for the definition of the most realistic parameters that describe a robust, yet simultaneously flexible operation of the coupled-clock pacemaker cell system. The systems approach to exploring cardiac pacemaker function will guide development of new therapies, such as biological pacemakers for treating insufficient cardiac pacemaker function that becomes especially prevalent with advancing age. PMID:24748434

N-Cadherin and Fibroblast Growth Factor Receptors crosstalk in the control of developmental and cancer cell migrations.

PubMed

Nguyen, Thao; Mège, René Marc

2016-11-01

Cell migrations are diverse. They constitutemajor morphogenetic driving forces during embryogenesis, but they contribute also to the loss of tissue homeostasis and cancer growth. Capabilities of cells to migrate as single cells or as collectives are controlled by internal and external signalling, leading to the reorganisation of their cytoskeleton as well as by the rebalancing of cell-matrix and cell-cell adhesions. Among the genes altered in numerous cancers, cadherins and growth factor receptors are of particular interest for cell migration regulation. In particular, cadherins such as N-cadherin and a class of growth factor receptors, namely FGFRs cooperate to regulate embryonic and cancer cell behaviours. In this review, we discuss on reciprocal crosstalk between N-cadherin and FGFRs during cell migration. Finally, we aim at clarifying the synergy between N-cadherin and FGFR signalling that ensure cellular reorganization during cell movements, mainly during cancer cell migration and metastasis but also during developmental processes. Copyright © 2016 Elsevier GmbH. All rights reserved.

Activation of Gαq subunits up-regulates the expression of the tumor suppressor Fhit.

PubMed

Zuo, Hao; Chan, Anthony S L; Ammer, Hermann; Wong, Yung H

2013-12-01

The tumor suppressor Fhit protein is defective or absent in many tumor cells due to methylation, mutation or deletion of the FHIT gene. Despite numerous attempts to unravel the functions of Fhit, the mechanisms by which the function and expression of Fhit are regulated remain poorly understood. We have recently shown that activated Gαq subunits interact directly with Fhit and enhance its inhibitory effect on cell growth. Here we investigated the regulation of Fhit expression by Gq. Our results showed that Fhit was up-regulated specifically by activating Gα subunits of the Gq subfamily but not by those of the other G protein subfamilies. This up-regulation effect was mediated by a PKC/MEK pathway independent of Src-mediated Fhit Tyr(114) phosphorylation. We further demonstrated that elevated Fhit expression was due to the specific regulation of Fhit protein synthesis in the ribosome by activated Gαq, where the regulations of cap-dependent protein synthesis were apparently not required. Moreover, we showed that activated Gαq could increase cell-cell adhesion through Fhit. These findings provide a possible handle to modulate the level of the Fhit tumor suppressor by manipulating the activity of Gq-coupled receptors. © 2013. Published by Elsevier Inc. All rights reserved.

Quantitative High-Throughput Screening and Orthogonal Assays to Identify Modulators of the Vitamin D Receptor (SETAC)

EPA Science Inventory

The Vitamin D nuclear receptor (VDR) is a selective, ligand-inducible transcription factor involved in numerous biological processes such as cell proliferation, differentiation, detoxification, calcium homeostasis, neurodevelopment, immune system regulation, cardiovascular functi...

Yeast as a model to study apoptosis?

PubMed

Fleury, Christophe; Pampin, Mathieu; Tarze, Agathe; Mignotte, Bernard

2002-02-01

Programmed cell death (PCD) serves as a major mechanism for the precise regulation of cell numbers, and as a defense mechanism to remove unwanted and potentially dangerous cells. Despite the striking heterogeneity of cell death induction pathways, the execution of the death program is often associated with characteristic morphological and biochemical changes termed apoptosis. Although for a long time the absence of mitochondrial changes was considered as a hallmark of apoptosis, mitochondria appear today as the central executioner of programmed cell death. This crucial position of mitochondria in programmed cell death control is not due to a simple loss of function (deficit in energy supplying), but rather to an active process in the regulation of effector mechanisms. The large diversity of regulators of apoptosis in mammals and their numerous interactions complicate the analysis of their individual functions. Yeast, eukaryotic but unicellular organism, lack the main regulators of apoptosis (caspases, Bcl-2 family members, ...) found in mammals. This absence render them a powerful tool for heterologous expression, functional studies, and even cloning of new regulators of apoptosis. Great advances have thus been made in our understanding of the molecular mechanisms of Bcl-2 family members interactions with themselves and other cellular proteins, specially thanks to the two hybrid system and the easy manipulation of yeast (molecular biology and genetics). This review will focus on the use of yeast as a tool to identify new regulators and study function of mammalian apoptosis regulators.

Architecture of GnRH-Gonadotrope-Vasculature Reveals a Dual Mode of Gonadotropin Regulation in Fish.

PubMed

Golan, Matan; Zelinger, Einat; Zohar, Yonathan; Levavi-Sivan, Berta

2015-11-01

The function and components of the hypothalamic-pituitary axis are conserved among vertebrates; however, in fish, a neuroglandular mode of delivery (direct contact between axons and endocrine cells) was considered dominant, whereas in tetrapods hypothalamic signals are relayed to their targets via the hypophysial portal blood system (neurovascular delivery mode). By using a transgenic zebrafish model we studied the functional and anatomical aspects of gonadotrope regulation thus revisiting the existing model. FSH cells were found to be situated close to the vasculature whereas the compact organization of LH cells prevented direct contact of all cells with the circulation. GnRH3 fibers formed multiple boutons upon reaching the pituitary, but most of these structures were located in the neurohypophysis rather than adjacent to gonadotropes. A close association was observed between FSH cells and GnRH3 boutons, but only a fifth of the LH cells were in direct contact with GnRH3 axons, suggesting that FSH cells are more directly regulated than LH cells. GnRH3 fibers closely followed the vasculature in the neurohypophysis and formed numerous boutons along these tracts. These vessels were found to be permeable to relatively large molecules, suggesting the uptake of GnRH3 peptides. Our findings have important implications regarding the differential regulation of LH and FSH and contradict the accepted notion that fish pituitary cells are mostly regulated directly by hypothalamic fibers. Instead, we provide evidence that zebrafish apply a dual mode of gonadotrope regulation by GnRH3 that combines both neuroglandular and neurovascular components.

Eukaryotic elongation factor 2 kinase regulates the synthesis of microtubule-related proteins in neurons.

PubMed

Kenney, Justin W; Genheden, Maja; Moon, Kyung-Mee; Wang, Xuemin; Foster, Leonard J; Proud, Christopher G

2016-01-01

Modulation of the elongation phase of protein synthesis is important for numerous physiological processes in both neurons and other cell types. Elongation is primarily regulated via eukaryotic elongation factor 2 kinase (eEF2K). However, the consequence of altering eEF2K activity on the synthesis of specific proteins is largely unknown. Using both pharmacological and genetic manipulations of eEF2K combined with two protein-labeling techniques, stable isotope labeling of amino acids in cell culture and bio-orthogonal non-canonical amino acid tagging, we identified a subset of proteins whose synthesis is sensitive to inhibition of eEF2K in murine primary cortical neurons. Gene ontology (GO) analyses indicated that processes related to microtubules are particularly sensitive to eEF2K inhibition. Our findings suggest that eEF2K likely contributes to neuronal function by regulating the synthesis of microtubule-related proteins. Modulation of the elongation phase of protein synthesis is important for numerous physiological processes in neurons. Here, using labeling of new proteins coupled with proteomic techniques in primary cortical neurons, we find that the synthesis of microtubule-related proteins is up-regulated by inhibition of elongation. This suggests that translation elongation is a key regulator of cytoskeletal dynamics in neurons. © 2015 The Authors. Journal of Neurochemistry published by John Wiley & Sons Ltd on behalf of International Society for Neurochemistry.

mTORβ Splicing Isoform Promotes Cell Proliferation and Tumorigenesis*

PubMed Central

Panasyuk, Ganna; Nemazanyy, Ivan; Zhyvoloup, Aleksander; Filonenko, Valeriy; Davies, Derek; Robson, Mathew; Pedley, R. Barbara; Waterfield, Michael; Gout, Ivan

2009-01-01

The mTOR (mammalian target of rapamycin) promotes growth in response to nutrients and growth factors and is deregulated in numerous pathologies, including cancer. The mechanisms by which mTOR senses and regulates energy metabolism and cell growth are relatively well understood, whereas the molecular events underlining how it mediates survival and proliferation remain to be elucidated. Here, we describe the existence of the mTOR splicing isoform, TORβ, which, in contrast to the full-length protein (mTORα), has the potential to regulate the G1 phase of the cell cycle and to stimulate cell proliferation. mTORβ is an active protein kinase that mediates downstream signaling through complexing with Rictor and Raptor proteins. Remarkably, overexpression of mTORβ transforms immortal cells and is tumorigenic in nude mice and therefore could be a proto-oncogene. PMID:19726679

Reversible effect of all-trans-retinoic acid on AML12 hepatocyte proliferation and cell cycle progression

EPA Science Inventory

The role of all-trans-retinoic acid (atRA) in the regulation of cellular proliferation and differentiation is well documented. Numerous studies have established the cancer preventive propertiesofatRAwhichfunctionstoregulate levels ofcellcycleproteinsessentialfortheGliS transition...

The plasma membrane as a capacitor for energy and metabolism.

PubMed

Ray, Supriyo; Kassan, Adam; Busija, Anna R; Rangamani, Padmini; Patel, Hemal H

2016-02-01

When considering which components of the cell are the most critical to function and physiology, we naturally focus on the nucleus, the mitochondria that regulate energy and apoptotic signaling, or other organelles such as the endoplasmic reticulum, Golgi, ribosomes, etc. Few people will suggest that the membrane is the most critical element of a cell in terms of function and physiology. Those that consider the membrane critical will point to its obvious barrier function regulated by the lipid bilayer and numerous ion channels that regulate homeostatic gradients. What becomes evident upon closer inspection is that not all membranes are created equal and that there are lipid-rich microdomains that serve as platforms of signaling and a means of communication with the intracellular environment. In this review, we explore the evolution of membranes, focus on lipid-rich microdomains, and advance the novel concept that membranes serve as "capacitors for energy and metabolism." Within this framework, the membrane then is the primary and critical regulator of stress and disease adaptation of the cell.

AMPK governs lineage specification through Tfeb-dependent regulation of lysosomes.

PubMed

Young, Nathan P; Kamireddy, Anwesh; Van Nostrand, Jeanine L; Eichner, Lillian J; Shokhirev, Maxim Nikolaievich; Dayn, Yelena; Shaw, Reuben J

2016-03-01

Faithful execution of developmental programs relies on the acquisition of unique cell identities from pluripotent progenitors, a process governed by combinatorial inputs from numerous signaling cascades that ultimately dictate lineage-specific transcriptional outputs. Despite growing evidence that metabolism is integrated with many molecular networks, how pathways that control energy homeostasis may affect cell fate decisions is largely unknown. Here, we show that AMP-activated protein kinase (AMPK), a central metabolic regulator, plays critical roles in lineage specification. Although AMPK-deficient embryonic stem cells (ESCs) were normal in the pluripotent state, these cells displayed profound defects upon differentiation, failing to generate chimeric embryos and preferentially adopting an ectodermal fate at the expense of the endoderm during embryoid body (EB) formation. AMPK(-/-) EBs exhibited reduced levels of Tfeb, a master transcriptional regulator of lysosomes, leading to diminished endolysosomal function. Remarkably, genetic loss of Tfeb also yielded endodermal defects, while AMPK-null ESCs overexpressing this transcription factor normalized their differential potential, revealing an intimate connection between Tfeb/lysosomes and germ layer specification. The compromised endolysosomal system resulting from AMPK or Tfeb inactivation blunted Wnt signaling, while up-regulating this pathway restored expression of endodermal markers. Collectively, these results uncover the AMPK pathway as a novel regulator of cell fate determination during differentiation. © 2016 Young et al.; Published by Cold Spring Harbor Laboratory Press.

PARP-2 regulates cell cycle-related genes through histone deacetylation and methylation independently of poly(ADP-ribosyl)ation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liang, Ya-Chen; Hsu, Chiao-Yu; Yao, Ya-Li

2013-02-01

Highlights: ► PARP-2 acts as a transcription co-repressor independently of PARylation activity. ► PARP-2 recruits HDAC5, 7, and G9a and generates repressive chromatin. ► PARP-2 is recruited to the c-MYC promoter by DNA-binding factor YY1. ► PARP-2 represses cell cycle-related genes and alters cell cycle progression. -- Abstract: Poly(ADP-ribose) polymerase-2 (PARP-2) catalyzes poly(ADP-ribosyl)ation (PARylation) and regulates numerous nuclear processes, including transcription. Depletion of PARP-2 alters the activity of transcription factors and global gene expression. However, the molecular action of how PARP-2 controls the transcription of target promoters remains unclear. Here we report that PARP-2 possesses transcriptional repression activity independently ofmore » its enzymatic activity. PARP-2 interacts and recruits histone deacetylases HDAC5 and HDAC7, and histone methyltransferase G9a to the promoters of cell cycle-related genes, generating repressive chromatin signatures. Our findings propose a novel mechanism of PARP-2 in transcriptional regulation involving specific protein–protein interactions and highlight the importance of PARP-2 in the regulation of cell cycle progression.« less

Nuclear Lamins

PubMed Central

Dechat, Thomas; Adam, Stephen A.; Taimen, Pekka; Shimi, Takeshi; Goldman, Robert D.

2010-01-01

The nuclear lamins are type V intermediate filament proteins that are critically important for the structural properties of the nucleus. In addition, they are involved in the regulation of numerous nuclear processes, including DNA replication, transcription and chromatin organization. The developmentally regulated expression of lamins suggests that they are involved in cellular differentiation. Their assembly dynamic properties throughout the cell cycle, particularly in mitosis, are influenced by posttranslational modifications. Lamins may regulate nuclear functions by direct interactions with chromatin and determining the spatial organization of chromosomes within the nuclear space. They may also regulate chromatin functions by interacting with factors that epigenetically modify the chromatin or directly regulate replication or transcription. PMID:20826548

Interview: from Down's syndrome to basic epigenetics and back again.

PubMed

Lawrence, Jeanne; Telfer, Caroline

2013-12-01

Dr Jeanne Lawrence talks to Caroline Telfer, Commissioning Editor. Dr Jeanne Lawrence is an internationally recognized leader in the study of chromosome regulation by noncoding RNA and nuclear and genome organization. Her research bridges fundamental questions about genome regulation with clinical implications of recent advances in epigenetics. Her interest in chromosome structure and regulation has been a theme throughout her career and she has been honored for her work developing sensitive FISH technology for the detection of single copy genes, as well as RNAs. Her laboratory's publications include the initial demonstration of cell type-specific gene organization with nuclear subdomains; the novel biology of a noncoding RNA, XIST, which coats a whole X-chromosome to induce its silencing; and a new architectural role for a large noncoding RNA to scaffold a nuclear body. Her laboratory's work on epigenetic chromosome regulation in stem cells led to recent studies regarding unanticipated roles of repeat sequences in normal chromosome regulation and deregulation in cancer. Most recently, her laboratory has demonstrated a new approach to translate the basic mechanism of X-chromosome inactivation to correct a chromosomal dosage imbalance in patient-derived cells with trisomy 21 (Down's syndrome). Dr Lawrence has received awards from numerous agencies, including a Research Career Development Award from the National Center for Human Genome Research, career awards from the American Society of Cell Biology, the German Society for Biochemistry, the Muscular Dystrophy Association and a John Merck Fund Translational Research Award. She has served on the NIH National Advisory Council for Human Genome Research, numerous study sections and is currently a monitoring editor for the Journal of Cell Biology. Dr Lawrence has a BA in Biology and Music from Stephens College (MO, USA), a MS in Human Genetics and Genetic Counseling from Rutgers University (NJ, USA) and a PhD in Developmental Biology from Brown University (RI, USA). She is currently a Professor and Interim Chair of the Department of Cell and Developmental Biology at the University of Massachusetts Medical School (MA, USA).

Function of alternative splicing

PubMed Central

Kelemen, Olga; Convertini, Paolo; Zhang, Zhaiyi; Wen, Yuan; Shen, Manli; Falaleeva, Marina; Stamm, Stefan

2017-01-01

Almost all polymerase II transcripts undergo alternative pre-mRNA splicing. Here, we review the functions of alternative splicing events that have been experimentally determined. The overall function of alternative splicing is to increase the diversity of mRNAs expressed from the genome. Alternative splicing changes proteins encoded by mRNAs, which has profound functional effects. Experimental analysis of these protein isoforms showed that alternative splicing regulates binding between proteins, between proteins and nucleic acids as well as between proteins and membranes. Alternative splicing regulates the localization of proteins, their enzymatic properties and their interaction with ligands. In most cases, changes caused by individual splicing isoforms are small. However, cells typically coordinate numerous changes in ‘splicing programs’, which can have strong effects on cell proliferation, cell survival and properties of the nervous system. Due to its widespread usage and molecular versatility, alternative splicing emerges as a central element in gene regulation that interferes with almost every biological function analyzed. PMID:22909801

A role for the geomagnetic field in cell regulation.

PubMed

Liboff, A R

2010-08-01

We advance the hypothesis that biological systems utilize the geomagnetic field (GMF) for functional purposes by means of ion cyclotron resonance-like (ICR) mechanisms. Numerous ICR-designed experiments have demonstrated that living things are sensitive, in varying degrees, to magnetic fields that are equivalent to both changes in the general magnetostatic intensity of the GMF, as well as its temporal perturbations. We propose the existence of ICR-like cell regulation processes, homologous to the way that biochemical messengers alter the net biological state through competing processes of enhancement and inhibition. In like manner, combinations of different resonance frequencies all coupled to the same local magnetic field provide a unique means for cell regulation. Recent work on ultraweak ICR magnetic fields by Zhadin and others fits into our proposed framework if one assumes that cellular systems generate time-varying electric fields of the order 100 mV/cm with bandwidths that include relevant ICR frequencies.

Cell shape and negative links in regulatory motifs together control spatial information flow in signaling networks.

PubMed

Neves, Susana R; Tsokas, Panayiotis; Sarkar, Anamika; Grace, Elizabeth A; Rangamani, Padmini; Taubenfeld, Stephen M; Alberini, Cristina M; Schaff, James C; Blitzer, Robert D; Moraru, Ion I; Iyengar, Ravi

2008-05-16

The role of cell size and shape in controlling local intracellular signaling reactions, and how this spatial information originates and is propagated, is not well understood. We have used partial differential equations to model the flow of spatial information from the beta-adrenergic receptor to MAPK1,2 through the cAMP/PKA/B-Raf/MAPK1,2 network in neurons using real geometries. The numerical simulations indicated that cell shape controls the dynamics of local biochemical activity of signal-modulated negative regulators, such as phosphodiesterases and protein phosphatases within regulatory loops to determine the size of microdomains of activated signaling components. The model prediction that negative regulators control the flow of spatial information to downstream components was verified experimentally in rat hippocampal slices. These results suggest a mechanism by which cellular geometry, the presence of regulatory loops with negative regulators, and key reaction rates all together control spatial information transfer and microdomain characteristics within cells.

Critical protein GAPDH and its regulatory mechanisms in cancer cells

PubMed Central

Zhang, Jin-Ying; Zhang, Fan; Hong, Chao-Qun; Giuliano, Armando E.; Cui, Xiao-Jiang; Zhou, Guang-Ji; Zhang, Guo-Jun; Cui, Yu-Kun

2015-01-01

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH), initially identified as a glycolytic enzyme and considered as a housekeeping gene, is widely used as an internal control in experiments on proteins, mRNA, and DNA. However, emerging evidence indicates that GAPDH is implicated in diverse functions independent of its role in energy metabolism; the expression status of GAPDH is also deregulated in various cancer cells. One of the most common effects of GAPDH is its inconsistent role in the determination of cancer cell fate. Furthermore, studies have described GAPDH as a regulator of cell death; other studies have suggested that GAPDH participates in tumor progression and serves as a new therapeutic target. However, related regulatory mechanisms of its numerous cellular functions and deregulated expression levels remain unclear. GAPDH is tightly regulated at transcriptional and posttranscriptional levels, which are involved in the regulation of diverse GAPDH functions. Several cancer-related factors, such as insulin, hypoxia inducible factor-1 (HIF-1), p53, nitric oxide (NO), and acetylated histone, not only modulate GAPDH gene expression but also affect protein functions via common pathways. Moreover, posttranslational modifications (PTMs) occurring in GAPDH in cancer cells result in new activities unrelated to the original glycolytic function of GAPDH. In this review, recent findings related to GAPDH transcriptional regulation and PTMs are summarized. Mechanisms and pathways involved in GAPDH regulation and its different roles in cancer cells are also described. PMID:25859407

Cell-stimulation therapy of lateral epicondylitis with frequency-modulated low-intensity electric current.

PubMed

Aliyev, R M; Geiger, G

2012-03-01

In addition to the routine therapy, the patients with lateral epicondylitis included into experimental group were subjected to a 12-week cell-stimulation therapy with low-intensity frequency-modulated electric current. The control group received the same routine therapy and sham stimulation (the therapeutic apparatus was not energized). The efficiency of this microcurrent therapy was estimated by comparing medical indices before therapy and at the end of a 12-week therapeutic course using a 10-point pain severity numeric rating scale (NRS) and Roles-Maudsley pain score. The study revealed high therapeutic efficiency of cell-stimulation with low-intensity electric current resulting probably from up-regulation of intracellular transmitters, interleukins, and prostaglandins playing the key role in the regulation of inflammation.

Human population-specific gene expression and transcriptional network modification with polymorphic transposable elements

PubMed Central

Wang, Lu; Mariño-Ramírez, Leonardo

2017-01-01

Abstract Transposable element (TE) derived sequences are known to contribute to the regulation of the human genome. The majority of known TE-derived regulatory sequences correspond to relatively ancient insertions, which are fixed across human populations. The extent to which human genetic variation caused by recent TE activity leads to regulatory polymorphisms among populations has yet to be thoroughly explored. In this study, we searched for associations between polymorphic TE (polyTE) loci and human gene expression levels using an expression quantitative trait loci (eQTL) approach. We compared locus-specific polyTE insertion genotypes to B cell gene expression levels among 445 individuals from 5 human populations. Numerous human polyTE loci correspond to both cis and trans eQTL, and their regulatory effects are directly related to cell type-specific function in the immune system. PolyTE loci are associated with differences in expression between European and African population groups, and a single polyTE loci is indirectly associated with the expression of numerous genes via the regulation of the B cell-specific transcription factor PAX5. The polyTE-gene expression associations we found indicate that human TE genetic variation can have important phenotypic consequences. Our results reveal that TE-eQTL are involved in population-specific gene regulation as well as transcriptional network modification. PMID:27998931

Protein O-linked ß-N-acetylglucosamine: A novel effector of cardiomyocyte metabolism and function

PubMed Central

Darley-Usmar, Victor M.; Ball, Lauren E.; Chatham, John C.

2014-01-01

The post-translational modification of serine and threonine residues of nuclear and cytoplasmic proteins by the O-linked attachment of the monosaccharide ß-N-acetyl-glucosamine (O-GlcNAc) is emerging as an important mechanism for the regulation of numerous biological processes critical for normal cell function. Active synthesis of O-GlcNAc is essential for cell viability and acute activation of pathways resulting in increased protein O-GlcNAc levels improves the tolerance of cells to a wide range of stress stimuli. Conversely sustained increases in O-GlcNAc levels have been implicated in numerous chronic disease states, especially as a pathogenic contributor to diabetic complications. There has been increasing interest in the role of O-GlcNAc in the heart and vascular system and acute activation of O-GlcNAc levels have been shown to reduce ischemia/reperfusion injury attenuate vascular injury responses as well mediate some of the detrimental effects of diabetes and hypertension on cardiac and vascular function. Here we provide an overview of our current understanding of pathways regulating protein O-GlcNAcylation, summarize the different methodologies for identifying and characterizing O-GlcNAcylated proteins and subsequently focus on two emerging areas: 1) the role of O-GlcNAc as a potential regulator of cardiac metabolism and 2) the cross talk between O-GlcNAc and reactive oxygen species. PMID:21878340

The forkhead-like transcription factor (Fhl1p) maintains yeast replicative lifespan by regulating ribonucleotide reductase 1 (RNR1) gene transcription

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tai, Akiko; Kamei, Yuka; Mukai, Yukio

In eukaryotes, numerous genetic factors contribute to the lifespan including metabolic enzymes, signal transducers, and transcription factors. As previously reported, the forkhead-like transcription factor (FHL1) gene was required for yeast replicative lifespan and cell proliferation. To determine how Fhl1p regulates the lifespan, we performed a DNA microarray analysis of a heterozygous diploid strain deleted for FHL1. We discovered numerous Fhl1p-target genes, which were then screened for lifespan-regulating activity. We identified the ribonucleotide reductase (RNR) 1 gene (RNR1) as a regulator of replicative lifespan. RNR1 encodes a large subunit of the RNR complex, which consists of two large (Rnr1p/Rnr3p) and twomore » small (Rnr2p/Rnr4p) subunits. Heterozygous deletion of FHL1 reduced transcription of RNR1 and RNR3, but not RNR2 and RNR4. Chromatin immunoprecipitation showed that Fhl1p binds to the promoter regions of RNR1 and RNR3. Cells harboring an RNR1 deletion or an rnr1-C428A mutation, which abolishes RNR catalytic activity, exhibited a short lifespan. In contrast, cells with a deletion of the other RNR genes had a normal lifespan. Overexpression of RNR1, but not RNR3, restored the lifespan of the heterozygous FHL1 mutant to the wild-type (WT) level. The Δfhl1/FHL1 mutant conferred a decrease in dNTP levels and an increase in hydroxyurea (HU) sensitivity. These findings reveal that Fhl1p regulates RNR1 gene transcription to maintain dNTP levels, thus modulating longevity by protection against replication stress. - Highlights: • Fhl1p regulates replicative lifespan and transcription of RNR large subunit genes. • Rnr1p uniquely acts as a lifespan regulator independent of the RNR complex. • dNTP levels modulate longevity by protection against replication stress.« less

Insulin-regulated aminopeptidase immunoreactivity is abundantly present in human hypothalamus and posterior pituitary gland, with reduced expression in paraventricular and suprachiasmatic neurons in chronic schizophrenia.

PubMed

Bernstein, Hans-Gert; Müller, Susan; Dobrowolny, Hendrik; Wolke, Carmen; Lendeckel, Uwe; Bukowska, Alicja; Keilhoff, Gerburg; Becker, Axel; Trübner, Kurt; Steiner, Johann; Bogerts, Bernhard

2017-08-01

The vasopressin- and oxytocin-degrading enzyme insulin-regulated aminopeptidase (IRAP) is expressed in various organs including the brain. However, knowledge about its presence in human hypothalamus is fragmentary. Functionally, for a number of reasons (genetic linkage, hydrolysis of oxytocin and vasopressin, its role as angiotensin IV receptor in learning and memory and others) IRAP might play a role in schizophrenia. We studied the regional and cellular localization of IRAP in normal human brain with special emphasis on the hypothalamus and determined numerical densities of IRAP-expressing cells in the paraventricular, supraoptic and suprachiasmatic nuclei in schizophrenia patients and controls. By using immunohistochemistry and Western blot analysis, IRAP was immunolocalized in postmortem human brains. Cell countings were performed to estimate numbers and numerical densities of IRAP immunoreactive hypothalamic neurons in schizophrenia patients and control cases. Shape, size and regional distribution of IRAP-expressing cells, as well the lack of co-localization with the glia marker glutamine synthetase, show that IRAP is expressed in neurons. IRAP immunoreactive cells were observed in the hippocampal formation, cerebral cortex, thalamus, amygdala and, abundantly, hypothalamus. Double labeling experiments (IRAP and oxytocin/neurophysin 1, IRAP with vasopressin/neurophysin 2) revealed that IRAP is present in oxytocinergic and in vasopressinergic neurons. In schizophrenia patients, the numerical density of IRAP-expressing neurons in the paraventricular and the suprachiasmatic nuclei is significantly reduced, which might be associated with the reduction in neurophysin-containing neurons in these nuclei in schizophrenia. The pathophysiological role of lowered hypothalamic IRAP expression in schizophrenia remains to be established.

Natural Compounds as Modulators of Cell Cycle Arrest: Application for Anticancer Chemotherapies

PubMed Central

Bailon-Moscoso, Natalia; Cevallos-Solorzano, Gabriela; Romero-Benavides, Juan Carlos; Orellana, Maria Isabel Ramirez

2017-01-01

Natural compounds from various plants, microorganisms and marine species play an important role in the discovery novel components that can be successfully used in numerous biomedical applications, including anticancer therapeutics. Since uncontrolled and rapid cell division is a hallmark of cancer, unraveling the molecular mechanisms underlying mitosis is key to understanding how various natural compounds might function as inhibitors of cell cycle progression. A number of natural compounds that inhibit the cell cycle arrest have proven effective for killing cancer cells in vitro, in vivo and in clinical settings. Significant advances that have been recently made in the understanding of molecular mechanisms underlying the cell cycle regulation using the chemotherapeutic agents is of great importance for improving the efficacy of targeted therapeutics and overcoming resistance to anticancer drugs, especially of natural origin, which inhibit the activities of cyclins and cyclin-dependent kinases, as well as other proteins and enzymes involved in proper regulation of cell cycle leading to controlled cell proliferation. PMID:28367072

PHF6 regulates phenotypic plasticity through chromatin organization within lineage-specific genes.

PubMed

Soto-Feliciano, Yadira M; Bartlebaugh, Jordan M E; Liu, Yunpeng; Sánchez-Rivera, Francisco J; Bhutkar, Arjun; Weintraub, Abraham S; Buenrostro, Jason D; Cheng, Christine S; Regev, Aviv; Jacks, Tyler E; Young, Richard A; Hemann, Michael T

2017-05-15

Developmental and lineage plasticity have been observed in numerous malignancies and have been correlated with tumor progression and drug resistance. However, little is known about the molecular mechanisms that enable such plasticity to occur. Here, we describe the function of the plant homeodomain finger protein 6 (PHF6) in leukemia and define its role in regulating chromatin accessibility to lineage-specific transcription factors. We show that loss of Phf6 in B-cell leukemia results in systematic changes in gene expression via alteration of the chromatin landscape at the transcriptional start sites of B-cell- and T-cell-specific factors. Additionally, Phf6 KO cells show significant down-regulation of genes involved in the development and function of normal B cells, show up-regulation of genes involved in T-cell signaling, and give rise to mixed-lineage lymphoma in vivo. Engagement of divergent transcriptional programs results in phenotypic plasticity that leads to altered disease presentation in vivo, tolerance of aberrant oncogenic signaling, and differential sensitivity to frontline and targeted therapies. These findings suggest that active maintenance of a precise chromatin landscape is essential for sustaining proper leukemia cell identity and that loss of a single factor (PHF6) can cause focal changes in chromatin accessibility and nucleosome positioning that render cells susceptible to lineage transition. © 2017 Soto-Feliciano et al.; Published by Cold Spring Harbor Laboratory Press.

Interaction of RNA-binding protein HuR and miR-466i regulates GM-CSF expression.

PubMed

Chen, Jing; Adamiak, William; Huang, Ganlei; Atasoy, Ulus; Rostami, Abdolmohamad; Yu, Shiguang

2017-12-08

Granulocyte-macrophage colony-stimulating factor (GM-CSF) produced by T helper 17 (Th17) cells plays an essential role in autoimmune diseases. Transcriptional regulation of Th17 cell differentiation has been extensively studied, but post-transcriptional regulation of Th17 cell differentiation has remained less well characterized. The RNA-binding protein HuR functions to promote the stability of target mRNAs via binding the AU-rich elements of the 3' untranslated region (3'UTR) of numerous pro-inflammatory cytokines including IL-4, IL-13, IL-17 and TNF-α. However, whether HuR regulates GM-CSF expression in Th17 cells has not been fully investigated. Here we showed that HuR conditional knockout (KO) Th17 cells have decreased GM-CSF mRNA in comparison with wild-type (WT) Th17 cells, and that HuR binds directly to GM-CSF mRNA 3'UTR. Interestingly, HuR deficiency increased the levels of certain microRNA expression in Th17 cells; for example, miR-466i functioned to mediate GM-CSF and IL-17 mRNA decay, which was confirmed by in vitro luciferase assay. Furthermore, we found that HuR promoted Mxi1 expression to inhibit certain miRNA expression. Taken together, these findings indicate that interaction of HuR and miR-466i orchestrates GM-CSF expression in Th17 cells.

ADAM10 is essential for Notch2-dependent marginal zone B cell development and CD23 cleavage in vivo

PubMed Central

Gibb, David R.; El Shikh, Mohey; Kang, Dae-Joong; Rowe, Warren J.; El Sayed, Rania; Cichy, Joanna; Yagita, Hideo; Tew, John G.; Dempsey, Peter J.; Crawford, Howard C.

2010-01-01

The proteolytic activity of a disintegrin and metalloproteinase 10 (ADAM10) regulates cell-fate decisions in Drosophila and mouse embryos. However, in utero lethality of ADAM10−/− mice has prevented examination of ADAM10 cleavage events in lymphocytes. To investigate their role in B cell development, we generated B cell–specific ADAM10 knockout mice. Intriguingly, deletion of ADAM10 prevented development of the entire marginal zone B cell (MZB) lineage. Additionally, cleavage of the low affinity IgE receptor, CD23, was profoundly impaired, but subsequent experiments demonstrated that ADAM10 regulates CD23 cleavage and MZB development by independent mechanisms. Development of MZBs is dependent on Notch2 signaling, which requires proteolysis of the Notch2 receptor by a previously unidentified proteinase. Further experiments revealed that Notch2 signaling is severely impaired in ADAM10-null B cells. Thus, ADAM10 critically regulates MZB development by initiating Notch2 signaling. This study identifies ADAM10 as the in vivo CD23 sheddase and an important regulator of B cell development. Moreover, it has important implications for the treatment of numerous CD23- and Notch-mediated pathologies, ranging from allergy to cancer. PMID:20156974

The let-7 microRNA interfaces extensively with the translation machinery to regulate cell differentiation

PubMed Central

Ding, Xavier C.; Slack, Frank J.; Großhans, Helge

2010-01-01

MicroRNAs (miRNAs) are noncoding RNAs that regulate numerous target genes through a posttranscriptional mechanism and thus control major developmental pathways. The phylogenetically conserved let-7 miRNA regulates cell proliferation and differentiation, thus functioning as a key regulator of developmental timing in C. elegans and a tumor suppressor gene in humans. Using a reverse genetic screen, we have identified genetic interaction partners of C. elegans let-7, including known and novel potential target genes. Initial identification of several translation initiation factors as suppressors of a let-7 mutation led us to systematically examine genetic interaction between let-7 and the translational machinery, which we found to be widespread. In the presence of wild-type let-7, depletion of the translation initiation factor eIF3 resulted in precocious cell differentiation, suggesting that developmental timing is translationally regulated, possibly by let-7. As overexpression of eIF3 in humans promotes translation of mRNAs that are also targets of let-7-mediated repression, we suggest that eIF3 may directly or indirectly oppose let-7 activity. This might provide an explanation for the opposite functions of let-7 and eIF3 in regulating tumorigenesis. PMID:18818519

Cancer Is to Embryology as Mutation Is to Genetics: Hypothesis of the Cancer as Embryological Phenomenon

PubMed Central

Abdelhay, Eliana

2017-01-01

Despite numerous advances in cell biology, genetics, and developmental biology, cancer origin has been attributed to genetic mechanisms primarily involving mutations. Embryologists have expressed timidly cancer embryological origin with little success in leveraging the discussion that cancer could involve a set of conventional cellular processes used to build the embryo during morphogenesis. Thus, this “cancer process” allows the harmonious and coherent construction of the embryo structural base, and its implementation as the embryonic process involves joint regulation of differentiation, proliferation, cell invasion, and migration, enabling the human being recreation of every generation. On the other hand, “cancer disease” is the representation of an abnormal state of the cell that might happen in the stem cells of an adult person, in which the mechanism for joint gene regulating of differentiation, proliferation, cell invasion, and migration could be reactivated in an entirely inappropriate context. PMID:28553657

miRNomes of haematopoietic stem cells and dendritic cells identify miR-30b as a regulator of Notch1

PubMed Central

Su, Xiaoping; Qian, Cheng; Zhang, Qian; Hou, Jin; Gu, Yan; Han, Yanmei; Chen, Yongjian; Jiang, Minghong; Cao, Xuetao

2013-01-01

Dendritic cells (DCs) are critical to initiate the immune response and maintain tolerance, depending on different status and subsets. The expression profiles of microRNAs (miRNAs) in various DC subsets and haematopoietic stem cells (HSCs), which generate DCs, remain to be fully identified. Here we examine miRNomes of mouse bone marrow HSCs, immature DCs, mature DCs and IL-10/NO-producing regulatory DCs by deep sequencing. We identify numerous stage-specific miRNAs and histone modification in HSCs and DCs at different differentiation stages. miR-30b, significantly upregulated via a TGF-beta/Smad3-mediated epigenetic pathway in regulatory DCs, can target Notch1 to promote IL-10 and NO production, suggesting that miR-30b is a negative regulator of immune response. We also identify miRNomes of in vivo counterparts of mature DCs and regulatory DCs and systematically compare them with DCs cultured in vitro. These results provide a resource for studying roles of miRNAs in stem cell biology, development and functional regulation of DC subsets. PMID:24309499

Cell envelope stress in mycobacteria is regulated by the novel signal transduction ATPase IniR in response to trehalose

PubMed Central

van Winden, Vincent J. C.; Sparrius, Marion; van de Weerd, Robert; Speer, Alexander; Ummels, Roy; Sherman, David R.

2017-01-01

The cell envelope of mycobacteria is a highly unique and complex structure that is functionally equivalent to that of Gram-negative bacteria to protect the bacterial cell. Defects in the integrity or assembly of this cell envelope must be sensed to allow the induction of stress response systems. The promoter that is specifically and most strongly induced upon exposure to ethambutol and isoniazid, first line drugs that affect cell envelope biogenesis, is the iniBAC promoter. In this study, we set out to identify the regulator of the iniBAC operon in Mycobacterium marinum using an unbiased transposon mutagenesis screen in a constitutively iniBAC-expressing mutant background. We obtained multiple mutants in the mce1 locus as well as mutants in an uncharacterized putative transcriptional regulator (MMAR_0612). This latter gene was shown to function as the iniBAC regulator, as overexpression resulted in constitutive iniBAC induction, whereas a knockout mutant was unable to respond to the presence of ethambutol and isoniazid. Experiments with the M. tuberculosis homologue (Rv0339c) showed identical results. RNAseq experiments showed that this regulatory gene was exclusively involved in the regulation of the iniBAC operon. We therefore propose to name this dedicated regulator iniBAC Regulator (IniR). IniR belongs to the family of signal transduction ATPases with numerous domains, including a putative sugar-binding domain. Upon testing different sugars, we identified trehalose as an activator and metabolic cue for iniBAC activation, which could also explain the effect of the mce1 mutations. In conclusion, cell envelope stress in mycobacteria is regulated by IniR in a cascade that includes trehalose. PMID:29281637

TMBIM-mediated Ca 2+ homeostasis and cell death

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Qun

Ca 2+ is a ubiquitous intracellular messenger that regulates numerous physiological activities in humans, animals, plants, and bacteria. Cytosolic Ca 2+ is kept at a low level, but subcellular organelles such as the endoplasmic reticulum (ER) and Golgi Apparatus maintain high-concentration Ca 2+ stores. Under resting conditions, store Ca 2+ homeostasis is dynamically regulated to equilibrate between active Ca 2+ uptake and passive Ca 2+ leak processes. The evolutionarily conserved Transmembrane BAX Inhibitor-1 Motif-containing (TMBIM) proteins mediate Ca 2+ homeostasis and cell death. This review focuses on recent advances in functional and structural analysis of TMBIM proteins in regulation ofmore » the two related functions. The roles of TMBIM proteins in pathogen infection and cancer are also discussed with prospects for treatment.« less

TMBIM-mediated Ca 2+ homeostasis and cell death

DOE PAGES

Liu, Qun

2017-01-05

Ca 2+ is a ubiquitous intracellular messenger that regulates numerous physiological activities in humans, animals, plants, and bacteria. Cytosolic Ca 2+ is kept at a low level, but subcellular organelles such as the endoplasmic reticulum (ER) and Golgi Apparatus maintain high-concentration Ca 2+ stores. Under resting conditions, store Ca 2+ homeostasis is dynamically regulated to equilibrate between active Ca 2+ uptake and passive Ca 2+ leak processes. The evolutionarily conserved Transmembrane BAX Inhibitor-1 Motif-containing (TMBIM) proteins mediate Ca 2+ homeostasis and cell death. This review focuses on recent advances in functional and structural analysis of TMBIM proteins in regulation ofmore » the two related functions. The roles of TMBIM proteins in pathogen infection and cancer are also discussed with prospects for treatment.« less

Pathological and therapeutic roles of innate lymphoid cells in diverse diseases.

PubMed

Kim, Jisu; Kim, Geon; Min, Hyeyoung

2017-11-01

Innate lymphoid cells (ILCs) are a recently defined type of innate-immunity cells that belong to the lymphoid lineage and have lymphoid morphology but do not express an antigen-specific B cell or T-cell receptor. ILCs regulate immune functions prior to the formation of adaptive immunity and exert effector functions through a cytokine release. ILCs have been classified into three groups according to the transcription factors that regulate their development and function and the effector cytokines they produce. Of note, ILCs resemble T helper (Th) cells, such as Th1, Th2, and Th17 cells, and show a similar dependence on transcription factors and distinct cytokine production. Despite their short history in immunology, ILCs have received much attention, and numerous studies have revealed biological functions of ILCs including host defense against pathogens, inflammation, tissue repair, and metabolic homeostasis. Here, we describe recent findings about the roles of ILCs in the pathogenesis of various diseases and potential therapeutic targets.

Specifying pancreatic endocrine cell fates.

PubMed

Collombat, Patrick; Hecksher-Sørensen, Jacob; Serup, Palle; Mansouri, Ahmed

2006-07-01

Cell replacement therapy could represent an attractive alternative to insulin injections for the treatment of diabetes. However, this approach requires a thorough understanding of the molecular switches controlling the specification of the different pancreatic cell-types in vivo. These are derived from an apparently identical pool of cells originating from the early gut endoderm, which are successively specified towards the pancreatic, endocrine, and hormone-expressing cell lineages. Numerous studies have outlined the crucial roles exerted by transcription factors in promoting the cell destiny, defining the cell identity and maintaining a particular cell fate. This review focuses on the mechanisms regulating the morphogenesis of the pancreas with particular emphasis on recent findings concerning the transcription factor hierarchy orchestrating endocrine cell fate allocation.

Molecular Programs Underlying Asymmetric Stem Cell Division and Their Disruption in Malignancy.

PubMed

Mukherjee, Subhas; Brat, Daniel J

2017-01-01

Asymmetric division of stem cells is a highly conserved and tightly regulated process by which a single stem cell produces two unequal daughter cells. One retains its stem cell identity while the other becomes specialized through a differentiation program and loses stem cell properties. Coordinating these events requires control over numerous intra- and extracellular biological processes and signaling networks. In the initial stages, critical events include the compartmentalization of fate determining proteins within the mother cell and their subsequent passage to the appropriate daughter cell in order to direct their destiny. Disturbance of these events results in an altered dynamic of self-renewing and differentiation within the cell population, which is highly relevant to the growth and progression of cancer. Other critical events include proper asymmetric spindle assembly, extrinsic regulation through micro-environmental cues, and non-canonical signaling networks that impact cell division and fate determination. In this review, we discuss mechanisms that maintain the delicate balance of asymmetric cell division in normal tissues and describe the current understanding how some of these mechanisms are deregulated in cancer.

Pim1 kinase regulates c-Kit gene translation.

PubMed

An, Ningfei; Cen, Bo; Cai, Houjian; Song, Jin H; Kraft, Andrew; Kang, Yubin

2016-01-01

Receptor tyrosine kinase, c-Kit (CD117) plays a pivotal role in the maintenance and expansion of hematopoietic stem/progenitor cells (HSPCs). Additionally, over-expression and/or mutational activation of c-Kit have been implicated in numerous malignant diseases including acute myeloid leukemia. However, the translational regulation of c-Kit expression remains largely unknown. We demonstrated that loss of Pim1 led to specific down-regulation of c-Kit expression in HSPCs of Pim1 -/- mice and Pim1 -/- 2 -/- 3 -/- triple knockout (TKO) mice, and resulted in attenuated ERK and STAT3 signaling in response to stimulation with stem cell factor. Transduction of c-Kit restored the defects in colony forming capacity seen in HSPCs from Pim1 -/- and TKO mice. Pharmacologic inhibition and genetic modification studies using human megakaryoblastic leukemia cells confirmed the regulation of c-Kit expression by Pim1 kinase: i.e., Pim1-specific shRNA knockdown down-regulated the expression of c-Kit whereas overexpression of Pim1 up-regulated the expression of c-Kit. Mechanistically, inhibition or knockout of Pim1 kinase did not affect the transcription of c-Kit gene. Pim1 kinase enhanced c-Kit 35 S methionine labeling and increased the incorporation of c-Kit mRNAs into the polysomes and monosomes, demonstrating that Pim1 kinase regulates c-Kit expression at the translational level. Our study provides the first evidence that Pim1 regulates c-Kit gene translation and has important implications in hematopoietic stem cell transplantation and cancer treatment.

Intracellular signaling by phospholipase D as a therapeutic target.

PubMed

Steed, P M; Chow, A H

2001-09-01

The pharmaceutical industry has recently focused on intracellular signaling as a means to integrate the multiple facets of complex disease states, such as inflammation, because these pathways respond to numerous extracellular signals and coordinate a collection of cell responses contributing to pathology. One critical aspect of intracellular signaling is regulation of key cell functions by lipid mediators, in particular the generation of a key mediator, phosphatidic acid (PA) via the hydrolysis of phosphatidylcholine by phospholipase D (PLD). Research in this field has intensified, due in part to the recent cloning and partial characterization of the two PLD isoforms in mammalian cells, and this work has contributed significantly to our understanding of events downstream of PA generation. It is these effector functions of PLD activity that make this pathway attractive as a therapeutic target while the biochemical properties of the PLD isozymes make them amenable to small molecule intervention. Recent studies indicate that PA, and its immediate metabolites diacylglycerol and lyso-PA, affect numerous cellular pathways including ligand-mediated secretion, cytoskeletal reorganisations, respiratory burst, prostaglandin release, cell migration, cytokine release, and mitogenesis. This review summarises the data implicating signaling via PLD in these cell functions, obtained from: (i) molecular analyses of PLD/effector interactions, (ii) correlation between PA production and cell responses, (iii) experimental manipulation of PA levels, (iv) inhibition of PLD regulators, and (v) direct inhibition of PA production. The utility of targeting PLD signaling for the treatment of acute/chronic inflammation and other indications is discussed in light of these data.

Low-molecular-weight fucoidan regulates myogenic differentiation through the mitogen-activated protein kinase pathway in C2C12 cells.

PubMed

Kim, Kui-Jin; Lee, Ok-Hwan; Lee, Boo-Yong

2011-12-01

Low-molecular-weight fucoidan (LMWF) has been broadly studied in recent years due to its numerous biological properties. Nevertheless, there have been no reports about the effects of LMWF on myogenic differentiation (MyoD). The objective of the present study was to demonstrate the impact of LMWF on myogenesis in C2C12 cells. The ultimate aim was to establish whether LMWF regulates myogenesis similar to heparin as a partial regulator of myogenesis. LMWF was prepared at a minimal size by ultra-filtration compared with crude fucoidan. We treated C2C12 cells with LMWF and/or heparin during MyoD. The data from the present study are the first to suggest that LMWF suppresses the expression of the myogenic regulatory factors and the myocyte enhancer factors as well as the morphological changes that occur during differentiation. Additionally, the expression of the mitogen-activated protein kinase (MAPK) family was significantly inhibited by LMWF when compared with controls. The LMWF-treated group showed significantly decreased expression of reactive oxygen species (ROS) enzymes compared with control cells. Heparin was used as a positive control because it has been reported to activate MyoD. Taken together, these results suggest that LMWF might regulate MyoD through the MAPK pathway and by regulating ROS activity in C2C12 cells.

ROLES OF CELL-INTRINSIC AND MICROENVIRONMENTAL FACTORS IN PHOTORECEPTOR CELL DIFFERENTIATION

PubMed Central

Bradford, Rebecca L.; Wang, Chenwei; Zack, Donald J.; Adler, Ruben

2005-01-01

Photoreceptor differentiation requires the coordinated expression of numerous genes. It is unknown whether those genes share common regulatory mechanisms or are independently regulated by distinct mechanisms. To distinguish between these scenarios, we have used in situ hybridization, RT-PCR and real time PCR to analyze the expression of visual pigments and other photoreceptor-specific genes during chick embryo retinal development in ovo, as well as in retinal cell cultures treated with molecules that regulate the expression of particular visual pigments. In ovo, onset of gene expression was asynchronous, becoming detectable at the time of photoreceptor generation (ED 5–8) for some photoreceptor genes, but only around the time of outer segment formation (ED 14–16) for others. Treatment of retinal cell cultures with activin, staurosporine or CNTF selectively induced or down-regulated specific visual pigment genes, but many cognate rod- or cone-specific genes were not affected by the treatments. These results indicate that many photoreceptor genes are independently regulated during development, are consistent with the existence of at least two distinct stages of gene expression during photoreceptor differentiation, suggest that intrinsic, coordinated regulation of a cascade of gene expression triggered by a commitment to the photoreceptor fate is not a general mechanism of photoreceptor differentiation, and imply that using a single photoreceptor-specific “marker” as a proxy to identify photoreceptor cell fate is problematic. PMID:16120439

Glucagon-like peptide 2 and its beneficial effects on gut function and health in production animals

USDA-ARS?s Scientific Manuscript database

Numerous endocrine cell subtypes exist within the intestinal mucosa and produce peptides contributing to the regulation of critical physiological processes including appetite, energy metabolism, gut function, and gut health. The mechanisms of action and the extent of the physiological effects of the...

Glial Cells in the Genesis and Regulation of Circadian Rhythms

PubMed Central

Chi-Castañeda, Donají; Ortega, Arturo

2018-01-01

Circadian rhythms are biological oscillations with a period of ~24 h. These rhythms are orchestrated by a circadian timekeeper in the suprachiasmatic nucleus of the hypothalamus, the circadian “master clock,” which exactly adjusts clock outputs to solar time via photic synchronization. At the molecular level, circadian rhythms are generated by the interaction of positive and negative feedback loops of transcriptional and translational processes of the so-called “clock genes.” A large number of clock genes encode numerous proteins that regulate their own transcription and that of other genes, collectively known as “clock-controlled genes.” In addition to the sleep/wake cycle, many cellular processes are regulated by circadian rhythms, including synaptic plasticity in which an exquisite interplay between neurons and glial cells takes place. In particular, there is compelling evidence suggesting that glial cells participate in and regulate synaptic plasticity in a circadian fashion, possibly representing the missing cellular and physiological link between circadian rhythms with learning and cognition processes. Here we review recent studies in support of this hypothesis, focusing on the interplay between glial cells, synaptic plasticity, and circadian rhythmogenesis. PMID:29483880

The Murine Ortholog of Notchless, a Direct Regulator of the Notch Pathway in Drosophila melanogaster, Is Essential for Survival of Inner Cell Mass Cells

PubMed Central

Cormier, Sarah; Le Bras, Stéphanie; Souilhol, Céline; Vandormael-Pournin, Sandrine; Durand, Béatrice; Babinet, Charles; Baldacci, Patricia; Cohen-Tannoudji, Michel

2006-01-01

Notch signaling is an evolutionarily conserved pathway involved in intercellular communication and is essential for proper cell fate choices. Numerous genes participate in the modulation of the Notch signaling pathway activity. Among them, Notchless (Nle) is a direct regulator of the Notch activity identified in Drosophila melanogaster. Here, we characterized the murine ortholog of Nle and demonstrated that it has conserved the ability to modulate Notch signaling. We also generated mice deficient for mouse Nle (mNle) and showed that its disruption resulted in embryonic lethality shortly after implantation. In late mNle−/− blastocysts, inner cell mass (ICM) cells died through a caspase 3-dependent apoptotic process. Most deficient embryos exhibited a delay in the temporal down-regulation of Oct4 expression in the trophectoderm (TE). However, mNle-deficient TE was able to induce decidual swelling in vivo and properly differentiated in vitro. Hence, our results indicate that mNle is mainly required in ICM cells, being instrumental for their survival, and raise the possibility that the death of mNle-deficient embryos might result from abnormal Notch signaling during the first steps of development. PMID:16611995

The murine ortholog of notchless, a direct regulator of the notch pathway in Drosophila melanogaster, is essential for survival of inner cell mass cells.

PubMed

Cormier, Sarah; Le Bras, Stéphanie; Souilhol, Céline; Vandormael-Pournin, Sandrine; Durand, Béatrice; Babinet, Charles; Baldacci, Patricia; Cohen-Tannoudji, Michel

2006-05-01

Notch signaling is an evolutionarily conserved pathway involved in intercellular communication and is essential for proper cell fate choices. Numerous genes participate in the modulation of the Notch signaling pathway activity. Among them, Notchless (Nle) is a direct regulator of the Notch activity identified in Drosophila melanogaster. Here, we characterized the murine ortholog of Nle and demonstrated that it has conserved the ability to modulate Notch signaling. We also generated mice deficient for mouse Nle (mNle) and showed that its disruption resulted in embryonic lethality shortly after implantation. In late mNle(-/-) blastocysts, inner cell mass (ICM) cells died through a caspase 3-dependent apoptotic process. Most deficient embryos exhibited a delay in the temporal down-regulation of Oct4 expression in the trophectoderm (TE). However, mNle-deficient TE was able to induce decidual swelling in vivo and properly differentiated in vitro. Hence, our results indicate that mNle is mainly required in ICM cells, being instrumental for their survival, and raise the possibility that the death of mNle-deficient embryos might result from abnormal Notch signaling during the first steps of development.

Regulatory RNA Key Player in p53-Mediated Apoptosis in Embryonic Stem Cells | Center for Cancer Research

Cancer.gov

Embryonic stem cells (ESCs) must maintain the integrity of their genomes or risk passing potentially deleterious mutations on to numerous tissues. Thus, ESCs have a unique genome surveillance system and easily undergo apoptosis or differentiation when DNA damage is detected. The protein p53 is known to promote differentiation in mouse ESCs (mESCs), but its role in DNA damage-induced apoptosis (DIA) is unclear. p53 may have a pro-apoptotic function since it can regulate apoptotic genes in embryonal cells. Given that ESCs have a distinct transcriptional program, Jing Huang, Ph.D., of CCR’s Laboratory of Cancer Biology and Genetics, and his colleagues wondered whether p53 might regulate DIA in ESCs by utilizing the ESC-specific expression program.

Novel interactions between erythroblast macrophage protein and cell migration.

PubMed

Javan, Gulnaz T; Can, Ismail; Yeboah, Fred; Lee, Youngil; Soni, Shivani

2016-09-01

Erythroblast macrophage protein is a novel protein known to mediate attachment of erythroid cells to macrophages to form erythroblastic islands in bone marrow during erythropoiesis. Emp-null macrophages are small with round morphologies, and lack cytoplasmic projections which imply immature structure. The role of Emp in macrophage development and function is not fully elucidated. Macrophages perform varied functions (e.g. homeostasis, erythropoiesis), and are implicated in numerous pathophysiological conditions such as cellular malignancy. The objective of the current study is to investigate the interaction of Emp with cytoskeletal- and cell migration-associated proteins involved in macrophage functions. A short hairpin RNA lentiviral system was use to down-regulate the expression of Emp in macrophage cells. A cell migration assay revealed that the relocation of macrophages was significantly inhibited when Emp expression was decreased. To further analyze changes in gene expression related to cell motility, PCR array was performed by down-regulating Emp expression. The results indicated that expression of mitogen-activated protein kinase 1 and thymoma viral proto-oncogene 1 were significantly higher when Emp was down-regulated. The results implicate Emp in abnormal cell motility, thus, warrants to assess its role in cancer where tumor cell motility is required for invasion and metastasis. Copyright © 2016 Elsevier Inc. All rights reserved.

Regulation of T Cell Differentiation and Function by EZH2

PubMed Central

Karantanos, Theodoros; Christofides, Anthos; Bardhan, Kankana; Li, Lequn; Boussiotis, Vassiliki A.

2016-01-01

The enhancer of zeste homolog 2 (EZH2), one of the polycomb-group proteins, is the catalytic subunit of Polycomb-repressive complex 2 (PRC2) and induces the trimethylation of the histone H3 lysine 27 (H3K27me3) promoting epigenetic gene silencing. EZH2 contains a SET domain promoting the methyltransferase activity, while the three other protein components of PRC2, namely EED, SUZ12, and RpAp46/48, induce compaction of the chromatin permitting EZH2 enzymatic activity. Numerous studies highlight the role of this evolutionary conserved protein as a master regulator of differentiation in humans involved in the repression of the homeotic gene and the inactivation of X-chromosome. Through its effects in the epigenetic regulation of critical genes, EZH2 has been strongly linked to cell cycle progression, stem cell pluripotency, and cancer biology, being currently at the cutting edge of research. Most recently, EZH2 has been associated with hematopoietic stem cell proliferation and differentiation, thymopoiesis and lymphopoiesis. Several studies have evaluated the role of EZH2 in the regulation of T cell differentiation and plasticity as well as its implications in the development of autoimmune diseases and graft-versus-host disease (GVHD). The aim of this review is to summarize the current knowledge regarding the role of EZH2 in the regulation of the differentiation and function of T cells focusing on possible applications in various immune-mediated conditions, including autoimmune disorders and GVHD. PMID:27199994

Impact of implementation choices on quantitative predictions of cell-based computational models

NASA Astrophysics Data System (ADS)

Kursawe, Jochen; Baker, Ruth E.; Fletcher, Alexander G.

2017-09-01

'Cell-based' models provide a powerful computational tool for studying the mechanisms underlying the growth and dynamics of biological tissues in health and disease. An increasing amount of quantitative data with cellular resolution has paved the way for the quantitative parameterisation and validation of such models. However, the numerical implementation of cell-based models remains challenging, and little work has been done to understand to what extent implementation choices may influence model predictions. Here, we consider the numerical implementation of a popular class of cell-based models called vertex models, which are often used to study epithelial tissues. In two-dimensional vertex models, a tissue is approximated as a tessellation of polygons and the vertices of these polygons move due to mechanical forces originating from the cells. Such models have been used extensively to study the mechanical regulation of tissue topology in the literature. Here, we analyse how the model predictions may be affected by numerical parameters, such as the size of the time step, and non-physical model parameters, such as length thresholds for cell rearrangement. We find that vertex positions and summary statistics are sensitive to several of these implementation parameters. For example, the predicted tissue size decreases with decreasing cell cycle durations, and cell rearrangement may be suppressed by large time steps. These findings are counter-intuitive and illustrate that model predictions need to be thoroughly analysed and implementation details carefully considered when applying cell-based computational models in a quantitative setting.

Trp-Asp (WD) Repeat Domain 1 Is Essential for Mouse Peri-implantation Development and Regulates Cofilin Phosphorylation.

PubMed

Xiao, Yi; Ma, Haixia; Wan, Ping; Qin, Dandan; Wang, Xiaoxiao; Zhang, Xiaoxin; Xiang, Yunlong; Liu, Wenbo; Chen, Jiong; Yi, Zhaohong; Li, Lei

2017-01-27

Trp-Asp (WD) repeat domain 1 (WDR1) is a highly conserved actin-binding protein across all eukaryotes and is involved in numerous actin-based processes by accelerating Cofilin severing actin filament. However, the function and the mechanism of WDR1 in mammalian early development are still largely unclear. We now report that WDR1 is essential for mouse peri-implantation development and regulates Cofilin phosphorylation in mouse cells. The disruption of maternal WDR1 does not obviously affect ovulation and female fertility. However, depletion of zygotic WDR1 results in embryonic lethality at the peri-implantation stage. In WDR1 knock-out cells, we found that WDR1 regulates Cofilin phosphorylation. Interestingly, WDR1 is overdosed to regulate Cofilin phosphorylation in mouse cells. Furthermore, we showed that WDR1 interacts with Lim domain kinase 1 (LIMK1), a well known phosphorylation kinase of Cofilin. Altogether, our results provide new insights into the role and mechanism of WDR1 in physiological conditions. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Epigenetic regulation of miRNA-Cancer Stem Cells nexus by Nutraceuticals

PubMed Central

Ahmad, Aamir; Li, Yiwei; Bao, Bin; Kong, Dejuan; Sarkar, Fazlul H.

2014-01-01

Nutraceuticals, the bioactive food components represented by many naturally occurring dietary compounds, have been investigated for a few decades for their numerous beneficial effects, including their anticancer properties. The initial interest in the cancer-preventing/therapeutic ability of these agents was based on their ability to affect multiple signaling pathways that are deregulated in cancer cells. With a shift in the focus of cancer research to the emerging areas such as epigenetic regulation, microRNAs (miRNAs) and the cancer stem cells (CSCs), nutraceuticals initially appeared out of place. However, research investigations over the last several years have slowly but firmly presented evidence that supports a relevance of these agents in modern day research. While nutraceuticals are increasingly being realized to alter miRNA/CSCs expression and function, the molecular mechanism(s) are not very clearly understood. Epigenetic regulation is one mechanism by which these agents exert their anticancer effects. In this focused mini review, we summarize our current understanding of epigenetic regulation of miRNAs and CSCs by nutraceuticals. We discuss both direct and indirect evidences that support such an activity of these compounds. PMID:24272883

The plasma membrane as a capacitor for energy and metabolism

PubMed Central

Ray, Supriyo; Kassan, Adam; Busija, Anna R.; Rangamani, Padmini

2016-01-01

When considering which components of the cell are the most critical to function and physiology, we naturally focus on the nucleus, the mitochondria that regulate energy and apoptotic signaling, or other organelles such as the endoplasmic reticulum, Golgi, ribosomes, etc. Few people will suggest that the membrane is the most critical element of a cell in terms of function and physiology. Those that consider the membrane critical will point to its obvious barrier function regulated by the lipid bilayer and numerous ion channels that regulate homeostatic gradients. What becomes evident upon closer inspection is that not all membranes are created equal and that there are lipid-rich microdomains that serve as platforms of signaling and a means of communication with the intracellular environment. In this review, we explore the evolution of membranes, focus on lipid-rich microdomains, and advance the novel concept that membranes serve as “capacitors for energy and metabolism.” Within this framework, the membrane then is the primary and critical regulator of stress and disease adaptation of the cell. PMID:26771520

Extracellular small RNAs: what, where, why?

PubMed Central

Hoy, Anna M.; Buck, Amy H.

2012-01-01

miRNAs (microRNAs) are a class of small RNA that regulate gene expression by binding to mRNAs and modulating the precise amount of proteins that get expressed in a cell at a given time. This form of gene regulation plays an important role in developmental systems and is critical for the proper function of numerous biological pathways. Although miRNAs exert their functions inside the cell, these and other classes of RNA are found in body fluids in a cell-free form that is resistant to degradation by RNases. A broad range of cell types have also been shown to secrete miRNAs in association with components of the RISC (RNA-induced silencing complex) and/or encapsulation within vesicles, which can be taken up by other cells. In the present paper, we provide an overview of the properties of extracellular miRNAs in relation to their capacity as biomarkers, stability against degradation and mediators of cell–cell communication. PMID:22817753

Protein tyrosine phosphatase-PEST (PTP-PEST) regulates mast cell-activating signals in PTP activity-dependent and -independent manners.

PubMed

Motohashi, Satoru; Koizumi, Karen; Honda, Reika; Maruyama, Atsuko; Palmer, Helen E F; Mashima, Keisuke

2014-01-01

Aggregation of the high-affinity IgE receptor (FcεRI) in mast cells leads to degranulation and production of numerous cytokines and lipid mediators that promote allergic inflammation. Tyrosine phosphorylation of proteins in response to FcεRI aggregation has been implicated in mast cell activation. Here, we determined the role of PTP-PEST (encoded by PTPN12) in the regulation of mast cell activation using the RBL-2H3 rat basophilic leukemia cell line as a model. PTP-PEST expression was significantly induced upon FcεRI-crosslinking, and aggregation of FcεRI induced the phosphorylation of PTP-PEST at Ser39, thus resulting in the suppression of PTP activity. By overexpressing a phosphatase-dead mutant (PTP-PEST CS) and a constitutively active mutant (PTP-PEST SA) in RBL-2H3 cells, we showed that PTP-PEST decreased degranulation and enhanced IL-4 and IL-13 transcription in FcεRI-crosslinked RBL-2H3 cells, but PTP activity of PTP-PEST was not necessary for this regulation. However, FcεRI-induced TNF-α transcription was increased by the overexpression of PTP-PEST SA and suppressed by the overexpression of PTP-PEST CS. Taken together, these results suggest that PTP-PEST is involved in the regulation of FcεRI-mediated mast cell activation through at least two different processes represented by PTP activity-dependent and -independent pathways. Copyright © 2014 Elsevier Inc. All rights reserved.

Methods to study maternal regulation of germ cell specification in zebrafish

PubMed Central

Kaufman, O.H.; Marlow, F.L.

2016-01-01

The process by which the germ line is specified in the zebrafish embryo is under the control of maternal gene products that were produced during oogenesis. Zebrafish are highly amenable to microscopic observation of the processes governing maternal germ cell specification because early embryos are transparent, and the germ line is specified rapidly (within 4–5 h post fertilization). Advantages of zebrafish over other models used to study vertebrate germ cell formation include their genetic tractability, the large numbers of progeny, and the easily manipulable genome, all of which make zebrafish an ideal system for studying the genetic regulators and cellular basis of germ cell formation and maintenance. Classical molecular biology techniques, including expression analysis through in situ hybridization and forward genetic screens, have laid the foundation for our understanding of germ cell development in zebrafish. In this chapter, we discuss some of these classic techniques, as well as recent cutting-edge methodologies that have improved our ability to visualize the process of germ cell specification and differentiation, and the tracking of specific molecules involved in these processes. Additionally, we discuss traditional and novel technologies for manipulating the zebrafish genome to identify new components through loss-of-function studies of putative germ cell regulators. Together with the numerous aforementioned advantages of zebrafish as a genetic model for studying development, we believe these new techniques will continue to advance zebrafish to the forefront for investigation of the molecular regulators of germ cell specification and germ line biology. PMID:27312489

Tracking single mRNA molecules in live cells

NASA Astrophysics Data System (ADS)

Moon, Hyungseok C.; Lee, Byung Hun; Lim, Kiseong; Son, Jae Seok; Song, Minho S.; Park, Hye Yoon

2016-06-01

mRNAs inside cells interact with numerous RNA-binding proteins, microRNAs, and ribosomes that together compose a highly heterogeneous population of messenger ribonucleoprotein (mRNP) particles. Perhaps one of the best ways to investigate the complex regulation of mRNA is to observe individual molecules. Single molecule imaging allows the collection of quantitative and statistical data on subpopulations and transient states that are otherwise obscured by ensemble averaging. In addition, single particle tracking reveals the sequence of events that occur in the formation and remodeling of mRNPs in real time. Here, we review the current state-of-the-art techniques in tagging, delivery, and imaging to track single mRNAs in live cells. We also discuss how these techniques are applied to extract dynamic information on the transcription, transport, localization, and translation of mRNAs. These studies demonstrate how single molecule tracking is transforming the understanding of mRNA regulation in live cells.

Quantitative Analysis of Human Pluripotency and Neural Specification by In-Depth (Phospho)Proteomic Profiling.

PubMed

Singec, Ilyas; Crain, Andrew M; Hou, Junjie; Tobe, Brian T D; Talantova, Maria; Winquist, Alicia A; Doctor, Kutbuddin S; Choy, Jennifer; Huang, Xiayu; La Monaca, Esther; Horn, David M; Wolf, Dieter A; Lipton, Stuart A; Gutierrez, Gustavo J; Brill, Laurence M; Snyder, Evan Y

2016-09-13

Controlled differentiation of human embryonic stem cells (hESCs) can be utilized for precise analysis of cell type identities during early development. We established a highly efficient neural induction strategy and an improved analytical platform, and determined proteomic and phosphoproteomic profiles of hESCs and their specified multipotent neural stem cell derivatives (hNSCs). This quantitative dataset (nearly 13,000 proteins and 60,000 phosphorylation sites) provides unique molecular insights into pluripotency and neural lineage entry. Systems-level comparative analysis of proteins (e.g., transcription factors, epigenetic regulators, kinase families), phosphorylation sites, and numerous biological pathways allowed the identification of distinct signatures in pluripotent and multipotent cells. Furthermore, as predicted by the dataset, we functionally validated an autocrine/paracrine mechanism by demonstrating that the secreted protein midkine is a regulator of neural specification. This resource is freely available to the scientific community, including a searchable website, PluriProt. Published by Elsevier Inc.

Nuclear Lipids in the Nervous System: What they do in Health and Disease.

PubMed

Garcia-Gil, Mercedes; Albi, Elisabetta

2017-02-01

In the last 20 years it has been widely demonstrated that cell nucleus contains neutral and polar lipids localized in nuclear membranes, nucleoli, nuclear matrix and chromatin. Nuclear lipids may show specific organization forming nuclear lipid microdomains and have both structural and functional roles. Depending on their localization, nuclear lipids play different roles such as the regulation of nuclear membrane and nuclear matrix fluidity but they also can act as platforms for vitamin and hormone function, for active chromatin anchoring, and for the regulation of gene expression, DNA duplication and transcription. Crosstalk among different kinds of lipid signalling pathways influence the physiopathology of numerous cell types. In neural cells the nuclear lipids are involved in cell proliferation, differentiation, inflammation, migration and apoptosis. Abnormal metabolism of nuclear lipids might be closely associated with tumorigenesis and neurodegenerative diseases such as Alzheimer disease and Parkinson disease among others.

Homeostatic regulation of meiotic DSB formation by ATM/ATR

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cooper, Tim J.; Wardell, Kayleigh; Garcia, Valerie

2014-11-15

Ataxia–telangiectasia mutated (ATM) and RAD3-related (ATR) are widely known as being central players in the mitotic DNA damage response (DDR), mounting responses to DNA double-strand breaks (DSBs) and single-stranded DNA (ssDNA) respectively. The DDR signalling cascade couples cell cycle control to damage-sensing and repair processes in order to prevent untimely cell cycle progression while damage still persists [1]. Both ATM/ATR are, however, also emerging as essential factors in the process of meiosis; a specialised cell cycle programme responsible for the formation of haploid gametes via two sequential nuclear divisions. Central to achieving accurate meiotic chromosome segregation is the introduction ofmore » numerous DSBs spread across the genome by the evolutionarily conserved enzyme, Spo11. This review seeks to explore and address how cells utilise ATM/ATR pathways to regulate Spo11-DSB formation, establish DSB homeostasis and ensure meiosis is completed unperturbed.« less

Midkine-A functions upstream of Id2a to regulate cell cycle kinetics in the developing vertebrate retina

PubMed Central

2012-01-01

Background Midkine is a small heparin binding growth factor expressed in numerous tissues during development. The unique midkine gene in mammals has two paralogs in zebrafish: midkine-a (mdka) and midkine-b (mdkb). In the zebrafish retina, during both larval development and adult photoreceptor regeneration, mdka is expressed in retinal stem and progenitor cells and functions as a molecular component of the retina’s stem cell niche. In this study, loss-of-function and conditional overexpression were used to investigate the function of Mdka in the retina of the embryonic zebrafish. Results The results show that during early retinal development Mdka functions to regulate cell cycle kinetics. Following targeted knockdown of Mdka synthesis, retinal progenitors cycle more slowly, and this results in microphthalmia, a diminished rate of cell cycle exit and a temporal delay of cell cycle exit and neuronal differentiation. In contrast, Mdka overexpression results in acceleration of the cell cycle and retinal overgrowth. Mdka gain-of-function, however, does not temporally advance cell cycle exit. Experiments to identify a potential Mdka signaling pathway show that Mdka functions upstream of the HLH regulatory protein, Id2a. Gene expression analysis shows Mdka regulates id2a expression, and co-injection of Mdka morpholinos and id2a mRNA rescues the Mdka loss-of-function phenotype. Conclusions These data show that in zebrafish, Mdka resides in a shared Id2a pathway to regulate cell cycle kinetics in retinal progenitors. This is the first study to demonstrate the function of Midkine during retinal development and adds Midkine to the list of growth factors that transcriptionally regulate Id proteins. PMID:23111152

Midkine-A functions upstream of Id2a to regulate cell cycle kinetics in the developing vertebrate retina.

PubMed

Luo, Jing; Uribe, Rosa A; Hayton, Sarah; Calinescu, Anda-Alexandra; Gross, Jeffrey M; Hitchcock, Peter F

2012-10-30

Midkine is a small heparin binding growth factor expressed in numerous tissues during development. The unique midkine gene in mammals has two paralogs in zebrafish: midkine-a (mdka) and midkine-b (mdkb). In the zebrafish retina, during both larval development and adult photoreceptor regeneration, mdka is expressed in retinal stem and progenitor cells and functions as a molecular component of the retina's stem cell niche. In this study, loss-of-function and conditional overexpression were used to investigate the function of Mdka in the retina of the embryonic zebrafish. The results show that during early retinal development Mdka functions to regulate cell cycle kinetics. Following targeted knockdown of Mdka synthesis, retinal progenitors cycle more slowly, and this results in microphthalmia, a diminished rate of cell cycle exit and a temporal delay of cell cycle exit and neuronal differentiation. In contrast, Mdka overexpression results in acceleration of the cell cycle and retinal overgrowth. Mdka gain-of-function, however, does not temporally advance cell cycle exit. Experiments to identify a potential Mdka signaling pathway show that Mdka functions upstream of the HLH regulatory protein, Id2a. Gene expression analysis shows Mdka regulates id2a expression, and co-injection of Mdka morpholinos and id2a mRNA rescues the Mdka loss-of-function phenotype. These data show that in zebrafish, Mdka resides in a shared Id2a pathway to regulate cell cycle kinetics in retinal progenitors. This is the first study to demonstrate the function of Midkine during retinal development and adds Midkine to the list of growth factors that transcriptionally regulate Id proteins.

The metazoan Mediator co-activator complex as an integrative hub for transcriptional regulation.

PubMed

Malik, Sohail; Roeder, Robert G

2010-11-01

The Mediator is an evolutionarily conserved, multiprotein complex that is a key regulator of protein-coding genes. In metazoan cells, multiple pathways that are responsible for homeostasis, cell growth and differentiation converge on the Mediator through transcriptional activators and repressors that target one or more of the almost 30 subunits of this complex. Besides interacting directly with RNA polymerase II, Mediator has multiple functions and can interact with and coordinate the action of numerous other co-activators and co-repressors, including those acting at the level of chromatin. These interactions ultimately allow the Mediator to deliver outputs that range from maximal activation of genes to modulation of basal transcription to long-term epigenetic silencing.

TRX-1 Regulates SKN-1 Nuclear Localization Cell Non-autonomously in Caenorhabditis elegans

PubMed Central

McCallum, Katie C.; Liu, Bin; Fierro-González, Juan Carlos; Swoboda, Peter; Arur, Swathi; Miranda-Vizuete, Antonio; Garsin, Danielle A.

2016-01-01

The Caenorhabditis elegans oxidative stress response transcription factor, SKN-1, is essential for the maintenance of redox homeostasis and is a functional ortholog of the Nrf family of transcription factors. The numerous levels of regulation that govern these transcription factors underscore their importance. Here, we add a thioredoxin, encoded by trx-1, to the expansive list of SKN-1 regulators. We report that loss of trx-1 promotes nuclear localization of intestinal SKN-1 in a redox-independent, cell non-autonomous fashion from the ASJ neurons. Furthermore, this regulation is not general to the thioredoxin family, as two other C. elegans thioredoxins, TRX-2 and TRX-3, do not play a role in this process. Moreover, TRX-1-dependent regulation requires signaling from the p38 MAPK-signaling pathway. However, while TRX-1 regulates SKN-1 nuclear localization, classical SKN-1 transcriptional activity associated with stress response remains largely unaffected. Interestingly, RNA-Seq analysis revealed that loss of trx-1 elicits a general, organism-wide down-regulation of several classes of genes; those encoding for collagens and lipid transport being most prevalent. Together, these results uncover a novel role for a thioredoxin in regulating intestinal SKN-1 nuclear localization in a cell non-autonomous manner, thereby contributing to the understanding of the processes involved in maintaining redox homeostasis throughout an organism. PMID:26920757

TRX-1 Regulates SKN-1 Nuclear Localization Cell Non-autonomously in Caenorhabditis elegans.

PubMed

McCallum, Katie C; Liu, Bin; Fierro-González, Juan Carlos; Swoboda, Peter; Arur, Swathi; Miranda-Vizuete, Antonio; Garsin, Danielle A

2016-05-01

The Caenorhabditis elegans oxidative stress response transcription factor, SKN-1, is essential for the maintenance of redox homeostasis and is a functional ortholog of the Nrf family of transcription factors. The numerous levels of regulation that govern these transcription factors underscore their importance. Here, we add a thioredoxin, encoded by trx-1, to the expansive list of SKN-1 regulators. We report that loss of trx-1 promotes nuclear localization of intestinal SKN-1 in a redox-independent, cell non-autonomous fashion from the ASJ neurons. Furthermore, this regulation is not general to the thioredoxin family, as two other C. elegans thioredoxins, TRX-2 and TRX-3, do not play a role in this process. Moreover, TRX-1-dependent regulation requires signaling from the p38 MAPK-signaling pathway. However, while TRX-1 regulates SKN-1 nuclear localization, classical SKN-1 transcriptional activity associated with stress response remains largely unaffected. Interestingly, RNA-Seq analysis revealed that loss of trx-1 elicits a general, organism-wide down-regulation of several classes of genes; those encoding for collagens and lipid transport being most prevalent. Together, these results uncover a novel role for a thioredoxin in regulating intestinal SKN-1 nuclear localization in a cell non-autonomous manner, thereby contributing to the understanding of the processes involved in maintaining redox homeostasis throughout an organism. Copyright © 2016 by the Genetics Society of America.

RNA-Binding Protein FXR1 Regulates p21 and TERC RNA to Bypass p53-Mediated Cellular Senescence in OSCC

PubMed Central

Majumder, Mrinmoyee; House, Reniqua; Palanisamy, Nallasivam; Qie, Shuo; Day, Terrence A.; Neskey, David; Diehl, J. Alan

2016-01-01

RNA-binding proteins (RBP) regulate numerous aspects of co- and post-transcriptional gene expression in cancer cells. Here, we demonstrate that RBP, fragile X-related protein 1 (FXR1), plays an essential role in cellular senescence by utilizing mRNA turnover pathway. We report that overexpressed FXR1 in head and neck squamous cell carcinoma targets (G-quadruplex (G4) RNA structure within) both mRNA encoding p21 (Cyclin-Dependent Kinase Inhibitor 1A (CDKN1A, Cip1) and the non-coding RNA Telomerase RNA Component (TERC), and regulates their turnover to avoid senescence. Silencing of FXR1 in cancer cells triggers the activation of Cyclin-Dependent Kinase Inhibitors, p53, increases DNA damage, and ultimately, cellular senescence. Overexpressed FXR1 binds and destabilizes p21 mRNA, subsequently reduces p21 protein expression in oral cancer cells. In addition, FXR1 also binds and stabilizes TERC RNA and suppresses the cellular senescence possibly through telomerase activity. Finally, we report that FXR1-regulated senescence is irreversible and FXR1-depleted cells fail to form colonies to re-enter cellular proliferation. Collectively, FXR1 displays a novel mechanism of controlling the expression of p21 through p53-dependent manner to bypass cellular senescence in oral cancer cells. PMID:27606879

A genome landscape of SRSF3-regulated splicing events and gene expression in human osteosarcoma U2OS cells

PubMed Central

Ajiro, Masahiko; Jia, Rong; Yang, Yanqin; Zhu, Jun; Zheng, Zhi-Ming

2016-01-01

Alternative RNA splicing is an essential process to yield proteomic diversity in eukaryotic cells, and aberrant splicing is often associated with numerous human diseases and cancers. We recently described serine/arginine-rich splicing factor 3 (SRSF3 or SRp20) being a proto-oncogene. However, the SRSF3-regulated splicing events responsible for its oncogenic activities remain largely unknown. By global profiling of the SRSF3-regulated splicing events in human osteosarcoma U2OS cells, we found that SRSF3 regulates the expression of 60 genes including ERRFI1, ANXA1 and TGFB2, and 182 splicing events in 164 genes, including EP300, PUS3, CLINT1, PKP4, KIF23, CHK1, SMC2, CKLF, MAP4, MBNL1, MELK, DDX5, PABPC1, MAP4K4, Sp1 and SRSF1, which are primarily associated with cell proliferation or cell cycle. Two SRSF3-binding motifs, CCAGC(G)C and A(G)CAGCA, are enriched to the alternative exons. An SRSF3-binding site in the EP300 exon 14 is essential for exon 14 inclusion. We found that the expression of SRSF1 and SRSF3 are mutually dependent and coexpressed in normal and tumor tissues/cells. SRSF3 also significantly regulates the expression of at least 20 miRNAs, including a subset of oncogenic or tumor suppressive miRNAs. These data indicate that SRSF3 affects a global change of gene expression to maintain cell homeostasis. PMID:26704980

Induction of virulence factors in Giardia duodenalis independent of host attachment

PubMed Central

Emery, Samantha J.; Mirzaei, Mehdi; Vuong, Daniel; Pascovici, Dana; Chick, Joel M.; Lacey, Ernest; Haynes, Paul A.

2016-01-01

Giardia duodenalis is responsible for the majority of parasitic gastroenteritis in humans worldwide. Host-parasite interaction models in vitro provide insights into disease and virulence and help us to understand pathogenesis. Using HT-29 intestinal epithelial cells (IEC) as a model we have demonstrated that initial sensitisation by host secretions reduces proclivity for trophozoite attachment, while inducing virulence factors. Host soluble factors triggered up-regulation of membrane and secreted proteins, including Tenascins, Cathepsin-B precursor, cystatin, and numerous Variant-specific Surface Proteins (VSPs). By comparison, host-cell attached trophozoites up-regulated intracellular pathways for ubiquitination, reactive oxygen species (ROS) detoxification and production of pyridoxal phosphate (PLP). We reason that these results demonstrate early pathogenesis in Giardia involves two independent host-parasite interactions. Motile trophozoites respond to soluble secreted signals, which deter attachment and induce expression of virulence factors. Trophozoites attached to host cells, in contrast, respond by up-regulating intracellular pathways involved in clearance of ROS, thus anticipating the host defence response. PMID:26867958

MiR224-3p inhibits hypoxia-induced autophagy by targeting autophagy-related genes in human glioblastoma cells.

PubMed

Guo, Xing; Xue, Hao; Guo, Xiaofan; Gao, Xiao; Xu, Shugang; Yan, Shaofeng; Han, Xiao; Li, Tong; Shen, Jie; Li, Gang

2015-12-08

Human glioblastoma multiforme (GBM) is a malignant solid tumor characterized by severe hypoxia. Autophagy plays a protective role in cancer cells under hypoxia. However, the microRNA (miRNA)-related molecular mechanisms underlying hypoxia-reduced autophagy remain poorly understood in GBM. In this study, we performed a miRNA microarray analysis on GBM cells and found that numerous miRNAs were differentially expressed under hypoxic conditions. Further research showed that miR224-3p, one of the significantly down-regulated miRNAs, was involved in regulating hypoxia-induced autophagy in GBM cells. Overexpression of miR224-3p abolished hypoxia-induced autophagy, whereas knocking down endogenous miR224-3p increased autophagic activity under normoxia. In addition, we demonstrated that miR224-3p inhibited autophagy by directly suppressing the expression of two autophagy-related genes (ATGs), ATG5 and FAK family-interacting protein of 200 kDa (FIP200). Furthermore, in vitro, miR224-3p attenuated cell proliferation and promoted hypoxia-induced apoptosis, and in vivo, overexpression of miR224-3p inhibited tumorigenesis of GBM cells. Collectively, our study identified a novel hypoxia-down-regulated miRNA, miR224-3p, as a key modulator of autophagy by inhibiting ATGs in GBM cells.

The modulation of Dicer regulates tumor immunogenicity in melanoma

PubMed Central

Hoffend, Nicholas C.; Magner, William J.; Tomasi, Thomas B.

2016-01-01

MicroRNAs (miRs) are small non-coding RNAs that regulate most cellular protein networks by targeting mRNAs for translational inhibition or degradation. Dicer, a type III endoribonuclease, is a critical component in microRNA biogenesis and is required for mature microRNA production. Abnormal Dicer expression occurs in numerous cancer types and correlates with poor patient prognosis. For example, increased Dicer expression in melanoma is associated with more aggressive tumors (higher tumor mitotic index and depth of invasion) and poor patient prognosis. However, the role that Dicer plays in melanoma development and immune evasion remains unclear. Here, we report on a newly discovered relationship between Dicer expression and tumor immunogenicity. To investigate Dicer's role in regulating melanoma immunogenicity, Dicer knockdown studies were performed. We found that B16F0-Dicer deficient cells exhibited decreased tumor growth compared to control cells and were capable of inducing anti-tumor immunity. The decrease in tumor growth was abrogated in immunodeficient NSG mice and was shown to be dependent upon CD8+ T cells. Dicer knockdown also induced a more responsive immune gene profile in melanoma cells. Further studies demonstrated that CD8+ T cells preferentially killed Dicer knockdown tumor cells compared to control cells. Taken together, we present evidence which links Dicer expression to tumor immunogenicity in melanoma. PMID:27356752

The modulation of Dicer regulates tumor immunogenicity in melanoma.

PubMed

Hoffend, Nicholas C; Magner, William J; Tomasi, Thomas B

2016-07-26

MicroRNAs (miRs) are small non-coding RNAs that regulate most cellular protein networks by targeting mRNAs for translational inhibition or degradation. Dicer, a type III endoribonuclease, is a critical component in microRNA biogenesis and is required for mature microRNA production. Abnormal Dicer expression occurs in numerous cancer types and correlates with poor patient prognosis. For example, increased Dicer expression in melanoma is associated with more aggressive tumors (higher tumor mitotic index and depth of invasion) and poor patient prognosis. However, the role that Dicer plays in melanoma development and immune evasion remains unclear. Here, we report on a newly discovered relationship between Dicer expression and tumor immunogenicity. To investigate Dicer's role in regulating melanoma immunogenicity, Dicer knockdown studies were performed. We found that B16F0-Dicer deficient cells exhibited decreased tumor growth compared to control cells and were capable of inducing anti-tumor immunity. The decrease in tumor growth was abrogated in immunodeficient NSG mice and was shown to be dependent upon CD8+ T cells. Dicer knockdown also induced a more responsive immune gene profile in melanoma cells. Further studies demonstrated that CD8+ T cells preferentially killed Dicer knockdown tumor cells compared to control cells. Taken together, we present evidence which links Dicer expression to tumor immunogenicity in melanoma.

Versatile roles of extracellular vesicles in cancer

PubMed Central

Kosaka, Nobuyoshi; Yoshioka, Yusuke; Fujita, Yu

2016-01-01

Numerous studies have shown that non–cell-autonomous regulation of cancer cells is an important aspect of tumorigenesis. Cancer cells need to communicate with stromal cells by humoral factors such as VEGF, FGFs, and Wnt in order to survive. Recently, extracellular vesicles (EVs) have also been shown to be involved in cell-cell communication between cancer cells and the surrounding microenvironment and to be important for the development of cancer. In addition, these EVs contain small noncoding RNAs, including microRNAs (miRNAs), which contribute to the malignancy of cancer cells. Here, we provide an overview of current research on EVs, especially miRNAs in EVs. We also propose strategies to treat cancers by targeting EVs around cancer cells. PMID:26974161

Mechanical regulation of stem-cell differentiation by the stretch-activated Piezo channel.

PubMed

He, Li; Si, Guangwei; Huang, Jiuhong; Samuel, Aravinthan D T; Perrimon, Norbert

2018-03-01

Somatic stem cells constantly adjust their self-renewal and lineage commitment by integrating various environmental cues to maintain tissue homeostasis. Although numerous chemical and biological signals have been identified that regulate stem-cell behaviour, whether stem cells can directly sense mechanical signals in vivo remains unclear. Here we show that mechanical stress regulates stem-cell differentiation in the adult Drosophila midgut through the stretch-activated ion channel Piezo. We find that Piezo is specifically expressed in previously unidentified enteroendocrine precursor cells, which have reduced proliferation ability and are destined to become enteroendocrine cells. Loss of Piezo activity reduces the generation of enteroendocrine cells in the adult midgut. In addition, ectopic expression of Piezo in all stem cells triggers both cell proliferation and enteroendocrine cell differentiation. Both the Piezo mutant and overexpression phenotypes can be rescued by manipulation of cytosolic Ca 2+ levels, and increases in cytosolic Ca 2+ resemble the Piezo overexpression phenotype, suggesting that Piezo functions through Ca 2+ signalling. Further studies suggest that Ca 2+ signalling promotes stem-cell proliferation and differentiation through separate pathways. Finally, Piezo is required for both mechanical activation of stem cells in a gut expansion assay and the increase of cytosolic Ca 2+ in response to direct mechanical stimulus in a gut compression assay. Thus, our study demonstrates the existence of a specific group of stem cells in the fly midgut that can directly sense mechanical signals through Piezo.

miR-28 regulates the germinal center reaction and blocks tumor growth in preclinical models of non-Hodgkin lymphoma

PubMed Central

Bartolomé-Izquierdo, Nahikari; Mur, Sonia M.

2017-01-01

Non-Hodgkin lymphoma comprises a variety of neoplasms, many of which arise from germinal center (GC)-experienced B cells. microRNA-28 (miR-28) is a GC-specific miRNA whose expression is lost in numerous mature B-cell neoplasms. Here we show that miR-28 regulates the GC reaction in primary B cells by impairing class switch recombination and memory B and plasma cell differentiation. Deep quantitative proteomics combined with transcriptome analysis identified miR-28 targets involved in cell-cycle and B-cell receptor signaling. Accordingly, we found that miR-28 expression diminished proliferation in primary and lymphoma cells in vitro. Importantly, miR-28 reexpression in human Burkitt (BL) and diffuse large B-cell lymphoma (DLBCL) xenografts blocked tumor growth, both when delivered in viral vectors or as synthetic, clinically amenable, molecules. Further, the antitumoral effect of miR-28 is conserved in a primary murine in vivo model of BL. Thus, miR-28 replacement is uncovered as a novel therapeutic strategy for DLBCL and BL treatment. PMID:28188132

Evolution-development congruence in pattern formation dynamics: Bifurcations in gene expression and regulation of networks structures.

PubMed

Kohsokabe, Takahiro; Kaneko, Kunihiko

2016-01-01

Search for possible relationships between phylogeny and ontogeny is important in evolutionary-developmental biology. Here we uncover such relationships by numerical evolution and unveil their origin in terms of dynamical systems theory. By representing developmental dynamics of spatially located cells with gene expression dynamics with cell-to-cell interaction under external morphogen gradient, gene regulation networks are evolved under mutation and selection with the fitness to approach a prescribed spatial pattern of expressed genes. For most numerical evolution experiments, evolution of pattern over generations and development of pattern by an evolved network exhibit remarkable congruence. Both in the evolution and development pattern changes consist of several epochs where stripes are formed in a short time, while for other temporal regimes, pattern hardly changes. In evolution, these quasi-stationary regimes are generations needed to hit relevant mutations, while in development, they are due to some gene expression that varies slowly and controls the pattern change. The morphogenesis is regulated by combinations of feedback or feedforward regulations, where the upstream feedforward network reads the external morphogen gradient, and generates a pattern used as a boundary condition for the later patterns. The ordering from up to downstream is common in evolution and development, while the successive epochal changes in development and evolution are represented as common bifurcations in dynamical-systems theory, which lead to the evolution-development congruence. Mechanism of exceptional violation of the congruence is also unveiled. Our results provide a new look on developmental stages, punctuated equilibrium, developmental bottlenecks, and evolutionary acquisition of novelty in morphogenesis. © 2015 The Authors. Journal of Experimental Zoology Part B: Molecular and Developmental Evolution Published by Wiley Periodicals, Inc.

Trichostatin A Sensitizes Hepatocellular Carcinoma Cells to Enhanced NK Cell-mediated Killing by Regulating Immune-related Genes.

PubMed

Shin, Sangsu; Kim, Miok; Lee, Seon-Jin; Park, Kang-Seo; Lee, Chang Hoon

2017-01-01

Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related death worldwide. The ability of HCC to avoid immune detection is considered one of the main factors making it difficult to cure. Abnormal histone deacetylation is thought to be one of the mechanisms for HCC immune escape, making histone deacetylases (HDACs) attractive targets for HCC treatment. Here, we investigated the effect of trichostatin A (TSA), a highly potent HDAC inhibitor, on HCC (HepG2) gene expression and function. A genome wide-transcriptional microarray was used to identify genes regulated by TSA in HepG2 cells. Gene Ontology was used to identify pathways regulated by TSA, and these changes were confirmed by qPCR. The effect of TSA on natural killer (NK) cell-mediated killing of HCC cell lines were analyzed by both flow cytometry and LDH cytotoxicity assay. A study was also conducted in a Balb/c nude mice xenograft model to assess the anti-tumor activity of TSA. TSA regulated the transcription of numerous innate immunity & tumor antigen recognition-associated genes, such as ULBP1 and RAET1G, in HCC cells. In vivo, TSA reduced tumor cell growth in an NK cell-dependent manner. In vitro, TSA treatment of HepG2 cells rendered them more susceptible to NK cell-mediated killing while increasing the expression of NKGD2 ligands, including ULBP1/2/3 and MICA/B. TSA also induced direct killing of HCC cells by stimulating apoptosis. TSA likely increases killing of HCC cells indirectly by increasing NK cell-directed killing and directly by increasing apoptosis. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

Trichostatin A Sensitizes Hepatocellular Carcinoma Cells to Enhanced NK Cell-mediated Killing by Regulating Immune-related Genes

PubMed Central

SHIN, SANGSU; KIM, MIOK; LEE, SEON-JIN; PARK, KANG-SEO

2017-01-01

Background/Aim: Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related death worldwide. The ability of HCC to avoid immune detection is considered one of the main factors making it difficult to cure. Abnormal histone deacetylation is thought to be one of the mechanisms for HCC immune escape, making histone deacetylases (HDACs) attractive targets for HCC treatment. Here, we investigated the effect of trichostatin A (TSA), a highly potent HDAC inhibitor, on HCC (HepG2) gene expression and function. Materials and Methods: A genome wide-transcriptional microarray was used to identify genes regulated by TSA in HepG2 cells. Gene Ontology was used to identify pathways regulated by TSA, and these changes were confirmed by qPCR. The effect of TSA on natural killer (NK) cell-mediated killing of HCC cell lines were analyzed by both flow cytometry and LDH cytotoxicity assay. A study was also conducted in a Balb/c nude mice xenograft model to assess the anti-tumor activity of TSA. Results: TSA regulated the transcription of numerous innate immunity & tumor antigen recognition-associated genes, such as ULBP1 and RAET1G, in HCC cells. In vivo, TSA reduced tumor cell growth in an NK cell-dependent manner. In vitro, TSA treatment of HepG2 cells rendered them more susceptible to NK cell-mediated killing while increasing the expression of NKGD2 ligands, including ULBP1/2/3 and MICA/B. TSA also induced direct killing of HCC cells by stimulating apoptosis. Conclusion: TSA likely increases killing of HCC cells indirectly by increasing NK cell-directed killing and directly by increasing apoptosis. PMID:28871002

Repression of myoblast proliferation and fibroblast growth factor receptor 1 promoter activity by KLF10 protein.

PubMed

Parakati, Rajini; DiMario, Joseph X

2013-05-10

FGFR1 gene expression regulates myoblast proliferation and differentiation, and its expression is controlled by Krüppel-like transcription factors. KLF10 interacts with the FGFR1 promoter, repressing its activity and cell proliferation. KLF10 represses FGFR1 promoter activity and thereby myoblast proliferation. A model of transcriptional control of chicken FGFR1 gene regulation during myogenesis is presented. Skeletal muscle development is controlled by regulation of myoblast proliferation and differentiation into muscle fibers. Growth factors such as fibroblast growth factors (FGFs) and their receptors (FGFRs) regulate cell proliferation and differentiation in numerous tissues, including skeletal muscle. Transcriptional regulation of FGFR1 gene expression is developmentally regulated by the Sp1 transcription factor, a member of the Krüppel-like factor (KLF) family of transcriptional regulators. Here, we show that another KLF transcription factor, KLF10, also regulates myoblast proliferation and FGFR1 promoter activity. Expression of KLF10 reduced myoblast proliferation by 86%. KLF10 expression also significantly reduced FGFR1 promoter activity in myoblasts and Sp1-mediated FGFR1 promoter activity in Drosophila SL2 cells. Southwestern blot, electromobility shift, and chromatin immunoprecipitation assays demonstrated that KLF10 bound to the proximal Sp factor binding site of the FGFR1 promoter and reduced Sp1 complex formation with the FGFR1 promoter at that site. These results indicate that KLF10 is an effective repressor of myoblast proliferation and represses FGFR1 promoter activity in these cells via an Sp1 binding site.

Quantitative Evaluation of CART-Containing Cells in Urinary Bladder of Rats with Renovascular Hypertension

PubMed Central

Janiuk, I.; Kasacka, I.

2015-01-01

Recent biological advances make it possible to discover new peptides associated with hypertension. The cocaine- and amphetamine-regulated transcript (CART) is a known factor in appetite and feeding behaviour. Various lines of evidence suggest that this peptide participates not only in control of feeding behaviour but also in the regulation of the cardiovascular and sympathetic systems and blood pressure. The role of CART in blood pressure regulation led us to undertake a study aimed at analysing quantitative changes in CART-containing cells in urinary bladders (UB) of rats with renovascular hypertension. We used the Goldblatt model of arterial hypertension (two-kidney, one clip) to evaluate quantitative changes. This model provides researchers with a commonly used tool to analyse the renin-angiotensin system of blood pressure control and, eventually, to develop drugs for the treatment of chronic hypertension. The study was performed on sections of urinary bladders of rats after 3-, 14-, 28-, 42 and 91 days from hypertension induction. Immunohistochemical identification of CART cells was performed on paraffin for the UBs of all the study animals. CART was detected in the endocrine cells, especially numerous in the submucosa and muscularis layers, with a few found in the transitional epithelium and only occasionally in serosa. Hypertension significantly increased the number of CART-positive cells in the rat UBs. After 3 and 42 days following the procedure, statistically significantly higher numbers of CART-positive cells were identified in comparison with the control animals. The differences between the hypertensive rats and the control animals concerned not only the number density of CART-immunoreactive cells but also their localization. After a 6-week period, each of the rats subjected to the renal artery clipping procedure developed stable hypertension. CART appeared in numerous transitional epithelium cells. As this study provides novel findings, the question appears about the type of connection between hypertension and the functioning and activity of CART in the urinary tract (UT). The study gives rise to the assumption that high blood pressure can be a factor that intensifies CART secretion. In conclusion, the endocrine system of the urinary tract is modified by renovascular hypertension. This may affect the production of hormones and biologically active substances and contribute to the development of possible hypertension complications. In order to fully comprehend the role of the CART peptide in blood pressure regulation, further analyses are necessary. PMID:26150151

Quantitative evaluation of CART-containing cells in urinary bladder of rats with renovascular hypertension.

PubMed

Janiuk, I; Kasacka, I

2015-04-13

Recent biological advances make it possible to discover new peptides associated with hypertension. The cocaine- and amphetamine-regulated transcript (CART) is a known factor in appetite and feeding behaviour. Various lines of evidence suggest that this peptide participates not only in control of feeding behaviour but also in the regulation of the cardiovascular and sympathetic systems and blood pressure. The role of CART in blood pressure regulation led us to undertake a study aimed at analysing quantitative changes in CART-containing cells in urinary bladders (UB) of rats with renovascular hypertension. We used the Goldblatt model of arterial hypertension (two-kidney, one clip) to evaluate quantitative changes. This model provides researchers with a commonly used tool to analyse the renin-angiotensin system of blood pressure control and, eventually, to develop drugs for the treatment of chronic hypertension. The study was performed on sections of urinary bladders of rats after 3-, 14-, 28-, 42 and 91 days from hypertension induction. Immunohistochemical identification of CART cells was performed on paraffin for the UBs of all the study animals. CART was detected in the endocrine cells, especially numerous in the submucosa and muscularis layers, with a few found in the transitional epithelium and only occasionally in serosa. Hypertension significantly increased the number of CART-positive cells in the rat UBs. After 3 and 42 days following the procedure, statistically significantly higher numbers of CART-positive cells were identified in comparison with the control animals. The differences between the hypertensive rats and the control animals concerned not only the number density of CART-immunoreactive cells but also their localization. After a 6-week period, each of the rats subjected to the renal artery clipping procedure developed stable hypertension. CART appeared in numerous transitional epithelium cells. As this study provides novel findings, the question appears about the type of connection between hypertension and the functioning and activity of CART in the urinary tract (UT). The study gives rise to the assumption that high blood pressure can be a factor that intensifies CART secretion. In conclusion, the endocrine system of the urinary tract is modified by renovascular hypertension. This may affect the production of hormones and biologically active substances and contribute to the development of possible hypertension complications. In order to fully comprehend the role of the CART peptide in blood pressure regulation, further analyses are necessary.

Acute Viral Respiratory Infection Rapidly Induces a CD8+ T Cell Exhaustion-like Phenotype.

PubMed

Erickson, John J; Lu, Pengcheng; Wen, Sherry; Hastings, Andrew K; Gilchuk, Pavlo; Joyce, Sebastian; Shyr, Yu; Williams, John V

2015-11-01

Acute viral infections typically generate functional effector CD8(+) T cells (TCD8) that aid in pathogen clearance. However, during acute viral lower respiratory infection, lung TCD8 are functionally impaired and do not optimally control viral replication. T cells also become unresponsive to Ag during chronic infections and cancer via signaling by inhibitory receptors such as programmed cell death-1 (PD-1). PD-1 also contributes to TCD8 impairment during viral lower respiratory infection, but how it regulates TCD8 impairment and the connection between this state and T cell exhaustion during chronic infections are unknown. In this study, we show that PD-1 operates in a cell-intrinsic manner to impair lung TCD8. In light of this, we compared global gene expression profiles of impaired epitope-specific lung TCD8 to functional spleen TCD8 in the same human metapneumovirus-infected mice. These two populations differentially regulate hundreds of genes, including the upregulation of numerous inhibitory receptors by lung TCD8. We then compared the gene expression of TCD8 during human metapneumovirus infection to those in acute or chronic lymphocytic choriomeningitis virus infection. We find that the immunophenotype of lung TCD8 more closely resembles T cell exhaustion late into chronic infection than do functional effector T cells arising early in acute infection. Finally, we demonstrate that trafficking to the infected lung alone is insufficient for TCD8 impairment or inhibitory receptor upregulation, but that viral Ag-induced TCR signaling is also required. Our results indicate that viral Ag in infected lungs rapidly induces an exhaustion-like state in lung TCD8 characterized by progressive functional impairment and upregulation of numerous inhibitory receptors. Copyright © 2015 by The American Association of Immunologists, Inc.

Estrogens and human papilloma virus oncogenes regulate human ether-à-go-go-1 potassium channel expression.

PubMed

Díaz, Lorenza; Ceja-Ochoa, Irais; Restrepo-Angulo, Iván; Larrea, Fernando; Avila-Chávez, Euclides; García-Becerra, Rocío; Borja-Cacho, Elizabeth; Barrera, David; Ahumada, Elías; Gariglio, Patricio; Alvarez-Rios, Elizabeth; Ocadiz-Delgado, Rodolfo; Garcia-Villa, Enrique; Hernández-Gallegos, Elizabeth; Camacho-Arroyo, Ignacio; Morales, Angélica; Ordaz-Rosado, David; García-Latorre, Ethel; Escamilla, Juan; Sánchez-Peña, Luz Carmen; Saqui-Salces, Milena; Gamboa-Dominguez, Armando; Vera, Eunice; Uribe-Ramírez, Marisela; Murbartián, Janet; Ortiz, Cindy Sharon; Rivera-Guevara, Claudia; De Vizcaya-Ruiz, Andrea; Camacho, Javier

2009-04-15

Ether-à-go-go-1 (Eag1) potassium channels are potential tools for detection and therapy of numerous cancers. Here, we show human Eag1 (hEag1) regulation by cancer-associated factors. We studied hEag1 gene expression and its regulation by estradiol, antiestrogens, and human papillomavirus (HPV) oncogenes (E6/E7). Primary cultures from normal placentas and cervical cancer tissues; tumor cell lines from cervix, choriocarcinoma, keratinocytes, and lung; and normal cell lines from vascular endothelium, keratinocytes, and lung were used. Reverse transcription-PCR (RT-PCR) experiments and Southern blot analysis showed Eag1 expression in all of the cancer cell types, normal trophoblasts, and vascular endothelium, in contrast to normal keratinocytes and lung cells. Estradiol and antiestrogens regulated Eag1 in a cell type-dependent manner. Real-time RT-PCR experiments in HeLa cells showed that Eag1 estrogenic regulation was strongly associated with the expression of estrogen receptor-alpha. Eag1 protein was detected by monoclonal antibodies in normal placenta and placental blood vessels. Patch-clamp recordings in normal trophoblasts treated with estradiol exhibited potassium currents resembling Eag1 channel activity. Eag1 gene expression in keratinocytes depended either on cellular immortalization or the presence of HPV oncogenes. Eag1 protein was found in keratinocytes transfected with E6/E7 HPV oncogenes. Cell proliferation of E6/E7 keratinocytes was decreased by Eag1 antibodies inhibiting channel activity and by the nonspecific Eag1 inhibitors imipramine and astemizole; the latter also increased apoptosis. Our results propose novel oncogenic mechanisms of estrogen/antiestrogen use and HPV infection. We also suggest Eag1 as an early indicator of cell proliferation leading to malignancies and a therapeutic target at early stages of cellular hyperproliferation.

A Novel Combination of Withaferin A and Sulforaphane Inhibits Epigenetic Machinery, Cellular Viability and Induces Apoptosis of Breast Cancer Cells

PubMed Central

Royston, Kendra J.; Udayakumar, Neha; Lewis, Kayla; Tollefsbol, Trygve O.

2017-01-01

With cancer often classified as a disease that has an important epigenetic component, natural compounds that have the ability to regulate the epigenome become ideal candidates for study. Humans have a complex diet, which illustrates the need to elucidate the mechanisms of interaction between these bioactive compounds in combination. The natural compounds withaferin A (WA), from the Indian winter cherry, and sulforaphane (SFN), from cruciferous vegetables, have numerous anti-cancer effects and some report their ability to regulate epigenetic processes. Our study is the first to investigate the combinatorial effects of low physiologically achievable concentrations of WA and SFN on breast cancer cell proliferation, histone deacetylase1 (HDAC1) and DNA methyltransferases (DNMTs). No adverse effects were observed on control cells at optimal concentrations. There was synergistic inhibition of cellular viability in MCF-7 cells and a greater induction of apoptosis with the combinatorial approach than with either compound administered alone in both MDA-MB-231 and MCF-7 cells. HDAC expression was down-regulated at multiple levels. Lastly, we determined the combined effects of these bioactive compounds on the pro-apoptotic BAX and anti-apoptotic BCL-2 and found decreases in BCL-2 and increases in BAX. Taken together, our findings demonstrate the ability of low concentrations of combinatorial WA and SFN to promote cancer cell death and regulate key epigenetic modifiers in human breast cancer cells. PMID:28534825

A Novel Combination of Withaferin A and Sulforaphane Inhibits Epigenetic Machinery, Cellular Viability and Induces Apoptosis of Breast Cancer Cells.

PubMed

Royston, Kendra J; Udayakumar, Neha; Lewis, Kayla; Tollefsbol, Trygve O

2017-05-19

With cancer often classified as a disease that has an important epigenetic component, natural compounds that have the ability to regulate the epigenome become ideal candidates for study. Humans have a complex diet, which illustrates the need to elucidate the mechanisms of interaction between these bioactive compounds in combination. The natural compounds withaferin A (WA), from the Indian winter cherry, and sulforaphane (SFN), from cruciferous vegetables, have numerous anti-cancer effects and some report their ability to regulate epigenetic processes. Our study is the first to investigate the combinatorial effects of low physiologically achievable concentrations of WA and SFN on breast cancer cell proliferation, histone deacetylase1 (HDAC1) and DNA methyltransferases (DNMTs). No adverse effects were observed on control cells at optimal concentrations. There was synergistic inhibition of cellular viability in MCF-7 cells and a greater induction of apoptosis with the combinatorial approach than with either compound administered alone in both MDA-MB-231 and MCF-7 cells. HDAC expression was down-regulated at multiple levels. Lastly, we determined the combined effects of these bioactive compounds on the pro-apoptotic BAX and anti-apoptotic BCL-2 and found decreases in BCL-2 and increases in BAX . Taken together, our findings demonstrate the ability of low concentrations of combinatorial WA and SFN to promote cancer cell death and regulate key epigenetic modifiers in human breast cancer cells.

p53 mediated transcriptional regulation of long non-coding RNA by 1-hydroxy-1-norresistomycin triggers intrinsic apoptosis in adenocarcinoma lung cancer.

PubMed

Ramalingam, Vaikundamoorthy; Varunkumar, Krishnamoorthy; Ravikumar, Vilwanathan; Rajaram, Rajendran

2018-05-01

Over a few decades, systemic chemotherapy and surgery are the only treatment options for lung cancer. Due to limited efficacy and overall poor survival of patients, it is necessary to develop a newer therapeutic strategy which specifically targets cancer cell proliferation pathway. Deciphering the role of long non-coding RNAs (lncRNAs) in tumorigenesis and pathogenesis of cancer cells has recently emerged. In the present study, marine actinomycetes derived 1-hydroxy-1-norresistomycin (HNM) was used to enhance the expression of lncRNAs through p53 transcriptional regulation and induced intrinsic apoptosis in non-small cell lung cancer cells. Initially, concentration dependent treatment with HNM has increased the ROS generation in mitochondria and sensitizes the mitochondrial membrane potential. Further, HNM downregulates the numerous oncogenes which regulate cancer cell proliferation, metastasis and invasion and tumor suppressor genes which are involved in intrinsic apoptosis confirmed with adopting techniques such as RT-PCR and western blot analysis. Moreover, ChIP assay results showed that HNM upregulates the p53 mediated transcriptional regulation of lncRNAs lead to apoptosis of cancer cells through cell cycle arrest and inhibition of proliferation. In conclusion, HNM found to be a potential therapeutic agent for treatment of lung cancer via suppression of oncogenes and expression of wide range of tumor suppressor genes are might have significant implications in cancer treatment and drug development. Copyright © 2018 Elsevier B.V. All rights reserved.

Vasohibin-1 is identified as a master-regulator of endothelial cell apoptosis using gene network analysis

PubMed Central

2013-01-01

Background Apoptosis is a critical process in endothelial cell (EC) biology and pathology, which has been extensively studied at protein level. Numerous gene expression studies of EC apoptosis have also been performed, however few attempts have been made to use gene expression data to identify the molecular relationships and master regulators that underlie EC apoptosis. Therefore, we sought to understand these relationships by generating a Bayesian gene regulatory network (GRN) model. Results ECs were induced to undergo apoptosis using serum withdrawal and followed over a time course in triplicate, using microarrays. When generating the GRN, this EC time course data was supplemented by a library of microarray data from EC treated with siRNAs targeting over 350 signalling molecules. The GRN model proposed Vasohibin-1 (VASH1) as one of the candidate master-regulators of EC apoptosis with numerous downstream mRNAs. To evaluate the role played by VASH1 in EC, we used siRNA to reduce the expression of VASH1. Of 10 mRNAs downstream of VASH1 in the GRN that were examined, 7 were significantly up- or down-regulated in the direction predicted by the GRN.Further supporting an important biological role of VASH1 in EC, targeted reduction of VASH1 mRNA abundance conferred resistance to serum withdrawal-induced EC death. Conclusion We have utilised Bayesian GRN modelling to identify a novel candidate master regulator of EC apoptosis. This study demonstrates how GRN technology can complement traditional methods to hypothesise the regulatory relationships that underlie important biological processes. PMID:23324451

Sirt2 suppresses glioma cell growth through targeting NF-κB–miR-21 axis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Ya’nan; Dai, Dongwei; Lu, Qiong

Highlights: •Sirt2 expression is down-regulated in human glioma tissues and cell lines. •Sirt2 regresses glioma cell growth and colony formation via inducing apoptosis. •miR-21 is essential for the functions of Sirt2 in glioma cells. •Sirt2 deacetylates p65 to decrease miR-21 expression. -- Abstract: Sirtuins are NAD{sup +}-dependent deacetylases that regulate numerous cellular processes including aging, DNA repair, cell cycle, metabolism, and survival under stress conditions. The roles of sirtuin family members are widely studied in carcinogenesis. However, their roles in glioma remain unclear. Here we report that Sir2 was under expressed in human glioma tissues and cell lines. We foundmore » that Sirt2 overexpression decreased cell proliferation and colony formation capacity. In addition, Sirt2 overexpression induced cellular apoptosis via up-regulating cleaved caspase 3 and Bax, and down-regulating anti-apoptotic protein Bcl-2. Sirt2 knockdown obtained opposing results. We showed that Sirt2 overexpression inhibited miR-21 expression, and Sirt2 was not sufficient to reduce cell proliferation and colony formation as well as to induce apoptosis when miR-21 was knocked down in glioma cells. Mechanically, we demonstrated that Sirt2 deacetylated p65 at K310 and blocked p65 binding to the promoter region of miR-21, thus regressing the transcription of miR-21. In summary, Sirt2 is critical in human glioma via NF-κB–miR-21 pathway and Sirt2 activator may serve as candidate drug for glioma therapy.« less

Apoptosis in Drosophila: which role for mitochondria?

PubMed

Clavier, Amandine; Rincheval-Arnold, Aurore; Colin, Jessie; Mignotte, Bernard; Guénal, Isabelle

2016-03-01

It is now well established that the mitochondrion is a central regulator of mammalian cell apoptosis. However, the importance of this organelle in non-mammalian apoptosis has long been regarded as minor, mainly because of the absence of a crucial role for cytochrome c in caspase activation. Recent results indicate that the control of caspase activation and cell death in Drosophila occurs at the mitochondrial level. Numerous proteins, including RHG proteins and proteins of the Bcl-2 family that are key regulators of Drosophila apoptosis, constitutively or transiently localize in mitochondria. These proteins participate in the cell death process at different levels such as degradation of Diap1, a Drosophila IAP, production of mitochondrial reactive oxygen species or stimulation of the mitochondrial fission machinery. Here, we review these mitochondrial events that might have their counterpart in human.

TES inhibits colorectal cancer progression through activation of p38.

PubMed

Li, Huili; Huang, Kun; Gao, Lu; Wang, Lixia; Niu, Yanfeng; Liu, Hongli; Wang, Zheng; Wang, Lin; Wang, Guobin; Wang, Jiliang

2016-07-19

The human TESTIN (TES) gene has been identified as a candidate tumor suppressor based on its location at a common fragile site - a region where loss of heterozygosity has been detected in numerous types of tumors. To investigate its role in colorectal cancer (CRC), we examined TES protein levels in CRC tissue samples and cell lines. We observed that TES was markedly reduced in both CRC tissue and cell lines. Additionally, overexpression of TES significantly inhibited cell proliferation, migration, and invasion, while increasing cell apoptosis in colon cancer cells. By contrast, shRNA-mediated TES knockdown elicited the opposite effects. TES inhibited the progression of CRC by up-regulating pro-apoptotic proteins, down-regulating anti-apoptotic proteins, and simultaneously activating p38 mitogen-activated protein kinase (MAPK) signaling pathways. Collectively, these data indicate that TES functions as a necessary suppressor of CRC progression by activating p38-MAPK signaling pathways. This suggests that TES may have a potential application in CRC diagnosis and targeted gene therapy.

TES inhibits colorectal cancer progression through activation of p38

PubMed Central

Gao, Lu; Wang, Lixia; Niu, Yanfeng; Liu, Hongli; Wang, Zheng; Wang, Lin; Wang, Guobin; Wang, Jiliang

2016-01-01

The human TESTIN (TES) gene has been identified as a candidate tumor suppressor based on its location at a common fragile site – a region where loss of heterozygosity has been detected in numerous types of tumors. To investigate its role in colorectal cancer (CRC), we examined TES protein levels in CRC tissue samples and cell lines. We observed that TES was markedly reduced in both CRC tissue and cell lines. Additionally, overexpression of TES significantly inhibited cell proliferation, migration, and invasion, while increasing cell apoptosis in colon cancer cells. By contrast, shRNA-mediated TES knockdown elicited the opposite effects. TES inhibited the progression of CRC by up-regulating pro-apoptotic proteins, down-regulating anti-apoptotic proteins, and simultaneously activating p38 mitogen-activated protein kinase (MAPK) signaling pathways. Collectively, these data indicate that TES functions as a necessary suppressor of CRC progression by activating p38-MAPK signaling pathways. This suggests that TES may have a potential application in CRC diagnosis and targeted gene therapy. PMID:27323777

The 26S Proteasome Complex: An Attractive Target for Cancer Therapy

PubMed Central

Frankland-Searby, Sarah; Bhaumik, Sukesh R.

2011-01-01

The 26S proteasome complex engages in an ATP-dependent proteolytic degradation of a variety of oncoproteins, transcription factors, cell cycle specific cyclins, cyclin-dependent kinase inhibitors, ornithine decarboxylase, and other key regulatory cellular proteins. Thus, the proteasome regulates either directly or indirectly many important cellular processes. Altered regulation of these cellular events is linked to the development of cancer. Therefore, the proteasome has become an attractive target for the treatment of numerous cancers. Several proteasome inhibitors that target the proteolytic active sites of the 26S proteasome complex have been developed and tested for anti-tumor activities. These proteasome inhibitors have displayed impressive anti-tumor functions by inducing apoptosis in different tumor types. Further, the proteasome inhibitors have been shown to induce cell cycle arrest, and inhibit angiogenesis, cell-cell adhesion, cell migration, immune and inflammatory responses, and DNA repair response. A number of proteasome inhibitors are now in clinical trials to treat multiple myeloma and solid tumors. Many other proteasome inhibitors with different efficiencies are being developed and tested for anti-tumor activities. Several proteasome inhibitors currently in clinical trials have shown significantly improved anti-tumor activities when combined with other drugs such as histone deacetylase (HDAC) inhibitors, Akt (protein kinase B) inhibitors, DNA damaging agents, Hsp90 (heat shock protein 90) inhibitors, and lenalidomide. The proteasome inhibitor bortezomib is now in the clinic to treat multiple myeloma and mantle cell lymphoma. Here, we discuss the 26S proteasome complex in carcinogenesis and different proteasome inhibitors with their potential therapeutic applications in treatment of numerous cancers. PMID:22037302

Circadian locomotor output cycles kaput affects the proliferation and migration of breast cancer cells by regulating the expression of E-cadherin via IQ motif containing GTPase activating protein 1.

PubMed

Li, Xiaoxue; Wang, Siyang; Yang, Shuhong; Ying, Junjie; Yu, Hang; Yang, Chunlei; Liu, Yanyou; Wang, Yuhui; Cheng, Shuting; Xiao, Jing; Guo, Huiling; Jiang, Zhou; Wang, Zhengrong

2018-05-01

The circadian rhythm regulates numerous physiological activities, including sleep and wakefulness, behavior, immunity and metabolism. Previous studies have demonstrated that circadian rhythm disorder is associated with the occurrence of tumors. Responsible for regulating a number of functions, the Circadian locomotor output cycles kaput ( Clock ) gene is one of the core regulatory genes of circadian rhythm. The Clock gene has also been implicated in the occurrence and development of tumors in previously studies. The present study evaluated the role of the Clock gene in the proliferation and migration of mouse breast cancer 4T1 cells, and investigated its possible regulatory pathways and mechanisms. It was reported that downregulation of Clock facilitated the proliferation and migration of breast cancer cells. Further investigation revealed the involvement of IQ motif containing GTPase activating protein 1 (IQGAP1) protein expression in the Clock regulatory pathway, further influencing the expression of E-cadherin, a known proprietor of tumor cell migration and invasion. To the best of our knowledge, the present study is the first to report that Clock , acting through the regulation of the scaffolding protein IQGAP1, regulates the downstream expression of E-cadherin, thereby affecting tumor cell structure and motility. These results confirmed the role of Clock in breast cancer tumor etiology and provide insight regarding the molecular avenues of its regulatory nature, which may translate beyond breast cancer into other known functions of the gene.

The interdependence of the Rho GTPases and apicobasal cell polarity.

PubMed

Mack, Natalie Ann; Georgiou, Marios

2014-01-01

Signaling via the Rho GTPases provides crucial regulation of numerous cell polarization events, including apicobasal (AB) polarity, polarized cell migration, polarized cell division and neuronal polarity. Here we review the relationships between the Rho family GTPases and epithelial AB polarization events, focusing on the 3 best-characterized members: Rho, Rac and Cdc42. We discuss a multitude of processes that are important for AB polarization, including lumen formation, apical membrane specification, cell-cell junction assembly and maintenance, as well as tissue polarity. Our discussions aim to highlight the immensely complex regulatory mechanisms that encompass Rho GTPase signaling during AB polarization. More specifically, in this review we discuss several emerging common themes, that include: 1) the need for Rho GTPase activities to be carefully balanced in both a spatial and temporal manner through a multitude of mechanisms; 2) the existence of signaling feedback loops and crosstalk to create robust cellular responses; and 3) the frequent multifunctionality that exists among AB polarity regulators. Regarding this latter theme, we provide further discussion of the potential plasticity of the cell polarity machinery and as a result the possible implications for human disease.

Gene expression networks underlying ovarian development in wild largemouth bass (Micropterus salmoides).

PubMed

Martyniuk, Christopher J; Prucha, Melinda S; Doperalski, Nicholas J; Antczak, Philipp; Kroll, Kevin J; Falciani, Francesco; Barber, David S; Denslow, Nancy D

2013-01-01

Oocyte maturation in fish involves numerous cell signaling cascades that are activated or inhibited during specific stages of oocyte development. The objectives of this study were to characterize molecular pathways and temporal gene expression patterns throughout a complete breeding cycle in wild female largemouth bass to improve understanding of the molecular sequence of events underlying oocyte maturation. Transcriptomic analysis was performed on eight morphologically diverse stages of the ovary, including primary and secondary stages of oocyte growth, ovulation, and atresia. Ovary histology, plasma vitellogenin, 17β-estradiol, and testosterone were also measured to correlate with gene networks. Global expression patterns revealed dramatic differences across ovarian development, with 552 and 2070 genes being differentially expressed during both ovulation and atresia respectively. Gene set enrichment analysis (GSEA) revealed that early primary stages of oocyte growth involved increases in expression of genes involved in pathways of B-cell and T-cell receptor-mediated signaling cascades and fibronectin regulation. These pathways as well as pathways that included adrenergic receptor signaling, sphingolipid metabolism and natural killer cell activation were down-regulated at ovulation. At atresia, down-regulated pathways included gap junction and actin cytoskeleton regulation, gonadotrope and mast cell activation, and vasopressin receptor signaling and up-regulated pathways included oxidative phosphorylation and reactive oxygen species metabolism. Expression targets for luteinizing hormone signaling were low during vitellogenesis but increased 150% at ovulation. Other networks found to play a significant role in oocyte maturation included those with genes regulated by members of the TGF-beta superfamily (activins, inhibins, bone morphogenic protein 7 and growth differentiation factor 9), neuregulin 1, retinoid X receptor, and nerve growth factor family. This study offers novel insight into the gene networks underlying vitellogenesis, ovulation and atresia and generates new hypotheses about the cellular pathways regulating oocyte maturation.

EBNA3C regulates p53 through induction of Aurora kinase B

PubMed Central

Jha, Hem C.; Yang, Karren; El-Naccache, Darine W.; Sun, Zhiguo; Robertson, Erle S.

2015-01-01

In multicellular organisms p53 maintains genomic integrity through activation of DNA repair, and apoptosis. EBNA3C can down regulate p53 transcriptional activity. Aurora kinase (AK) B phosphorylates p53, which leads to degradation of p53. Aberrant expression of AK-B is a hallmark of numerous human cancers. Therefore changes in the activities of p53 due to AK-B and EBNA3C expression is important for understanding EBV-mediated cell transformation. Here we show that the activities of p53 and its homolog p73 are dysregulated in EBV infected primary cells which can contribute to increased cell transformation. Further, we showed that the ETS-1 binding site is crucial for EBNA3C-mediated up-regulation of AK-B transcription. Further, we determined the Ser 215 residue of p53 is critical for functional regulation by AK-B and EBNA3C and that the kinase domain of AK-B which includes amino acid residues 106, 111 and 205 was important for p53 regulation. AK-B with a mutation at residue 207 was functionally similar to wild type AK-B in terms of its kinase activities and knockdown of AK-B led to enhanced p73 expression independent of p53. This study explores an additional mechanism by which p53 is regulated by AK-B and EBNA3C contributing to EBV-induced B-cell transformation. PMID:25691063

Transcriptome-wide analysis of the Trypanosoma cruzi proliferative cycle identifies the periodically expressed mRNAs and their multiple levels of control

PubMed Central

Chávez, Santiago; Eastman, Guillermo; Smircich, Pablo; Becco, Lorena Lourdes; Oliveira-Rizzo, Carolina; Fort, Rafael; Potenza, Mariana; Garat, Beatriz; Sotelo-Silveira, José Roberto

2017-01-01

Trypanosoma cruzi is the protozoan parasite causing American trypanosomiasis or Chagas disease, a neglected parasitosis with important human health impact in Latin America. The efficacy of current therapy is limited, and its toxicity is high. Since parasite proliferation is a fundamental target for rational drug design, we sought to progress into its understanding by applying a genome-wide approach. Treating a TcI linage strain with hydroxyurea, we isolated epimastigotes in late G1, S and G2/M cell cycle stages at 70% purity. The sequencing of each phase identified 305 stage-specific transcripts (1.5-fold change, p≤0.01), coding for conserved cell cycle regulated proteins and numerous proteins whose cell cycle dependence has not been recognized before. Comparisons with the parasite T. brucei and the human host reveal important differences. The meta-analysis of T. cruzi transcriptomic and ribonomic data indicates that cell cycle regulated mRNAs are subject to sub-cellular compartmentalization. Compositional and structural biases of these genes- including CAI, GC content, UTR length, and polycistron position- may contribute to their regulation. To discover nucleotide motifs responsible for the co-regulation of cell cycle regulated genes, we looked for overrepresented motifs at their UTRs and found a variant of the cell cycle sequence motif at the 3' UTR of most of the S and G2 stage genes. We additionally identified hairpin structures at the 5' UTRs of a high proportion of the transcripts, suggesting that periodic gene expression might also rely on translation initiation in T. cruzi. In summary, we report a comprehensive list of T. cruzi cell cycle regulated genes, including many previously unstudied proteins, we show evidence favoring a multi-step control of their expression, and we identify mRNA motifs that may mediate their regulation. Our results provide novel information of the T. cruzi proliferative proteins and the integrated levels of their gene expression control. PMID:29182646

MicroRNA-20b-5p inhibits platelet-derived growth factor-induced proliferation of human fetal airway smooth muscle cells by targeting signal transducer and activator of transcription 3.

PubMed

Tang, Jin; Luo, Lingying

2018-06-01

Pediatric asthma is still a health threat to the pediatric population in recent years. The airway remodeling induced by abnormal airway smooth muscle (ASM) cell proliferation is an important cause of asthma. MicroRNAs (miRNAs) are important regulators of ASM cell proliferation. Numerous studies have reported that miR-20b-5p is a critical regulator for cell proliferation. However, whether miR-20b-5p is involved in regulating ASM cell proliferation remains unknown. In this study, we aimed to investigate the potential role of miR-20b-5p in regulating the proliferation of fetal ASM cell induced by platelet-derived growth factor (PDGF). Here, we showed that miR-20b-5p was significantly decreased in fetal ASM cells treated with PDGF. Biological experiments showed that the overexpression of miR-20b-5p inhibited the proliferation while miR-20b-5p inhibition markedly promoted the proliferation of fetal ASM cells. Bioinformatics analysis and luciferase reporter assay showed that miR-20b-5p directly targeted the 3'-UTR of signal transducer and activator of transcription 3 (STAT3). Further data showed that miR-20b-5p negatively regulated the expression of STAT3 in fetal ASM cells. Moreover, miR-20b-5p regulates the transcriptional activity of STAT3 in fetal ASM cells. Overexpression of STAT3 reversed the inhibitory effect of miR-20b-5p overexpression on fetal ASM cell proliferation while the knockdown of STAT3 abrogated the promoted effect of miR-20b-5p inhibition on fetal ASM cell proliferation. Overall, our results show that miR-20b-5p impedes PDGF-induced proliferation of fetal ASM cells through targeting STAT3. Our study suggests that miR-20b-5p may play an important role in airway remodeling during asthma and suggests that miR-20b-5p may serve as a potential therapeutic target for pediatric asthma. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

BRN2, a POUerful driver of melanoma phenotype switching and metastasis.

PubMed

Fane, Mitchell E; Chhabra, Yash; Smith, Aaron G; Sturm, Richard A

2018-05-21

The POU domain family of transcription factors play a central role in embryogenesis and are highly expressed in neural crest cells and the developing brain. BRN2 is a class III POU domain protein that is a key mediator of neuroendocrine and melanocytic development and differentiation. While BRN2 is a central regulator in numerous developmental programs, it has also emerged as a major player in the biology of tumourigenesis. In melanoma, BRN2 has been implicated as one of the master regulators of the acquisition of invasive behavior within the phenotype-switching model of progression. As a mediator of melanoma cell phenotype-switching it co-ordinates the transition to a de-differentiated, slow cycling and highly motile cell type. Its inverse expression relationship with MITF is believed to mediate tumour progression and metastasis within this model. Recent evidence has now outlined a potential epigenetic switching mechanism in melanoma cells driven by BRN2 expression that induces melanoma cell invasion. We summarise the role of BRN2 in tumour cell dissemination and metastasis in melanoma, while also examining it as a potential metastatic regulator in other tumour models. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

The highly conserved MraZ protein is a transcriptional regulator in Escherichia coli

DOE Office of Scientific and Technical Information (OSTI.GOV)

Eraso, Jesus M.; Markillie, Lye Meng; Mitchell, Hugh D.

2014-05-05

The mraZ and mraW genes are highly conserved in bacteria, both in sequence and location at the head of the division and cell wall (dcw) gene cluster. Although MraZ has structural similarity to the AbrB transition state regulator and the MazE antitoxin, and MraW is known to methylate ribosomal RNA, mraZ and mraW null mutants have no detectable growth phenotype in any species tested to date, hampering progress in understanding their physiological role. Here we show that overproduction of Escherichia coli MraZ perturbs cell division and the cell envelope, is more lethal at high levels or in minimal growth medium,more » and that MraW antagonizes these effects. MraZGFP localizes to the nucleoid, suggesting that it binds DNA. Indeed, purified MraZ directly binds a region upstream from its own promoter containing three direct repeats to regulate its own expression and that of downstream cell division and cell wall genes. MraZ-LacZ fusions are repressed by excess MraZ but not when DNA binding by MraZ is inhibited. RNAseq analysis indicates that MraZ is a global transcriptional regulator with numerous targets in addition to dcw genes. One of these targets, mioC, is directly bound by MraZ in a region with three direct repeats.« less

Concise Review: Plasma and Nuclear Membranes Convey Mechanical Information to Regulate Mesenchymal Stem Cell Lineage.

PubMed

Uzer, Gunes; Fuchs, Robyn K; Rubin, Janet; Thompson, William R

2016-06-01

Numerous factors including chemical, hormonal, spatial, and physical cues determine stem cell fate. While the regulation of stem cell differentiation by soluble factors is well-characterized, the role of mechanical force in the determination of lineage fate is just beginning to be understood. Investigation of the role of force on cell function has largely focused on "outside-in" signaling, initiated at the plasma membrane. When interfaced with the extracellular matrix, the cell uses integral membrane proteins, such as those found in focal adhesion complexes to translate force into biochemical signals. Akin to these outside-in connections, the internal cytoskeleton is physically linked to the nucleus, via proteins that span the nuclear membrane. Although structurally and biochemically distinct, these two forms of mechanical coupling influence stem cell lineage fate and, when disrupted, often lead to disease. Here we provide an overview of how mechanical coupling occurs at the plasma and nuclear membranes. We also discuss the role of force on stem cell differentiation, with focus on the biochemical signals generated at the cell membrane and the nucleus, and how those signals influence various diseases. While the interaction of stem cells with their physical environment and how they respond to force is complex, an understanding of the mechanical regulation of these cells is critical in the design of novel therapeutics to combat diseases associated with aging, cancer, and osteoporosis. Stem Cells 2016;34:1455-1463. © 2016 AlphaMed Press.

Deregulation of Apoptosis - Is it Still an Important Issue in Pathogenesis of Chronic Lymphocytic Leukemia?

PubMed

Podhorecka, Monika; Macheta, Arkadiusz; Bozko, Maria; Bozko, Andrzej; Malek, Nisar P; Bozko, Przemyslaw

2016-01-01

Chronic lymphocytic leukemia (CLL), a clonal expansion of B CD5+ cells, is the most common type of adult leukemia in western countries. The accumulation of neoplastic B-cells is primarily caused by prolonged life-span of these cells due to deregulation of apoptosis, and only marginally due to a higher proliferation rate. In spite of numerous reports characterizing particular mechanisms of B-CLL cell apoptosis, still relatively little is known about the complex regulation of this process. Therefore, more detailed research is required to understand the complicated mechanisms and regulatory processes of apoptosis in neoplastic B lymphocytes.

Dancing with Hormones: A Current Perspective of Nitrate Signaling and Regulation in Arabidopsis

PubMed Central

Guan, Peizhu

2017-01-01

In nature and agriculture, nitrate availability is a main environmental cue for plant growth, development and stress responses. Nitrate signaling and regulation are hence at the center of communications between plant intrinsic programs and the environment. It is also well known that endogenous phytohormones play numerous critical roles in integrating extrinsic cues and intrinsic responses, regulating and refining almost all aspects of plant growth, development and stress responses. Therefore, interaction between nitrate and phytohormones, such as auxins, cytokinins, abscisic acid, gibberellins, and ethylene, is prevalent. The growing evidence indicates that biosynthesis, de-conjugation, transport, and signaling of hormones are partly controlled by nitrate signaling. Recent advances with nitrate signaling and transcriptional regulation in Arabidopsis give rise to new paradigms. Given the comprehensive nitrate transport, sensing, signaling and regulations at the level of the cell and organism, nitrate itself is a local and long-distance signal molecule, conveying N status at the whole-plant level. A direct molecular link between nitrate signaling and cell cycle progression was revealed with TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR1-20 (TCP20) – NIN-LIKE PROTEIN 6/7 (NLP6/7) regulatory nexus. NLPs are key regulators of nitrogen responses in plants. TCPs function as the main regulators of plant morphology and architecture, with the emerging role as integrators of plant developmental responses to the environment. By analogy with auxin being proposed as a plant morphogen, nitrate may be an environmental morphogen. The morphogen-gradient-dependent and cell-autonomous mechanisms of nitrate signaling and regulation are an integral part of cell growth and cell identification. This is especially true in root meristem growth that is regulated by intertwined nitrate, phytohormones, and glucose-TOR signaling pathways. Furthermore, the nitrate transcriptional hierarchy is emerging. Nitrate regulators in primary nitrate signaling can individually and combinatorially control downstream transcriptional networks and hormonal pathways for signal propagation and amplification. Under the new paradigms, nitrate-induced hormone metabolism and signaling deserve fresh examination. The close interplay and convergent regulation of nitrate and hormonal signaling at morphological, physiological, and molecular levels have significant effects on important agronomic traits, especially nutrient-dependent adaptive root system growth and architecture. PMID:29033968

Diverse structures, functions and uses of FK506 binding proteins.

PubMed

Bonner, Julia Maeve; Boulianne, Gabrielle L

2017-10-01

FK506 (Tacrolimus), isolated from Streptomyces tsukubaenis is a powerful immunosuppressant shown to inhibit T cell activation. FK506 mediated immunosuppression requires the formation of a complex between FK506, a FK506 binding protein (FKBP) and calcineurin. Numerous FKBPs have been identified in a wide range of species, from single celled organisms to humans. FKBPs show peptidylprolyl cis/trans isomerase (PPIase) activity and have been shown to affect a wide range of cellular processes including protein folding, receptor signaling and apoptosis. FKBPs also affect numerous biological functions in addition to immunosuppression including regulation of cardiac function, neuronal function and development and have been implicated in several diseases including cardiac disease, cancer and neurodegenerative diseases such as Alzheimer's disease. More recently, FKBPs have proven useful as molecular tools for studying protein interactions, localization and functions. This review provides an overview of the current state of knowledge of FKBPs and their numerous biological functions and uses. Copyright © 2017 Elsevier Inc. All rights reserved.

Mentha piperita as a pivotal neuro-protective agent against gamma irradiation induced DNA fragmentation and apoptosis : Mentha extract as a neuroprotective against gamma irradiation.

PubMed

Hassan, Hanaa A; Hafez, Hani S; Goda, Mona S

2013-01-01

Ionizing radiation is classified as a potent carcinogen, and its injury to living cells, in particular to DNA, is due to oxidative stress enhancing apoptotic cell death. Our present study aimed to characterize and semi-quantify the radiation-induced apoptosis in CNS and the activity of Mentha extracts as neuron-protective agent. Our results through flow cytometry exhibited the significant disturbance and arrest in cell cycle in % of M1: SubG1 phase, M2: G0/1 phase of diploid cycle, M3: S phase and M4: G2/M phase of cell cycle in brain tissue (p < 0.05). Significant increase in % of apoptosis and P53 protein expression as apoptotic biomarkers were coincided with significant decrease in Bcl(2) as an anti-apoptotic marker. The biochemical analysis recorded a significant decrease in the levels of reduced glutathione, superoxide dismutase, deoxyribonucleic acid (DNA) and ribonucleic acid contents. Moreover, numerous histopathological alterations were detected in brain tissues of gamma irradiated mice such as signs of chromatolysis in pyramidal cells of cortex, nuclear vacuolation, numerous apoptotic cell, and neural degeneration. On the other hand, gamma irradiated mice pretreated with Mentha extract showed largely an improvement in all the above tested parameters through a homeostatic state for the content of brain apoptosis and stabilization of DNA cycle with a distinct improvement in cell cycle analysis and antioxidant defense system. Furthermore, the aforementioned effects of Mentha extracts through down-regulation of P53 expression and up-regulation of Bcl(2) domain protected brain structure from extensive damage. Therefore, Mentha extract seems to have a significant role to ameliorate the neuronal injury induced by gamma irradiation.

Uncovering a key to the process of metastasis in human cancers: a review of critical regulators of anoikis.

PubMed

Tan, Kevin; Goldstein, David; Crowe, Philip; Yang, Jia-Lin

2013-11-01

Anoikis ('homelessness' in Greek) is a form of apoptosis following the detachment of cells from the appropriate extracellular matrix (Chiarugi and Giannoni in Biochem Pharmacol 76:1352-1364, 2008). Resistance to anoikis is a critical mediator of metastasis in cancer by enabling cancer cells to survive during invasion and transport in the blood and lymph. Numerous regulators and mechanisms of anoikis in human cancer have been proposed to date. Consequently, the identification of key regulators of anoikis that can be targeted to at least partially restore anoikis sensitivity in cancer cells is important in the development of therapies to treat metastatic cancer. A literature search focusing on the regulators of anoikis in human cancer was performed on the Medline, Embase and Scopus databases. Mcl-1, Cav-1, Bcl-(xL), cFLIP, 14-3-3ζ and Bit1 appear to regulate anoikis in human cancer by participating in the intrinsic apoptotic pathway, extrinsic apoptotic pathway or caspase-independent pathways. Mcl-1, Cav-1, Bcl-(xL), cFLIP and 14-3-3ζ are suppressors of anoikis, and their upregulation confers anoikis resistance to cancer cells. Bit1 is a promoter of anoikis and is downregulated to confer anoikis resistance in metastatic cancer. Anoikis is a complex process involving the crosstalk between different signalling pathways. The dysregulated expression of key regulators of anoikis that participate in these signalling pathways promotes anoikis resistance in human cancer. These regulators of anoikis might therefore be the targets for developing therapies to overcome anoikis resistance in metastatic cancer.

The merged basins of signal transduction pathways in spatiotemporal cell biology.

PubMed

Hou, Yingchun; Hou, Yang; He, Siyu; Ma, Caixia; Sun, Mengyao; He, Huimin; Gao, Ning

2014-03-01

Numerous evidences have indicated that a signal system is composed by signal pathways, each pathway is composed by sub-pathways, and the sub-pathway is composed by the original signal terminals initiated with a protein/gene. We infer the terminal signals merged signal transduction system as "signal basin". In this article, we discussed the composition and regulation of signal basins, and the relationship between the signal basin control and triple W of spatiotemporal cell biology. Finally, we evaluated the importance of the systemic regulation to gene expression by signal basins under triple W. We hope our discussion will be the beginning to cause the attention for this area from the scientists of life science. © 2013 Wiley Periodicals, Inc.

Aquaporin-2 Regulation in Health and Disease

PubMed Central

Radin, M. Judith; Yu, Ming-Jiun; Stoedkilde, Lene; Miller, R. Lance; Hoffert, Jason D.; Frokiaer, Jorgen; Pisitkun, Trairak; Knepper, Mark A.

2012-01-01

Aquaporin-2 (AQP2), the vasopressin-regulated water channel of the renal collecting duct, is dysregulated in numerous water balance disorders in humans and animals including those associated with polyuria (e.g. urinary tract obstruction, hypokalemia, inflammation, and lithium toxicity) and dilutional hyponatremia (e.g. SIADH). Normal regulation of AQP2 by vasopressin involves two independent regulatory mechanisms: 1) short-term regulation of AQP2 trafficking to and from the apical plasma membrane, and 2) long term regulation of the total abundance of the AQP2 protein in the cells. Most water balance disorders are the result of dysregulation of processes that regulate the total abundance of AQP2 in collecting duct cells. In general, the level of AQP2 in a collecting duct cell is determined by a balance between production via translation of AQP2 mRNA and removal via degradation and/or secretion into the urine in exosomes. AQP2 abundance increases in response to vasopressin chiefly due to increased translation subsequent to increases in AQP2 mRNA. Vasopressin-mediated regulation AQP2 gene transcription is poorly understood, although several transcription factor binding elements in the 5’ flanking region of the AQP2 gene have been identified and candidate transcription factors corresponding to these element have been discovered in proteomics studies. Here we review progress in this area and discuss elements of vasopressin signaling in the collecting duct that may impinge on regulation of AQP2 in health and in the context of examples of polyuric diseases. PMID:23130944

Neutral endopeptidase regulates neurogenic inflammatory responses induced by stimulation of human oral keratinocytes with bacterial lipopolysaccharide and nicotine.

PubMed

Nakata, Motoki; Awano, Shuji; Kinoshita, Naomasa; Yoshida, Akihiro; Ansai, Toshihiro

2013-10-01

Neutral endopeptidase (NEP) is present on various epithelial cells and inactivates numerous physiologically active peptides. Neutral endopeptidase may regulate proinflammatory signals in oral mucosal epithelium. However, the function of NEP in oral mucosal epithelium is unknown. The present study investigated the action of NEP upon proinflammatory signals on human oral keratinocytes and the influence of endothelin-converting enzyme (ECE)-1, an enzyme similar to NEP, on the functions of NEP. Oral keratinocytes were cultured in medium containing inflammatory inducers [lipopolysaccharide (LPS) and nicotine], NEP inhibitors, and ECE-1/NEP inhibitors, either alone or in combination. The concentrations of substance P (SP) and interleukin-1β (IL-1β) were measured in the supernatant. Additionally, the concentrations of SP and IL-1β were measured in the supernatant of cells incubated with LPS or nicotine after transfection with NEP small interfering RNA (siRNA). The concentrations of SP and IL-1β were significantly increased in cells incubated with NEP inhibitors and, to a lesser extent, in cells incubated with ECE-1/NEP inhibitors, compared with controls (cells incubated with LPS or nicotine alone). The concentrations of SP and IL-1β in cells transfected with NEP siRNA were significantly augmented compared with controls. In conclusion, the present study demonstrated that NEP down-regulated the levels of SP and IL-1β produced from human oral keratinocytes, although ECE-1 may be partly related to the down-regulation. © 2013 Eur J Oral Sci.

Epigenetic regulation of miRNA-cancer stem cells nexus by nutraceuticals.

PubMed

Ahmad, Aamir; Li, Yiwei; Bao, Bin; Kong, Dejuan; Sarkar, Fazlul H

2014-01-01

Nutraceuticals, the bioactive food components represented by many naturally occurring dietary compounds, have been investigated for a few decades for their numerous beneficial effects, including their anticancer properties. The initial interest in the cancer-preventing/therapeutic ability of these agents was based on their ability to affect multiple signaling pathways that are deregulated in cancer cells. With a shift in the focus of cancer research to the emerging areas such as epigenetic regulation, microRNAs (miRNAs) and the cancer stem cells (CSCs), nutraceuticals initially appeared out of place. However, research investigations over the last several years have slowly but firmly presented evidence that supports a relevance of these agents in modern day research. While nutraceuticals are increasingly being realized to alter miRNA/CSCs expression and function, the molecular mechanism(s) are not very clearly understood. Epigenetic regulation is one mechanism by which these agents exert their anticancer effects. In this focused mini review, we summarize our current understanding of epigenetic regulation of miRNAs and CSCs by nutraceuticals. We discuss both direct and indirect evidences that support such an activity of these compounds. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Deficiency of the Arabidopsis Helicase RTEL1 Triggers a SOG1-Dependent Replication Checkpoint in Response to DNA Cross-Links

PubMed Central

Hu, Zhubing; Cools, Toon; Kalhorzadeh, Pooneh; Heyman, Jefri; De Veylder, Lieven

2015-01-01

To maintain genome integrity, DNA replication is executed and regulated by a complex molecular network of numerous proteins, including helicases and cell cycle checkpoint regulators. Through a systematic screening for putative replication mutants, we identified an Arabidopsis thaliana homolog of human Regulator of Telomere Length 1 (RTEL1), which functions in DNA replication, DNA repair, and recombination. RTEL1 deficiency retards plant growth, a phenotype including a prolonged S-phase duration and decreased cell proliferation. Genetic analysis revealed that rtel1 mutant plants show activated cell cycle checkpoints, specific sensitivity to DNA cross-linking agents, and increased homologous recombination, but a lack of progressive shortening of telomeres, indicating that RTEL1 functions have only been partially conserved between mammals and plants. Surprisingly, RTEL1 deficiency induces tolerance to the deoxynucleotide-depleting drug hydroxyurea, which could be mimicked by DNA cross-linking agents. This resistance does not rely on the essential replication checkpoint regulator WEE1 but could be blocked by a mutation in the SOG1 transcription factor. Taken together, our data indicate that RTEL1 is required for DNA replication and that its deficiency activates a SOG1-dependent replication checkpoint. PMID:25595823

Sodium 4-phenylbutyrate upregulates ENaC and sodium absorption in T84 cells.

PubMed

Iordache, Claudiu; Duszyk, Marek

2007-01-15

Butyrate and other short-chain fatty acids (SCFA), produced by colonic bacterial flora, affect numerous epithelial cell functions. To better understand how SCFA regulate ion transport, we investigated the effects of 4-phenylbutyrate (4-PBA) on Na(+) absorption in T84 cells. Under standard cell culture conditions, the short circuit current did not display any amiloride-sensitive Na(+) absorption and was wholly representative of Cl(-) secretion. However, when T84 cells were grown in the presence of 5 mM 4-PBA, a gradual appearance of amiloride-sensitive Na(+) channel (ENaC) activity was observed that reached a plateau after 24 h. Quantitative RT-PCR and Western blot studies of ENaC subunit expression indicated that 4-PBA stimulated alpha and gamma subunits. Trichostatin A, an inhibitor of histone deacetylase, mimicked the effects of 4-PBA, suggesting that 4-PBA affects ENaC expression by inhibiting deacetylases. 4-PBA had no effect on ENaC expression in airway epithelial cells indicating tissue-specific effect. We conclude that butyrate plays an important role in regulating colonic Na(+) absorption by increasing ENaC transcription and activity.

Initiation of DNA replication requires actin dynamics and formin activity.

PubMed

Parisis, Nikolaos; Krasinska, Liliana; Harker, Bethany; Urbach, Serge; Rossignol, Michel; Camasses, Alain; Dewar, James; Morin, Nathalie; Fisher, Daniel

2017-11-02

Nuclear actin regulates transcriptional programmes in a manner dependent on its levels and polymerisation state. This dynamics is determined by the balance of nucleocytoplasmic shuttling, formin- and redox-dependent filament polymerisation. Here, using Xenopus egg extracts and human somatic cells, we show that actin dynamics and formins are essential for DNA replication. In proliferating cells, formin inhibition abolishes nuclear transport and initiation of DNA replication, as well as general transcription. In replicating nuclei from transcriptionally silent Xenopus egg extracts, we identified numerous actin regulators, and disruption of actin dynamics abrogates nuclear transport, preventing NLS (nuclear localisation signal)-cargo release from RanGTP-importin complexes. Nuclear formin activity is further required to promote loading of cyclin-dependent kinase (CDK) and proliferating cell nuclear antigen (PCNA) onto chromatin, as well as initiation and elongation of DNA replication. Therefore, actin dynamics and formins control DNA replication by multiple direct and indirect mechanisms. © 2017 The Authors.

Mechanisms regulating enhanced HLA class II-mediated CD4+ T cell recognition of human B-cell lymphoma by resveratrol

PubMed Central

RADWAN, FAISAL F. Y.; ZHANG, LIXIA; HOSSAIN, AZIM; DOONAN, BENTLY P.; GOD, JASON; HAQUE, AZIZUL

2015-01-01

Malignant B-cells express measurable levels of HLA class II proteins, but often escape immune recognition by CD4+ T cells. Resveratrol (Resv) has been the focus of numerous investigations due to its potential chemopreventive and anti-cancer effects, but it has never been tested in the regulation of immune components in B-cell tumors. Here, we show for the first time that Resv treatment enhances HLA class II-mediated immune detection of B-cell lymphomas by altering immune components and class II presentation in tumor cells. Resv treatment induced an upregulation of both classical and non-classical HLA class II proteins (DR and DM) in B-lymphoma cells. Resv also altered endolysosomal cathepsins (Cat S, B and D) and a thiol reductase (GILT), increasing HLA class II-mediated antigen (Ag) processing in B-cell lymphomas and their subsequent recognition by CD4+ T cells. Mechanistic study demonstrated that Resv treatment activated the recycling class II pathway of Ag presentation through upregulation of Rab 4B protein expression in B-lymphoma cells. These findings suggest that HLA class II-mediated immune recognition of malignant B-cells can be improved by Resv treatment, thus encouraging its potential use in chemoimmunotherapy of B-cell lymphoma. PMID:21854084

Copper deficiency alters cell bioenergetics and induces mitochondrial fusion through up-regulation of MFN2 and OPA1 in erythropoietic cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bustos, Rodrigo I.; Jensen, Erik L.; Ruiz, Lina M.

2013-08-02

Highlights: •In copper deficiency, cell proliferation is not affected. In turn, cell differentiation is impaired. •Enlarged mitochondria are due to up-regulation of MNF2 and OPA1. •Mitochondria turn off respiratory chain and ROS production. •Energy metabolism switch from mitochondria to glycolysis. -- Abstract: Copper is essential in cell physiology, participating in numerous enzyme reactions. In mitochondria, copper is a cofactor for respiratory complex IV, the cytochrome c oxidase. Low copper content is associated with anemia and the appearance of enlarged mitochondria in erythropoietic cells. These findings suggest a connection between copper metabolism and bioenergetics, mitochondrial dynamics and erythropoiesis, which has notmore » been explored so far. Here, we describe that bathocuproine disulfonate-induced copper deficiency does not alter erythropoietic cell proliferation nor induce apoptosis. However it does impair erythroid differentiation, which is associated with a metabolic switch between the two main energy-generating pathways. That is, from mitochondrial function to glycolysis. Switching off mitochondria implies a reduction in oxygen consumption and ROS generation along with an increase in mitochondrial membrane potential. Mitochondrial fusion proteins MFN2 and OPA1 were up-regulated along with the ability of mitochondria to fuse. Morphometric analysis of mitochondria did not show changes in total mitochondrial biomass but rather bigger mitochondria because of increased fusion. Similar results were also obtained with human CD34+, which were induced to differentiate into red blood cells. In all, we have shown that adequate copper levels are important for maintaining proper mitochondrial function and for erythroid differentiation where the energy metabolic switch plus the up-regulation of fusion proteins define an adaptive response to copper deprivation to keep cells alive.« less

PDGF-regulated rab4-dependent recycling of alphavbeta3 integrin from early endosomes is necessary for cell adhesion and spreading.

PubMed

Roberts, M; Barry, S; Woods, A; van der Sluijs, P; Norman, J

2001-09-18

It has been postulated that the regulation of integrin vesicular traffic facilitates cell migration by internalizing integrins at the rear of the cell and transporting them forward within vesicles for exocytosis at the leading edge to form new contacts with the extracellular matrix. The rab family of GTPases control key targeting events in the endo/exocytic pathway; therefore, these GTPases may be involved in the regulation of cell-matrix contact assembly. The endo/exocytic cycle of alphavbeta3 and alpha5beta1 integrins was studied using mouse 3T3 fibroblast cell lines. In serum-starved cells, internalized integrins were transported through rab4-positive, early endosomes and arrived at the rab11-positive, perinuclear recycling compartment approximately 30 min after endocytosis. From the recycling compartment, integrins were recycled to the plasma membrane in a rab11-dependent fashion. Following treatment with PDGF, alphavbeta3 integrin, but not alpha5beta1, was rapidly recycled directly back to the plasma membrane from the early endosomes via a rab4-dependent mechanism without the involvement of rab11. This rapid recycling pathway directed alphavbeta3 to numerous small puncta distributed evenly across the dorsal surface of the cell, and the integrin only became localized into focal complexes at later times following PDGF addition. Interestingly, inhibition of PDGF-stimulated alphavbeta3 recycling using dominant-negative rab4 mutants compromised cell adhesion and spreading on vitronectin (a ligand for alphavbeta3), but adhesion to fibronectin (a ligand for alpha5beta1 and alphavbeta3) was unchanged. We propose that growth factor-regulated, rab4-dependent recycling of alphavbeta3 integrin from early endosomes to the plasma membrane is a critical upstream event in the assembly of cell-matrix contacts.

miR-142-3p is involved in CD25+ CD4 T cell proliferation by targeting the expression of glycoprotein A repetitions predominant.

PubMed

Zhou, Qihui; Haupt, Sonja; Prots, Iryna; Thümmler, Katja; Kremmer, Elisabeth; Lipsky, Peter E; Schulze-Koops, Hendrik; Skapenko, Alla

2013-06-15

Because of the numerous targets of microRNAs (miRNAs), functional dissection of specific miRNA/mRNA interactions is important to understand the complex miRNA regulatory mechanisms. Glycoprotein A repetitions predominant (GARP) is specifically expressed on regulatory CD25(+) CD4 T cells upon their activation. GARP has a long 3' untranslated region containing five highly conserved regions suggesting miRNA regulation of its expression. Although GARP is physiologically expressed on a cell subset characterized by stringent control of proliferation, amplification of the GARP gene has been found in many tumors characterized by uncontrolled proliferation. In this study, we investigated in detail miRNA regulation of GARP expression, in particular by miR-142-3p, and dissected the functional outcome of miR-142-3p/GARP mRNA interaction. We demonstrate that miR-142-3p binds directly to the 3' untranslated region of GARP and represses GARP protein expression by Argonaute 2-associated degradation of GARP mRNA. Functionally, miR-142-3p-mediated regulation of GARP is involved in the expansion of CD25(+) CD4 T cells in response to stimulation. The data indicate that miR-142-3p regulates GARP expression on CD25(+) CD4 T cells and, as a result, their expansion in response to activation. Our data provide novel insight into the molecular mechanisms controlling regulatory T cell expansion. They may also have implications for understanding tumor cell biology.

Anticancer activity and mediation of apoptosis in human HL-60 leukaemia cells by edible sea cucumber (Holothuria edulis) extract.

PubMed

Wijesinghe, W A J P; Jeon, You Jin; Ramasamy, Perumal; Wahid, Mohd Effendy A; Vairappan, Charles S

2013-08-15

Sea cucumbers have been a dietary delicacy and important ingredient in Asian traditional medicinal over many centuries. In this study, edible sea cucumber Holothuria edulis was evaluated for its in vitro anticancer potential. An aqueous fraction of the edible sea cucumber (ESC-AQ) has been shown to deliver a strong cytotoxic effect against the human HL-60 leukaemia cell line. An induction effect of apoptotic body formation in response to ESC-AQ treatment was confirmed in HL-60 cells stained with Hoechst 33342 and confirmed via flow cytometry analysis. The up regulation of Bax and caspase-3 protein expression was observed while the expression of Bcl-xL protein was down regulated in ESC-AQ treated HL-60 cells. Due to the profound anticancer activity, ESC-AQ appears to be an economically important biomass fraction that can be exploited in numerous industrial applications as a source of functional ingredients. Copyright © 2013 Elsevier Ltd. All rights reserved.

Cloning of the promoter region of a human gene, FOXL2, and its regulation by STAT3.

PubMed

Han, Yangyang; Wang, Tianxiao; Sun, Shudong; Zhai, Zhaohui; Tang, Shengjian

2017-09-01

Forkhead box L2 (FOXL2) is a transcription factor, which is involved in blepharophimosis, ptosis, and epicanthus in versus syndrome (BPES), premature ovarian failure (POF), as well as almost all stages of ovarian development and function. FOXL2 has various target genes, which are implicated in numerous processes, including sex determination, cell cycle regulation and apoptosis and stress response regulation in mammals. However, studies regarding the upstream regulation of FOXL2 are limited. In the present study, the promoter of FOXL2 was successfully cloned and registered in Gen Bank, and a dual luciferase reporter (DLR) analysis demonstrated that the luciferase activity was significantly induced by the promoter of FOXL2. Subsequently, bioinformatics analysis indicated that FOXL2 may be regulated by STAT3, and this was confirmed by a DLR analysis and western blotting, using STAT3 inhibitors. Further study using real‑time cellular analysis indicated that the viability of He La cells was markedly suppressed by STAT3 inhibitors. The present study demonstrated novel findings regarding the upstream regulation of FOXL2 expression and provide a new perspective for future studies in the field.

Roles of mineralocorticoid and glucocorticoid receptors in the regulation of progenitor proliferation in the adult hippocampus.

PubMed

Wong, Edmund Y H; Herbert, Joe

2005-08-01

New neurons are produced continually in the dentate gyrus of the hippocampus. Numerous factors modulate the rate of neuron production. One of the most important is the adrenal-derived corticoids. Raised levels of corticoids suppress proliferation of progenitor cells, while removal of corticoids by adrenalectomy reverses this. The exact mechanisms by which corticoids mediate such regulation are unknown, but corticoids are believed to act through the receptors for mineralocorticoids (MR) and glucocorticoids (GR). Previous reports regarding the roles of these receptors in regulating cell proliferation came to contrasting conclusions. Here we use both agonists and antagonists to these receptors in adult male rats to investigate and clarify their roles. Blockade of MR with spironolactone in adrenalectomised male rats implanted with a corticosterone pellet to reproduce basal levels enhanced proliferation, whereas treatment with the GR antagonist mifepristone had no effect. However, mifepristone reversed the suppressive effect of additional corticosterone in intact rats. Both aldosterone and RU362, agonists of MR and GR, respectively, reduced proliferation in adrenalectomised rats, and combined treatment with both agonists had an additional suppressive action. These results clearly show that occupancies of both receptors act in the same direction on progenitor proliferation. The existence of two receptors with different affinities for corticoids may ensure that proliferation of progenitor cells in the adult dentate gyrus is regulated across the range of adrenal corticoid activity, including both basal and stressful contexts. Although a small proportion of newly formed cells may express GR and MR, corticosterone probably regulates proliferation indirectly through other local cells.

Epithelial heparan sulfate regulates Sonic Hedgehog signaling in lung development.

PubMed

He, Hua; Huang, Meina; Sun, Shenfei; Wu, Yihui; Lin, Xinhua

2017-08-01

The tree-like structure of the mammalian lung is generated from branching morphogenesis, a reiterative process that is precisely regulated by numerous factors. How the cell surface and extra cellular matrix (ECM) molecules regulate this process is still poorly understood. Herein, we show that epithelial deletion of Heparan Sulfate (HS) synthetase Ext1 resulted in expanded branching tips and reduced branching number, associated with several mesenchymal developmental defects. We further demonstrate an expanded Fgf10 expression and increased FGF signaling activity in Ext1 mutant lungs, suggesting a cell non-autonomous mechanism. Consistent with this, we observed reduced levels of SHH signaling which is responsible for suppressing Fgf10 expression. Moreover, reactivating SHH signaling in mutant lungs rescued the tip dilation phenotype and attenuated FGF signaling. Importantly, the reduced SHH signaling activity did not appear to be caused by decreased Shh expression or protein stability; instead, biologically active form of SHH proteins were reduced in both the Ext1 mutant epithelium and surrounding wild type mesenchymal cells. Together, our study highlights the epithelial HS as a key player for dictating SHH signaling critical for lung morphogenesis.

Evaluation of MicroRNAs Regulating Anoikis Pathways and Its Therapeutic Potential

PubMed Central

Malagobadan, Sharan

2015-01-01

Dysregulation of microRNAs (miRNAs) has been implicated in almost every known survival mechanisms utilized by cancer cells. One of such mechanisms, anoikis resistance, plays a pivotal role in enabling metastasis by allowing cancer cells to circumvent cell death induced by lack of attachment. Understanding how miRNAs regulate the various anoikis pathways has become the research question of increasing number of studies published in the past years. Through these studies, a growing list of miRNAs has been identified to be important players in promoting either anoikis or resistance to anoikis. In this review, we will be focusing on these miRNAs and how the findings from those studies can contribute to novel therapeutic strategies against cancer progression. We will be examining miRNAs that have been found to promote anoikis sensitivity in numerous cancer types followed by miRNAs that inhibit anoikis. In addition, we will also be taking a look at major signaling pathways involved in the action of the each of these miRNAs to gain a better understanding on how miRNAs regulate anoikis. PMID:26587543

Systems Modeling of Molecular Mechanisms Controlling Cytokine-driven CD4+ T Cell Differentiation and Phenotype Plasticity

PubMed Central

Carbo, Adria; Hontecillas, Raquel; Kronsteiner, Barbara; Viladomiu, Monica; Pedragosa, Mireia; Lu, Pinyi; Philipson, Casandra W.; Hoops, Stefan; Marathe, Madhav; Eubank, Stephen; Bisset, Keith; Wendelsdorf, Katherine; Jarrah, Abdul; Mei, Yongguo; Bassaganya-Riera, Josep

2013-01-01

Differentiation of CD4+ T cells into effector or regulatory phenotypes is tightly controlled by the cytokine milieu, complex intracellular signaling networks and numerous transcriptional regulators. We combined experimental approaches and computational modeling to investigate the mechanisms controlling differentiation and plasticity of CD4+ T cells in the gut of mice. Our computational model encompasses the major intracellular pathways involved in CD4+ T cell differentiation into T helper 1 (Th1), Th2, Th17 and induced regulatory T cells (iTreg). Our modeling efforts predicted a critical role for peroxisome proliferator-activated receptor gamma (PPARγ) in modulating plasticity between Th17 and iTreg cells. PPARγ regulates differentiation, activation and cytokine production, thereby controlling the induction of effector and regulatory responses, and is a promising therapeutic target for dysregulated immune responses and inflammation. Our modeling efforts predict that following PPARγ activation, Th17 cells undergo phenotype switch and become iTreg cells. This prediction was validated by results of adoptive transfer studies showing an increase of colonic iTreg and a decrease of Th17 cells in the gut mucosa of mice with colitis following pharmacological activation of PPARγ. Deletion of PPARγ in CD4+ T cells impaired mucosal iTreg and enhanced colitogenic Th17 responses in mice with CD4+ T cell-induced colitis. Thus, for the first time we provide novel molecular evidence in vivo demonstrating that PPARγ in addition to regulating CD4+ T cell differentiation also plays a major role controlling Th17 and iTreg plasticity in the gut mucosa. PMID:23592971

Discovery of Transcription Factors Novel to Mouse Cerebellar Granule Cell Development Through Laser-Capture Microdissection.

PubMed

Zhang, Peter G Y; Yeung, Joanna; Gupta, Ishita; Ramirez, Miguel; Ha, Thomas; Swanson, Douglas J; Nagao-Sato, Sayaka; Itoh, Masayoshi; Kawaji, Hideya; Lassmann, Timo; Daub, Carsten O; Arner, Erik; de Hoon, Michiel; Carninci, Piero; Forrest, Alistair R R; Hayashizaki, Yoshihide; Goldowitz, Dan

2018-06-01

Laser-capture microdissection was used to isolate external germinal layer tissue from three developmental periods of mouse cerebellar development: embryonic days 13, 15, and 18. The cerebellar granule cell-enriched mRNA library was generated with next-generation sequencing using the Helicos technology. Our objective was to discover transcriptional regulators that could be important for the development of cerebellar granule cells-the most numerous neuron in the central nervous system. Through differential expression analysis, we have identified 82 differentially expressed transcription factors (TFs) from a total of 1311 differentially expressed genes. In addition, with TF-binding sequence analysis, we have identified 46 TF candidates that could be key regulators responsible for the variation in the granule cell transcriptome between developmental stages. Altogether, we identified 125 potential TFs (82 from differential expression analysis, 46 from motif analysis with 3 overlaps in the two sets). From this gene set, 37 TFs are considered novel due to the lack of previous knowledge about their roles in cerebellar development. The results from transcriptome-wide analyses were validated with existing online databases, qRT-PCR, and in situ hybridization. This study provides an initial insight into the TFs of cerebellar granule cells that might be important for development and provide valuable information for further functional studies on these transcriptional regulators.

Identification of Wnt Pathway Target Genes Regulating the Division and Differentiation of Larval Seam Cells and Vulval Precursor Cells in Caenorhabditis elegans.

PubMed

Gorrepati, Lakshmi; Krause, Michael W; Chen, Weiping; Brodigan, Thomas M; Correa-Mendez, Margarita; Eisenmann, David M

2015-06-05

The evolutionarily conserved Wnt/β-catenin signaling pathway plays a fundamental role during metazoan development, regulating numerous processes including cell fate specification, cell migration, and stem cell renewal. Wnt ligand binding leads to stabilization of the transcriptional effector β-catenin and upregulation of target gene expression to mediate a cellular response. During larval development of the nematode Caenorhabditis elegans, Wnt/β-catenin pathways act in fate specification of two hypodermal cell types, the ventral vulval precursor cells (VPCs) and the lateral seam cells. Because little is known about targets of the Wnt signaling pathways acting during larval VPC and seam cell differentiation, we sought to identify genes regulated by Wnt signaling in these two hypodermal cell types. We conditionally activated Wnt signaling in larval animals and performed cell type-specific "mRNA tagging" to enrich for VPC and seam cell-specific mRNAs, and then used microarray analysis to examine gene expression compared to control animals. Two hundred thirty-nine genes activated in response to Wnt signaling were identified, and we characterized 50 genes further. The majority of these genes are expressed in seam and/or vulval lineages during normal development, and reduction of function for nine genes caused defects in the proper division, fate specification, fate execution, or differentiation of seam cells and vulval cells. Therefore, the combination of these techniques was successful at identifying potential cell type-specific Wnt pathway target genes from a small number of cells and at increasing our knowledge of the specification and behavior of these C. elegans larval hypodermal cells. Copyright © 2015 Gorrepati et al.

The Formin Diaphanous Regulates Myoblast Fusion through Actin Polymerization and Arp2/3 Regulation

PubMed Central

Deng, Su; Bothe, Ingo; Baylies, Mary K.

2015-01-01

The formation of multinucleated muscle cells through cell-cell fusion is a conserved process from fruit flies to humans. Numerous studies have shown the importance of Arp2/3, its regulators, and branched actin for the formation of an actin structure, the F-actin focus, at the fusion site. This F-actin focus forms the core of an invasive podosome-like structure that is required for myoblast fusion. In this study, we find that the formin Diaphanous (Dia), which nucleates and facilitates the elongation of actin filaments, is essential for Drosophila myoblast fusion. Following cell recognition and adhesion, Dia is enriched at the myoblast fusion site, concomitant with, and having the same dynamics as, the F-actin focus. Through analysis of Dia loss-of-function conditions using mutant alleles but particularly a dominant negative Dia transgene, we demonstrate that reduction in Dia activity in myoblasts leads to a fusion block. Significantly, no actin focus is detected, and neither branched actin regulators, SCAR or WASp, accumulate at the fusion site when Dia levels are reduced. Expression of constitutively active Dia also causes a fusion block that is associated with an increase in highly dynamic filopodia, altered actin turnover rates and F-actin distribution, and mislocalization of SCAR and WASp at the fusion site. Together our data indicate that Dia plays two roles during invasive podosome formation at the fusion site: it dictates the level of linear F-actin polymerization, and it is required for appropriate branched actin polymerization via localization of SCAR and WASp. These studies provide new insight to the mechanisms of cell-cell fusion, the relationship between different regulators of actin polymerization, and invasive podosome formation that occurs in normal development and in disease. PMID:26295716

miR-1271 promotes non-small-cell lung cancer cell proliferation and invasion via targeting HOXA5

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Yongfang; Xu, Lianhong; Jiang, Lixin, E-mail: jianglx66766@163.com

2015-03-13

MicroRNAs (miRNAs) are short, non-coding RNAs (∼22 nt) that play important roles in the pathogenesis of human diseases by negatively regulating numerous target genes at posttranscriptional level. However, the role of microRNAs in lung cancer, particularly non-small-cell lung cancer (NSCLC), has remained elusive. In this study, two microRNAs, miR-1271 and miR-628, and their predicted target genes were identified differentially expressed in NSCLC by analyzing the miRNA and mRNA expression data from NSCLC tissues and their matching normal controls. miR-1271 and its target gene HOXA5 were selected for further investigation. CCK-8 proliferation assay showed that the cell proliferation was promoted by miR-1271more » in NSCLC cells, while miR-1271 inhibitor could significantly inhibited the proliferation of NSCLC cells. Interestingly, migration and invasion assay indicated that overexpression of miR-1271 could significantly promoted the migration and invasion of NSCLC cells, whereas miR-1271 inhibitor could inhibited both cell migration and invasion of NSCLC cells. Western blot showed that miR-1271 suppressed the protein level of HOXA5, and luciferase assays confirmed that miR-1271 directly bound to the 3'untranslated region of HOXA5. This study indicated indicate that miR-1271 regulates NSCLC cell proliferation and invasion, via the down-regulation of HOXA5. Thus, miR-1271 may represent a potential therapeutic target for NSCLC intervention. - Highlights: • Overexpression of miR-1271 promoted proliferation and invasion of NSCLC cells. • miR-1271 inhibitor inhibited the proliferation and invasion of NSCLC cells. • miR-1271 targets 3′ UTR of HOXA5 in NSCLC cells. • miR-1271 negatively regulates HOXA5 in NSCLC cells.« less

Multi-Faceted Functions of Secretory IgA at Mucosal Surfaces

PubMed Central

Corthésy, Blaise

2013-01-01

Secretory IgA (SIgA) plays an important role in the protection and homeostatic regulation of intestinal, respiratory, and urogenital mucosal epithelia separating the outside environment from the inside of the body. This primary function of SIgA is referred to as immune exclusion, a process that limits the access of numerous microorganisms and mucosal antigens to these thin and vulnerable mucosal barriers. SIgA has been shown to be involved in avoiding opportunistic pathogens to enter and disseminate in the systemic compartment, as well as tightly controlling the necessary symbiotic relationship existing between commensals and the host. Clearance by peristalsis appears thus as one of the numerous mechanisms whereby SIgA fulfills its function at mucosal surfaces. Sampling of antigen-SIgA complexes by microfold (M) cells, intimate contact occurring with Peyer’s patch dendritic cells (DC), down-regulation of inflammatory processes, modulation of epithelial, and DC responsiveness are some of the recently identified processes to which the contribution of SIgA has been underscored. This review aims at presenting, with emphasis at the biochemical level, how the molecular complexity of SIgA can serve these multiple and non-redundant modes of action. PMID:23874333

Homogeneous Time-resolved Förster Resonance Energy Transfer-based Assay for Detection of Insulin Secretion.

PubMed

Aslanoglou, Despoina; George, Emily W; Freyberg, Zachary

2018-05-10

The detection of insulin secretion is critical for elucidating mechanisms of regulated secretion as well as in studies of metabolism. Though numerous insulin assays have existed for decades, the recent advent of homogeneous time-resolved Förster Resonance Energy Transfer (HTRF) technology has significantly simplified these measurements. This is a rapid, cost-effective, reproducible, and robust optical assay reliant upon antibodies conjugated to bright fluorophores with long lasting emission which facilitates time-resolved Förster Resonance Energy Transfer. Moreover, HTRF insulin detection is amenable for the development of high-throughput screening assays. Here we use HTRF to detect insulin secretion in INS-1E cells, a rat insulinoma-derived cell line. This allows us to estimate basal levels of insulin and their changes in response to glucose stimulation. In addition, we use this insulin detection system to confirm the role of dopamine as a negative regulator of glucose-stimulated insulin secretion (GSIS). In a similar manner, other dopamine D2-like receptor agonists, quinpirole, and bromocriptine, reduce GSIS in a concentration-dependent manner. Our results highlight the utility of the HTRF insulin assay format in determining the role of numerous drugs in GSIS and their pharmacological profiles.

Identification of Wnt Pathway Target Genes Regulating the Division and Differentiation of Larval Seam Cells and Vulval Precursor Cells in Caenorhabditis elegans

PubMed Central

Gorrepati, Lakshmi; Krause, Michael W.; Chen, Weiping; Brodigan, Thomas M.; Correa-Mendez, Margarita; Eisenmann, David M.

2015-01-01

The evolutionarily conserved Wnt/β-catenin signaling pathway plays a fundamental role during metazoan development, regulating numerous processes including cell fate specification, cell migration, and stem cell renewal. Wnt ligand binding leads to stabilization of the transcriptional effector β-catenin and upregulation of target gene expression to mediate a cellular response. During larval development of the nematode Caenorhabditis elegans, Wnt/β-catenin pathways act in fate specification of two hypodermal cell types, the ventral vulval precursor cells (VPCs) and the lateral seam cells. Because little is known about targets of the Wnt signaling pathways acting during larval VPC and seam cell differentiation, we sought to identify genes regulated by Wnt signaling in these two hypodermal cell types. We conditionally activated Wnt signaling in larval animals and performed cell type–specific "mRNA tagging" to enrich for VPC and seam cell–specific mRNAs, and then used microarray analysis to examine gene expression compared to control animals. Two hundred thirty-nine genes activated in response to Wnt signaling were identified, and we characterized 50 genes further. The majority of these genes are expressed in seam and/or vulval lineages during normal development, and reduction of function for nine genes caused defects in the proper division, fate specification, fate execution, or differentiation of seam cells and vulval cells. Therefore, the combination of these techniques was successful at identifying potential cell type–specific Wnt pathway target genes from a small number of cells and at increasing our knowledge of the specification and behavior of these C. elegans larval hypodermal cells. PMID:26048561

Natural Compounds As Modulators of Non-apoptotic Cell Death in Cancer Cells

PubMed Central

Guamán-Ortiz, Luis Miguel; Orellana, Maria Isabel Ramirez; Ratovitski, Edward A.

2017-01-01

Cell death is an innate capability of cells to be removed from microenvironment, if and when they are damaged by multiple stresses. Cell death is often regulated by multiple molecular pathways and mechanism, including apoptosis, autophagy, and necroptosis. The molecular network underlying these processes is often intertwined and one pathway can dynamically shift to another one acquiring certain protein components, in particular upon treatment with various drugs. The strategy to treat human cancer ultimately relies on the ability of anticancer therapeutics to induce tumor-specific cell death, while leaving normal adjacent cells undamaged. However, tumor cells often develop the resistance to the drug-induced cell death, thus representing a great challenge for the anticancer approaches. Numerous compounds originated from the natural sources and biopharmaceutical industries are applied today in clinics showing advantageous results. However, some exhibit serious toxic side effects. Thus, novel effective therapeutic approaches in treating cancers are continued to be developed. Natural compounds with anticancer activity have gained a great interest among researchers and clinicians alike since they have shown more favorable safety and efficacy then the synthetic marketed drugs. Numerous studies in vitro and in vivo have found that several natural compounds display promising anticancer potentials. This review underlines certain information regarding the role of natural compounds from plants, microorganisms and sea life forms, which are able to induce non-apoptotic cell death in tumor cells, namely autophagy and necroptosis. PMID:28367073

MiR-145 regulates PAK4 via the MAPK pathway and exhibits an antitumor effect in human colon cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Zhigang; Zhang, Xiaoping; Yang, Zhili

2012-10-26

Highlights: Black-Right-Pointing-Pointer MiR-145 targets a putative binding site in the 3 Prime UTR of PAK4. Black-Right-Pointing-Pointer MiR-145 played an important role in inhibiting cell growth by directly targeting PAK4. Black-Right-Pointing-Pointer MiR-145 may function as tumor suppressors. -- Abstract: MicroRNAs (miRNAs) are regulators of numerous cellular events; accumulating evidence indicates that miRNAs play a key role in a wide range of biological functions, such as cellular proliferation, differentiation, and apoptosis in cancer. Down-regulated expression of miR-145 has been reported in colon cancer tissues and cell lines. The molecular mechanisms underlying miR-145 and the regulation of colon carcinogenesis remain unclear. In thismore » study, we investigated the levels of miR-145 in human colon cancer cells using qRT-PCR and found markedly decreased levels compared to normal epithelial cells. We identified PAK4 as a novel target of miR-145 using informatics screening. Additionally, we demonstrated that miR-145 targets a putative binding site in the 3 Prime UTR of PAK4 and that its abundance is inversely associated with miR-145 expression in colon cancer cells; we confirmed this relationship using the luciferase reporter assay. Furthermore, restoration of miR-145 by mimics in SW620 cells significantly attenuated cell growth in vitro, in accordance with the inhibitory effects induced by siRNA mediated knockdown of PAK4. Taken together, these findings demonstrate that miR-145 downregulates P-ERK expression by targeting PAK4 and leads to inhibition of tumor growth.« less

Heterologous expression of anti-apoptotic human 14-3-3β/α enhances iron-mediated programmed cell death in yeast

PubMed Central

Eid, Rawan; Zhou, David R.; Arab, Nagla T. T.; Boucher, Eric; Young, Paul G.; Mandato, Craig A.

2017-01-01

The induction of Programmed Cell Death (PCD) requires the activation of complex responses involving the interplay of a variety of different cellular proteins, pathways, and processes. Uncovering the mechanisms regulating PCD requires an understanding of the different processes that both positively and negatively regulate cell death. Here we have examined the response of normal as well as PCD resistant yeast cells to different PCD inducing stresses. As expected cells expressing the pro-survival human 14-3-3β/α sequence show increased resistance to numerous stresses including copper and rapamycin. In contrast, other stresses including iron were more lethal in PCD resistant 14-3-3β/α expressing cells. The increased sensitivity to PCD was not iron and 14-3-3β/α specific since it was also observed with other stresses (hydroxyurea and zinc) and other pro-survival sequences (human TC-1 and H-ferritin). Although microscopical examination revealed little differences in morphology with iron or copper stresses, cells undergoing PCD in response to high levels of prolonged copper treatment were reduced in size. This supports the interaction some forms of PCD have with the mechanisms regulating cell growth. Analysis of iron-mediated effects in yeast mutant strains lacking key regulators suggests that a functional vacuole is required to mediate the synergistic effects of iron and 14-3-3β/α on yeast PCD. Finally, mild sub-lethal levels of copper were found to attenuate the observed inhibitory effects of iron. Taken together, we propose a model in which a subset of stresses like iron induces a complex process that requires the cross-talk of two different PCD inducing pathways. PMID:28854230

Cell therapy worldwide: an incipient revolution.

PubMed

Rao, Mahendra; Mason, Chris; Solomon, Susan

2015-01-01

The regenerative medicine field is large, diverse and active worldwide. A variety of different organizational and product models have been successful, and pioneering entrepreneurs have shown both what can work and, critically, what does not. Evolving regulations, novel funding mechanisms combined with new technological breakthroughs are keeping the field in a state of flux. The field struggles to cope with the lack of infrastructure and investment, it nevertheless has evolved from its roots in human stem cell therapy and tissue and organ transplants to a field composed of a variety of products from multiple cell sources with approval for use in numerous countries. Currently, tens of thousands of patients have been treated with some kind of cell therapy.

Pro‐migratory and TGF‐β‐activating functions of αvβ6 integrin in pancreatic cancer are differentially regulated via an Eps8‐dependent GTPase switch

PubMed Central

Tod, Jo; Hanley, Christopher J; Morgan, Mark R; Rucka, Marta; Mellows, Toby; Lopez, Maria‐Antoinette; Kiely, Philip; Moutasim, Karwan A; Frampton, Steven J; Sabnis, Durgagauri; Fine, David R; Johnson, Colin; Marshall, John F; Scita, Giorgio; Jenei, Veronika

2017-01-01

Abstract The integrin αvβ6 is up‐regulated in numerous carcinomas, where expression commonly correlates with poor prognosis. αvβ6 promotes tumour invasion, partly through regulation of proteases and cell migration, and is also the principal mechanism by which epithelial cells activate TGF‐β1; this latter function complicates therapeutic targeting of αvβ6, since TGF‐β1 has both tumour‐promoting and ‐suppressive effects. It is unclear how these different αvβ6 functions are linked; both require actin cytoskeletal reorganization, and it is suggested that tractive forces generated during cell migration activate TGF‐β1 by exerting mechanical tension on the ECM‐bound latent complex. We examined the functional relationship between cell invasion and TGF‐β1 activation in pancreatic ductal adenocarcinoma (PDAC) cells, and confirmed that both processes are αvβ6‐dependent. Surprisingly, we found that cellular functions could be biased towards either motility or TGF‐β1 activation depending on the presence or absence of epidermal growth factor receptor pathway substrate 8 (Eps8), a regulator of actin remodelling, endocytosis, and GTPase activation. Similar to αvβ6, we found that Eps8 was up‐regulated in >70% of PDACs. In complex with Abi1/Sos1, Eps8 regulated αvβ6‐dependent cell migration through activation of Rac1. Down‐regulation of Eps8, Sos1 or Rac1 suppressed cell movement, while simultaneously increasing αvβ6‐dependent TGF‐β1 activation. This latter effect was modulated through increased cell tension, regulated by Rho activation. Thus, the Eps8/Abi1/Sos1 tricomplex acts as a key molecular switch altering the balance between Rac1 and Rho activation; its presence or absence in PDAC cells modulates αvβ6‐dependent functions, resulting in a pro‐migratory (Rac1‐dependent) or a pro‐TGF‐β1 activation (Rho‐dependent) functional phenotype, respectively. © 2017 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland. PMID:28608476

Nerve growth factor in the adult brain of a teleostean model for aging research: Nothobranchius furzeri.

PubMed

D'Angelo, L; Castaldo, L; Cellerino, A; de Girolamo, P; Lucini, C

2014-07-01

Nerve growth factor (NGF) acts on central nervous system neurons, regulating naturally occurring cell death, synaptic connectivity, fiber guidance and dendritic morphology. The dynamically regulated production of NGF beginning in development, extends throughout adult life and aging, exerting numerous roles through a surprising variety of neurons and glial cells. This study analyzes the localization of NGF in the brain of the teleost fish Nothobranchius furzeri, an emerging model for aging research due to its short lifespan. Immunochemical and immunohistochemical experiments were performed by employing an antibody mapping at the N-terminus of the mature chain human origin NGF. Western blot analysis revealed an intense and well defined band of 20 kDa, which corresponds to proNGF of N. furzeri. Immunohistochemistry revealed NGF immunoreactivity (IR) diffused throughout all regions of telencephalon, diencephalon, mesencephalon and rhomboencephalon. It was detected in neurons and in glial cells, the latter mostly lining the mesencephalic and rhomboencephalic ventricles. Particularly in neurons, NGF IR was localized in perikarya and, to a less extent, in fibers. The widespread distribution of proNGF suggests that it might modulate numerous physiological functions in the adult brain of N. furzeri. The present survey constitutes a baseline study to enhance the understanding of the mechanisms underlying the role of NGF during aging processes. Copyright © 2014 Elsevier GmbH. All rights reserved.

NF-κB Signalling in Glioblastoma

PubMed Central

Soubannier, Vincent; Stifani, Stefano

2017-01-01

Nuclear factor-κB (NF-κB) is a transcription factor regulating a wide array of genes mediating numerous cellular processes such as proliferation, differentiation, motility and survival, to name a few. Aberrant activation of NF-κB is a frequent event in numerous cancers, including glioblastoma, the most common and lethal form of brain tumours of glial cell origin (collectively termed gliomas). Glioblastoma is characterized by high cellular heterogeneity, resistance to therapy and almost inevitable recurrence after surgery and treatment. NF-κB is aberrantly activated in response to a variety of stimuli in glioblastoma, where its activity has been implicated in processes ranging from maintenance of cancer stem-like cells, stimulation of cancer cell invasion, promotion of mesenchymal identity, and resistance to radiotherapy. This review examines the mechanisms of NF-κB activation in glioblastoma, the involvement of NF-κB in several mechanisms underlying glioblastoma propagation, and discusses some of the important questions of future research into the roles of NF-κB in glioblastoma. PMID:28598356

Mathematical modeling and numerical simulation of the mitotic spindle orientation system.

PubMed

Ibrahim, Bashar

2018-05-21

The mitotic spindle orientation and position is crucial for the fidelity of chromosome segregation during asymmetric cell division to generate daughter cells with different sizes or fates. This mechanism is best understood in the budding yeast Saccharomyces cerevisiae, named the spindle position checkpoint (SPOC). The SPOC inhibits cells from exiting mitosis until the mitotic spindle is properly oriented along the mother-daughter polarity axis. Despite many experimental studies, the mechanisms underlying SPOC regulation remains elusive and unexplored theoretically. Here, a minimal mathematical is developed to describe SPOC activation and silencing having autocatalytic feedback-loop. Numerical simulations of the nonlinear ordinary differential equations (ODEs) model accurately reproduce the phenotype of SPOC mechanism. Bifurcation analysis of the nonlinear ODEs reveals the orientation dependency on spindle pole bodies, and how this dependence is altered by parameter values. These results provide for systems understanding on the molecular organization of spindle orientation system via mathematical modeling. The presented mathematical model is easy to understand and, within the above mentioned context, can be used as a base for further development of quantitative models in asymmetric cell-division. Copyright © 2018. Published by Elsevier Inc.

Phloem-mobile signals affecting flowers: applications for crop breeding.

PubMed

McGarry, Roisin C; Kragler, Friedrich

2013-04-01

Transport of endogenous macromolecules within and between tissues serves as a signaling pathway to regulate numerous aspects of plant growth. The florigenic FT gene product moves via the phloem from leaves to apical tissues and induces the flowering program in meristems. Similarly, short interfering RNA (siRNA) signals produced in source or sink tissues move cell-to-cell and long distance via the phloem to apical tissues. Recent advances in identifying these mobile signals regulating flowering or the epigenetic status of targeted tissues can be applicable to crop-breeding programs. In this review, we address the identity of florigen, the mechanism of allocation, and how virus-induced flowering and grafting of transgenes producing siRNA signals affecting meiosis can produce transgene-free progenies useful for agriculture. Copyright © 2013 Elsevier Ltd. All rights reserved.

Measurement and Analysis of in vitro Actin Polymerization

PubMed Central

Doolittle, Lynda K.; Rosen, Michael K.; Padrick, Shae B.

2014-01-01

Summary The polymerization of actin underlies force generation in numerous cellular processes. While actin polymerization can occur spontaneously, cells maintain control over this important process by preventing actin filament nucleation and then allowing stimulated polymerization and elongation by several regulated factors. Actin polymerization, regulated nucleation and controlled elongation activities can be reconstituted in vitro, and used to probe the signaling cascades cells use to control when and where actin polymerization occurs. Introducing a pyrene fluorophore allows detection of filament formation by an increase in pyrene fluorescence. This method has been used for many years and continues to be broadly used, owing to its simplicity and flexibility. Here we describe how to perform and analyze these in vitro actin polymerization assays, with an emphasis on extracting useful descriptive parameters from kinetic data. PMID:23868594

Matrigel Basement Membrane Matrix influences expression of microRNAs in cancer cell lines

DOE Office of Scientific and Technical Information (OSTI.GOV)

Price, Karina J.; School of Medicine and Pharmacology, University of Western Australia, Nedlands, WA 6008; Tsykin, Anna

2012-10-19

Highlights: Black-Right-Pointing-Pointer Matrigel alters cancer cell line miRNA expression relative to culture on plastic. Black-Right-Pointing-Pointer Many identified Matrigel-regulated miRNAs are implicated in cancer. Black-Right-Pointing-Pointer miR-1290, -210, -32 and -29b represent a Matrigel-induced miRNA signature. Black-Right-Pointing-Pointer miR-32 down-regulates Integrin alpha 5 (ITGA5) mRNA. -- Abstract: Matrigel is a medium rich in extracellular matrix (ECM) components used for three-dimensional cell culture and is known to alter cellular phenotypes and gene expression. microRNAs (miRNAs) are small, non-coding RNAs that regulate gene expression and have roles in cancer. While miRNA profiles of numerous cell lines cultured on plastic have been reported, the influence ofmore » Matrigel-based culture on cancer cell miRNA expression is largely unknown. This study investigated the influence of Matrigel on the expression of miRNAs that might facilitate ECM-associated cancer cell growth. We performed miRNA profiling by microarray using two colon cancer cell lines (SW480 and SW620), identifying significant differential expression of miRNAs between cells cultured in Matrigel and on plastic. Many of these miRNAs have previously been implicated in cancer-related processes. A common Matrigel-induced miRNA signature comprised of up-regulated miR-1290 and miR-210 and down-regulated miR-29b and miR-32 was identified using RT-qPCR across five epithelial cancer cell lines (SW480, SW620, HT-29, A549 and MDA-MB-231). Experimental modulation of these miRNAs altered expression of their known target mRNAs involved in cell adhesion, proliferation and invasion, in colon cancer cell lines. Furthermore, ITGA5 was identified as a novel putative target of miR-32 that may facilitate cancer cell interactions with the ECM. We propose that culture of cancer cell lines in Matrigel more accurately recapitulates miRNA expression and function in cancer than culture on plastic and thus is a valuable approach to the in vitro study of miRNAs.« less

Epigenetic regulation of cancer biology and anti-tumor immunity by EZH2.

PubMed

Christofides, Anthos; Karantanos, Theodoros; Bardhan, Kankana; Boussiotis, Vassiliki A

2016-12-20

Polycomb group proteins regulate chromatin structure and have an important regulatory role on gene expression in various cell types. Two polycomb group complexes (Polycomb repressive complex 1 (PRC1) and 2 (PRC2)) have been identified in mammalian cells. Both PRC1 and PRC2 compact chromatin, and also catalyze histone modifications. PRC1 mediates monoubiquitination of histone H2A, whereas PRC2 catalyzes methylation of histone H3 on lysine 27. These alterations of histones can lead to altered gene expression patterns by regulating chromatin structure. Numerous studies have highlighted the role of the PRC2 catalytic component enhancer of zeste homolog 2 (EZH2) in neoplastic development and progression, and EZH2 mutations have been identified in various malignancies. Through modulating the expression of critical genes, EZH2 is actively involved in fundamental cellular processes such as cell cycle progression, cell proliferation, differentiation and apoptosis. In addition to cancer cells, EZH2 also has a decisive role in the differentiation and function of T effector and T regulatory cells. In this review we summarize the recent progress regarding the role of EZH2 in human malignancies, highlight the molecular mechanisms by which EZH2 aberrations promote the pathogenesis of cancer, and discuss the anti-tumor effects of EZH2 targeting via activating direct anti-cancer mechanisms and anti-tumor immunity.

Epigenetic regulation of cancer biology and anti-tumor immunity by EZH2

PubMed Central

Bardhan, Kankana; Boussiotis, Vassiliki A.

2016-01-01

Polycomb group proteins regulate chromatin structure and have an important regulatory role on gene expression in various cell types. Two polycomb group complexes (Polycomb repressive complex 1 (PRC1) and 2 (PRC2)) have been identified in mammalian cells. Both PRC1 and PRC2 compact chromatin, and also catalyze histone modifications. PRC1 mediates monoubiquitination of histone H2A, whereas PRC2 catalyzes methylation of histone H3 on lysine 27. These alterations of histones can lead to altered gene expression patterns by regulating chromatin structure. Numerous studies have highlighted the role of the PRC2 catalytic component enhancer of zeste homolog 2 (EZH2) in neoplastic development and progression, and EZH2 mutations have been identified in various malignancies. Through modulating the expression of critical genes, EZH2 is actively involved in fundamental cellular processes such as cell cycle progression, cell proliferation, differentiation and apoptosis. In addition to cancer cells, EZH2 also has a decisive role in the differentiation and function of T effector and T regulatory cells. In this review we summarize the recent progress regarding the role of EZH2 in human malignancies, highlight the molecular mechanisms by which EZH2 aberrations promote the pathogenesis of cancer, and discuss the anti-tumor effects of EZH2 targeting via activating direct anti-cancer mechanisms and anti-tumor immunity. PMID:27793053

Redox signaling, Nox5 and vascular remodeling in hypertension.

PubMed

Montezano, Augusto C; Tsiropoulou, Sofia; Dulak-Lis, Maria; Harvey, Adam; Camargo, Livia De Lucca; Touyz, Rhian M

2015-09-01

Extensive data indicate a role for reactive oxygen species (ROS) and redox signaling in vascular damage in hypertension. However, molecular mechanisms underlying these processes remain unclear, but oxidative post-translational modification of vascular proteins is critical. This review discusses how proteins are oxidatively modified and how redox signaling influences vascular smooth muscle cell growth and vascular remodeling in hypertension. We also highlight Nox5 as a novel vascular ROS-generating oxidase. Oxidative stress in hypertension leads to oxidative imbalance that affects vascular cell function through redox signaling. Many Nox isoforms produce ROS in the vascular wall, and recent findings show that Nox5 may be important in humans. ROS regulate signaling by numerous processes including cysteine oxidative post-translational modification such as S-nitrosylation, S-glutathionylation and sulfydration. In vascular smooth muscle cells, this influences cellular responses to oxidative stimuli promoting changes from a contractile to a proliferative phenotype. In hypertension, Nox-induced ROS production is increased, leading to perturbed redox signaling through oxidative modifications of vascular proteins. This influences mitogenic signaling and cell cycle regulation, leading to altered cell growth and vascular remodeling in hypertension.

Regulation of metabolism by the Mediator complex.

PubMed

Youn, Dou Yeon; Xiaoli, Alus M; Pessin, Jeffrey E; Yang, Fajun

2016-01-01

The Mediator complex was originally discovered in yeast, but it is conserved in all eukaryotes. Its best-known function is to regulate RNA polymerase II-dependent gene transcription. Although the mechanisms by which the Mediator complex regulates transcription are often complicated by the context-dependent regulation, this transcription cofactor complex plays a pivotal role in numerous biological pathways. Biochemical, molecular, and physiological studies using cancer cell lines or model organisms have established the current paradigm of the Mediator functions. However, the physiological roles of the mammalian Mediator complex remain poorly defined, but have attracted a great interest in recent years. In this short review, we will summarize some of the reported functions of selective Mediator subunits in the regulation of metabolism. These intriguing findings suggest that the Mediator complex may be an important player in nutrient sensing and energy balance in mammals.

Identification of a new gene regulatory circuit involving B cell receptor activated signaling using a combined analysis of experimental, clinical and global gene expression data

PubMed Central

Schrader, Alexandra; Meyer, Katharina; Walther, Neele; Stolz, Ailine; Feist, Maren; Hand, Elisabeth; von Bonin, Frederike; Evers, Maurits; Kohler, Christian; Shirneshan, Katayoon; Vockerodt, Martina; Klapper, Wolfram; Szczepanowski, Monika; Murray, Paul G.; Bastians, Holger; Trümper, Lorenz; Spang, Rainer; Kube, Dieter

2016-01-01

To discover new regulatory pathways in B lymphoma cells, we performed a combined analysis of experimental, clinical and global gene expression data. We identified a specific cluster of genes that was coherently expressed in primary lymphoma samples and suppressed by activation of the B cell receptor (BCR) through αIgM treatment of lymphoma cells in vitro. This gene cluster, which we called BCR.1, includes numerous cell cycle regulators. A reduced expression of BCR.1 genes after BCR activation was observed in different cell lines and also in CD10+ germinal center B cells. We found that BCR activation led to a delayed entry to and progression of mitosis and defects in metaphase. Cytogenetic changes were detected upon long-term αIgM treatment. Furthermore, an inverse correlation of BCR.1 genes with c-Myc co-regulated genes in distinct groups of lymphoma patients was observed. Finally, we showed that the BCR.1 index discriminates activated B cell-like and germinal centre B cell-like diffuse large B cell lymphoma supporting the functional relevance of this new regulatory circuit and the power of guided clustering for biomarker discovery. PMID:27166259

Research Resource: Preovulatory LH Surge Effects on Follicular Theca and Granulosa Transcriptomes

PubMed Central

Gunewardena, Sumedha; Hong, Xiaoman; Spitschak, Marion; Baufeld, Anja

2013-01-01

The molecular mechanisms that regulate the pivotal transformation processes observed in the follicular wall following the preovulatory LH surge, are still not established, particularly for cells of the thecal layer. To elucidate thecal cell (TC) and granulosa cell (GC) type-specific biologic functions and signaling pathways, large dominant bovine follicles were collected before and 21 hours after an exogenous GnRH-induced LH surge. Antral GCs (aGCs; aspirated by follicular puncture) and membrane-associated GCs (mGCs; scraped from the follicular wall) were compared with TC expression profiles determined by mRNA microarrays. Of the approximately 11 000 total genes expressed in the periovulatory follicle, only 2% of thecal vs 25% of the granulosa genes changed in response to the LH surge. The majority of the 203 LH-regulated thecal genes were also LH regulated in GCs, leaving a total of 57 genes as LH-regulated TC-specific genes. Of the 57 thecal-specific LH-regulated genes, 74% were down-regulated including CYP17A1 and NR5A1, whereas most other genes are being identified for the first time within theca. Many of the newly identified up-regulated thecal genes (eg, PTX3, RND3, PPP4R4) were also up-regulated in granulosa. Minimal expression differences were observed between aGCs and mGCs; however, transcripts encoding extracellular proteins (NID2) and matrix modulators (ADAMTS1, SASH1) dominated these differences. We also identified large numbers of unknown LH-regulated GC genes and discuss their putative roles in ovarian function. This Research Resource provides an easy-to-access global evaluation of LH regulation in TCs and GCs that implicates numerous molecular pathways heretofore unknown within the follicle. PMID:23716604

Research resource: preovulatory LH surge effects on follicular theca and granulosa transcriptomes.

PubMed

Christenson, Lane K; Gunewardena, Sumedha; Hong, Xiaoman; Spitschak, Marion; Baufeld, Anja; Vanselow, Jens

2013-07-01

The molecular mechanisms that regulate the pivotal transformation processes observed in the follicular wall following the preovulatory LH surge, are still not established, particularly for cells of the thecal layer. To elucidate thecal cell (TC) and granulosa cell (GC) type-specific biologic functions and signaling pathways, large dominant bovine follicles were collected before and 21 hours after an exogenous GnRH-induced LH surge. Antral GCs (aGCs; aspirated by follicular puncture) and membrane-associated GCs (mGCs; scraped from the follicular wall) were compared with TC expression profiles determined by mRNA microarrays. Of the approximately 11 000 total genes expressed in the periovulatory follicle, only 2% of thecal vs 25% of the granulosa genes changed in response to the LH surge. The majority of the 203 LH-regulated thecal genes were also LH regulated in GCs, leaving a total of 57 genes as LH-regulated TC-specific genes. Of the 57 thecal-specific LH-regulated genes, 74% were down-regulated including CYP17A1 and NR5A1, whereas most other genes are being identified for the first time within theca. Many of the newly identified up-regulated thecal genes (eg, PTX3, RND3, PPP4R4) were also up-regulated in granulosa. Minimal expression differences were observed between aGCs and mGCs; however, transcripts encoding extracellular proteins (NID2) and matrix modulators (ADAMTS1, SASH1) dominated these differences. We also identified large numbers of unknown LH-regulated GC genes and discuss their putative roles in ovarian function. This Research Resource provides an easy-to-access global evaluation of LH regulation in TCs and GCs that implicates numerous molecular pathways heretofore unknown within the follicle.

Live-Cell Imaging of Mitochondria and the Actin Cytoskeleton in Budding Yeast.

PubMed

Higuchi-Sanabria, Ryo; Swayne, Theresa C; Boldogh, Istvan R; Pon, Liza A

2016-01-01

Maintenance and regulation of proper mitochondrial dynamics and functions are necessary for cellular homeostasis. Numerous diseases, including neurodegeneration and muscle myopathies, and overall cellular aging are marked by declining mitochondrial function and subsequent loss of multiple other cellular functions. For these reasons, optimized protocols are needed for visualization and quantification of mitochondria and their function and fitness. In budding yeast, mitochondria are intimately associated with the actin cytoskeleton and utilize actin for their movement and inheritance. This chapter describes optimal approaches for labeling mitochondria and the actin cytoskeleton in living budding yeast cells, for imaging the labeled cells, and for analyzing the resulting images.

tRNA wobble modifications and protein homeostasis

PubMed Central

Ranjan, Namit; Rodnina, Marina V.

2016-01-01

Abstract tRNA is a central component of the protein synthesis machinery in the cell. In living cells, tRNAs undergo numerous post-transcriptional modifications. In particular, modifications at the anticodon loop play an important role in ensuring efficient protein synthesis, maintaining protein homeostasis, and helping cell adaptation and survival. Hypo-modification of the wobble position of the tRNA anticodon loop is of particular relevance for translation regulation and is implicated in various human diseases. In this review we summarize recent evidence of how methyl and thiol modifications in eukaryotic tRNA at position 34 affect cellular fitness and modulate regulatory circuits at normal conditions and under stress. PMID:27335723

Riding the Waves: How Our Cells Send Signals | Center for Cancer Research

Cancer.gov

The ability of cells to perceive and respond to their environment is critical in order to maintain basic cellular functions such as development, tissue repair, and response to stress. This process happens through a complex system of communication, called cell signaling, which governs basic cellular activities and coordinates cell actions. Errors in cell signaling have been linked to numerous diseases, including cancer. NF-κB is a protein complex that plays a critical role in many cell signaling pathways by controlling gene activation. It is widely used by cells to regulate cell growth and survival and helps to protect the cell from conditions that would otherwise cause it to die. Many tumor cells have mutations in genes that cause NF-κB to become overactive. Blocking NF-κB could cause tumor cells to stop growing, die, or become more sensitive to therapeutics.

MEF2 Cooperates With Forskolin/cAMP and GATA4 to Regulate Star Gene Expression in Mouse MA-10 Leydig Cells.

PubMed

Daems, Caroline; Di-Luoffo, Mickaël; Paradis, Élise; Tremblay, Jacques J

2015-07-01

In Leydig cells, steroidogenic acute regulatory protein (STAR) participates in cholesterol shuttling from the outer to the inner mitochondrial membrane, the rate-limiting step in steroidogenesis. Steroid hormone biosynthesis and steroidogenic gene expression are regulated by LH, which activates various signaling pathways and transcription factors, including cAMP/Ca(2+)/CAMK (Ca(2+)/calmodulin-dependent kinase)-myocyte enhancer factor 2 (MEF2). The 4 MEF2 transcription factors are essential regulators of cell differentiation and organogenesis in numerous tissues. Recently, MEF2 was identified in Sertoli and Leydig cells of the testis. Here, we report that MEF2 regulates steroidogenesis in mouse MA-10 Leydig cells by acting on the Star gene. In MA-10 cells depleted of MEF2 using siRNAs (small interfering RNAs), STAR protein levels, Star mRNA levels, and promoter activity were significantly decreased. On its own, MEF2 did not activate the mouse Star promoter but was found to cooperate with forskolin/cAMP. By chromatin immunoprecipitation and DNA precipitation assays, we confirmed MEF2 binding to a consensus element located at -232 bp of the Star promoter. Mutation or deletion of the MEF2 element reduced but did not abrogate the MEF2/cAMP cooperation, indicating that MEF2 cooperates with other DNA-bound transcription factor(s). We identified GATA4 (GATA binding protein 4) as a partner for MEF2 in Leydig cells, because mutation of the GATA element abrogated the MEF2/cAMP cooperation on a reporter lacking a MEF2 element. MEF2 and GATA4 interact as revealed by coimmunoprecipitation, and MEF2 and GATA4 transcriptionally cooperate on the Star promoter. Altogether, our results define MEF2 as a novel regulator of steroidogenesis and Star transcription in Leydig cells and identify GATA4 as a key partner for MEF2-mediated action.

Mediator: A key regulator of plant development.

PubMed

Buendía-Monreal, Manuel; Gillmor, C Stewart

2016-11-01

Mediator is a multiprotein complex that regulates transcription at the level of RNA pol II assembly, as well as through regulation of chromatin architecture, RNA processing and recruitment of epigenetic marks. Though its modular structure is conserved in eukaryotes, its subunit composition has diverged during evolution and varies in response to environmental and tissue-specific inputs, suggesting different functions for each subunit and/or Mediator conformation. In animals, Mediator has been implicated in the control of differentiation and morphogenesis through modulation of numerous signaling pathways. In plants, studies have revealed roles for Mediator in regulation of cell division, cell fate and organogenesis, as well as developmental timing and hormone responses. We begin this review with an overview of biochemical mechanisms of yeast and animal Mediator that are likely to be conserved in all eukaryotes, as well as a brief discussion of the role of Mediator in animal development. We then present a comprehensive review of studies of the role of Mediator in plant development. Finally, we point to important questions for future research on the role of Mediator as a master coordinator of development. Copyright © 2016 Elsevier Inc. All rights reserved.

Stochastic multi-scale models of competition within heterogeneous cellular populations: Simulation methods and mean-field analysis.

PubMed

Cruz, Roberto de la; Guerrero, Pilar; Spill, Fabian; Alarcón, Tomás

2016-10-21

We propose a modelling framework to analyse the stochastic behaviour of heterogeneous, multi-scale cellular populations. We illustrate our methodology with a particular example in which we study a population with an oxygen-regulated proliferation rate. Our formulation is based on an age-dependent stochastic process. Cells within the population are characterised by their age (i.e. time elapsed since they were born). The age-dependent (oxygen-regulated) birth rate is given by a stochastic model of oxygen-dependent cell cycle progression. Once the birth rate is determined, we formulate an age-dependent birth-and-death process, which dictates the time evolution of the cell population. The population is under a feedback loop which controls its steady state size (carrying capacity): cells consume oxygen which in turn fuels cell proliferation. We show that our stochastic model of cell cycle progression allows for heterogeneity within the cell population induced by stochastic effects. Such heterogeneous behaviour is reflected in variations in the proliferation rate. Within this set-up, we have established three main results. First, we have shown that the age to the G1/S transition, which essentially determines the birth rate, exhibits a remarkably simple scaling behaviour. Besides the fact that this simple behaviour emerges from a rather complex model, this allows for a huge simplification of our numerical methodology. A further result is the observation that heterogeneous populations undergo an internal process of quasi-neutral competition. Finally, we investigated the effects of cell-cycle-phase dependent therapies (such as radiation therapy) on heterogeneous populations. In particular, we have studied the case in which the population contains a quiescent sub-population. Our mean-field analysis and numerical simulations confirm that, if the survival fraction of the therapy is too high, rescue of the quiescent population occurs. This gives rise to emergence of resistance to therapy since the rescued population is less sensitive to therapy. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

INHIBITION OF ERN1 SIGNALING ENZYME AFFECTS HYPOXIC REGULATION OF THE EXPRESSION OF E2F8, EPAS1, HOXC6, ATF3, TBX3 AND FOXF1 GENES IN U87 GLIOMA CELLS.

PubMed

Minchenko, O H; Tsymbal, D O; Minchenko, D O; Kovalevska, O V; Karbovskyi, L L; Bikfalvi, A

2015-01-01

Hypoxia as well as the endoplasmic reticulum stress are important factors of malignant tumor growth and control of the expression of genes, which regulate numerous metabolic processes and cell proliferation. Furthermore, blockade of ERN1 (endoplasmic reticulum to nucleus 1) suppresses cell proliferation and tumor growth. We studied the effect of hypoxia on the expression of genes encoding the transcription factors such as E2F8 (E2F transcription factor 8), EPAS1 (endothelial PAS domain protein 1), TBX3 (T-box 3), ATF3 (activating transcription factor 3), FOXF1 (forkhead box F), and HOXC6 (homeobox C6) in U87 glioma cells with and without ERN1 signaling enzyme function. We have established that hypoxia enhances the expression of HOXC6, E2F8, ATF3, and EPAS1 genes but does not change TBX3 and FOXF1 gene expression in glioma cells with ERNI function. At the same time, the expression level of all studied genes is strongly decreased, except for TBX3 gene, in glioma cells without ERN1 function. Moreover, the inhibition of ERN1 signaling enzyme function significantly modifies the effect of hypoxia on the expression of these transcription factor genes. removes or introduces this regulation as well as changes a direction or magnitude of hypoxic regulation. Present study demonstrates that fine-tuning of the expression of proliferation related genes depends upon hypoxia and ERN1-mediated endoplasmic reticulum stress signaling and correlates with slower proliferation rate of glioma cells without ERN1 function.

Deficiency of the Arabidopsis helicase RTEL1 triggers a SOG1-dependent replication checkpoint in response to DNA cross-links.

PubMed

Hu, Zhubing; Cools, Toon; Kalhorzadeh, Pooneh; Heyman, Jefri; De Veylder, Lieven

2015-01-01

To maintain genome integrity, DNA replication is executed and regulated by a complex molecular network of numerous proteins, including helicases and cell cycle checkpoint regulators. Through a systematic screening for putative replication mutants, we identified an Arabidopsis thaliana homolog of human Regulator of Telomere Length 1 (RTEL1), which functions in DNA replication, DNA repair, and recombination. RTEL1 deficiency retards plant growth, a phenotype including a prolonged S-phase duration and decreased cell proliferation. Genetic analysis revealed that rtel1 mutant plants show activated cell cycle checkpoints, specific sensitivity to DNA cross-linking agents, and increased homologous recombination, but a lack of progressive shortening of telomeres, indicating that RTEL1 functions have only been partially conserved between mammals and plants. Surprisingly, RTEL1 deficiency induces tolerance to the deoxynucleotide-depleting drug hydroxyurea, which could be mimicked by DNA cross-linking agents. This resistance does not rely on the essential replication checkpoint regulator WEE1 but could be blocked by a mutation in the SOG1 transcription factor. Taken together, our data indicate that RTEL1 is required for DNA replication and that its deficiency activates a SOG1-dependent replication checkpoint. © 2015 American Society of Plant Biologists. All rights reserved.

Evidence of a local negative role for cocaine and amphetamine regulated transcript (CART), inhibins and low molecular weight insulin like growth factor binding proteins in regulation of granulosa cell estradiol production during follicular waves in cattle

PubMed Central

Kobayashi, Yasuhiro; Jimenez-Krassel, Fermin; Ireland, James J; Smith, George W

2006-01-01

The ability of ovarian follicles to produce large amounts of estradiol is a hallmark of follicle health status. Estradiol producing capacity is lost in ovarian follicles before morphological signs of atresia. A prominent wave like pattern of growth of antral follicles is characteristic of monotocous species such as cattle, horses and humans. While our knowledge of the role of pituitary gonadotropins in support of antral follicle growth and development is well established, the intrinsic factors that suppress estradiol production and may help promote atresia during follicular waves are not well understood. Numerous growth factors and cytokines have been reported to suppress granulosa cell estradiol production in vitro, but the association of expression of many such factors in vivo with follicle health status and their physiological significance are not clear. The purpose of this review is to discuss the in vivo and in vitro evidence supporting a local physiological role for cocaine and amphetamine regulated transcript, inhibins and low molecular weight insulin like growth factor binding proteins in negative regulation of granulosa cell estradiol production, with emphasis on evidence from the bovine model system. PMID:16611367

The Eukaryote-Like Serine/Threonine Kinase STK Regulates the Growth and Metabolism of Zoonotic Streptococcus suis

PubMed Central

Zhang, Chunyan; Sun, Wen; Tan, Meifang; Dong, Mengmeng; Liu, Wanquan; Gao, Ting; Li, Lu; Xu, Zhuofei; Zhou, Rui

2017-01-01

Like eukaryotes, bacteria express one or more serine/threonine kinases (STKs) that initiate diverse signaling networks. The STK from Streptococcus suis is encoded by a single-copy stk gene, which is crucial in stress response and virulence. To further understand the regulatory mechanism of STK in S. suis, a stk deletion strain (Δstk) and its complementary strain (CΔstk) were constructed to systematically decode STK characteristics by applying whole transcriptome RNA sequencing (RNA-Seq) and phosphoproteomic analysis. Numerous genes were differentially expressed in Δstk compared with the wild-type parental strain SC-19, including 320 up-regulated and 219 down-regulated genes. Particularly, 32 virulence-associated genes (VAGs) were significantly down-regulated in Δstk. Seven metabolic pathways relevant to bacterial central metabolism and translation are significantly repressed in Δstk. Phosphoproteomic analysis further identified 12 phosphoproteins that exhibit differential phosphorylation in Δstk. These proteins are associated with cell growth and division, glycolysis, and translation. Consistently, phenotypic assays confirmed that the Δstk strain displayed deficient growth and attenuated pathogenicity. Thus, STK is a central regulator that plays an important role in cell growth and division, as well as S. suis metabolism. PMID:28326294

Pro-apoptotic BIM is an essential initiator of physiological endothelial cell death independent of regulation by FOXO3.

PubMed

Koenig, M N; Naik, E; Rohrbeck, L; Herold, M J; Trounson, E; Bouillet, P; Thomas, T; Voss, A K; Strasser, A; Coultas, L

2014-11-01

The growth of new blood vessels by angiogenesis is essential for normal development, but can also cause or contribute to the pathology of numerous diseases. Recent studies have shown that BIM, a pro-apoptotic BCL2-family protein, is required for endothelial cell apoptosis in vivo, and can contribute to the anti-angiogenic effect of VEGF-A inhibitors in certain tumor models. Despite its importance, the extent to which BIM is autonomously required for physiological endothelial apoptosis remains unknown and its regulation under such conditions is poorly defined. While the transcription factor FOXO3 has been proposed to induce Bim in response to growth factor withdrawal, evidence for this function is circumstantial. We report that apoptosis was reduced in Bim(-/-) primary endothelial cells, demonstrating a cell-autonomous role for BIM in endothelial death following serum and growth factor withdrawal. In conflict with in vitro studies, BIM-dependent endothelial death in vivo did not require FOXO3. Moreover, endothelial apoptosis proceeded normally in mice lacking FOXO-binding sites in the Bim promoter. Bim mRNA was upregulated in endothelial cells starved of serum and growth factors and this was accompanied by the downregulation of miRNAs of the miR-17∼92 cluster. Bim mRNA levels were also elevated in miR-17∼92(+/-) endothelial cells cultured under steady-state conditions, suggesting that miR-17∼92 cluster miRNAs may contribute to regulating overall Bim mRNA levels in endothelial cells.

Gene Expression Networks Underlying Ovarian Development in Wild Largemouth Bass (Micropterus salmoides)

PubMed Central

Martyniuk, Christopher J.; Prucha, Melinda S.; Doperalski, Nicholas J.; Antczak, Philipp; Kroll, Kevin J.; Falciani, Francesco; Barber, David S.; Denslow, Nancy D.

2013-01-01

Background Oocyte maturation in fish involves numerous cell signaling cascades that are activated or inhibited during specific stages of oocyte development. The objectives of this study were to characterize molecular pathways and temporal gene expression patterns throughout a complete breeding cycle in wild female largemouth bass to improve understanding of the molecular sequence of events underlying oocyte maturation. Methods Transcriptomic analysis was performed on eight morphologically diverse stages of the ovary, including primary and secondary stages of oocyte growth, ovulation, and atresia. Ovary histology, plasma vitellogenin, 17β-estradiol, and testosterone were also measured to correlate with gene networks. Results Global expression patterns revealed dramatic differences across ovarian development, with 552 and 2070 genes being differentially expressed during both ovulation and atresia respectively. Gene set enrichment analysis (GSEA) revealed that early primary stages of oocyte growth involved increases in expression of genes involved in pathways of B-cell and T-cell receptor-mediated signaling cascades and fibronectin regulation. These pathways as well as pathways that included adrenergic receptor signaling, sphingolipid metabolism and natural killer cell activation were down-regulated at ovulation. At atresia, down-regulated pathways included gap junction and actin cytoskeleton regulation, gonadotrope and mast cell activation, and vasopressin receptor signaling and up-regulated pathways included oxidative phosphorylation and reactive oxygen species metabolism. Expression targets for luteinizing hormone signaling were low during vitellogenesis but increased 150% at ovulation. Other networks found to play a significant role in oocyte maturation included those with genes regulated by members of the TGF-beta superfamily (activins, inhibins, bone morphogenic protein 7 and growth differentiation factor 9), neuregulin 1, retinoid X receptor, and nerve growth factor family. Conclusions This study offers novel insight into the gene networks underlying vitellogenesis, ovulation and atresia and generates new hypotheses about the cellular pathways regulating oocyte maturation. PMID:23527095

Biochemistry and biology: heart-to-heart to investigate cardiac progenitor cells.

PubMed

Chimenti, Isotta; Forte, Elvira; Angelini, Francesco; Messina, Elisa; Giacomello, Alessandro

2013-02-01

Cardiac regenerative medicine is a rapidly evolving field, with promising future developments for effective personalized treatments. Several stem/progenitor cells are candidates for cardiac cell therapy, and emerging evidence suggests how multiple metabolic and biochemical pathways strictly regulate their fate and renewal. In this review, we will explore a selection of areas of common interest for biology and biochemistry concerning stem/progenitor cells, and in particular cardiac progenitor cells. Numerous regulatory mechanisms have been identified that link stem cell signaling and functions to the modulation of metabolic pathways, and vice versa. Pharmacological treatments and culture requirements may be exploited to modulate stem cell pluripotency and self-renewal, possibly boosting their regenerative potential for cell therapy. Mitochondria and their many related metabolites and messengers, such as oxygen, ROS, calcium and glucose, have a crucial role in regulating stem cell fate and the balance of their functions, together with many metabolic enzymes. Furthermore, protein biochemistry and proteomics can provide precious clues on the definition of different progenitor cell populations, their physiology and their autocrine/paracrine regulatory/signaling networks. Interdisciplinary approaches between biology and biochemistry can provide productive insights on stem/progenitor cells, allowing the development of novel strategies and protocols for effective cardiac cell therapy clinical translation. This article is part of a Special Issue entitled Biochemistry of Stem Cells. Copyright © 2012 Elsevier B.V. All rights reserved.

Coexistence of proguanylin (1-15) and somatostatin in the gastrointestinal tract.

PubMed

Ieda, H; Naruse, S; Furuya, S; Ozaki, T; Ando, E; Nokihara, K; Hori, S; Kitagawa, M; Hayakawa, T

1998-12-01

In order to identify proguanylin-secreting cells, we have raised an antiserum against the synthetic fragment of human proguanylin (1-15) and have examined the proguanylin-positive cells in the human and rat gastrointestinal tract by immunohistochemical methods. Numerous proguanylin (1-15)-immunoreactive cells were found in the gastrointestinal tract. They were either pyramidal or spindle shaped in the stomach. Spindle-shaped cells, frequently possessing long slender processes, were located at the base of the pyloric epithelium and did not extend to the lumen. In the duodenum and jejunum, these cells were mostly pyramidal in shape and often had a slender process towards the lumen. The immunostaining was completely blocked by the human proguanylin (1-15) fragment. Paneth and goblet cells were negative against this antiserum. The number of serotonin-positive cells was much larger than that of proguanylin-positive cells in all the segments tested. The number of proguanylin-positive cells decreased from the jejunum to the ileum and very few cells were observed in the colon. In contrast to serotonin-positive cells, most somatostatin-positive cells were also positive for proguanylin. Thus, proguanylin (1-15) or its related protein appears to coexist with somatostatin in intestinal endocrine D cells which may be a source of circulating proguanylin. Proguanylin, like somatostatin, may also regulate intestinal function as a local regulator.

Effects of Different Zinc Species on Cellar Zinc Distribution, Cell Cycle, Apoptosis and Viability in MDAMB231 Cells.

PubMed

Wang, Yan-hong; Zhao, Wen-jie; Zheng, Wei-juan; Mao, Li; Lian, Hong-zhen; Hu, Xin; Hua, Zi-chun

2016-03-01

Intracellular metal elements exist in mammalian cells with the concentration range from picomoles per litre to micromoles per litre and play a considerable role in various biological procedures. Element provided by different species can influence the availability and distribution of the element in a cell and could lead to different biological effects on the cell's growth and function. Zinc as an abundant and widely distributed essential trace element, is involved in numerous and relevant physiological functions. Zinc homeostasis in cells, which is regulated by metallothioneins, zinc transporter/SLC30A, Zrt-/Irt-like proteins/SLC39A and metal-response element-binding transcription factor-1 (MTF-1), is crucial for normal cellular functioning. In this study, we investigated the influences of different zinc species, zinc sulphate, zinc gluconate and bacitracin zinc, which represented inorganic, organic and biological zinc species, respectively, on cell cycle, viability and apoptosis in MDAMB231 cells. It was found that the responses of cell cycle, apoptosis and death to different zinc species in MDAMB231 cells are different. Western blot analysis of the expression of several key proteins in regulating zinc-related transcription, cell cycle, apoptosis, including MTF-1, cyclin B1, cyclin D1, caspase-8 and caspase-9 in treated cells further confirmed the observed results on cell level.

ADAM10 and γ-secretase regulate sensory regeneration in the avian vestibular organs.

PubMed

Warchol, Mark E; Stone, Jennifer; Barton, Matthew; Ku, Jeffrey; Veile, Rose; Daudet, Nicolas; Lovett, Michael

2017-08-01

The loss of sensory hair cells from the inner ear is a leading cause of hearing and balance disorders. The mammalian ear has a very limited ability to replace lost hair cells, but the inner ears of non-mammalian vertebrates can spontaneously regenerate hair cells after injury. Prior studies have shown that replacement hair cells are derived from epithelial supporting cells and that the differentiation of new hair cells is regulated by the Notch signaling pathway. The present study examined molecular influences on regeneration in the avian utricle, which has a particularly robust regenerative ability. Chicken utricles were placed in organotypic culture and hair cells were lesioned by application of the ototoxic antibiotic streptomycin. Cultures were then allowed to regenerate in vitro for seven days. Some specimens were treated with small molecule inhibitors of γ-secretase or ADAM10, proteases which are essential for transmission of Notch signaling. As expected, treatment with both inhibitors led to increased numbers of replacement hair cells. However, we also found that inhibition of both proteases resulted in increased regenerative proliferation. Subsequent experiments showed that inhibition of γ-secretase or ADAM10 could also trigger proliferation in undamaged utricles. To better understand these phenomena, we used RNA-Seq profiling to characterize changes in gene expression following γ-secretase inhibition. We observed expression patterns that were consistent with Notch pathway inhibition, but we also found that the utricular sensory epithelium contains numerous γ-secretase substrates that might regulate cell cycle entry and possibly supporting cell-to-hair cell conversion. Together, our data suggest multiple roles for γ-secretase and ADAM10 in vestibular hair cell regeneration. Copyright © 2017. Published by Elsevier Inc.

RBFOX2 protein domains and cellular activities.

PubMed

Arya, Anurada D; Wilson, David I; Baralle, Diana; Raponi, Michaela

2014-08-01

RBFOX2 (RNA-binding protein, Fox-1 homologue 2)/RBM9 (RNA-binding-motif protein 9)/RTA (repressor of tamoxifen action)/HNRBP2 (hexaribonucleotide-binding protein 2) encodes an RNA-binding protein involved in tissue specific alternative splicing regulation and steroid receptors transcriptional activity. Its ability to regulate specific splicing profiles depending on context has been related to different expression levels of the RBFOX2 protein itself and that of other splicing regulatory proteins involved in the shared modulation of specific genes splicing. However, this cannot be the sole explanation as to why RBFOX2 plays a widespread role in numerous cellular mechanisms from development to cell survival dependent on cell/tissue type. RBFOX2 isoforms with altered protein domains exist. In the present article, we describe the main RBFOX2 protein domains, their importance in the context of splicing and transcriptional regulation and we propose that RBFOX2 isoform distribution may play a fundamental role in RBFOX2-specific cellular effects.

Analysis of Cysteine Redox Post-Translational Modifications in Cell Biology and Drug Pharmacology.

PubMed

Wani, Revati; Murray, Brion W

2017-01-01

Reversible cysteine oxidation is an emerging class of protein post-translational modification (PTM) that regulates catalytic activity, modulates conformation, impacts protein-protein interactions, and affects subcellular trafficking of numerous proteins. Redox PTMs encompass a broad array of cysteine oxidation reactions with different half-lives, topographies, and reactivities such as S-glutathionylation and sulfoxidation. Recent studies from our group underscore the lesser known effect of redox protein modifications on drug binding. To date, biological studies to understand mechanistic and functional aspects of redox regulation are technically challenging. A prominent issue is the lack of tools for labeling proteins oxidized to select chemotype/oxidant species in cells. Predictive computational tools and curated databases of oxidized proteins are facilitating structural and functional insights into regulation of the network of oxidized proteins or redox proteome. In this chapter, we discuss analytical platforms for studying protein oxidation, suggest computational tools currently available in the field to determine redox sensitive proteins, and begin to illuminate roles of cysteine redox PTMs in drug pharmacology.

ADAM-17: The Enzyme That Does It All

PubMed Central

Gooz, Monika

2010-01-01

This review focuses on the role of ADAM-17 in disease. Since its debut as the tumor necrosis factor converting enzyme or TACE, ADAM-17 has been reported to be an indispensible regulator of almost every cellular event from proliferation to migration. The central role of ADAM-17 in cell regulation is rooted in its diverse array of substrates: cytokines, growth factors, and their receptors as well as adhesion molecules are activated or inactivated by their cleavage with ADAM-17. It is therefore not surprising that ADAM-17 is implicated in numerous human diseases including cancer, heart disease, diabetes, rheumatoid arthritis, kidney fibrosis, Alzheimer’s disease, and is a promising target for future treatments. The specific role of ADAM-17 in the pathophysiology of these diseases is very complex and depends on the cellular context. To exploit the therapeutic potential of ADAM-17, it is important to understand how its activity is regulated and how specific organs and cells can be targeted to inactivate or activate the enzyme. PMID:20184396

Modulation of flagellum attachment zone protein FLAM3 and regulation of the cell shape in Trypanosoma brucei life cycle transitions

PubMed Central

Sunter, Jack D.; Benz, Corinna; Andre, Jane; Whipple, Sarah; McKean, Paul G.; Gull, Keith; Ginger, Michael L.; Lukeš, Julius

2015-01-01

ABSTRACT The cell shape of Trypanosoma brucei is influenced by flagellum-to-cell-body attachment through a specialised structure – the flagellum attachment zone (FAZ). T. brucei exhibits numerous morphological forms during its life cycle and, at each stage, the FAZ length varies. We have analysed FLAM3, a large protein that localises to the FAZ region within the old and new flagellum. Ablation of FLAM3 expression causes a reduction in FAZ length; however, this has remarkably different consequences in the tsetse procyclic form versus the mammalian bloodstream form. In procyclic form cells FLAM3 RNAi results in the transition to an epimastigote-like shape, whereas in bloodstream form cells a severe cytokinesis defect associated with flagellum detachment is observed. Moreover, we demonstrate that the amount of FLAM3 and its localisation is dependent on ClpGM6 expression and vice versa. This evidence demonstrates that FAZ is a key regulator of trypanosome shape, with experimental perturbations being life cycle form dependent. An evolutionary cell biology explanation suggests that these differences are a reflection of the division process, the cytoskeleton and intrinsic structural plasticity of particular life cycle forms. PMID:26148511

Musashi RNA-Binding Proteins as Cancer Drivers and Novel Therapeutic Targets.

PubMed

Kudinov, Alexander E; Karanicolas, John; Golemis, Erica A; Boumber, Yanis

2017-05-01

Aberrant gene expression that drives human cancer can arise from epigenetic dysregulation. Although much attention has focused on altered activity of transcription factors and chromatin-modulating proteins, proteins that act posttranscriptionally can potently affect expression of oncogenic signaling proteins. The RNA-binding proteins (RBP) Musashi-1 (MSI1) and Musashi-2 (MSI2) are emerging as regulators of multiple critical biological processes relevant to cancer initiation, progression, and drug resistance. Following identification of Musashi as a regulator of progenitor cell identity in Drosophila , the human Musashi proteins were initially linked to control of maintenance of hematopoietic stem cells, then stem cell compartments for additional cell types. More recently, the Musashi proteins were found to be overexpressed and prognostic of outcome in numerous cancer types, including colorectal, lung, and pancreatic cancers; glioblastoma; and several leukemias. MSI1 and MSI2 bind and regulate the mRNA stability and translation of proteins operating in essential oncogenic signaling pathways, including NUMB/Notch, PTEN/mTOR, TGFβ/SMAD3, MYC, cMET, and others. On the basis of these activities, MSI proteins maintain cancer stem cell populations and regulate cancer invasion, metastasis, and development of more aggressive cancer phenotypes, including drug resistance. Although RBPs are viewed as difficult therapeutic targets, initial efforts to develop MSI-specific inhibitors are promising, and RNA interference-based approaches to inhibiting these proteins have had promising outcomes in preclinical studies. In the interim, understanding the function of these translational regulators may yield insight into the relationship between mRNA expression and protein expression in tumors, guiding tumor-profiling analysis. This review provides a current overview of Musashi as a cancer driver and novel therapeutic target. Clin Cancer Res; 23(9); 2143-53. ©2017 AACR . ©2017 American Association for Cancer Research.

The transcription factor EGR1 localizes to the nucleolus and is linked to suppression of ribosomal precursor synthesis.

PubMed

Ponti, Donatella; Bellenchi, Gian Carlo; Puca, Rosa; Bastianelli, Daniela; Maroder, Marella; Ragona, Giuseppe; Roussel, Pascal; Thiry, Marc; Mercola, Dan; Calogero, Antonella

2014-01-01

EGR1 is an immediate early gene with a wide range of activities as transcription factor, spanning from regulation of cell growth to differentiation. Numerous studies show that EGR1 either promotes the proliferation of stimulated cells or suppresses the tumorigenic growth of transformed cells. Upon interaction with ARF, EGR1 is sumoylated and acquires the ability to bind to specific targets such as PTEN and in turn to regulate cell growth. ARF is mainly localized to the periphery of nucleolus where is able to negatively regulate ribosome biogenesis. Since EGR1 colocalizes with ARF under IGF-1 stimulation we asked the question of whether EGR1 also relocate to the nucleolus to interact with ARF. Here we show that EGR1 colocalizes with nucleolar markers such as fibrillarin and B23 in the presence of ARF. Western analysis of nucleolar extracts from HeLa cells was used to confirm the presence of EGR1 in the nucleolus mainly as the 100 kDa sumoylated form. We also show that the level of the ribosomal RNA precursor 47S is inversely correlated to the level of EGR1 transcripts. The EGR1 iseffective to regulate the synthesis of the 47S rRNA precursor. Then we demonstrated that EGR1 binds to the Upstream Binding Factor (UBF) leading us to hypothesize that the regulating activity of EGR1 is mediated by its interaction within the transcriptional complex of RNA polymerase I. These results confirm the presence of EGR1 in the nucleolus and point to a role for EGR1 in the control of nucleolar metabolism.

Predicting network modules of cell cycle regulators using relative protein abundance statistics.

PubMed

Oguz, Cihan; Watson, Layne T; Baumann, William T; Tyson, John J

2017-02-28

Parameter estimation in systems biology is typically done by enforcing experimental observations through an objective function as the parameter space of a model is explored by numerical simulations. Past studies have shown that one usually finds a set of "feasible" parameter vectors that fit the available experimental data equally well, and that these alternative vectors can make different predictions under novel experimental conditions. In this study, we characterize the feasible region of a complex model of the budding yeast cell cycle under a large set of discrete experimental constraints in order to test whether the statistical features of relative protein abundance predictions are influenced by the topology of the cell cycle regulatory network. Using differential evolution, we generate an ensemble of feasible parameter vectors that reproduce the phenotypes (viable or inviable) of wild-type yeast cells and 110 mutant strains. We use this ensemble to predict the phenotypes of 129 mutant strains for which experimental data is not available. We identify 86 novel mutants that are predicted to be viable and then rank the cell cycle proteins in terms of their contributions to cumulative variability of relative protein abundance predictions. Proteins involved in "regulation of cell size" and "regulation of G1/S transition" contribute most to predictive variability, whereas proteins involved in "positive regulation of transcription involved in exit from mitosis," "mitotic spindle assembly checkpoint" and "negative regulation of cyclin-dependent protein kinase by cyclin degradation" contribute the least. These results suggest that the statistics of these predictions may be generating patterns specific to individual network modules (START, S/G2/M, and EXIT). To test this hypothesis, we develop random forest models for predicting the network modules of cell cycle regulators using relative abundance statistics as model inputs. Predictive performance is assessed by the areas under receiver operating characteristics curves (AUC). Our models generate an AUC range of 0.83-0.87 as opposed to randomized models with AUC values around 0.50. By using differential evolution and random forest modeling, we show that the model prediction statistics generate distinct network module-specific patterns within the cell cycle network.

The Multifaceted Roles of Neutrophil Gelatinase Associated Lipocalin (NGAL) In Inflammation and Cancer

PubMed Central

Chakraborty, Subhankar; Kaur, Sukhwinder; Guha, Sushovan; Batra, Surinder K.

2012-01-01

Neutrophil gelatinase associated lipocalin (NGAL), also known as oncogene 24p3, uterocalin, siderocalin or lipocalin 2, is a 24 kDa secreted glycoprotein originally purified from a culture of mouse kidney cells infected with simian virus 40 (SV-40). Subsequent investigations have revealed that it is a member of the lipocalin family of proteins that transport small, hydrophobic ligands. Since then, NGAL expression has been reported in several normal tissues where it serves to provide protection against bacterial infection and modulate oxidative stress. Its expression is also dysregulated in several benign and malignant diseases. Its small size, secreted nature and relative stability have led to it being investigated as a diagnostic and prognostic biomarker in numerous diseases including inflammation and cancer. Functional studies, conducted primarily on lipocalin 2 (Lcn2), the mouse homologue of human NGAL have revealed that Lcn2 has a strong affinity for iron complexed to both bacterial siderophores (iron binding proteins) and certain human proteins like norepinephrine. By sequestering iron-laden siderophores, Lcn2 deprives bacteria of a vital nutrient and thus inhibits their growth (bacteriostatic effect). In malignant cells, its proposed functions range from inhibiting apoptosis (in thyroid cancer cells), invasion and angiogenesis (in pancreatic cancer) to increasing proliferation and metastasis (in breast and colon cancer). Ectopic expression of Lcn2 also promotes BCR-ABL induced chronic myelogenous leukemia in murine models. By transporting iron into and out of the cell, NGAL also regulates iron responsive genes. Further, it stabilizes the proteolytic enzyme matrix metalloprotease-9 (MMP-9) by forming a complex with it, and thereby prevents its autodegradation. The factors regulating NGAL expression are numerous and range from pro-inflammatory cytokines like interleukins, tumor necrosis factor-α and interferons to vitamins like retinoic acid. The purpose of this review article is to examine the expression, structure, regulation and biological role of NGAL and critically assess its potential as a novel diagnostic and prognostic marker in both benign and malignant human diseases. PMID:22513004

Treating atherosclerosis with regulatory T cells.

PubMed

Foks, Amanda C; Lichtman, Andrew H; Kuiper, Johan

2015-02-01

Regulatory T cells (Tregs) play an important role in the regulation of T-cell-mediated immune responses through suppression of T-cell proliferation and secretion of inhibitory cytokines, such as interleukin-10 and transforming growth factor-β. Impaired Treg numbers and function have been associated with numerous diseases, and an imbalance between proinflammatory/proatherogenic cells and Tregs promotes atherosclerotic disease. Restoration of this balance by inducing Tregs has great therapeutic potential to prevent cardiovascular disease. In addition to suppressing differentiation and function of effector T cells, Tregs have been shown to induce anti-inflammatory macrophages, inhibit foam cell formation and to influence cholesterol metabolism. Furthermore, Tregs suppress immune responses of endothelial cells and innate lymphoid cells. In this review, we focus on the recent knowledge on Treg subsets, their activity and function in atherosclerosis, and discuss promising strategies to use Tregs as a therapeutic tool to prevent cardiovascular disease. © 2014 American Heart Association, Inc.

Coordinating Regulation of Gene Expression in Cardiovascular Disease: Interactions between Chromatin Modifiers and Transcription Factors

PubMed Central

Bauer, Ashley J.; Martin, Kathleen A.

2017-01-01

Cardiovascular disease is a leading cause of death with increasing economic burden. The pathogenesis of cardiovascular diseases is complex, but can arise from genetic and/or environmental risk factors. This can lead to dysregulated gene expression in numerous cell types including cardiomyocytes, endothelial cells, vascular smooth muscle cells, and inflammatory cells. While initial studies addressed transcriptional control of gene expression, epigenetics has been increasingly appreciated to also play an important role in this process through alterations in chromatin structure and gene accessibility. Chromatin-modifying proteins including enzymes that modulate DNA methylation, histone methylation, and histone acetylation can influence gene expression in numerous ways. These chromatin modifiers and their marks can promote or prevent transcription factor recruitment to regulatory regions of genes through modifications to DNA, histones, or the transcription factors themselves. This review will focus on the emerging question of how epigenetic modifiers and transcription factors interact to coordinately regulate gene expression in cardiovascular disease. While most studies have addressed the roles of either epigenetic or transcriptional control, our understanding of the integration of these processes is only just beginning. Interrogating these interactions is challenging, and improved technical approaches will be needed to fully dissect the temporal and spatial relationships between transcription factors, chromatin modifiers, and gene expression in cardiovascular disease. We summarize the current state of the field and provide perspectives on limitations and future directions. Through studies of epigenetic and transcriptional interactions, we can advance our understanding of the basic mechanisms of cardiovascular disease pathogenesis to develop novel therapeutics. PMID:28428957

Smooth muscle-protein translocation and tissue function.

PubMed

Eddinger, Thomas J

2014-09-01

Smooth muscle (SM) tissue is a complex organization of multiple cell types and is regulated by numerous signaling molecules (neurotransmitters, hormones, cytokines, etc.). SM contractile function can be regulated via expression and distribution of the contractile and cytoskeletal proteins, and activation of any of the second messenger pathways that regulate them. Spatial-temporal changes in the contractile, cytoskeletal or regulatory components of SM cells (SMCs) have been proposed to alter SM contractile activity. Ca(2+) sensitization/desensitization can occur as a result of changes at any of these levels, and specific pathways have been identified at all of these levels. Understanding when and how proteins can translocate within the cytoplasm, or to-and-from the plasmalemma and the cytoplasm to alter contractile activity is critical. Numerous studies have reported translocation of proteins associated with the adherens junction and G protein-coupled receptor activation pathways in isolated SMC systems. Specific examples of translocation of vinculin to and from the adherens junction and protein kinase C (PKC) and 17 kDa PKC-potentiated inhibitor of myosin light chain phosphatase (CPI-17) to and from the plasmalemma in isolated SMC systems but not in intact SM tissues are discussed. Using both isolated SMC systems and SM tissues in parallel to pursue these studies will advance our understanding of both the role and mechanism of these pathways as well as their possible significance for Ca(2+) sensitization in intact SM tissues and organ systems. © 2014 Wiley Periodicals, Inc.

A Special Connection between γδ T Cells and Natural Antibodies?

PubMed Central

Born, Willi K.; Huang, Yafei; Zeng, Wanjiang; Torres, Raul M.; O’Brien, Rebecca L.

2017-01-01

Natural antibodies (NAbs) play an important role in early host defense, autophagy and tissue remodeling, and in immune regulation. They arise spontaneously (without specific immunization), and are already present at birth. NAbs are produced by B1 B cells, MZ B cells and other B cell types. They include all major Ig subclasses but IgM antibodies are prevalent, especially early in development. NAbs may be poly-specific, recognize particular auto-antigens, or detect neo-determinants such as those exposed during apoptosis or generated by oxidation. NAbs do not require cognate T cell help but depend on soluble mediators produced by T cells. Our recent studies suggest that γδ T cells may have a special relationship with NAbs, and play a prominent role in their regulation, in part through the fine-tuning of IL-4-levels. The spontaneously activated state of these cells likely enables their cytokine production and other functions in the absence of external stimulation. Ontogenetically, the earlier arising γδ T cells are better positioned than αβ T cells to shape the developing repertoire of NAbs. Intriguingly, ligand specificities of NAbs and γδ T cell receptors appear to be overlapping, perhaps allowing γδ cognate help for certain NAb specificities. Via NAbs, γδ T cells could exert a regulatory influence on numerous processes in health and disease. PMID:27235134

A Special Connection between γδ T Cells and Natural Antibodies?

PubMed

Born, Willi K; Huang, Yafei; Zeng, Wanjiang; Torres, Raul M; O'Brien, Rebecca L

2016-12-01

Natural antibodies (NAbs) play an important role in early host defense, autophagy and tissue remodeling, and in immune regulation. They arise spontaneously (without specific immunization), and are already present at birth. NAbs are produced by B1 B cells, MZ B cells and other B cell types. They include all major Ig subclasses but IgM antibodies are prevalent, especially early in development. NAbs may be poly-specific, recognize particular auto-antigens, or detect neo-determinants such as those exposed during apoptosis or generated by oxidation. NAbs do not require cognate T cell help but depend on soluble mediators produced by T cells. Our recent studies suggest that γδ T cells may have a special relationship with NAbs, and play a prominent role in their regulation, in part through the fine-tuning of IL-4 levels. The spontaneously activated state of these cells likely enables their cytokine production and other functions in the absence of external stimulation. Ontogenetically, the earlier arising γδ T cells are better positioned than αβ T cells to shape the developing repertoire of NAbs. Intriguingly, ligand specificities of NAbs and γδ T cell receptors appear to be overlapping, perhaps allowing γδ cognate help for certain NAb specificities. Via NAbs, γδ T cells could exert a regulatory influence on numerous processes in health and disease.

In vivo Host Environment Alters Pseudomonas aeruginosa Susceptibility to Aminoglycoside Antibiotics

PubMed Central

Pan, Xiaolei; Dong, Yuanyuan; Fan, Zheng; Liu, Chang; Xia, Bin; Shi, Jing; Bai, Fang; Jin, Yongxin; Cheng, Zhihui; Jin, Shouguang; Wu, Weihui

2017-01-01

During host infection, Pseudomonas aeruginosa coordinately regulates the expression of numerous genes to adapt to the host environment while counteracting host clearance mechanisms. As infected patients take antibiotics, the invading bacteria encounter antibiotics in the host milieu. P. aeruginosa is highly resistant to antibiotics due to multiple chromosomally encoded resistant determinants. And numerous in vitro studies have demonstrated the regulatory mechanisms of antibiotic resistance related genes in response to antibiotics. However, it is not well-known how host environment affects bacterial response to antibiotics. In this study, we found that P. aeruginosa cells directly isolated from mice lungs displayed higher susceptibility to tobramycin than in vitro cultured bacteria. In vitro experiments demonstrated that incubation with A549 and differentiated HL60 (dHL60) cells sensitized P. aeruginosa to tobramycin. Further studies revealed that reactive oxygen species produced by the host cells contributed to the increased bacterial susceptibility. At the same concentration of tobramycin, presence of A549 and dHL60 cells resulted in higher expression of heat shock proteins, which are known inducible by tobramycin. Further analyses revealed decreased membrane potential upon incubation with the host cells and modification of lipopolysaccharide, which contributed to the increased susceptibility to tobramycin. Therefore, our results demonstrate that contact with host cells increased bacterial susceptibility to tobramycin. PMID:28352614

Upregulation of MicroRNA-4262 Targets Kaiso (ZBTB33) to Inhibit the Proliferation and EMT of Cervical Cancer Cells.

PubMed

Feng, Jing

2017-08-11

More and more studies have reported that dysregulation of microRNAs (miRNAs) lead to the proliferation and EMT of multiple cancers. Recently, several reports have demonstrated that dysregulation of miR-4262 is in numerous cancers. However, its role and precise mechanism in human cervical cancer (CC) have not been well clarified. Hence, my study was aim to explore the biological roles and precise mechanisms of miR-4262 in CC cell lines. In my study, I found that the level of miR-4262 is significantly decreased in CC tissues and cell lines. Moreover, decreased expression of miR-4262 was closely related to increased expression of Kaiso (ZBTB33) that belongs to the BTB/POZ family in CC tissues and cell lines. The proliferation and EMT of CC cells were inhibited by miR-4262 mimic. However, down-regulation of miR-4262 enhanced the proliferation and EMT of CC cells. Next, bioinformatics analysis predicted that miR-4262 might directly target the Kaiso gene. Besides, luciferase reporter assay had confirmed this result. Moreover, introduction of Kaiso in CC cells partially blocked the effects of miR-4262 mimic. In conclusion, miR-4262 suppressed the proliferation and EMT of CC cells by directly down-regulation of Kaiso.

Changes in global gene expression during in vitro decidualization of rat endometrial stromal cells

PubMed Central

Vallejo, Griselda; Maschi, Darío; Citrinovitz, Ana Cecilia Mestre; Aiba, Kazuhiro; Maronna, Ricardo; Yohai, Victor; Ko, Minoru S. H.; Beato, Miguel; Saragüeta, Patricia

2009-01-01

During the preimplantation phase of pregnancy the endometrial stroma differentiates into decidua, a process that implies numerous morphological changes and is an example of physiological transdifferentiation. Here we show that UIII rat endometrial stromal cells cultured in the presence of calf serum acquired morphological features of decidual cells and expressed decidual markers. To identify genes involved in decidualization we compared gene expression patterns of control and decidualized UIII cells using cDNA microarray. We found 322 annotated genes exhibiting significant differences in expression (>3 fold, FDR > 0.005), of which 312 have not been previously related to decidualization. Analysis of overrepresented functions revealed that protein synthesis, gene expression and chromatin architecture and remodeling are the most relevant modified functions during decidualization. Relevant genes are also found in the functional terms differentiation, cell proliferation, signal transduction, and matrix/structural proteins. Several of these new genes involved in decidualization (Csdc2, Trim27, Eef1a1, Bmp1, Wt1, Aes, Gna12, and Men1) are shown to be also regulated in uterine decidua during normal pregnancy. Thus, the UIII cell culture model will allow future mechanistic studies to define the transcriptional network regulating reprogramming of stromal cells into decidual cells. PMID:19780023

A Short-Term Advantage for Syngamy in the Origin of Eukaryotic Sex: Effects of Cell Fusion on Cell Cycle Duration and Other Effects Related to the Duration of the Cell Cycle-Relationship between Cell Growth Curve and the Optimal Size of the Species, and Circadian Cell Cycle in Photosynthetic Unicellular Organisms.

PubMed

Mancebo Quintana, J M; Mancebo Quintana, S

2012-01-01

The origin of sex is becoming a vexatious issue for Evolutionary Biology. Numerous hypotheses have been proposed, based on the genetic effects of sex, on trophic effects or on the formation of cysts and syncytia. Our approach addresses the change in cell cycle duration which would cause cell fusion. Several results are obtained through graphical and mathematical analysis and computer simulations. (1) In poor environments, cell fusion would be an advantageous strategy, as fusion between cells of different size shortens the cycle of the smaller cell (relative to the asexual cycle), and the majority of mergers would occur between cells of different sizes. (2) The easiest-to-evolve regulation of cell proliferation (sexual/asexual) would be by modifying the checkpoints of the cell cycle. (3) A regulation of this kind would have required the existence of the G2 phase, and sex could thus be the cause of the appearance of this phase. Regarding cell cycle, (4) the exponential curve is the only cell growth curve that has no effect on the optimal cell size in unicellular species; (5) the existence of a plateau with no growth at the end of the cell cycle explains the circadian cell cycle observed in unicellular algae.

A Short-Term Advantage for Syngamy in the Origin of Eukaryotic Sex: Effects of Cell Fusion on Cell Cycle Duration and Other Effects Related to the Duration of the Cell Cycle—Relationship between Cell Growth Curve and the Optimal Size of the Species, and Circadian Cell Cycle in Photosynthetic Unicellular Organisms

PubMed Central

Mancebo Quintana, J. M.; Mancebo Quintana, S.

2012-01-01

The origin of sex is becoming a vexatious issue for Evolutionary Biology. Numerous hypotheses have been proposed, based on the genetic effects of sex, on trophic effects or on the formation of cysts and syncytia. Our approach addresses the change in cell cycle duration which would cause cell fusion. Several results are obtained through graphical and mathematical analysis and computer simulations. (1) In poor environments, cell fusion would be an advantageous strategy, as fusion between cells of different size shortens the cycle of the smaller cell (relative to the asexual cycle), and the majority of mergers would occur between cells of different sizes. (2) The easiest-to-evolve regulation of cell proliferation (sexual/asexual) would be by modifying the checkpoints of the cell cycle. (3) A regulation of this kind would have required the existence of the G2 phase, and sex could thus be the cause of the appearance of this phase. Regarding cell cycle, (4) the exponential curve is the only cell growth curve that has no effect on the optimal cell size in unicellular species; (5) the existence of a plateau with no growth at the end of the cell cycle explains the circadian cell cycle observed in unicellular algae. PMID:22666626

Live-cell imaging of mammalian RNAs with Spinach2.

PubMed

Strack, Rita L; Jaffrey, Samie R

2015-01-01

The ability to monitor RNAs of interest in living cells is crucial to understanding the function, dynamics, and regulation of this important class of molecules. In recent years, numerous strategies have been developed with the goal of imaging individual RNAs of interest in living cells, each with their own advantages and limitations. This chapter provides an overview of current methods of live-cell RNA imaging, including a detailed discussion of genetically encoded strategies for labeling RNAs in mammalian cells. This chapter then focuses on the development and use of "RNA mimics of GFP" or Spinach technology for tagging mammalian RNAs and includes a detailed protocol for imaging 5S and CGG60 RNA with the recently described Spinach2 tag. © 2015 Elsevier Inc. All rights reserved.

The norepinephrine transporter and its regulation.

PubMed

Mandela, Prashant; Ordway, Gregory A

2006-04-01

For many years, the norepinephrine transporter (NET) was considered a 'static' protein that contributed to the termination of the action of norepinephrine in the synapse of noradrenergic neurons. The concept that the NET is dynamically regulated, adjusting noradrenergic transmission by changing its function and/or expression, was considered initially in the mid 1980s. Since that time, a plethora of studies demonstrate that the NET is regulated by several intracellular and extracellular signaling molecules, and that phosphorylation of the NET is a major pathway regulating its cell surface expression and thereby its function. The NET is a target of action of a number of drugs that are used long-term therapeutically or abused chronically. This has driven numerous investigations of how the NET and its function are regulated by long-term exposure to drugs. While repeated exposure to many drugs has been shown to affect NET function and expression, the intracellular mechanisms for these effects remains elusive.

Regulation of body growth by microRNAs.

PubMed

Lui, Julian C

2017-11-15

Regulation of body growth remains a fascinating and unresolved biological mystery. One key component of body growth is skeletal and longitudinal bone growth. Children grow taller because their bones grew longer, and the predominant driver of longitudinal bone growth is a cartilaginous structure found near the ends of long bone named the growth plate. Numerous recent studies have started to unveil the importance of microRNAs in regulation of growth plate functions, therefore contributing to regulation of linear growth. In addition to longitudinal growth, other organs in our body need to increase in size and cell number as we grow, and the regulation of organ growth involves both systemic factors like hormones; and other intrinsic mechanisms, which we are just beginning to understand. This review aims to summarize some recent important findings on how microRNAs are involved in both of these processes: the regulation of longitudinal bone growth, and the regulation of organs and overall body growth. Published by Elsevier B.V.

Energy management of fuel cell/solar cell/supercapacitor hybrid power source

NASA Astrophysics Data System (ADS)

Thounthong, Phatiphat; Chunkag, Viboon; Sethakul, Panarit; Sikkabut, Suwat; Pierfederici, Serge; Davat, Bernard

This study presents an original control algorithm for a hybrid energy system with a renewable energy source, namely, a polymer electrolyte membrane fuel cell (PEMFC) and a photovoltaic (PV) array. A single storage device, i.e., a supercapacitor (ultracapacitor) module, is in the proposed structure. The main weak point of fuel cells (FCs) is slow dynamics because the power slope is limited to prevent fuel starvation problems, improve performance and increase lifetime. The very fast power response and high specific power of a supercapacitor complements the slower power output of the main source to produce the compatibility and performance characteristics needed in a load. The energy in the system is balanced by d.c.-bus energy regulation (or indirect voltage regulation). A supercapacitor module functions by supplying energy to regulate the d.c.-bus energy. The fuel cell, as a slow dynamic source in this system, supplies energy to the supercapacitor module in order to keep it charged. The photovoltaic array assists the fuel cell during daytime. To verify the proposed principle, a hardware system is realized with analog circuits for the fuel cell, solar cell and supercapacitor current control loops, and with numerical calculation (dSPACE) for the energy control loops. Experimental results with small-scale devices, namely, a PEMFC (1200 W, 46 A) manufactured by the Ballard Power System Company, a photovoltaic array (800 W, 31 A) manufactured by the Ekarat Solar Company and a supercapacitor module (100 F, 32 V) manufactured by the Maxwell Technologies Company, illustrate the excellent energy-management scheme during load cycles.

EMMPRIN/CD147 up-regulates urokinase-type plasminogen activator: implications in oral tumor progression

PubMed Central

2012-01-01

Backgrounds An elevated level of EMMPRIN in cancer tissues have been correlated with tumor invasion in numerous cancers including oral cavity and larynx. Although EMMPRIN's effect has been generally attributed to its MMP inducing activity, we have previously demonstrated in breast cancer model that EMMPRIN can also enhance invasion by upregulating uPA. In this study, the role of EMMPRIN in regulating uPA and invasion was investigated in oral squamous cell carcinoma (OSCC) progression. Methods Precancerous and invasive oral tumoral tissues were used as well as the corresponding cell lines, DOK and SCC-9 respectively. The paracrine regulation of uPA by EMMPRIN was investigated by treating culture cells with EMMPRIN-enriched membrane vesicles. UPA expression was analyzed by qPCR and immunostaining and the consequence on the invasion capacity was studied using modified Boyden chamber assay, in the presence or absence of EMMPRIN blocking antibody, the uPA inhibitor amiloride or the MMP inhibitor marimastat. Results OSCC tumors were shown to express more EMMPRIN and uPA compared to dysplastic lesions. The corresponding cell models, SCC-9 and DOK cells, displayed similar expression pattern. In both cell types EMMPRIN upregulated the expression of uPA as well as that of MMP-2 and MMP-9. EMMPRIN treatment led to a significant increase in cell invasion both in the invasive SCC-9 and in the less invasive dysplastic DOK cells, in an MMP and uPA dependent manner. Conclusions Our results suggest that the upregulation of uPA contributes to EMMPRIN's effect in promoting oral tumor invasion. PMID:22443116

EMMPRIN/CD147 up-regulates urokinase-type plasminogen activator: implications in oral tumor progression.

PubMed

Lescaille, Géraldine; Menashi, Suzanne; Cavelier-Balloy, Bénédicte; Khayati, Farah; Quemener, Cathy; Podgorniak, Marie Pierre; Naïmi, Benyoussef; Calvo, Fabien; Lebbe, Céleste; Mourah, Samia

2012-03-23

An elevated level of EMMPRIN in cancer tissues have been correlated with tumor invasion in numerous cancers including oral cavity and larynx. Although EMMPRIN's effect has been generally attributed to its MMP inducing activity, we have previously demonstrated in breast cancer model that EMMPRIN can also enhance invasion by upregulating uPA. In this study, the role of EMMPRIN in regulating uPA and invasion was investigated in oral squamous cell carcinoma (OSCC) progression. Precancerous and invasive oral tumoral tissues were used as well as the corresponding cell lines, DOK and SCC-9 respectively. The paracrine regulation of uPA by EMMPRIN was investigated by treating culture cells with EMMPRIN-enriched membrane vesicles. UPA expression was analyzed by qPCR and immunostaining and the consequence on the invasion capacity was studied using modified Boyden chamber assay, in the presence or absence of EMMPRIN blocking antibody, the uPA inhibitor amiloride or the MMP inhibitor marimastat. OSCC tumors were shown to express more EMMPRIN and uPA compared to dysplastic lesions. The corresponding cell models, SCC-9 and DOK cells, displayed similar expression pattern. In both cell types EMMPRIN upregulated the expression of uPA as well as that of MMP-2 and MMP-9. EMMPRIN treatment led to a significant increase in cell invasion both in the invasive SCC-9 and in the less invasive dysplastic DOK cells, in an MMP and uPA dependent manner. Our results suggest that the upregulation of uPA contributes to EMMPRIN's effect in promoting oral tumor invasion.

Evolution‐development congruence in pattern formation dynamics: Bifurcations in gene expression and regulation of networks structures

PubMed Central

Kohsokabe, Takahiro

2016-01-01

ABSTRACT Search for possible relationships between phylogeny and ontogeny is important in evolutionary‐developmental biology. Here we uncover such relationships by numerical evolution and unveil their origin in terms of dynamical systems theory. By representing developmental dynamics of spatially located cells with gene expression dynamics with cell‐to‐cell interaction under external morphogen gradient, gene regulation networks are evolved under mutation and selection with the fitness to approach a prescribed spatial pattern of expressed genes. For most numerical evolution experiments, evolution of pattern over generations and development of pattern by an evolved network exhibit remarkable congruence. Both in the evolution and development pattern changes consist of several epochs where stripes are formed in a short time, while for other temporal regimes, pattern hardly changes. In evolution, these quasi‐stationary regimes are generations needed to hit relevant mutations, while in development, they are due to some gene expression that varies slowly and controls the pattern change. The morphogenesis is regulated by combinations of feedback or feedforward regulations, where the upstream feedforward network reads the external morphogen gradient, and generates a pattern used as a boundary condition for the later patterns. The ordering from up to downstream is common in evolution and development, while the successive epochal changes in development and evolution are represented as common bifurcations in dynamical‐systems theory, which lead to the evolution‐development congruence. Mechanism of exceptional violation of the congruence is also unveiled. Our results provide a new look on developmental stages, punctuated equilibrium, developmental bottlenecks, and evolutionary acquisition of novelty in morphogenesis. J. Exp. Zool. (Mol. Dev. Evol.) 326B:61–84, 2016. © 2015 The Authors. Journal of Experimental Zoology Part B: Molecular and Developmental Evolution Published by Wiley Periodicals, Inc. PMID:26678220

Structural correlates of the creatine transporter function regulation: the undiscovered country.

PubMed

Santacruz, Lucia; Jacobs, Danny O

2016-08-01

Creatine (Cr) and phosphocreatine constitute an energy shuttle that links ATP production in mitochondria to subcellular locations of ATP consumption. Cells in tissues that are reliant on this energy shuttle, such as myocytes and neurons, appear to have very limited ability to synthesize creatine. Therefore, these cells depend on Cr uptake across the cell membrane by a specialized creatine transporter (CrT solute carrier SLC6A8) in order to maintain intracellular creatine levels. Cr supplementation has been shown to have a beneficial effect in numerous in vitro and in vivo models, particularly in cases of oxidative stress, and is also widely used by athletes as a performance enhancement nutraceutical. Intracellular creatine content is maintained within narrow limits. However, the physiological and cellular mechanisms that mediate Cr transport during health and disease (such as cardiac failure) are not understood. In this narrative mini-review, we summarize the last three decades of research on CrT structure, function and regulation.

RNA search engines empower the bacterial intranet.

PubMed

Dendooven, Tom; Luisi, Ben F

2017-08-15

RNA acts not only as an information bearer in the biogenesis of proteins from genes, but also as a regulator that participates in the control of gene expression. In bacteria, small RNA molecules (sRNAs) play controlling roles in numerous processes and help to orchestrate complex regulatory networks. Such processes include cell growth and development, response to stress and metabolic change, transcription termination, cell-to-cell communication, and the launching of programmes for host invasion. All these processes require recognition of target messenger RNAs by the sRNAs. This review summarizes recent results that have provided insights into how bacterial sRNAs are recruited into effector ribonucleoprotein complexes that can seek out and act upon target transcripts. The results hint at how sRNAs and their protein partners act as pattern-matching search engines that efficaciously regulate gene expression, by performing with specificity and speed while avoiding off-target effects. The requirements for efficient searches of RNA patterns appear to be common to all domains of life. © 2017 The Author(s).

Angiopoietin–Tie signalling in the cardiovascular and lymphatic systems

PubMed Central

Eklund, Lauri; Kangas, Jaakko; Saharinen, Pipsa

2016-01-01

Endothelial cells that form the inner layer of blood and lymphatic vessels are important regulators of vascular functions and centrally involved in the pathogenesis of vascular diseases. In addition to the vascular endothelial growth factor (VEGF) receptor pathway, the angiopoietin (Ang)–Tie system is a second endothelial cell specific ligand–receptor signalling system necessary for embryonic cardiovascular and lymphatic development. The Ang–Tie system also regulates postnatal angiogenesis, vessel remodelling, vascular permeability and inflammation to maintain vascular homoeostasis in adult physiology. This system is implicated in numerous diseases where the vasculature has an important contribution, such as cancer, sepsis, diabetes, atherosclerosis and ocular diseases. Furthermore, mutations in the TIE2 signalling pathway cause defects in vascular morphogenesis, resulting in venous malformations and primary congenital glaucoma. Here, we review recent advances in the understanding of the Ang–Tie signalling system, including cross-talk with the vascular endothelial protein tyrosine phosphatase (VE-PTP) and the integrin cell adhesion receptors, focusing on the Ang–Tie system in vascular development and pathogenesis of vascular diseases. PMID:27941161

RNA search engines empower the bacterial intranet

PubMed Central

Dendooven, Tom

2017-01-01

RNA acts not only as an information bearer in the biogenesis of proteins from genes, but also as a regulator that participates in the control of gene expression. In bacteria, small RNA molecules (sRNAs) play controlling roles in numerous processes and help to orchestrate complex regulatory networks. Such processes include cell growth and development, response to stress and metabolic change, transcription termination, cell-to-cell communication, and the launching of programmes for host invasion. All these processes require recognition of target messenger RNAs by the sRNAs. This review summarizes recent results that have provided insights into how bacterial sRNAs are recruited into effector ribonucleoprotein complexes that can seek out and act upon target transcripts. The results hint at how sRNAs and their protein partners act as pattern-matching search engines that efficaciously regulate gene expression, by performing with specificity and speed while avoiding off-target effects. The requirements for efficient searches of RNA patterns appear to be common to all domains of life. PMID:28710287

The mTOR signalling pathway in cancer and the potential mTOR inhibitory activities of natural phytochemicals.

PubMed

Tan, Heng Kean; Moad, Ahmed Ismail Hassan; Tan, Mei Lan

2014-01-01

The mammalian target of rapamycin (mTOR) kinase plays an important role in regulating cell growth and cell cycle progression in response to cellular signals. It is a key regulator of cell proliferation and many upstream activators and downstream effectors of mTOR are known to be deregulated in various types of cancers. Since the mTOR signalling pathway is commonly activated in human cancers, many researchers are actively developing inhibitors that target key components in the pathway and some of these drugs are already on the market. Numerous preclinical investigations have also suggested that some herbs and natural phytochemicals, such as curcumin, resveratrol, timosaponin III, gallic acid, diosgenin, pomegranate, epigallocatechin gallate (EGCC), genistein and 3,3'-diindolylmethane inhibit the mTOR pathway either directly or indirectly. Some of these natural compounds are also in the clinical trial stage. In this review, the potential anti-cancer and chemopreventive activities and the current status of clinical trials of these phytochemicals are discussed.

Cranial neural crest recycle surface integrins in a substratum-dependent manner to promote rapid motility.

PubMed

Strachan, Lauren R; Condic, Maureen L

2004-11-08

Cell migration is essential for proper development of numerous structures derived from embryonic neural crest cells (NCCs). Although the migratory pathways of NCCs have been determined, the molecular mechanisms regulating NCC motility remain unclear. NCC migration is integrin dependent, and recent work has shown that surface expression levels of particular integrin alpha subunits are important determinants of NCC motility in vitro. Here, we provide evidence that rapid cranial NCC motility on laminin requires integrin recycling. NCCs showed both ligand- and receptor-specific integrin regulation in vitro. On laminin, NCCs accumulated internalized laminin but not fibronectin receptors over 20 min, whereas on fibronectin neither type of receptor accumulated internally beyond 2 min. Internalized laminin receptors colocalized with receptor recycling vesicles and were subsequently recycled back to the cell surface. Blocking receptor recycling with bafilomycin A inhibited NCC motility on laminin, indicating that substratum-dependent integrin recycling is essential for rapid cranial neural crest migration.

Plasmodesmata: channels for intercellular signaling during plant growth and development.

PubMed

Sevilem, Iris; Yadav, Shri Ram; Helariutta, Ykä

2015-01-01

Plants have evolved strategies for short- and long-distance communication to coordinate plant development and to adapt to changing environmental conditions. Plasmodesmata (PD) are intercellular nanochannels that provide an effective pathway for both selective and nonselective movement of various molecules that function in diverse biological processes. Numerous non-cell-autonomous proteins (NCAP) and small RNAs have been identified that have crucial roles in cell fate determination and organ patterning during development. Both the density and aperture size of PD are developmentally regulated, allowing formation of spatial symplastic domains for establishment of tissue-specific developmental programs. The PD size exclusion limit (SEL) is controlled by reversible deposition of callose, as well as by some PD-associated proteins. Although a large number of PD-associated proteins have been identified, many of their functions remain unknown. Despite the fact that PD are primarily membranous structures, surprisingly very little is known about their lipid composition. Thus, future studies in PD biology will provide deeper insights into the high-resolution structure and tightly regulated functions of PD and the evolution of PD-mediated cell-to-cell communication in plants.

Carbon nanotube multilayered nanocomposites as multifunctional substrates for actuating neuronal differentiation and functions of neural stem cells.

PubMed

Shao, Han; Li, Tingting; Zhu, Rong; Xu, Xiaoting; Yu, Jiandong; Chen, Shengfeng; Song, Li; Ramakrishna, Seeram; Lei, Zhigang; Ruan, Yiwen; He, Liumin

2018-08-01

Carbon nanotubes (CNTs) have shown potential applications in neuroscience as growth substrates owing to their numerous unique properties. However, a key concern in the fabrication of homogeneous composites is the serious aggregation of CNTs during incorporation into the biomaterial matrix. Moreover, the regulation mechanism of CNT-based substrates on neural differentiation remains unclear. Here, a novel strategy was introduced for the construction of CNT nanocomposites via layer-by-layer assembly of negatively charged multi-walled CNTs and positively charged poly(dimethyldiallylammonium chloride). Results demonstrated that the CNT-multilayered nanocomposites provided a potent regulatory signal over neural stem cells (NSCs), including cell adhesion, viability, differentiation, neurite outgrowth, and electrophysiological maturation of NSC-derived neurons. Importantly, the dynamic molecular mechanisms in the NSC differentiation involved the integrin-mediated interactions between NSCs and CNT multilayers, thereby activating focal adhesion kinase, subsequently triggering downstream signaling events to regulate neuronal differentiation and synapse formation. This study provided insights for future applications of CNT-multilayered nanomaterials in neural fields as potent modulators of stem cell behavior. Copyright © 2018 Elsevier Ltd. All rights reserved.

Interleukin-22: immunobiology and pathology

PubMed Central

Dudakov, Jarrod A.; Hanash, Alan M.; van den Brink, Marcel R.M.

2015-01-01

Interleukin-22 (IL-22) is a recently described IL-10 family cytokine that is produced by T-helper (Th)-17 cells, γδ T cells, NKT cells and newly described innate lymphoid cells (ILCs). Knowledge of IL-22 biology has rapidly evolved since its discovery in 2000, and a role for IL-22 has been identified in numerous tissues including the intestines, lung, liver, kidney, thymus, pancreas and skin. IL-22 primarily targets non-hematopoietic epithelial and stromal cells where it can promote proliferation and play a role in tissue regeneration. In addition, IL-22 regulates host defense at barrier surfaces. However, IL-22 has also been linked to several conditions involving inflammatory tissue pathology. In this review, we will assess the current understanding of this cytokine, including its physiologic and pathologic effects on epithelial cell function. PMID:25706098

ICAM-1-expressing neutrophils exhibit enhanced effector functions in murine models of endotoxemia.

PubMed

Woodfin, Abigail; Beyrau, Martina; Voisin, Mathieu-Benoit; Ma, Bin; Whiteford, James R; Hordijk, Peter L; Hogg, Nancy; Nourshargh, Sussan

2016-02-18

Intracellular adhesion molecule-1 (ICAM-1) is a transmembrane glycoprotein expressed on the cell surface of numerous cell types such as endothelial and epithelial cells, vascular smooth muscle cells, and certain leukocyte subsets. With respect to the latter, ICAM-1 has been detected on neutrophils in several clinical and experimental settings, but little is known about the regulation of expression or function of neutrophil ICAM-1. In this study, we report on the de novo induction of ICAM-1 on the cell surface of murine neutrophils by lipopolysaccharide (LPS), tumor necrosis factor, and zymosan particles in vitro. The induction of neutrophil ICAM-1 was associated with enhanced phagocytosis of zymosan particles and reactive oxygen species (ROS) generation. Conversely, neutrophils from ICAM-1-deficient mice were defective in these effector functions. Mechanistically, ICAM-1-mediated intracellular signaling appeared to support neutrophil ROS generation and phagocytosis. In vivo, LPS-induced inflammation in the mouse cremaster muscle and peritoneal cavity led to ICAM-1 expression on intravascular and locally transmigrated neutrophils. The use of chimeric mice deficient in ICAM-1 on myeloid cells demonstrated that neutrophil ICAM-1 was not required for local neutrophil transmigration, but supported optimal intravascular and extravascular phagocytosis of zymosan particles. Collectively, the present results shed light on regulation of expression and function of ICAM-1 on neutrophils and identify it as an additional regulator of neutrophil effector responses in host defense. © 2016 by The American Society of Hematology.

Gastric peptides and their regulation of hunger and satiety.

PubMed

Stengel, Andreas; Taché, Yvette

2012-12-01

Ingestion of food affects the secretion of hormones from specialized endocrine cells scattered within the intestinal mucosa. Upon release, these hormones mostly decrease food intake by signaling information to the brain. Although enteroendocrine cells in the small intestine were thought to represent the predominant gut-brain regulators of food intake, recent advances also established a major role for gastric hormones in these regulatory pathways. First and foremost, the gastric endocrine X/A-like cell was in the focus of many studies due to the production of ghrelin, which is until now the only known orexigenic hormone that is peripherally produced and centrally acting. Although X/A-cells were initially thought to only release one hormone that stimulates food intake, this view has changed with the identification of additional peptide products also derived from this cell, namely desacyl ghrelin, obestatin, and nesfatin-1. Desacyl ghrelin may play a counter-regulatory role to the food intake stimulatory effect of ghrelin. The same property was suggested for obestatin; however, this hypothesis could not be confirmed in numerous subsequent studies. Moreover, the description of the stomach as the major source of the novel anorexigenic hormone nesfatin-1 derived from the NUCB2 gene further corroborated the assumption that the gastric X/A-like cell products are not only stimulant but also inhibitors of feeding, thereby acting as so far unique dual regulator of food intake located in a logistically important place where the gastrointestinal tract has initial contact with food.

A discrete in continuous mathematical model of cardiac progenitor cells formation and growth as spheroid clusters (Cardiospheres).

PubMed

Di Costanzo, Ezio; Giacomello, Alessandro; Messina, Elisa; Natalini, Roberto; Pontrelli, Giuseppe; Rossi, Fabrizio; Smits, Robert; Twarogowska, Monika

2018-03-14

We propose a discrete in continuous mathematical model describing the in vitro growth process of biophsy-derived mammalian cardiac progenitor cells growing as clusters in the form of spheres (Cardiospheres). The approach is hybrid: discrete at cellular scale and continuous at molecular level. In the present model, cells are subject to the self-organizing collective dynamics mechanism and, additionally, they can proliferate and differentiate, also depending on stochastic processes. The two latter processes are triggered and regulated by chemical signals present in the environment. Numerical simulations show the structure and the development of the clustered progenitors and are in a good agreement with the results obtained from in vitro experiments.

MiR-34a regulates the invasive capacity of canine osteosarcoma cell lines

PubMed Central

Lopez, Cecilia M.; Yu, Peter Y.; Zhang, Xiaoli; Yilmaz, Ayse Selen; London, Cheryl A.

2018-01-01

Background Osteosarcoma (OSA) is the most common bone tumor in children and dogs; however, no substantial improvement in clinical outcome has occurred in either species over the past 30 years. MicroRNAs (miRNAs) are small non-coding RNAs that regulate gene expression and play a fundamental role in cancer. The purpose of this study was to investigate the potential contribution of miR-34a loss to the biology of canine OSA, a well-established spontaneous model of the human disease. Methodology and principal findings RT-qPCR demonstrated that miR-34a expression levels were significantly reduced in primary canine OSA tumors and canine OSA cell lines as compared to normal canine osteoblasts. In canine OSA cell lines stably transduced with empty vector or pre-miR-34a lentiviral constructs, overexpression of miR-34a inhibited cellular invasion and migration but had no effect on cell proliferation or cell cycle distribution. Transcriptional profiling of canine OSA8 cells possessing enforced miR-34a expression demonstrated dysregulation of numerous genes, including significant down-regulation of multiple putative targets of miR-34a. Moreover, gene ontology analysis of down-regulated miR-34a target genes showed enrichment of several biological processes related to cell invasion and motility. Lastly, we validated changes in miR-34a putative target gene expression, including decreased expression of KLF4, SEM3A, and VEGFA transcripts in canine OSA cells overexpressing miR-34a and identified KLF4 and VEGFA as direct target genes of miR-34a. Concordant with these data, primary canine OSA tumor tissues demonstrated increased expression levels of putative miR-34a target genes. Conclusions These data demonstrate that miR-34a contributes to invasion and migration in canine OSA cells and suggest that loss of miR-34a may promote a pattern of gene expression contributing to the metastatic phenotype in canine OSA. PMID:29293555

MiR-34a regulates the invasive capacity of canine osteosarcoma cell lines.

PubMed

Lopez, Cecilia M; Yu, Peter Y; Zhang, Xiaoli; Yilmaz, Ayse Selen; London, Cheryl A; Fenger, Joelle M

2018-01-01

Osteosarcoma (OSA) is the most common bone tumor in children and dogs; however, no substantial improvement in clinical outcome has occurred in either species over the past 30 years. MicroRNAs (miRNAs) are small non-coding RNAs that regulate gene expression and play a fundamental role in cancer. The purpose of this study was to investigate the potential contribution of miR-34a loss to the biology of canine OSA, a well-established spontaneous model of the human disease. RT-qPCR demonstrated that miR-34a expression levels were significantly reduced in primary canine OSA tumors and canine OSA cell lines as compared to normal canine osteoblasts. In canine OSA cell lines stably transduced with empty vector or pre-miR-34a lentiviral constructs, overexpression of miR-34a inhibited cellular invasion and migration but had no effect on cell proliferation or cell cycle distribution. Transcriptional profiling of canine OSA8 cells possessing enforced miR-34a expression demonstrated dysregulation of numerous genes, including significant down-regulation of multiple putative targets of miR-34a. Moreover, gene ontology analysis of down-regulated miR-34a target genes showed enrichment of several biological processes related to cell invasion and motility. Lastly, we validated changes in miR-34a putative target gene expression, including decreased expression of KLF4, SEM3A, and VEGFA transcripts in canine OSA cells overexpressing miR-34a and identified KLF4 and VEGFA as direct target genes of miR-34a. Concordant with these data, primary canine OSA tumor tissues demonstrated increased expression levels of putative miR-34a target genes. These data demonstrate that miR-34a contributes to invasion and migration in canine OSA cells and suggest that loss of miR-34a may promote a pattern of gene expression contributing to the metastatic phenotype in canine OSA.

NUDT2 Disruption Elevates Diadenosine Tetraphosphate (Ap4A) and Down-Regulates Immune Response and Cancer Promotion Genes

PubMed Central

Marriott, Andrew S.; Vasieva, Olga; Fang, Yongxiang; Copeland, Nikki A.; McLennan, Alexander G.; Jones, Nigel J.

2016-01-01

Regulation of gene expression is one of several roles proposed for the stress-induced nucleotide diadenosine tetraphosphate (Ap4A). We have examined this directly by a comparative RNA-Seq analysis of KBM-7 chronic myelogenous leukemia cells and KBM-7 cells in which the NUDT2 Ap4A hydrolase gene had been disrupted (NuKO cells), causing a 175-fold increase in intracellular Ap4A. 6,288 differentially expressed genes were identified with P < 0.05. Of these, 980 were up-regulated and 705 down-regulated in NuKO cells with a fold-change ≥ 2. Ingenuity® Pathway Analysis (IPA®) was used to assign these genes to known canonical pathways and functional networks. Pathways associated with interferon responses, pattern recognition receptors and inflammation scored highly in the down-regulated set of genes while functions associated with MHC class II antigens were prominent among the up-regulated genes, which otherwise showed little organization into major functional gene sets. Tryptophan catabolism was also strongly down-regulated as were numerous genes known to be involved in tumor promotion in other systems, with roles in the epithelial-mesenchymal transition, proliferation, invasion and metastasis. Conversely, some pro-apoptotic genes were up-regulated. Major upstream factors predicted by IPA® for gene down-regulation included NFκB, STAT1/2, IRF3/4 and SP1 but no major factors controlling gene up-regulation were identified. Potential mechanisms for gene regulation mediated by Ap4A and/or NUDT2 disruption include binding of Ap4A to the HINT1 co-repressor, autocrine activation of purinoceptors by Ap4A, chromatin remodeling, effects of NUDT2 loss on transcript stability, and inhibition of ATP-dependent regulatory factors such as protein kinases by Ap4A. Existing evidence favors the last of these as the most probable mechanism. Regardless, our results suggest that the NUDT2 protein could be a novel cancer chemotherapeutic target, with its inhibition potentially exerting strong anti-tumor effects via multiple pathways involving metastasis, invasion, immunosuppression and apoptosis. PMID:27144453

NUDT2 Disruption Elevates Diadenosine Tetraphosphate (Ap4A) and Down-Regulates Immune Response and Cancer Promotion Genes.

PubMed

Marriott, Andrew S; Vasieva, Olga; Fang, Yongxiang; Copeland, Nikki A; McLennan, Alexander G; Jones, Nigel J

2016-01-01

Regulation of gene expression is one of several roles proposed for the stress-induced nucleotide diadenosine tetraphosphate (Ap4A). We have examined this directly by a comparative RNA-Seq analysis of KBM-7 chronic myelogenous leukemia cells and KBM-7 cells in which the NUDT2 Ap4A hydrolase gene had been disrupted (NuKO cells), causing a 175-fold increase in intracellular Ap4A. 6,288 differentially expressed genes were identified with P < 0.05. Of these, 980 were up-regulated and 705 down-regulated in NuKO cells with a fold-change ≥ 2. Ingenuity® Pathway Analysis (IPA®) was used to assign these genes to known canonical pathways and functional networks. Pathways associated with interferon responses, pattern recognition receptors and inflammation scored highly in the down-regulated set of genes while functions associated with MHC class II antigens were prominent among the up-regulated genes, which otherwise showed little organization into major functional gene sets. Tryptophan catabolism was also strongly down-regulated as were numerous genes known to be involved in tumor promotion in other systems, with roles in the epithelial-mesenchymal transition, proliferation, invasion and metastasis. Conversely, some pro-apoptotic genes were up-regulated. Major upstream factors predicted by IPA® for gene down-regulation included NFκB, STAT1/2, IRF3/4 and SP1 but no major factors controlling gene up-regulation were identified. Potential mechanisms for gene regulation mediated by Ap4A and/or NUDT2 disruption include binding of Ap4A to the HINT1 co-repressor, autocrine activation of purinoceptors by Ap4A, chromatin remodeling, effects of NUDT2 loss on transcript stability, and inhibition of ATP-dependent regulatory factors such as protein kinases by Ap4A. Existing evidence favors the last of these as the most probable mechanism. Regardless, our results suggest that the NUDT2 protein could be a novel cancer chemotherapeutic target, with its inhibition potentially exerting strong anti-tumor effects via multiple pathways involving metastasis, invasion, immunosuppression and apoptosis.

Kynurenine promotes the goblet cell differentiation of HT-29 colon carcinoma cells by modulating Wnt, Notch and AhR signals.

PubMed

Park, Joo-Hung; Lee, Jeong-Min; Lee, Eun-Jin; Kim, Da-Jeong; Hwang, Won-Bhin

2018-04-01

Various amino acids regulate cell growth and differentiation. In the present study, we examined the ability of HT-29 cells to differentiate into goblet cells in RPMI and DMEM which are largely different in the amounts of numerous amino acids. Most of the HT-29 cells differentiated into goblet cells downregulating the stem cell marker Lgr5 when cultured in DMEM, but remained undifferentiated in RPMI. The goblet cell differentiation in DMEM was inhibited by 1-methyl-tryptophan (1-MT), an inhibitor of indoleamine 2,3 dioxygenase-1 which is the initial enzyme in tryptophan metabolism along the kynurenine (KN) pathway, whereas tryptophan and KN induced goblet cell differentiation in RPMI. The levels of Notch1 and its activation product Notch intracytoplasmic domain in HT-29 cells were lower in DMEM than those in RPMI and were increased by 1-MT in both media. HT-29 cells grown in both media expressed β-catenin at the same level on day 2 when goblet cell differentiation was not observed. β-catenin expression, which was increased by 1-MT in both media, was decreased by KN. DMEM reduced Hes1 expression while enhancing Hath1 expression. Finally, aryl hydrocarbon receptor (AhR) activation moderately induced goblet cell differentiation. Our results suggest that KN promotes goblet cell differentiation by regulating Wnt, Notch, and AhR signals and expression of Hes1 and Hath1.

Regulation of cellular function via electromagnetic field frequency and extracellular environment: A theoretical- experimental approach

NASA Astrophysics Data System (ADS)

Taghian, Toloo; Sheikh, Abdul; Narmoneva, Daria; Kogan, Andrei

2015-03-01

Application of external electric field (EF) as a non-pharmacological, non-invasive tool to control cell function is of great therapeutic interest. We developed a theoretical-experimental approach to investigate the biophysical mechanisms of EF interaction with cells in electrode-free physiologically-relevant configuration. Our numerical results demonstrated that EF frequency is the major parameter to control cell response to EF. Non-oscillating or low-frequency EF leads to charge accumulation on the cell surface membrane that may mediate membrane initiated cell responses. In contrast, high-frequency EF penetrates the cell membrane and reaches cell cytoplasm, where it may directly activate intracellular responses. The theoretical predictions were confirmed in our experimental studies of the effects of applied EF on vascular cell function. Results show that non-oscillating EF increases vascular endothelial growth factor (VEGF) expression while field polarity controls cell adhesion rate. High-frequency, but not low frequency, EF provides differential regulation of cytoplasmic focal adhesion kinase and VEGF expression depending on the substrate, with increased expression in cells cultured on RGD-rich synthetic hydrogels, and decreased expression for matrigel culture. The authors acknowledge the financial support from the NSF (DMR-1206784 & DMR-0804199 to AK); the NIH (1R21 DK078814-01A1 to DN) and the University of Cincinnati (Interdisciplinary Faculty Research Support Grant to DN and AK).

Reduction of CD147 surface expression on primary T cells leads to enhanced cell proliferation.

PubMed

Biegler, Brian; Kasinrerk, Watchara

2012-12-01

CD147 is a ubiquitously expressed membrane glycoprotein that has numerous functional associations in health and disease. However, the molecular mechanisms by which CD147 participates in these processes are unclear. Establishing physiologically relevant silencing of CD147 in primary T cells could provide clues essential for elucidating some aspects of CD147 biology. To date, achieving the knockdown of CD147 in primary T cells has remained elusive. Utilizing RNA interference and the Nucleofector transfection system, we were able to reduce the expression of CD147 in primary T cells. Comparison of basic functions, such as proliferation and CD25 expression, were then made between control populations and populations with reduced expression. Up-regulation of CD147 was found upon T-cell activation, indicating a role in T-cell responses. To better understand the possible importance of this up-regulation, we knocked down the expression of CD147 using RNA interference. When compared to control populations the CD147 knockdown populations exhibited increased proliferation. This alteration of cell proliferation, however, was not linked to a change in CD25 expression. We achieved reduction of CD147 surface expression in primary T cells by siRNA-mediated gene silencing. Our results point to CD147 having a possible negative regulatory role in T cell-mediated immune responses.

Extracellular calcium sensing and extracellular calcium signaling

NASA Technical Reports Server (NTRS)

Brown, E. M.; MacLeod, R. J.; O'Malley, B. W. (Principal Investigator)

2001-01-01

The cloning of a G protein-coupled extracellular Ca(2+) (Ca(o)(2+))-sensing receptor (CaR) has elucidated the molecular basis for many of the previously recognized effects of Ca(o)(2+) on tissues that maintain systemic Ca(o)(2+) homeostasis, especially parathyroid chief cells and several cells in the kidney. The availability of the cloned CaR enabled the development of DNA and antibody probes for identifying the CaR's mRNA and protein, respectively, within these and other tissues. It also permitted the identification of human diseases resulting from inactivating or activating mutations of the CaR gene and the subsequent generation of mice with targeted disruption of the CaR gene. The characteristic alterations in parathyroid and renal function in these patients and in the mice with "knockout" of the CaR gene have provided valuable information on the CaR's physiological roles in these tissues participating in mineral ion homeostasis. Nevertheless, relatively little is known about how the CaR regulates other tissues involved in systemic Ca(o)(2+) homeostasis, particularly bone and intestine. Moreover, there is evidence that additional Ca(o)(2+) sensors may exist in bone cells that mediate some or even all of the known effects of Ca(o)(2+) on these cells. Even more remains to be learned about the CaR's function in the rapidly growing list of cells that express it but are uninvolved in systemic Ca(o)(2+) metabolism. Available data suggest that the receptor serves numerous roles outside of systemic mineral ion homeostasis, ranging from the regulation of hormonal secretion and the activities of various ion channels to the longer term control of gene expression, programmed cell death (apoptosis), and cellular proliferation. In some cases, the CaR on these "nonhomeostatic" cells responds to local changes in Ca(o)(2+) taking place within compartments of the extracellular fluid (ECF) that communicate with the outside environment (e.g., the gastrointestinal tract). In others, localized changes in Ca(o)(2+) within the ECF can originate from several mechanisms, including fluxes of calcium ions into or out of cellular or extracellular stores or across epithelium that absorb or secrete Ca(2+). In any event, the CaR and other receptors/sensors for Ca(o)(2+) and probably for other extracellular ions represent versatile regulators of numerous cellular functions and may serve as important therapeutic targets.

Recent Progress in Understanding Subtype Specific Regulation of NMDA Receptors by G Protein Coupled Receptors (GPCRs)

PubMed Central

Yang, Kai; Jackson, Michael F.; MacDonald, John F.

2014-01-01

G Protein Coupled Receptors (GPCRs) are the largest family of receptors whose ligands constitute nearly a third of prescription drugs in the market. They are widely involved in diverse physiological functions including learning and memory. NMDA receptors (NMDARs), which belong to the ionotropic glutamate receptor family, are likewise ubiquitously expressed in the central nervous system (CNS) and play a pivotal role in learning and memory. Despite its critical contribution to physiological and pathophysiological processes, few pharmacological interventions aimed directly at regulating NMDAR function have been developed to date. However, it is well established that NMDAR function is precisely regulated by cellular signalling cascades recruited downstream of G protein coupled receptor (GPCR) stimulation. Accordingly, the downstream regulation of NMDARs likely represents an important determinant of outcome following treatment with neuropsychiatric agents that target selected GPCRs. Importantly, the functional consequence of such regulation on NMDAR function varies, based not only on the identity of the GPCR, but also on the cell type in which relevant receptors are expressed. Indeed, the mechanisms responsible for regulating NMDARs by GPCRs involve numerous intracellular signalling molecules and regulatory proteins that vary from one cell type to another. In the present article, we highlight recent findings from studies that have uncovered novel mechanisms by which selected GPCRs regulate NMDAR function and consequently NMDAR-dependent plasticity. PMID:24562329

Increased Bone Mass in Female Mice Lacking Mast Cell Chymase

PubMed Central

Lind, Thomas; Gustafson, Ann-Marie; Calounova, Gabriela; Hu, Lijuan; Rasmusson, Annica; Jonsson, Kenneth B.; Wernersson, Sara; Åbrink, Magnus; Andersson, Göran; Larsson, Sune; Melhus, Håkan; Pejler, Gunnar

2016-01-01

Here we addressed the potential impact of chymase, a mast-cell restricted protease, on mouse bone phenotype. We show that female mice lacking the chymase Mcpt4 acquired a persistent expansion of diaphyseal bone in comparison with wild type controls, reaching a 15% larger diaphyseal cross sectional area at 12 months of age. Mcpt4-/- mice also showed increased levels of a bone anabolic serum marker and higher periosteal bone formation rate. However, they were not protected from experimental osteoporosis, suggesting that chymase regulates normal bone homeostasis rather than the course of osteoporosis. Further, the absence of Mcpt4 resulted in age-dependent upregulation of numerous genes important for bone formation but no effects on osteoclast activity. In spite of the latter, Mcpt4-/- bones had increased cortical porosity and reduced endocortical mineralization. Mast cells were found periosteally and, notably, bone-proximal mast cells in Mcpt4-/- mice were degranulated to a larger extent than in wild type mice. Hence, chymase regulates degranulation of bone mast cells, which could affect the release of mast cell-derived factors influencing bone remodelling. Together, these findings reveal a functional impact of mast cell chymase on bone. Further studies exploring the possibility of using chymase inhibitors as a strategy to increase bone volume may be warranted. PMID:27936149

Peroxisomes in Different Skeletal Cell Types during Intramembranous and Endochondral Ossification and Their Regulation during Osteoblast Differentiation by Distinct Peroxisome Proliferator-Activated Receptors

PubMed Central

Qian, Guofeng; Karnati, Srikanth; Baumgart-Vogt, Eveline

2015-01-01

Ossification defects leading to craniofacial dysmorphism or rhizomelia are typical phenotypes in patients and corresponding knockout mouse models with distinct peroxisomal disorders. Despite these obvious skeletal pathologies, to date no careful analysis exists on the distribution and function of peroxisomes in skeletal tissues and their alterations during ossification. Therefore, we analyzed the peroxisomal compartment in different cell types of mouse cartilage and bone as well as in primary cultures of calvarial osteoblasts. The peroxisome number and metabolism strongly increased in chondrocytes during endochondral ossification from the reserve to the hypertrophic zone, whereas in bone, metabolically active osteoblasts contained a higher numerical abundance of this organelle than osteocytes. The high abundance of peroxisomes in these skeletal cell types is reflected by high levels of Pex11β gene expression. During culture, calvarial pre-osteoblasts differentiated into secretory osteoblasts accompanied by peroxisome proliferation and increased levels of peroxisomal genes and proteins. Since many peroxisomal genes contain a PPAR-responsive element, we analyzed the gene expression of PPARɑ/ß/ɣ in calvarial osteoblasts and MC3T3-E1 cells, revealing higher levels for PPARß than for PPARɑ and PPARɣ. Treatment with different PPAR agonists and antagonists not only changed the peroxisomal compartment and associated gene expression, but also induced complex alterations of the gene expression patterns of the other PPAR family members. Studies in M3CT3-E1 cells showed that the PPARß agonist GW0742 activated the PPRE-mediated luciferase expression and up-regulated peroxisomal gene transcription (Pex11, Pex13, Pex14, Acox1 and Cat), whereas the PPARß antagonist GSK0660 led to repression of the PPRE and a decrease of the corresponding mRNA levels. In the same way, treatment of calvarial osteoblasts with GW0742 increased in peroxisome number and related gene expression and accelerated osteoblast differentiation. Taken together, our results suggest that PPARß regulates the numerical abundance and metabolic function of peroxisomes via Pex11ß in parallel to osteoblast differentiation. PMID:26630504

Regulation of G-protein-coupled signaling pathways in allergic inflammation

PubMed Central

2009-01-01

Abstract Allergic diseases such as asthma are elicited by maladaptive activation of immune cells such as mast cells and lymphocytes by otherwise innocuous allergens. The numerous mediators secreted by such cells promote both acute inflammation and, in many instances, chronic tissue remodeling. Most of these compounds exert their effects on end-organ targets such as epithelial and endothelial cells and airway smooth muscle by activating G-protein-coupled receptors (GPCRs), which are by far the most abundant type of cell surface receptor. Since GPCRs are also the most common target of allergy therapeutics, a better understanding of their intracellular signaling mechanisms is vital to improve the efficacy of such drugs or to develop new targets. In this review, we focus on some of the new regulatory elements that control the duration and amplitude of GPCR signal transduction pathways in immune effector cells and end-organ structural cells affected by allergic inflammation. PMID:18810336

Genetic and Epigenetic Regulation of Human Cardiac Reprogramming and Differentiation in Regenerative Medicine

PubMed Central

Burridge, Paul W.; Sharma, Arun; Wu, Joseph C.

2016-01-01

Regeneration or replacement of lost cardiomyocytes within the heart has the potential to revolutionize cardiovascular medicine. Numerous methodologies have been used to achieve this aim, including the engraftment of bone marrow- and heart-derived cells as well as the identification of modulators of adult cardiomyocyte proliferation. Recently, the conversion of human somatic cells into induced pluripotent stem cells and induced cardiomyocyte-like cells has transformed potential approaches toward this goal, and the engraftment of cardiac progenitors derived from human embryonic stem cells into patients is now feasible. Here we review recent advances in our understanding of the genetic and epigenetic control of human cardiogenesis, cardiac differentiation, and the induced reprogramming of somatic cells to cardiomyocytes. We also cover genetic programs for inducing the proliferation of endogenous cardiomyocytes and discuss the genetic state of cells used in cardiac regenerative medicine. PMID:26631515

Development of the intrinsic and extrinsic innervation of the gut.

PubMed

Uesaka, Toshihiro; Young, Heather M; Pachnis, Vassilis; Enomoto, Hideki

2016-09-15

The gastrointestinal (GI) tract is innervated by intrinsic enteric neurons and by extrinsic efferent and afferent nerves. The enteric (intrinsic) nervous system (ENS) in most regions of the gut consists of two main ganglionated layers; myenteric and submucosal ganglia, containing numerous types of enteric neurons and glial cells. Axons arising from the ENS and from extrinsic neurons innervate most layers of the gut wall and regulate many gut functions. The majority of ENS cells are derived from vagal neural crest cells (NCCs), which proliferate, colonize the entire gut, and first populate the myenteric region. After gut colonization by vagal NCCs, the extrinsic nerve fibers reach the GI tract, and Schwann cell precursors (SCPs) enter the gut along the extrinsic nerves. Furthermore, a subpopulation of cells in myenteric ganglia undergoes a radial (inward) migration to form the submucosal plexus, and the intrinsic and extrinsic innervation to the mucosal region develops. Here, we focus on recent progress in understanding the developmental processes that occur after the gut is colonized by vagal ENS precursors, and provide an up-to-date overview of molecular mechanisms regulating the development of the intrinsic and extrinsic innervation of the GI tract. Copyright © 2016 Elsevier Inc. All rights reserved.

Pharmacological activation of lysophosphatidic acid receptors regulates erythropoiesis

PubMed Central

Lin, Kuan-Hung; Ho, Ya-Hsuan; Chiang, Jui-Chung; Li, Meng-Wei; Lin, Shi-Hung; Chen, Wei-Min; Chiang, Chi-Ling; Lin, Yu-Nung; Yang, Ya-Jan; Chen, Chiung-Nien; Lu, Jenher; Huang, Chang-Jen; Tigyi, Gabor; Yao, Chao-Ling; Lee, Hsinyu

2016-01-01

Lysophosphatidic acid (LPA), a growth factor-like phospholipid, regulates numerous physiological functions, including cell proliferation and differentiation. In a previous study, we have demonstrated that LPA activates erythropoiesis by activating the LPA 3 receptor subtype (LPA3) under erythropoietin (EPO) induction. In the present study, we applied a pharmacological approach to further elucidate the functions of LPA receptors during red blood cell (RBC) differentiation. In K562 human erythroleukemia cells, knockdown of LPA2 enhanced erythropoiesis, whereas knockdown of LPA3 inhibited RBC differentiation. In CD34+ human hematopoietic stem cells (hHSC) and K526 cells, the LPA3 agonist 1-oleoyl-2-methyl-sn-glycero-3-phosphothionate (2S-OMPT) promoted erythropoiesis, whereas the LPA2 agonist dodecyl monophosphate (DMP) and the nonlipid specific agonist GRI977143 (GRI) suppressed this process. In zebrafish embryos, hemoglobin expression was significantly increased by 2S-OMPT treatment but was inhibited by GRI. Furthermore, GRI treatment decreased, whereas 2S-OMPT treatment increased RBC counts and amount of hemoglobin level in adult BALB/c mice. These results indicate that LPA2 and LPA3 play opposing roles during RBC differentiation. The pharmacological activation of LPA receptor subtypes represent a novel strategies for augmenting or inhibiting erythropoiesis. PMID:27244685

Pharmacological activation of lysophosphatidic acid receptors regulates erythropoiesis.

PubMed

Lin, Kuan-Hung; Ho, Ya-Hsuan; Chiang, Jui-Chung; Li, Meng-Wei; Lin, Shi-Hung; Chen, Wei-Min; Chiang, Chi-Ling; Lin, Yu-Nung; Yang, Ya-Jan; Chen, Chiung-Nien; Lu, Jenher; Huang, Chang-Jen; Tigyi, Gabor; Yao, Chao-Ling; Lee, Hsinyu

2016-05-31

Lysophosphatidic acid (LPA), a growth factor-like phospholipid, regulates numerous physiological functions, including cell proliferation and differentiation. In a previous study, we have demonstrated that LPA activates erythropoiesis by activating the LPA 3 receptor subtype (LPA3) under erythropoietin (EPO) induction. In the present study, we applied a pharmacological approach to further elucidate the functions of LPA receptors during red blood cell (RBC) differentiation. In K562 human erythroleukemia cells, knockdown of LPA2 enhanced erythropoiesis, whereas knockdown of LPA3 inhibited RBC differentiation. In CD34(+) human hematopoietic stem cells (hHSC) and K526 cells, the LPA3 agonist 1-oleoyl-2-methyl-sn-glycero-3-phosphothionate (2S-OMPT) promoted erythropoiesis, whereas the LPA2 agonist dodecyl monophosphate (DMP) and the nonlipid specific agonist GRI977143 (GRI) suppressed this process. In zebrafish embryos, hemoglobin expression was significantly increased by 2S-OMPT treatment but was inhibited by GRI. Furthermore, GRI treatment decreased, whereas 2S-OMPT treatment increased RBC counts and amount of hemoglobin level in adult BALB/c mice. These results indicate that LPA2 and LPA3 play opposing roles during RBC differentiation. The pharmacological activation of LPA receptor subtypes represent a novel strategies for augmenting or inhibiting erythropoiesis.

Satellite Cells and the Muscle Stem Cell Niche

PubMed Central

Yin, Hang; Price, Feodor

2013-01-01

Adult skeletal muscle in mammals is a stable tissue under normal circumstances but has remarkable ability to repair after injury. Skeletal muscle regeneration is a highly orchestrated process involving the activation of various cellular and molecular responses. As skeletal muscle stem cells, satellite cells play an indispensible role in this process. The self-renewing proliferation of satellite cells not only maintains the stem cell population but also provides numerous myogenic cells, which proliferate, differentiate, fuse, and lead to new myofiber formation and reconstitution of a functional contractile apparatus. The complex behavior of satellite cells during skeletal muscle regeneration is tightly regulated through the dynamic interplay between intrinsic factors within satellite cells and extrinsic factors constituting the muscle stem cell niche/microenvironment. For the last half century, the advance of molecular biology, cell biology, and genetics has greatly improved our understanding of skeletal muscle biology. Here, we review some recent advances, with focuses on functions of satellite cells and their niche during the process of skeletal muscle regeneration. PMID:23303905

Growth phase-dependent control of R27 conjugation is mediated by the interplay between the plasmid-encoded regulatory circuit TrhR/TrhY-HtdA and the cAMP regulon.

PubMed

Gibert, Marta; Paytubi, Sonia; Beltrán, Sergi; Juárez, Antonio; Balsalobre, Carlos; Madrid, Cristina

2016-12-01

Plasmids of the incompatibility group HI1 (IncHI1) have been isolated from several Gram-negative pathogens and are associated with the spread of multidrug resistance. Their conjugation is tightly regulated and it is inhibited at temperatures higher than 30°C, indicating that conjugation occurs outside warm-blooded hosts. Using R27, the prototype of IncHI1 plasmids, we report that plasmid transfer efficiency in E. coli strongly depends on the physiological state of the donor cells. Conjugation frequency is high when cells are actively growing, dropping sharply when cells enter the stationary phase of growth. Accordingly, our transcriptomic assays show significant downregulation of numerous R27 genes during the stationary phase, including several tra (transfer) genes. Growth phase-dependent regulation of tra genes transcription is independent of H-NS, a silencer of horizontal gene transfer, and ppGpp and RpoS, regulators of the stationary phase, but highly dependent on the plasmid-encoded regulatory circuit TrhR/TrhY-HtdA. The metabolic sensor cAMP, whose synthesis is chromosomally encoded, is also involved in the growth phase regulation of R27 conjugation by modulating htdA expression. Our data suggest that the involvement of regulators encoded by both chromosome and plasmid are required for efficient physiological control of IncHI1 plasmid conjugation. © 2016 Society for Applied Microbiology and John Wiley & Sons Ltd.

A review of natural and synthetic antioxidants important for health and longevity.

PubMed

Wojcik, M; Burzynska-Pedziwiatr, I; Wozniak, L A

2010-01-01

Reactive oxygen species (ROS) generated in the presence of O(2) by mitochondria, phagocytic cells, peroxisomes, and cytochrome P450 enzymes under physiological conditions, may play a dual function in the human organism. On the one hand, they participate in cell signal transduction cascades, leading to the activation of some transcription factors responsible for regulating of the expression of genes relevant for cell growth and differentiation. On the other hand, they cause oxidative damage of cellular DNA, protein and lipids, resulting in the initiation or development of numerous diseases such as cancer, cardiovascular diseases, type 2 diabetes mellitus, cataract, rheumatoid arthritis, or different neurodegenerative diseases. Both endogenous compounds (glutathione, ubiquinol, urate, bilirubin) and enzymes (superoxide dismutase, catalase, glutathione peroxidase) are engaged in the detoxification of ROS. In addition, numerous dietary components such as vitamin C, vitamin E, carotenoids, and polyphenols are thought to be involved in the antioxidant defense system. The present review article is focused on the summary and the assessment of research on the impact of dietary antioxidants in the prevention of chronic diseases, particularly cancer and cardiovascular diseases.

The Presence of Pretreated Lignocellulosic Solids from Birch during Saccharomyces cerevisiae Fermentations Leads to Increased Tolerance to Inhibitors--A Proteomic Study of the Effects.

PubMed

Koppram, Rakesh; Mapelli, Valeria; Albers, Eva; Olsson, Lisbeth

2016-01-01

The fermentation performance of Saccharomyces cerevisiae in the cellulose to ethanol conversion process is largely influenced by the components of pretreated biomass. The insoluble solids in pretreated biomass predominantly constitute cellulose, lignin, and -to a lesser extent- hemicellulose. It is important to understand the effects of water-insoluble solids (WIS) on yeast cell physiology and metabolism for the overall process optimization. In the presence of synthetic lignocellulosic inhibitors, we observed a reduced lag phase and enhanced volumetric ethanol productivity by S. cerevisiae CEN.PK 113-7D when the minimal medium was supplemented with WIS of pretreated birch or spruce and glucose as the carbon source. To investigate the underlying molecular reasons for the effects of WIS, we studied the response of WIS at the proteome level in yeast cells in the presence of acetic acid as an inhibitor. Comparisons were made with cells grown in the presence of acetic acid but without WIS in the medium. Altogether, 729 proteins were detected and quantified, of which 246 proteins were significantly up-regulated and 274 proteins were significantly down-regulated with a fold change≥1.2 in the presence of WIS compared to absence of WIS. The cells in the presence of WIS up-regulated several proteins related to cell wall, glycolysis, electron transport chain, oxidative stress response, oxygen and radical detoxification and unfolded protein response; and down-regulated most proteins related to biosynthetic pathways including amino acid, purine, isoprenoid biosynthesis, aminoacyl-tRNA synthetases and pentose phosphate pathway. Overall, the identified differentially regulated proteins may indicate that the likelihood of increased ATP generation in the presence of WIS was used to defend against acetic acid stress at the expense of reduced biomass formation. Although, comparative proteomics of cells with and without WIS in the acetic acid containing medium revealed numerous changes, a direct effect of WIS on cellular physiology remains to be investigated.

The Presence of Pretreated Lignocellulosic Solids from Birch during Saccharomyces cerevisiae Fermentations Leads to Increased Tolerance to Inhibitors – A Proteomic Study of the Effects

PubMed Central

Koppram, Rakesh; Mapelli, Valeria; Albers, Eva; Olsson, Lisbeth

2016-01-01

The fermentation performance of Saccharomyces cerevisiae in the cellulose to ethanol conversion process is largely influenced by the components of pretreated biomass. The insoluble solids in pretreated biomass predominantly constitute cellulose, lignin, and -to a lesser extent- hemicellulose. It is important to understand the effects of water-insoluble solids (WIS) on yeast cell physiology and metabolism for the overall process optimization. In the presence of synthetic lignocellulosic inhibitors, we observed a reduced lag phase and enhanced volumetric ethanol productivity by S. cerevisiae CEN.PK 113-7D when the minimal medium was supplemented with WIS of pretreated birch or spruce and glucose as the carbon source. To investigate the underlying molecular reasons for the effects of WIS, we studied the response of WIS at the proteome level in yeast cells in the presence of acetic acid as an inhibitor. Comparisons were made with cells grown in the presence of acetic acid but without WIS in the medium. Altogether, 729 proteins were detected and quantified, of which 246 proteins were significantly up-regulated and 274 proteins were significantly down-regulated with a fold change≥1.2 in the presence of WIS compared to absence of WIS. The cells in the presence of WIS up-regulated several proteins related to cell wall, glycolysis, electron transport chain, oxidative stress response, oxygen and radical detoxification and unfolded protein response; and down-regulated most proteins related to biosynthetic pathways including amino acid, purine, isoprenoid biosynthesis, aminoacyl-tRNA synthetases and pentose phosphate pathway. Overall, the identified differentially regulated proteins may indicate that the likelihood of increased ATP generation in the presence of WIS was used to defend against acetic acid stress at the expense of reduced biomass formation. Although, comparative proteomics of cells with and without WIS in the acetic acid containing medium revealed numerous changes, a direct effect of WIS on cellular physiology remains to be investigated. PMID:26849651

Stimulation of lactate receptor (HCAR1) affects cellular DNA repair capacity.

PubMed

Wagner, Waldemar; Kania, Katarzyna D; Ciszewski, Wojciech M

2017-04-01

Numerous G-protein coupled receptors have been reported to enhance cancer cell survival and resistance to clinically used chemotherapeutics. Recently, hydroxycarboxylic acid receptor 1 (HCAR1) was shown to drive lactate-dependent enhancement of cell survival and metastasis in pancreatic and breast cancers. Furthermore, our previous study confirmed the involvement of HCAR1 in lactate-related enhancement of DNA repair in cervical cancer cells. In the present study, we examined the possible mechanisms of HCAR1-mediated enhancement of DNA repair capacity. We observed that the HCAR1 agonist dihydroxybenzoic acid (DHBA) up-regulated BRCA1 (breast cancer type 1 susceptibility protein) and NBS1 (Nijmegen breakage syndrome 1) expression in HeLa cells. Moreover, HCAR1 silencing decreased mRNA and protein levels of BRCA1 by 30% and 20%, respectively. Immunocytochemical analyses of BRCA1, nibrin and DNA-PKcs indicated an increased accumulation of these proteins in cell nuclei after DHBA stimulation. Subsequently, these changes in the DNA repair protein levels translated into an enhanced DNA repair rate after doxorubicin treatment, as shown by γ-H2AX and comet assay experiments. In contrast, the down-regulation of HCAR1 decreased the efficiency of DNA repair. Finally, we observed the abrogation of DHBA-driven BRCA1 protein up-regulation and enhanced DNA repair following the preincubation of cells with the PKC inhibitor Gö6983. Taken together, our data indicate that lactate receptor/HCAR1 expression in cervical carcinoma cells may contribute to the modulation of cellular DNA repair mechanisms. Copyright © 2017 Elsevier B.V. All rights reserved.

Pro-apoptotic BIM is an essential initiator of physiological endothelial cell death independent of regulation by FOXO3

PubMed Central

Koenig, M N; Naik, E; Rohrbeck, L; Herold, M J; Trounson, E; Bouillet, P; Thomas, T; Voss, A K; Strasser, A; Coultas, L

2014-01-01

The growth of new blood vessels by angiogenesis is essential for normal development, but can also cause or contribute to the pathology of numerous diseases. Recent studies have shown that BIM, a pro-apoptotic BCL2-family protein, is required for endothelial cell apoptosis in vivo, and can contribute to the anti-angiogenic effect of VEGF-A inhibitors in certain tumor models. Despite its importance, the extent to which BIM is autonomously required for physiological endothelial apoptosis remains unknown and its regulation under such conditions is poorly defined. While the transcription factor FOXO3 has been proposed to induce Bim in response to growth factor withdrawal, evidence for this function is circumstantial. We report that apoptosis was reduced in Bim−/− primary endothelial cells, demonstrating a cell-autonomous role for BIM in endothelial death following serum and growth factor withdrawal. In conflict with in vitro studies, BIM-dependent endothelial death in vivo did not require FOXO3. Moreover, endothelial apoptosis proceeded normally in mice lacking FOXO-binding sites in the Bim promoter. Bim mRNA was upregulated in endothelial cells starved of serum and growth factors and this was accompanied by the downregulation of miRNAs of the miR-17∼92 cluster. Bim mRNA levels were also elevated in miR-17∼92+/− endothelial cells cultured under steady-state conditions, suggesting that miR-17∼92 cluster miRNAs may contribute to regulating overall Bim mRNA levels in endothelial cells. PMID:24971484

An alternative approach in regulation of expression of a transgene by endogenous miR-145 in carcinoma and normal breast cell lines.

PubMed

Ghanbari Safari, Maryam; Baesi, Kazem; Hosseinkhani, Saman

2017-03-01

MicroRNAs are small noncoding RNAs that regulate gene expression by repressing translation of target cellular transcripts. Increasing evidences indicate that miRNAs have different expression profiles and play crucial roles in numerous cellular processes. Delivery and expression of transgenes for cancer therapy must be specific for tumors to avoid killing of healthy tissues. Many investigators have shown that transgene expression can be suppressed in normal cells using vectors that are responsive to microRNA regulation. To overcome this problem, miR-145 that exhibits downregulation in many types of cancer cells was chosen for posttranscriptional regulatory systems mediated by microRNAs. In this study, a psiCHECK-145T vector carrying four tandem copies of target sequences of miR-145 into 3'-UTR of the Renilla luciferase gene was constructed. Renilla luciferase activity from the psiCHECK-145T vector was 57% lower in MCF10A cells with high miR-145 expression as compared to a control condition. Additionally, overexpression of miR-145 in MCF-7 cells with low expression level of miR-145 showed more than 76% reduction in the Renilla luciferase activity from the psiCHECK-145T vector. Inclusion of miR-145 target sequences into the 3'-UTR of the Renilla luciferase gene is a feasible strategy for restricting transgene expression in a breast cancer cell line while sparing a breast normal cell line. © 2015 International Union of Biochemistry and Molecular Biology, Inc.

Dynamics of re-constitution of the human nuclear proteome after cell division is regulated by NLS-adjacent phosphorylation

PubMed Central

Róna, Gergely; Borsos, Máté; Ellis, Jonathan J; Mehdi, Ahmed M; Christie, Mary; Környei, Zsuzsanna; Neubrandt, Máté; Tóth, Judit; Bozóky, Zoltán; Buday, László; Madarász, Emília; Bodén, Mikael; Kobe, Bostjan; Vértessy, Beáta G

2014-01-01

Phosphorylation by the cyclin-dependent kinase 1 (Cdk1) adjacent to nuclear localization signals (NLSs) is an important mechanism of regulation of nucleocytoplasmic transport. However, no systematic survey has yet been performed in human cells to analyze this regulatory process, and the corresponding cell-cycle dynamics have not yet been investigated. Here, we focused on the human proteome and found that numerous proteins, previously not identified in this context, are associated with Cdk1-dependent phosphorylation sites adjacent to their NLSs. Interestingly, these proteins are involved in key regulatory events of DNA repair, epigenetics, or RNA editing and splicing. This finding indicates that cell-cycle dependent events of genome editing and gene expression profiling may be controlled by nucleocytoplasmic trafficking. For in-depth investigations, we selected a number of these proteins and analyzed how point mutations, expected to modify the phosphorylation ability of the NLS segments, perturb nucleocytoplasmic localization. In each case, we found that mutations mimicking hyper-phosphorylation abolish nuclear import processes. To understand the mechanism underlying these phenomena, we performed a video microscopy-based kinetic analysis to obtain information on cell-cycle dynamics on a model protein, dUTPase. We show that the NLS-adjacent phosphorylation by Cdk1 of human dUTPase, an enzyme essential for genomic integrity, results in dynamic cell cycle-dependent distribution of the protein. Non-phosphorylatable mutants have drastically altered protein re-import characteristics into the nucleus during the G1 phase. Our results suggest a dynamic Cdk1-driven mechanism of regulation of the nuclear proteome composition during the cell cycle. PMID:25483092

Adapting in vitro embryonic stem cell differentiation to the study of locus control regions.

PubMed

Lahiji, Armin; Kučerová-Levisohn, Martina; Holmes, Roxanne; Zúñiga-Pflücker, Juan Carlos; Ortiz, Benjamin D

2014-05-01

Numerous locus control region (LCR) activities have been discovered in gene loci important to immune cell development and function. LCRs are a distinct class of cis-acting gene regulatory elements that appear to contain all the DNA sequence information required to establish an independently and predictably regulated gene expression program at any genomic site in native chromatin of a whole animal. As such, LCR-regulated transgenic reporter systems provide invaluable opportunities to investigate the mechanisms of gene regulatory DNA action during development. Furthermore the qualities of LCR-driven gene expression, including spatiotemporal specificity and "integration site-independence" would be highly desirable to incorporate into vectors used in therapeutic genetic engineering. Thus, advancement in the methods used to investigate LCRs is of considerable basic and translational significance. We study the LCR present in the mouse T cell receptor (TCR)-α gene locus. Until recently, transgenic mice provided the only experimental model capable of supporting the entire spectrum of LCR activities. We have recently reported complete manifestation of TCRα LCR function in T cells derived in vitro from mouse embryonic stem cells (ESC), thus validating a complete cell culture model for the full range of LCR activities seen in transgenic mice. Here we discuss the critical parameters involved in studying LCR-regulated gene expression during in vitro hematopoietic differentiation from ESCs. This advance provides an approach to speed progress in the LCR field, and facilitate the clinical application of its findings, particularly to the genetic engineering of T cells. Copyright © 2014 Elsevier B.V. All rights reserved.

Proteomic Analysis of Arginine Methylation Sites in Human Cells Reveals Dynamic Regulation During Transcriptional Arrest*

PubMed Central

Sylvestersen, Kathrine B.; Horn, Heiko; Jungmichel, Stephanie; Jensen, Lars J.; Nielsen, Michael L.

2014-01-01

The covalent attachment of methyl groups to the side-chain of arginine residues is known to play essential roles in regulation of transcription, protein function, and RNA metabolism. The specific N-methylation of arginine residues is catalyzed by a small family of gene products known as protein arginine methyltransferases; however, very little is known about which arginine residues become methylated on target substrates. Here we describe a proteomics methodology that combines single-step immunoenrichment of methylated peptides with high-resolution mass spectrometry to identify endogenous arginine mono-methylation (MMA) sites. We thereby identify 1027 site-specific MMA sites on 494 human proteins, discovering numerous novel mono-methylation targets and confirming the majority of currently known MMA substrates. Nuclear RNA-binding proteins involved in RNA processing, RNA localization, transcription, and chromatin remodeling are predominantly found modified with MMA. Despite this, MMA sites prominently are located outside RNA-binding domains as compared with the proteome-wide distribution of arginine residues. Quantification of arginine methylation in cells treated with Actinomycin D uncovers strong site-specific regulation of MMA sites during transcriptional arrest. Interestingly, several MMA sites are down-regulated after a few hours of transcriptional arrest. In contrast, the corresponding di-methylation or protein expression levels are not altered, confirming that MMA sites contain regulated functions on their own. Collectively, we present a site-specific MMA data set in human cells and demonstrate for the first time that MMA is a dynamic post-translational modification regulated during transcriptional arrest by a hitherto uncharacterized arginine demethylase. PMID:24563534

pH regulators in invadosomal functioning: proton delivery for matrix tasting.

PubMed

Brisson, Lucie; Reshkin, Stephan J; Goré, Jacques; Roger, Sébastien

2012-01-01

Invadosomes are actin-rich finger-like cellular structures sensing and interacting with the surrounding extracellular matrix (ECM) and involved in its proteolytic remodeling. Invadosomes are structures distinct from other adhesion complexes, and have been identified in normal cells that have to cross tissue barriers to fulfill their function such as leukocytes, osteoclasts and endothelial cells. They also represent features of highly aggressive cancer cells, allowing them to escape from the primary tumor, to invade surrounding tissues and to reach systemic circulation. They are localized to the ventral membrane of cells grown under 2-dimensional conditions and are supposed to be present all around cells grown in 3-dimensional matrices. Indeed invadosomes are key structures in physiological processes such as inflammation and the immune response, bone remodeling, tissue repair, but also in pathological conditions such as osteopetrosis and the development of metastases. Invadosomes are subdivided into podosomes, found in normal cells, and into invadopodia specific for cancer cells. While these two structures exhibit differences in organization, size, number and half-life, they share similarities in molecular composition, participation in cell-matrix adhesion and promoting matrix degradation. A key determinant in invadosomal function is the recruitment and release of proteases, such as matrix metalloproteinases (MMPs), serine proteases and cysteine cathepsins, together with their activation in a tightly controlled and highly acidic microenvironment. Therefore numerous pH regulators such as V-ATPases and Na(+)/H(+) exchangers, are found in invadosomes and are directly involved in their constitution as well as their functioning. This review focuses on the participation of pH regulators in invadosome function in physiological and pathological conditions, with a particular emphasis on ECM remodeling by osteoclasts during bone resorption and by cancer cells. Copyright © 2012 Elsevier GmbH. All rights reserved.

Insight into the Role of Long Non-coding RNAs During Osteogenesis in Mesenchymal Stem Cells.

PubMed

Huo, Sibei; Zhou, Yachuan; He, Xinyu; Wan, Mian; Du, Wei; Xu, Xin; Ye, Ling; Zhou, Xuedong; Zheng, Liwei

2018-01-01

Long non-coding RNAs (LncRNAs) are non-protein coding transcripts longer than 200 nucleotides in length. Instead of being "transcriptional noise", lncRNAs are emerging as a key modulator in various biological processes and disease development. Mesenchymal stem cells can be isolated from various adult tissues, such as bone marrow and dental tissues. The differentiation processes into multiple lineages, such as osteogenic differentiation, are precisely orchestrated by molecular signals in both genetic and epigenetic ways. Recently, several lines of evidence suggested the role of lncRNAs participating in cell differentiation through the regulation of gene transcriptions. And the involvement of lncRNAs may be associated with initiation and progression of mesenchymal stem cell-related diseases. We aimed at addressing the role of lncRNAs in the regulation of osteogenesis of mesenchymal stem cells derived from bone marrow and dental tissues, and discussing the potential utility of lncRNAs as biomarkers and therapeutic targets for mesenchymal stem cell-related diseases. Numerous lncRNAs were differentially expressed during osteogenesis or odontogenesis of mesenchymal stem cells, and some of them were confirmed to be able to regulate the differentiation processes through the modifications of chromatin, transcriptional and post-transcriptional processes. LncRNAs were also associated with some diseases related with pathologic differentiation of mesenchymal stem cells. LncRNAs involve in the osteogenic differentiation of bone marrow and dental tissuederived mesenchymal stem cells, and they could become promising therapeutic targets and prognosis parameters. However, the mechanisms of the role of lncRNAs are still enigmatic and require further investigation. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

REACTIVE OXYGEN SPECIES: IMPACT ON SKELETAL MUSCLE

PubMed Central

Powers, Scott K.; Ji, Li Li; Kavazis, Andreas N.; Jackson, Malcolm J.

2014-01-01

It is well established that contracting muscles produce both reactive oxygen and nitrogen species. Although the sources of oxidant production during exercise continue to be debated, growing evidence suggests that mitochondria are not the dominant source. Regardless of the sources of oxidants in contracting muscles, intense and prolonged exercise can result in oxidative damage to both proteins and lipids in the contracting myocytes. Further, oxidants regulate numerous cell signaling pathways and modulate the expression of many genes. This oxidant-mediated change in gene expression involves changes at transcriptional, mRNA stability, and signal transduction levels. Furthermore, numerous products associated with oxidant-modulated genes have been identified and include antioxidant enzymes, stress proteins, and mitochondrial electron transport proteins. Interestingly, low and physiological levels of reactive oxygen species are required for normal force production in skeletal muscle, but high levels of reactive oxygen species result in contractile dysfunction and fatigue. Ongoing research continues to explore the redox-sensitive targets in muscle that are responsible for both redox-regulation of muscle adaptation and oxidant-mediated muscle fatigue. PMID:23737208

Insulin signalling and the regulation of glucose and lipid metabolism

NASA Astrophysics Data System (ADS)

Saltiel, Alan R.; Kahn, C. Ronald

2001-12-01

The epidemic of type 2 diabetes and impaired glucose tolerance is one of the main causes of morbidity and mortality worldwide. In both disorders, tissues such as muscle, fat and liver become less responsive or resistant to insulin. This state is also linked to other common health problems, such as obesity, polycystic ovarian disease, hyperlipidaemia, hypertension and atherosclerosis. The pathophysiology of insulin resistance involves a complex network of signalling pathways, activated by the insulin receptor, which regulates intermediary metabolism and its organization in cells. But recent studies have shown that numerous other hormones and signalling events attenuate insulin action, and are important in type 2 diabetes.

Induction of hepatic haematopoiesis with fibronectin expression by EMT stromal cells during the second trimester of development.

PubMed

Lambropoulou, M; Tamiolakis, D; Venizelos, I; Alexiadis, G; Anastasopoulos, G; Limberis, V; Galazios, G; Tsikouras, P; Simopoulou, M; Nikolaidou, S; Petrakis, G; Papadopoulos, N

2007-09-01

In an initial period of vertebrate phylogeny (bone marrow-less vertebrates), lymphohaematopoiesis takes place in numerous organs containing a suitable microenvironment. Among other organs (i.e., gonads, kidney and spleen), the liver is apparently the most appropriate site for homing and differentiation of haematopoietic cell precursors. Interaction between haematopoietic cells and stromal cells is important for regulation of haematopoiesis. Numerous soluble and membrane-bound factors directly regulating haematopoiesis have been documented, but little is known about the effect of the foetal hepatic epithelial-to-mesenchymal transition (EMT) stromal cells' activity and their product-fibronectin, on foetal hepatic haematopoiesis. The binding of late-stage erythroid cells to FN has been well characterised and is believed to be critical for the terminal stages of erythroid differentiation. The intention of this article is to provide a quantitative overview of FN, produced by hepatic EMT stromal cells, in foetal hepatic haematopoiesis during the first and second trimester of development. Paraffin-embedded specimens from the liver of 30 human embryos in the first and second trimesters of gestation were investigated by conventional histology and immunohistology for the presence of FN and specific haematopoietic cell types. The staining intensity, and localisation of FN and haematopoietic markers in sequential sections were examined. Furthermore, double immunohistochemical staining was performed to assess simultaneous detection of FN and haematopoietic markers. FN was expressed in the EMT stromal cells of the hepatic portal triads more strongly during the second trimester than the first. Furthermore, an intense immunostaining for haematopoietic lineages, and especially for erythropoiesis, was observed in the second trimester compared to the first. The results of the double immunostaining disclosed an intimate co-expression of the FN and CD haematopoietic markers. Foetal hepatic EMT stromal cells provide a unique microenvironment that supports the emergence, expansion and maintenance of human foetal haematopoietic development during the mid-gestational stage. FN produced by the EMT stromal cells follows a time course parallel to that of haematopoiesis. We suggest that in foetal liver, phenotypic modifications of EMT stromal cells expressing FN concerning the cell adhesion capacity of the protein are associated with proliferation and differentiation of specific haematopoietic cell lineages during the second trimester of gestation, probably reflecting the increasing demand of the growing foetus for mature erythroid and myeloid cells.

Dissipation of contractile forces: the missing piece in cell mechanics.

PubMed

Kurzawa, Laetitia; Vianay, Benoit; Senger, Fabrice; Vignaud, Timothée; Blanchoin, Laurent; Théry, Manuel

2017-07-07

Mechanical forces are key regulators of cell and tissue physiology. The basic molecular mechanism of fiber contraction by the sliding of actin filament upon myosin leading to conformational change has been known for decades. The regulation of force generation at the level of the cell, however, is still far from elucidated. Indeed, the magnitude of cell traction forces on the underlying extracellular matrix in culture is almost impossible to predict or experimentally control. The considerable variability in measurements of cell-traction forces indicates that they may not be the optimal readout to properly characterize cell contractile state and that a significant part of the contractile energy is not transferred to cell anchorage but instead is involved in actin network dynamics. Here we discuss the experimental, numerical, and biological parameters that may be responsible for the variability in traction force production. We argue that limiting these sources of variability and investigating the dissipation of mechanical work that occurs with structural rearrangements and the disengagement of force transmission is key for further understanding of cell mechanics. © 2017 Kurzawa et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Minireview: Emerging Roles for Extracellular Vesicles in Diabetes and Related Metabolic Disorders

PubMed Central

Lakhter, Alexander J.

2015-01-01

Extracellular vesicles (EVs), membrane-contained vesicles released by most cell types, have attracted a large amount of research interest over the past decade. Because of their ability to transfer cargo via regulated processes, causing functional impacts on recipient cells, these structures may play important roles in cell-cell communication and have implications in the physiology of numerous organ systems. In addition, EVs have been described in most human biofluids and have wide potential as relatively noninvasive biomarkers of various pathologic conditions. Specifically, EVs produced by the pancreatic β-cell have been demonstrated to regulate physiologic and pathologic responses to β-cell stress, including β-cell proliferation and apoptosis. β-Cell EVs are also capable of interacting with immune cells and may contribute to the activation of autoimmune processes that trigger or propagate β-cell inflammation and destruction during the development of diabetes. EVs from adipose tissue have been shown to contribute to the development of the chronic inflammation and insulin resistance associated with obesity and metabolic syndrome via interactions with other adipose, liver, and muscle cells. Circulating EVs may also serve as biomarkers for metabolic derangements and complications associated with diabetes. This minireview describes the properties of EVs in general, followed by a more focused review of the literature describing EVs affecting the β-cell, β-cell autoimmunity, and the development of insulin resistance, which all have the potential to affect development of type 1 or type 2 diabetes. PMID:26393296

hZIP1 zinc uptake transporter down regulation and zinc depletion in prostate cancer

PubMed Central

Franklin, Renty B; Feng, Pei; Milon, B; Desouki, Mohamed M; Singh, Keshav K; Kajdacsy-Balla, André; Bagasra, Omar; Costello, Leslie C

2005-01-01

Background The genetic and molecular mechanisms responsible for and associated with the development and progression of prostate malignancy are largely unidentified. The peripheral zone is the major region of the human prostate gland where malignancy develops. The normal peripheral zone glandular epithelium has the unique function of accumulating high levels of zinc. In contrast, the ability to accumulate zinc is lost in the malignant cells. The lost ability of the neoplastic epithelial cells to accumulate zinc is a consistent factor in their development of malignancy. Recent studies identified ZIP1 (SLC39A1) as an important zinc transporter involved in zinc accumulation in prostate cells. Therefore, we investigated the possibility that down-regulation of hZIP1 gene expression might be involved in the inability of malignant prostate cells to accumulate zinc. To address this issue, the expression of hZIP1 and the depletion of zinc in malignant versus non-malignant prostate glands of prostate cancer tissue sections were analyzed. hZIP1 expression was also determined in malignant prostate cell lines. Results hZIP1 gene expression, ZIP1 transporter protein, and cellular zinc were prominent in normal peripheral zone glandular epithelium and in benign hyperplastic glands (also zinc accumulating glands). In contrast, hZIP1 gene expression and transporter protein were markedly down-regulated and zinc was depleted in adenocarcinomatous glands and in prostate intra-epithelial neoplastic foci (PIN). These changes occur early in malignancy and are sustained during its progression in the peripheral zone. hZIP1 is also expressed in the malignant cell lines LNCaP, PC-3, DU-145; and in the nonmalignant cell lines HPr-1 and BPH-1. Conclusion The studies clearly establish that hZIP1 gene expression is down regulated and zinc is depleted in adenocarcinomatous glands. The fact that all the malignant cell lines express hZIP1 indicates that the down-regulation in adenocarcinomatous glands is likely due to in situ gene silencing. These observations, coupled with the numerous and consistent reports of loss of zinc accumulation in malignant cells in prostate cancer, lead to the plausible proposal that down regulation of hZIP1 is a critical early event in the development prostate cancer. PMID:16153295

Analysis of Cell Division and Elongation Underlying the Developmental Acceleration of Root Growth in Arabidopsis thaliana1

PubMed Central

Beemster, Gerrit T.S.; Baskin, Tobias I.

1998-01-01

To investigate the relation between cell division and expansion in the regulation of organ growth rate, we used Arabidopsis thaliana primary roots grown vertically at 20°C with an elongation rate that increased steadily during the first 14 d after germination. We measured spatial profiles of longitudinal velocity and cell length and calculated parameters of cell expansion and division, including rates of local cell production (cells mm−1 h−1) and cell division (cells cell−1 h−1). Data were obtained for the root cortex and also for the two types of epidermal cell, trichoblasts and atrichoblasts. Accelerating root elongation was caused by an increasingly longer growth zone, while maximal strain rates remained unchanged. The enlargement of the growth zone and, hence, the accelerating root elongation rate, were accompanied by a nearly proportionally increased cell production. This increased production was caused by increasingly numerous dividing cells, whereas their rates of division remained approximately constant. Additionally, the spatial profile of cell division rate was essentially constant. The meristem was longer than generally assumed, extending well into the region where cells elongated rapidly. In the two epidermal cell types, meristem length and cell division rate were both very similar to that of cortical cells, and differences in cell length between the two epidermal cell types originated at the apex of the meristem. These results highlight the importance of controlling the number of dividing cells, both to generate tissues with different cell lengths and to regulate the rate of organ enlargement. PMID:9536070

Regulators of Cholangiocyte Proliferation.

PubMed

Hall, Chad; Sato, Keisaku; Wu, Nan; Zhou, Tianhao; Kyritsi, Konstantina; Meng, Fanyin; Glaser, Shannon; Alpini, Gianfranco

2017-02-10

Cholangiocytes, a small population of cells within the normal liver, have been the focus of a significant amount of research over the past two decades because of their involvement in cholangiopathies such as primary sclerosing cholangitis and primary biliary cholangitis. This article summarizes landmark studies in the field of cholangiocyte physiology and aims to provide an updated review of biliary pathogenesis. The historical approach of rodent extrahepatic bile duct ligation and the relatively recent utilization of transgenic mice have led to significant discoveries in cholangiocyte pathophysiology. Cholangiocyte physiology is a complex system based on heterogeneity within the biliary tree and a number of signaling pathways that serve to regulate bile composition. Studies have expanded the list of neuropeptides, neurotransmitters, and hormones that have been shown to be key regulators of proliferation and biliary damage. The peptide histamine and hormones, such as melatonin and angiotensin, angiotensin, as well as numerous sex hormones, have been implicated in cholangiocyte proliferation during cholestasis. Numerous pathways promote cholangiocyte proliferation during cholestasis, and there is growing evidence to suggest that cholangiocyte proliferation may promote hepatic fibrosis. These pathways may represent significant therapeutic potential for a subset of cholestatic liver diseases that currently lack effective therapies.

The mitosis-regulating and protein-protein interaction activities of astrin are controlled by aurora-A-induced phosphorylation.

PubMed

Chiu, Shao-Chih; Chen, Jo-Mei Maureen; Wei, Tong-You Wade; Cheng, Tai-Shan; Wang, Ya-Hui Candice; Ku, Chia-Feng; Lian, Chiao-Hsuan; Liu, Chun-Chih Jared; Kuo, Yi-Chun; Yu, Chang-Tze Ricky

2014-09-01

Cells display dramatic morphological changes in mitosis, where numerous factors form regulatory networks to orchestrate the complicated process, resulting in extreme fidelity of the segregation of duplicated chromosomes into two daughter cells. Astrin regulates several aspects of mitosis, such as maintaining the cohesion of sister chromatids by inactivating Separase and stabilizing spindle, aligning and segregating chromosomes, and silencing spindle assembly checkpoint by interacting with Src kinase-associated phosphoprotein (SKAP) and cytoplasmic linker-associated protein-1α (CLASP-1α). To understand how Astrin is regulated in mitosis, we report here that Astrin acts as a mitotic phosphoprotein, and Aurora-A phosphorylates Astrin at Ser(115). The phosphorylation-deficient mutant Astrin S115A abnormally activates spindle assembly checkpoint and delays mitosis progression, decreases spindle stability, and induces chromosome misalignment. Mechanistic analyses reveal that Astrin phosphorylation mimicking mutant S115D, instead of S115A, binds and induces ubiquitination and degradation of securin, which sequentially activates Separase, an enzyme required for the separation of sister chromatids. Moreover, S115A fails to bind mitosis regulators, including SKAP and CLASP-1α, which results in the mitotic defects observed in Astrin S115A-transfected cells. In conclusion, Aurora-A phosphorylates Astrin and guides the binding of Astrin to its cellular partners, which ensures proper progression of mitosis. Copyright © 2014 the American Physiological Society.

SND2, a NAC transcription factor gene, regulates genes involved in secondary cell wall development in Arabidopsis fibres and increases fibre cell area in Eucalyptus

PubMed Central

2011-01-01

Background NAC domain transcription factors initiate secondary cell wall biosynthesis in Arabidopsis fibres and vessels by activating numerous transcriptional regulators and biosynthetic genes. NAC family member SND2 is an indirect target of a principal regulator of fibre secondary cell wall formation, SND1. A previous study showed that overexpression of SND2 produced a fibre cell-specific increase in secondary cell wall thickness in Arabidopsis stems, and that the protein was able to transactivate the cellulose synthase8 (CesA8) promoter. However, the full repertoire of genes regulated by SND2 is unknown, and the effect of its overexpression on cell wall chemistry remains unexplored. Results We overexpressed SND2 in Arabidopsis and analyzed homozygous lines with regards to stem chemistry, biomass and fibre secondary cell wall thickness. A line showing upregulation of CesA8 was selected for transcriptome-wide gene expression profiling. We found evidence for upregulation of biosynthetic genes associated with cellulose, xylan, mannan and lignin polymerization in this line, in agreement with significant co-expression of these genes with native SND2 transcripts according to public microarray repositories. Only minor alterations in cell wall chemistry were detected. Transcription factor MYB103, in addition to SND1, was upregulated in SND2-overexpressing plants, and we detected upregulation of genes encoding components of a signal transduction machinery recently proposed to initiate secondary cell wall formation. Several homozygous T4 and hemizygous T1 transgenic lines with pronounced SND2 overexpression levels revealed a negative impact on fibre wall deposition, which may be indirectly attributable to excessive overexpression rather than co-suppression. Conversely, overexpression of SND2 in Eucalyptus stems led to increased fibre cross-sectional cell area. Conclusions This study supports a function for SND2 in the regulation of cellulose and hemicellulose biosynthetic genes in addition of those involved in lignin polymerization and signalling. SND2 seems to occupy a subordinate but central tier in the secondary cell wall transcriptional network. Our results reveal phenotypic differences in the effect of SND2 overexpression between woody and herbaceous stems and emphasize the importance of expression thresholds in transcription factor studies. PMID:22133261

Nucleolar protein trafficking in response to HIV-1 Tat: rewiring the nucleolus.

PubMed

Jarboui, Mohamed Ali; Bidoia, Carlo; Woods, Elena; Roe, Barbara; Wynne, Kieran; Elia, Giuliano; Hall, William W; Gautier, Virginie W

2012-01-01

The trans-activator Tat protein is a viral regulatory protein essential for HIV-1 replication. Tat trafficks to the nucleoplasm and the nucleolus. The nucleolus, a highly dynamic and structured membrane-less sub-nuclear compartment, is the site of rRNA and ribosome biogenesis and is involved in numerous cellular functions including transcriptional regulation, cell cycle control and viral infection. Importantly, transient nucleolar trafficking of both Tat and HIV-1 viral transcripts are critical in HIV-1 replication, however, the role(s) of the nucleolus in HIV-1 replication remains unclear. To better understand how the interaction of Tat with the nucleolar machinery contributes to HIV-1 pathogenesis, we investigated the quantitative changes in the composition of the nucleolar proteome of Jurkat T-cells stably expressing HIV-1 Tat fused to a TAP tag. Using an organellar proteomic approach based on mass spectrometry, coupled with Stable Isotope Labelling in Cell culture (SILAC), we quantified 520 proteins, including 49 proteins showing significant changes in abundance in Jurkat T-cell nucleolus upon Tat expression. Numerous proteins exhibiting a fold change were well characterised Tat interactors and/or known to be critical for HIV-1 replication. This suggests that the spatial control and subcellular compartimentaliation of these cellular cofactors by Tat provide an additional layer of control for regulating cellular machinery involved in HIV-1 pathogenesis. Pathway analysis and network reconstruction revealed that Tat expression specifically resulted in the nucleolar enrichment of proteins collectively participating in ribosomal biogenesis, protein homeostasis, metabolic pathways including glycolytic, pentose phosphate, nucleotides and amino acids biosynthetic pathways, stress response, T-cell signaling pathways and genome integrity. We present here the first differential profiling of the nucleolar proteome of T-cells expressing HIV-1 Tat. We discuss how these proteins collectively participate in interconnected networks converging to adapt the nucleolus dynamic activities, which favor host biosynthetic activities and may contribute to create a cellular environment supporting robust HIV-1 production.

Nucleolar Protein Trafficking in Response to HIV-1 Tat: Rewiring the Nucleolus

PubMed Central

Jarboui, Mohamed Ali; Bidoia, Carlo; Woods, Elena; Roe, Barbara; Wynne, Kieran; Elia, Giuliano; Hall, William W.; Gautier, Virginie W.

2012-01-01

The trans-activator Tat protein is a viral regulatory protein essential for HIV-1 replication. Tat trafficks to the nucleoplasm and the nucleolus. The nucleolus, a highly dynamic and structured membrane-less sub-nuclear compartment, is the site of rRNA and ribosome biogenesis and is involved in numerous cellular functions including transcriptional regulation, cell cycle control and viral infection. Importantly, transient nucleolar trafficking of both Tat and HIV-1 viral transcripts are critical in HIV-1 replication, however, the role(s) of the nucleolus in HIV-1 replication remains unclear. To better understand how the interaction of Tat with the nucleolar machinery contributes to HIV-1 pathogenesis, we investigated the quantitative changes in the composition of the nucleolar proteome of Jurkat T-cells stably expressing HIV-1 Tat fused to a TAP tag. Using an organellar proteomic approach based on mass spectrometry, coupled with Stable Isotope Labelling in Cell culture (SILAC), we quantified 520 proteins, including 49 proteins showing significant changes in abundance in Jurkat T-cell nucleolus upon Tat expression. Numerous proteins exhibiting a fold change were well characterised Tat interactors and/or known to be critical for HIV-1 replication. This suggests that the spatial control and subcellular compartimentaliation of these cellular cofactors by Tat provide an additional layer of control for regulating cellular machinery involved in HIV-1 pathogenesis. Pathway analysis and network reconstruction revealed that Tat expression specifically resulted in the nucleolar enrichment of proteins collectively participating in ribosomal biogenesis, protein homeostasis, metabolic pathways including glycolytic, pentose phosphate, nucleotides and amino acids biosynthetic pathways, stress response, T-cell signaling pathways and genome integrity. We present here the first differential profiling of the nucleolar proteome of T-cells expressing HIV-1 Tat. We discuss how these proteins collectively participate in interconnected networks converging to adapt the nucleolus dynamic activities, which favor host biosynthetic activities and may contribute to create a cellular environment supporting robust HIV-1 production. PMID:23166591

A role for sex and a common HFE gene variant in brain iron uptake.

PubMed

Duck, Kari A; Neely, Elizabeth B; Simpson, Ian A; Connor, James R

2018-03-01

HFE (high iron) is an essential protein for regulating iron transport into cells. Mutations of the HFE gene result in loss of this regulation causing accumulation of iron within the cell. The mutated protein has been found increasingly in numerous neurodegenerative disorders in which increased levels of iron in the brain are reported. Additionally, evidence that these mutations are associated with elevated brain iron challenges the paradigm that the brain is protected by the blood-brain barrier. While much has been studied regarding the role of HFE in cellular iron uptake, it has remained unclear what role the protein plays in the transport of iron into the brain. We investigated regulation of iron transport into the brain using a mouse model with a mutation in the HFE gene. We demonstrated that the rate of radiolabeled iron ( 59 Fe) uptake was similar between the two genotypes despite higher brain iron concentrations in the mutant. However, there were significant differences in iron uptake between males and females regardless of genotype. These data indicate that brain iron status is consistently maintained and tightly regulated at the level of the blood-brain barrier.

Post-transcriptional bursting in genes regulated by small RNA molecules

NASA Astrophysics Data System (ADS)

Rodrigo, Guillermo

2018-03-01

Gene expression programs in living cells are highly dynamic due to spatiotemporal molecular signaling and inherent biochemical stochasticity. Here we study a mechanism based on molecule-to-molecule variability at the RNA level for the generation of bursts of protein production, which can lead to heterogeneity in a cell population. We develop a mathematical framework to show numerically and analytically that genes regulated post transcriptionally by small RNA molecules can exhibit such bursts due to different states of translation activity (on or off), mostly revealed in a regime of few molecules. We exploit this framework to compare transcriptional and post-transcriptional bursting and also to illustrate how to tune the resulting protein distribution with additional post-transcriptional regulations. Moreover, because RNA-RNA interactions are predictable with an energy model, we define the kinetic constants of on-off switching as functions of the two characteristic free-energy differences of the system, activation and formation, with a nonequilibrium scheme. Overall, post-transcriptional bursting represents a distinctive principle linking gene regulation to gene expression noise, which highlights the importance of the RNA layer beyond the simple information transfer paradigm and significantly contributes to the understanding of the intracellular processes from a first-principles perspective.

Phospholipase D Signaling Pathways and Phosphatidic Acid as Therapeutic Targets in Cancer

PubMed Central

Bruntz, Ronald C.; Lindsley, Craig W.

2014-01-01

Phospholipase D is a ubiquitous class of enzymes that generates phosphatidic acid as an intracellular signaling species. The phospholipase D superfamily plays a central role in a variety of functions in prokaryotes, viruses, yeast, fungi, plants, and eukaryotic species. In mammalian cells, the pathways modulating catalytic activity involve a variety of cellular signaling components, including G protein–coupled receptors, receptor tyrosine kinases, polyphosphatidylinositol lipids, Ras/Rho/ADP-ribosylation factor GTPases, and conventional isoforms of protein kinase C, among others. Recent findings have shown that phosphatidic acid generated by phospholipase D plays roles in numerous essential cellular functions, such as vesicular trafficking, exocytosis, autophagy, regulation of cellular metabolism, and tumorigenesis. Many of these cellular events are modulated by the actions of phosphatidic acid, and identification of two targets (mammalian target of rapamycin and Akt kinase) has especially highlighted a role for phospholipase D in the regulation of cellular metabolism. Phospholipase D is a regulator of intercellular signaling and metabolic pathways, particularly in cells that are under stress conditions. This review provides a comprehensive overview of the regulation of phospholipase D activity and its modulation of cellular signaling pathways and functions. PMID:25244928

Phospholipase D signaling pathways and phosphatidic acid as therapeutic targets in cancer.

PubMed

Bruntz, Ronald C; Lindsley, Craig W; Brown, H Alex

2014-10-01

Phospholipase D is a ubiquitous class of enzymes that generates phosphatidic acid as an intracellular signaling species. The phospholipase D superfamily plays a central role in a variety of functions in prokaryotes, viruses, yeast, fungi, plants, and eukaryotic species. In mammalian cells, the pathways modulating catalytic activity involve a variety of cellular signaling components, including G protein-coupled receptors, receptor tyrosine kinases, polyphosphatidylinositol lipids, Ras/Rho/ADP-ribosylation factor GTPases, and conventional isoforms of protein kinase C, among others. Recent findings have shown that phosphatidic acid generated by phospholipase D plays roles in numerous essential cellular functions, such as vesicular trafficking, exocytosis, autophagy, regulation of cellular metabolism, and tumorigenesis. Many of these cellular events are modulated by the actions of phosphatidic acid, and identification of two targets (mammalian target of rapamycin and Akt kinase) has especially highlighted a role for phospholipase D in the regulation of cellular metabolism. Phospholipase D is a regulator of intercellular signaling and metabolic pathways, particularly in cells that are under stress conditions. This review provides a comprehensive overview of the regulation of phospholipase D activity and its modulation of cellular signaling pathways and functions. Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.

The role of the mitochondrial pyruvate carrier in substrate regulation

PubMed Central

Vacanti, Nathaniel M.; Divakaruni, Ajit S.; Green, Courtney R.; Parker, Seth J.; Henry, Robert R.; Ciaraldi, Theodore P.; Murphy, Anne N.; Metallo, Christian M.

2014-01-01

SUMMARY Pyruvate lies at a central biochemical node connecting carbohydrate, amino acid, and fatty acid metabolism, and the regulation of pyruvate flux into mitochondria represents a critical step in intermediary metabolism impacting numerous diseases. To characterize changes in mitochondrial substrate utilization in the context of compromised mitochondrial pyruvate transport, we applied 13C metabolic flux analysis (MFA) to cells after transcriptional or pharmacological inhibition of the mitochondrial pyruvate carrier (MPC). Despite profound suppression of both glucose and pyruvate oxidation, cell growth, oxygen consumption, and tricarboxylic acid (TCA) metabolism were surprisingly maintained. Oxidative TCA flux was achieved through enhanced reliance on glutaminolysis through malic enzyme and pyruvate dehydrogenase (PDH) as well as fatty acid and branched chain amino acid oxidation. Thus, in contrast to inhibition of complex I or PDH, suppression of pyruvate transport induces a form of metabolic flexibility associated with use of lipids and amino acids as catabolic and anabolic fuels. PMID:25458843

Genome-Wide Search Reveals the Existence of a Limited Number of Thyroid Hormone Receptor Alpha Target Genes in Cerebellar Neurons

PubMed Central

Chatonnet, Fabrice; Guyot, Romain; Picou, Frédéric; Bondesson, Maria; Flamant, Frederic

2012-01-01

Thyroid hormone (T3) has a major influence on cerebellum post-natal development. The major phenotypic landmark of exposure to low levels of T3 during development (hypothyroidism) in the cerebellum is the retarded inward migration of the most numerous cell type, granular neurons. In order to identify the direct genetic regulation exerted by T3 on cerebellar neurons and their precursors, we used microarray RNA hybridization to perform a time course analysis of T3 induced gene expression in primary cultures of cerebellar neuronal cell. These experiments suggest that we identified a small set of genes which are directly regulated, both in vivo and in vitro, during cerebellum post-natal development. These modest changes suggest that T3 does not acts directly on granular neurons and mainly indirectly influences the cellular interactions taking place during development. PMID:22586439

B Cell Receptor Activation Predominantly Regulates AKT-mTORC1/2 Substrates Functionally Related to RNA Processing

PubMed Central

Mohammad, Dara K.; Ali, Raja H.; Turunen, Janne J.; Nore, Beston F.; Smith, C. I. Edvard

2016-01-01

Protein kinase B (AKT) phosphorylates numerous substrates on the consensus motif RXRXXpS/T, a docking site for 14-3-3 interactions. To identify novel AKT-induced phosphorylation events following B cell receptor (BCR) activation, we performed proteomics, biochemical and bioinformatics analyses. Phosphorylated consensus motif-specific antibody enrichment, followed by tandem mass spectrometry, identified 446 proteins, containing 186 novel phosphorylation events. Moreover, we found 85 proteins with up regulated phosphorylation, while in 277 it was down regulated following stimulation. Up regulation was mainly in proteins involved in ribosomal and translational regulation, DNA binding and transcription regulation. Conversely, down regulation was preferentially in RNA binding, mRNA splicing and mRNP export proteins. Immunoblotting of two identified RNA regulatory proteins, RBM25 and MEF-2D, confirmed the proteomics data. Consistent with these findings, the AKT-inhibitor (MK-2206) dramatically reduced, while the mTORC-inhibitor PP242 totally blocked phosphorylation on the RXRXXpS/T motif. This demonstrates that this motif, previously suggested as an AKT target sequence, also is a substrate for mTORC1/2. Proteins with PDZ, PH and/or SH3 domains contained the consensus motif, whereas in those with an HMG-box, H15 domains and/or NF-X1-zinc-fingers, the motif was absent. Proteins carrying the consensus motif were found in all eukaryotic clades indicating that they regulate a phylogenetically conserved set of proteins. PMID:27487157

Drosophila: a model for studying genetic and molecular aspects of haematopoiesis and associated leukaemias

PubMed Central

Crozatier, Michèle; Vincent, Alain

2011-01-01

Vertebrate haematopoietic stem cells (HSCs) give rise to a hierarchically organised set of progenitors for erythroid, myeloid, lymphoid and megakaryocyte lineages, and are responsible for lifelong maintenance of the blood system. Dysregulation of the haematopoietic differentiation programme is at the origin of numerous pathologies, including leukaemias. With the discoveries that many transcriptional regulators and signalling pathways controlling blood cell development are conserved between humans and Drosophila melanogaster, the fruit fly has become a good model for investigating the mechanisms underlying the generation of blood cell lineages and blood cell homeostasis. In this review article, we discuss how genetic and molecular studies of Drosophila haematopoiesis can contribute to our understanding of the haematopoietic niche, as well as of the origin and/or progression of haematopoietic malignancies in humans. PMID:21669932

The Nav1.2 channel is regulated by GSK3

PubMed Central

James, Thomas F.; Nenov, Miroslav N.; Wildburger, Norelle C.; Lichti, Cheryl; Luisi, Jonathan; Vergara, Fernanda; Panova-Electronova, Neli I.; Nilsson, Carol L.; Rudra, Jai; Green, Thomas A.; Labate, Demetrio; Laezza, Fernanda

2015-01-01

Background Phosphorylation plays an essential role in regulating the voltage-gated sodium (Nav) channels and excitability. Yet, a surprisingly limited number of kinases have been identified as regulators of Nav channels. Herein, we posited that glycogen synthase kinase 3 (GSK3), a critical kinase found associated with numerous brain disorders, might directly regulate neuronal Nav channels. Methods We used patch-clamp electrophysiology to record sodium currents from Nav1.2 channels stably expressed in HEK-293 cells. mRNA and protein levels were quantified with RT-PCR, Western blot, or confocal microscopy, and in vitro phosphorylation and mass spectrometry to identify phosphorylated residues. Results We found that exposure of cells to GSK3 inhibitor XIII significantly potentiates the peak current density of Nav1.2, a phenotype reproduced by silencing GSK3 with siRNA. Contrarily, overexpression of GSK3β suppressed Nav1.2-encoded currents. Neither mRNA nor total protein expression were changed upon GSK3 inhibition. Cell surface labeling of CD4-chimeric constructs expressing intracellular domains of the Nav1.2 channel indicates that cell surface expression of CD4-Nav1.2-Ctail was up-regulated upon pharmacological inhibition of GSK3, resulting in an increase of surface puncta at the plasma membrane. Finally, using in vitro phosphorylation in combination with high resolution mass spectrometry, we further demonstrate that GSK3β phosphorylates T1966 at the C-terminal tail of Nav1.2. Conclusion These findings provide evidence for a new mechanism by which GSK3 modulate Nav channel function via its C-terminal tail. General Significance These findings provide fundamental knowledge in understanding signaling dysfunction common in several neuropsychiatric disorders. PMID:25615535

mRNA-Selective Translation Induced by FSH in Primary Sertoli Cells

PubMed Central

Musnier, Astrid; León, Kelly; Morales, Julia; Reiter, Eric; Boulo, Thomas; Costache, Vlad; Vourc'h, Patrick; Heitzler, Domitille; Oulhen, Nathalie; Poupon, Anne; Boulben, Sandrine; Cormier, Patrick

2012-01-01

FSH is a key hormonal regulator of Sertoli cell secretory activity, required to optimize sperm production. To fulfil its biological function, FSH binds a G protein-coupled receptor, the FSH-R. The FSH-R-transduced signaling network ultimately leads to the transcription or down-regulation of numerous genes. In addition, recent evidence has suggested that FSH might also regulate protein translation. However, this point has never been demonstrated conclusively yet. Here we have addressed this issue in primary rat Sertoli cells endogenously expressing physiological levels of FSH-R. We observed that, within 90 min of stimulation, FSH not only enhanced overall protein synthesis in a mammalian target of rapamycin-dependent manner but also increased the recruitment of mRNA to polysomes. m7GTP pull-down experiments revealed the functional recruitment of mammalian target of rapamycin and p70 S6 kinase to the 5′cap, further supported by the enhanced phosphorylation of one of p70 S6 kinase targets, the eukaryotic initiation factor 4B. Importantly, the scaffolding eukaryotic initiation factor 4G was also recruited, whereas eukaryotic initiation factor 4E-binding protein, the eukaryotic initiation factor 4E generic inhibitor, appeared to play a minor role in translational regulations induced by FSH, in contrast to what is generally observed in response to anabolic factors. This particular regulation of the translational machinery by FSH stimulation might support mRNA-selective translation, as shown here by quantitative RT-PCR amplification of the c-fos and vascular endothelial growth factor mRNA but not of all FSH target mRNA, in polysomal fractions. These findings add a new level of complexity to FSH biological roles in its natural target cells, which has been underappreciated so far. PMID:22383463

Prokineticin-1 (PROK1) modulates interleukin (IL)-11 expression via prokineticin receptor 1 (PROKR1) and the calcineurin/NFAT signalling pathway.

PubMed

Cook, Ian H; Evans, Jemma; Maldonado-Pérez, David; Critchley, Hilary O; Sales, Kurt J; Jabbour, Henry N

2010-03-01

Prokineticin-1 (PROK1) is a multifunctional secreted protein which signals via the G-protein coupled receptor, PROKR1. Previous data from our laboratory using a human genome survey microarray showed that PROK1-prokineticin receptor 1 (PROKR1) signalling regulates numerous genes important for establishment of early pregnancy, including the cytokine interleukin (IL)-11. Here, we have shown that PROK1-PROKR1 induces the expression of IL-11 in PROKR1 Ishikawa cells and first trimester decidua via the calcium-calcineurin signalling pathway in a guanine nucleotide-binding protein (G(q/11)), extracellular signal-regulated kinases, Ca(2+) and calcineurin-nuclear factor of activated T cells dependent manner. Conversely, treatment of human decidua with a lentiviral miRNA to abolish endogenous PROK1 expression results in a significant reduction in IL-11 expression and secretion. Importantly, we have also shown a regulatory role for the regulator of calcineurin 1 isoform 4 (RCAN1-4). Overexpression of RCAN1-4 in PROKR1 Ishikawa cells using an adenovirus leads to a reduction in PROK1 induced IL-11 indicating that RCAN1-4 is a negative regulator in the calcineurin-mediated signalling to IL-11. Finally, we have shown the potential for both autocrine and paracrine signalling in the human endometrium by co-localizing IL-11, IL-11Ralpha and PROKR1 within the stromal and glandular epithelial cells of non-pregnant endometrium and first trimester decidua. Overall we have identified and characterized the signalling components of a novel PROK1-PROKR1 signalling pathway regulating IL-11.

Prokineticin-1 (PROK1) modulates interleukin (IL)-11 expression via prokineticin receptor 1 (PROKR1) and the calcineurin/NFAT signalling pathway

PubMed Central

Cook, Ian H.; Evans, Jemma; Maldonado-Pérez, David; Critchley, Hilary O.; Sales, Kurt J.; Jabbour, Henry N.

2010-01-01

Prokineticin-1 (PROK1) is a multifunctional secreted protein which signals via the G-protein coupled receptor, PROKR1. Previous data from our laboratory using a human genome survey microarray showed that PROK1–prokineticin receptor 1 (PROKR1) signalling regulates numerous genes important for establishment of early pregnancy, including the cytokine interleukin (IL)-11. Here, we have shown that PROK1–PROKR1 induces the expression of IL-11 in PROKR1 Ishikawa cells and first trimester decidua via the calcium–calcineurin signalling pathway in a guanine nucleotide-binding protein (Gq/11), extracellular signal-regulated kinases, Ca2+ and calcineurin–nuclear factor of activated T cells dependent manner. Conversely, treatment of human decidua with a lentiviral miRNA to abolish endogenous PROK1 expression results in a significant reduction in IL-11 expression and secretion. Importantly, we have also shown a regulatory role for the regulator of calcineurin 1 isoform 4 (RCAN1-4). Overexpression of RCAN1-4 in PROKR1 Ishikawa cells using an adenovirus leads to a reduction in PROK1 induced IL-11 indicating that RCAN1-4 is a negative regulator in the calcineurin-mediated signalling to IL-11. Finally, we have shown the potential for both autocrine and paracrine signalling in the human endometrium by co-localizing IL-11, IL-11Rα and PROKR1 within the stromal and glandular epithelial cells of non-pregnant endometrium and first trimester decidua. Overall we have identified and characterized the signalling components of a novel PROK1–PROKR1 signalling pathway regulating IL-11. PMID:19801577

Mitochondrial O-GlcNAc Transferase (mOGT) Regulates Mitochondrial Structure, Function, and Survival in HeLa Cells*

PubMed Central

Sacoman, Juliana L.; Dagda, Raul Y.; Burnham-Marusich, Amanda R.; Dagda, Ruben K.; Berninsone, Patricia M.

2017-01-01

O-Linked N-acetylglucosamine transferase (OGT) catalyzes O-GlcNAcylation of target proteins and regulates numerous biological processes. OGT is encoded by a single gene that yields nucleocytosolic and mitochondrial isoforms. To date, the role of the mitochondrial isoform of OGT (mOGT) remains largely unknown. Using high throughput proteomics, we identified 84 candidate mitochondrial glycoproteins, of which 44 are novel. Notably, two of the candidate glycoproteins identified (cytochrome oxidase 2 (COX2) and NADH:ubiquinone oxidoreductase core subunit 4 (MT-ND4)) are encoded by mitochondrial DNA. Using siRNA in HeLa cells, we found that reducing endogenous mOGT expression leads to alterations in mitochondrial structure and function, including Drp1-dependent mitochondrial fragmentation, reduction in mitochondrial membrane potential, and a significant loss of mitochondrial content in the absence of mitochondrial ROS. These defects are associated with a compensatory increase in oxidative phosphorylation per mitochondrion. mOGT is also critical for cell survival; siRNA-mediated knockdown of endogenous mOGT protected cells against toxicity mediated by rotenone, a complex I inhibitor. Conversely, reduced expression of both nucleocytoplasmic (ncOGT) and mitochondrial (mOGT) OGT isoforms is associated with increased mitochondrial respiration and elevated glycolysis, suggesting that ncOGT is a negative regulator of cellular bioenergetics. Last, we determined that mOGT is probably involved in the glycosylation of a restricted set of mitochondrial targets. We identified four proteins implicated in mitochondrial biogenesis and metabolism regulation as candidate substrates of mOGT, including leucine-rich PPR-containing protein and mitochondrial aconitate hydratase. Our findings suggest that mOGT is catalytically active in vivo and supports mitochondrial structure, health, and survival, whereas ncOGT predominantly regulates cellular bioenergetics. PMID:28100784

Anchoring Junctions As Drug Targets: Role in Contraceptive Development

PubMed Central

Mruk, Dolores D.; Silvestrini, Bruno; Cheng, C. Yan

2010-01-01

In multicellular organisms, cell-cell interactions are mediated in part by cell junctions, which underlie tissue architecture. Throughout spermatogenesis, for instance, preleptotene leptotene spermatocytes residing in the basal compartment of the seminiferous epithelium must traverse the blood-testis barrier to enter the adluminal compartment for continued development. At the same time, germ cells must also remain attached to Sertoli cells, and numerous studies have reported extensive restructuring at the Sertoli-Sertoli and Sertoli-germ cell interface during germ cell movement across the seminiferous epithelium. Furthermore, the proteins and signaling cascades that regulate adhesion between testicular cells have been largely delineated. These findings have unveiled a number of potential “druggable” targets that can be used to induce premature release of germ cells from the seminiferous epithelium, resulting in transient infertility. Herein, we discuss a novel approach with the aim of developing a nonhormonal male contraceptive for future human use, one that involves perturbing adhesion between Sertoli and germ cells in the testis. PMID:18483144

Probing the biology of cell boundary conditions through confinement of Xenopus cell-free cytoplasmic extracts.

PubMed

Bermudez, Jessica G; Chen, Hui; Einstein, Lily C; Good, Matthew C

2017-01-01

Cell-free cytoplasmic extracts prepared from Xenopus eggs and embryos have for decades provided a biochemical system with which to interrogate complex cell biological processes in vitro. Recently, the application of microfabrication and microfluidic strategies in biology has narrowed the gap between in vitro and in vivo studies by enabling formation of cell-size compartments containing functional cytoplasm. These approaches provide numerous advantages over traditional biochemical experiments performed in a test tube. Most notably, the cell-free cytoplasm is confined using a two- or three-dimensional boundary, which mimics the natural configuration of a cell. This strategy enables characterization of the spatial organization of a cell, and the role that boundaries play in regulating intracellular assembly and function. In this review, we describe the marriage of Xenopus cell-free cytoplasm and confinement technologies to generate synthetic cell-like systems, the recent biological insights they have enabled, and the promise they hold for future scientific discovery. © 2017 Wiley Periodicals, Inc.

["Light" epithelial cells of swine and bovine oviducts].

PubMed

Suuroia, T; Aunapuu, M; Arend, A; Sépp, E

2002-01-01

The ultrastructure of oviduct epithelium of clinically healthy cows and 15 sows was investigated using scanning and transmission electron microscopy. In all parts of the oviduct, ciliated and non-ciliated epithelial cells are present, but their number varies in both the investigated animals in different regions of the oviduct, depending on the phase of the estrous cycle. In addition to ciliated cells with numerous cilia on their luminal surface, so-called pale ciliary cells were found in all parts of the oviduct of cows and sows. The cytoplasm of these cells is electron-lucent, their luminal surface carries few cilia and short microvilli. The apical cytoplasm contains species specific secretory granules, which means that these cells have features characteristic of both secretory and ciliated cells. It is suggested that the pale ciliated and non-ciliated secretory cells are functional stages of the same tubar epithelium cell, and that the transformation between these two cell types is regulated by functional requirements of the organ in different phases of the estrous cycle.

Mifepristone-inducible transgene expression in neural progenitor cells in vitro and in vivo

PubMed Central

Hjelm, BE; Grunseich, C; Gowing, G; Avalos, P; Tian, J; Shelley, BC; Mooney, M; Narwani, K; Shi, Y; Svendsen, CN; Wolfe, JH; Fischbeck, KH; Pierson, TM

2016-01-01

Numerous gene and cell therapy strategies are being developed for the treatment of neurodegenerative disorders. Many of these strategies use constitutive expression of therapeutic transgenic proteins, and although functional in animal models of disease, this method is less likely to provide adequate flexibility for delivering therapy to humans. Ligand-inducible gene expression systems may be more appropriate for these conditions, especially within the central nervous system (CNS). Mifepristone’s ability to cross the blood–brain barrier makes it an especially attractive ligand for this purpose. We describe the production of a mifepristone-inducible vector system for regulated expression of transgenes within the CNS. Our inducible system used a lentivirus-based vector platform for the ex vivo production of mifepristone-inducible murine neural progenitor cells that express our transgenes of interest. These cells were processed through a series of selection steps to ensure that the cells exhibited appropriate transgene expression in a dose-dependent and temporally controlled manner with minimal background activity. Inducible cells were then transplanted into the brains of rodents, where they exhibited appropriate mifepristone-inducible expression. These studies detail a strategy for regulated expression in the CNS for use in the development of safe and efficient gene therapy for neurological disorders. PMID:26863047

3-D Numerical Simulations of Biofilm Dynamics with Quorum Sensing in a Flow Cell

DTIC Science & Technology

2014-01-01

resistant mutants [?]. Inspired by experimental findings, researchers have come up with some mathematical models to study biofilm formation and function...develop a full 3D mathematical model to study how quorum sensing regulates biofilm formation and development as well as the pros and cons of quorum...have given an overview of current advances in mathematical modeling of biofilms. Concerning coupling biofilm growth with quorum sensing features

Novel Array-Based Target Identification for Synergistic Sensitization of Breast Cancer to Herceptin

DTIC Science & Technology

2010-05-01

cancer cell lines and expressed in human breast tumors. Oncotarget, (submitted). Abstract Farah Rahmatpanah, Zhenyu Jia, Tatsuya Azum, Eileen Adamson...Michael McClelland, Eileen Adamson, Dan Mercola. Egr1 regulates the coordinated expression of numerous EGF receptor target genes as identified by...ChIP on chip. Genome Biology 2008, 9:R166 [Epub ahead of print]. Jun Hayakawa, Shalu Mittal, Yipeng Wang, Kemal Korkmaz, Mashide Ohmichi, Eileen

pH sensing via bicarbonate-regulated “soluble” adenylyl cyclase (sAC)

PubMed Central

Rahman, Nawreen; Buck, Jochen; Levin, Lonny R.

2013-01-01

Soluble adenylyl cyclase (sAC) is a source of the second messenger cyclic adenosine 3′, 5′ monophosphate (cAMP). sAC is directly regulated by bicarbonate (HCO−3) ions. In living cells, HCO−3 ions are in nearly instantaneous equilibrium with carbon dioxide (CO2) and pH due to the ubiquitous presence of carbonic anhydrases. Numerous biological processes are regulated by CO2, HCO−3, and/or pH, and in a number of these, sAC has been shown to function as a physiological CO2/HCO3/pH sensor. In this review, we detail the known pH sensing functions of sAC, and we discuss two highly-studied, pH-dependent pathways in which sAC might play a role. PMID:24324443

Inhibition of hyaluronan synthesis by vesnarinone in cultured human myofibroblasts.

PubMed

Ueki, N; Taguchi, T; Takahashi, M; Adachi, M; Ohkawa, T; Amuro, Y; Hada, T; Higashino, K

2000-02-02

Hyaluronan (HA), which is a major component of the extracellular matrix (ECM), is regulated during myofibroproliferative responses to numerous forms of inflammatory stimuli. It is a key factor involved in cellular migration and adherence. The development of a potent and non-toxic inhibitor of HA synthesis would open up a new avenue for the treatment of fibrocontractive diseases such as pulmonary fibrosis and liver cirrhosis. In this study, the effects of vesnarinone (OPC-8212: 3,4-dihydro-6-[4-(3, 4-dimethoxybenzoyl)-1-piperazinyl]-2(1H)-quinolinone) on the secretion of HA in human myofibroblast cell lines (MRC-5 and LI90 cells, referred to as pulmonary and hepatic myofibroblasts, respectively) were examined. Vesnarinone specifically and dose-dependently inhibited HA secretion by myofibroblasts up-regulated by fetal calf serum (FCS). The treatment of vesnarinone did not modify the phenotype of myofibroblast cells in culture. Vesnarinone also potently inhibited the HA secretion by the two myofibroblast cell lines up-regulated by transforming growth factor-beta1 (TGF-beta1) or tumor necrosis factor-alpha (TNF-alpha). The addition of vesnarinone to myofibroblasts resulted in a significant decrease of HA synthase (HAS) activity, with or without the addition of FCS or either cytokine. These findings suggest that vesnarinone inhibits the secretion of HA in myofibroblasts by specifically suppressing HAS activity, and may therefore prove useful for the treatment of chronic inflammation and tissue fibrosis.

ATP: a vasoactive signal in the pericyte-containing microvasculature of the rat retina

PubMed Central

Kawamura, Hajime; Sugiyama, Tetsuya; Wu, David M; Kobayashi, Masato; Yamanishi, Shigeki; Katsumura, Kozo; Puro, Donald G

2003-01-01

In this study we tested the hypothesis that extracellular ATP regulates the function of the pericyte-containing retinal microvessels. Pericytes, which are more numerous in the retina than in any other tissue, are abluminally located cells that may adjust capillary perfusion by contracting and relaxing. At present, knowledge of the vasoactive molecules that regulate pericyte function is limited. Here, we focused on the actions of extracellular ATP because this nucleotide is a putative glial-to-vascular signal, as well as being a substance released by activated platelets and injured cells. In microvessels freshly isolated from the adult rat retina, we monitored ionic currents via perforated-patch pipettes, measured intracellular calcium levels with the use of fura-2, and visualized microvascular contractions with the aid of time-lapse photography. We found that ATP induced depolarizing changes in the ionic currents, increased calcium levels and caused pericytes to contract. P2X7 receptors and UTP-activated receptors mediated these effects. Consistent with ATP serving as a vasoconstrictor for the pericyte-containing microvasculature of the retina, the microvascular lumen narrowed when an adjacent pericyte contracted. In addition, the sustained activation of P2X7 receptors inhibited cell-to-cell electrotonic transmission within the microvascular networks. Thus, ATP not only affects the contractility of individual pericytes, but also appears to regulate the spatial and temporal dynamics of the vasomotor response. PMID:12876212

Exosome proteomics reveals transcriptional regulator proteins with potential to mediate downstream pathways.

PubMed

Ung, Timothy H; Madsen, Helen J; Hellwinkel, Justin E; Lencioni, Alex M; Graner, Michael W

2014-11-01

Exosomes are virus-sized, membrane-enclosed vesicles with origins in the cellular endosomal system, but are released extracellularly. As a population, these tiny vesicles carry relatively enormous amounts of information in their protein, lipid and nucleic acid content, and the vesicles can have profound impacts on recipient cells. This review employs publically-available data combined with gene ontology applications to propose a novel concept, that exosomes transport transcriptional and translational machinery that may have direct impacts on gene expression in recipient cells. Here, we examine the previously published proteomic contents of medulloblastoma-derived exosomes, focusing on transcriptional regulators; we found that there are numerous proteins that may have potential roles in transcriptional and translational regulation with putative influence on downstream, cancer-related pathways. We expanded this search to all of the proteins in the Vesiclepedia database; using gene ontology approaches, we see that these regulatory factors are implicated in many of the processes involved in cancer initiation and progression. This information suggests that some of the effects of exosomes on recipient cells may be due to the delivery of protein factors that can directly and fundamentally change the transcriptional landscape of the cells. Within a tumor environment, this has potential to tilt the advantage towards the cancer. © 2014 The Authors. Cancer Science published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Cancer Association.

Sfp1 and Rtg3 reciprocally modulate carbon source‐conditional stress adaptation in the pathogenic yeast Candida albicans

PubMed Central

Kastora, Stavroula L.; Herrero‐de‐Dios, Carmen; Avelar, Gabriela M.; Munro, Carol A.

2017-01-01

Summary The pathogenicity of the clinically important yeast, Candida albicans, is dependent on robust responses to host‐imposed stresses. These stress responses have generally been dissected in vitro at 30°C on artificial growth media that do not mimic host niches. Yet host inputs, such as changes in carbon source or temperature, are known to affect C. albicans stress adaptation. Therefore, we performed screens to identify novel regulators that promote stress resistance during growth on a physiologically relevant carboxylic acid and at elevated temperatures. These screens revealed that, under these ‘non‐standard’ growth conditions, numerous uncharacterised regulators are required for stress resistance in addition to the classical Hog1, Cap1 and Cta4 stress pathways. In particular, two transcription factors (Sfp1 and Rtg3) promote stress resistance in a reciprocal, carbon source‐conditional manner. SFP1 is induced in stressed glucose‐grown cells, whereas RTG3 is upregulated in stressed lactate‐grown cells. Rtg3 and Sfp1 regulate the expression of key stress genes such as CTA4, CAP1 and HOG1 in a carbon source‐dependent manner. These mechanisms underlie the stress sensitivity of C. albicans sfp1 cells during growth on glucose, and rtg3 cells on lactate. The data suggest that C. albicans exploits environmentally contingent regulatory mechanisms to retain stress resistance during host colonisation. PMID:28574606

SUMOylation is developmentally regulated and required for cell pairing during conjugation in Tetrahymena thermophila.

PubMed

Nasir, Amjad M; Yang, Qianyi; Chalker, Douglas L; Forney, James D

2015-02-01

The covalent attachment of small ubiquitin-like modifier (SUMO) to target proteins regulates numerous nuclear events in eukaryotes, including transcription, mitosis and meiosis, and DNA repair. Despite extensive interest in nuclear pathways within the field of ciliate molecular biology, there have been no investigations of the SUMO pathway in Tetrahymena. The developmental program of sexual reproduction of this organism includes cell pairing, micronuclear meiosis, and the formation of a new somatic macronucleus. We identified the Tetrahymena thermophila SMT3 (SUMO) and UBA2 (SUMO-activating enzyme) genes and demonstrated that the corresponding green fluorescent protein (GFP) tagged gene products are found predominantly in the somatic macronucleus during vegetative growth. Use of an anti-Smt3p antibody to perform immunoblot assays with whole-cell lysates during conjugation revealed a large increase in SUMOylation that peaked during formation of the new macronucleus. Immunofluorescence using the same antibody showed that the increase was localized primarily within the new macronucleus. To initiate functional analysis of the SUMO pathway, we created germ line knockout cell lines for both the SMT3 and UBA2 genes and found both are essential for cell viability. Conditional Smt3p and Uba2p cell lines were constructed by incorporation of the cadmium-inducible metallothionein promoter. Withdrawal of cadmium resulted in reduced cell growth and increased sensitivity to DNA-damaging agents. Interestingly, Smt3p and Uba2p conditional cell lines were unable to pair during sexual reproduction in the absence of cadmium, consistent with a function early in conjugation. Our studies are consistent with multiple roles for SUMOylation in Tetrahymena, including a dynamic regulation associated with the sexual life cycle. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Hypoxia-inducing factors as master regulators of stemness properties and altered metabolism of cancer- and metastasis-initiating cells

PubMed Central

Mimeault, Murielle; Batra, Surinder K

2013-01-01

Accumulating lines of experimental evidence have revealed that hypoxia-inducible factors, HIF-1α and HIF-2α, are key regulators of the adaptation of cancer- and metastasis-initiating cells and their differentiated progenies to oxygen and nutrient deprivation during cancer progression under normoxic and hypoxic conditions. Particularly, the sustained stimulation of epidermal growth factor receptor (EGFR), insulin-like growth factor-1 receptor (IGF-1R), stem cell factor (SCF) receptor KIT, transforming growth factor-β receptors (TGF-βRs) and Notch and their downstream signalling elements such as phosphatidylinositol 3′-kinase (PI3K)/Akt/molecular target of rapamycin (mTOR) may lead to an enhanced activity of HIFs. Moreover, the up-regulation of HIFs in cancer cells may also occur in the hypoxic intratumoral regions formed within primary and secondary neoplasms as well as in leukaemic cells and metastatic prostate and breast cancer cells homing in the hypoxic endosteal niche of bone marrow. The activated HIFs may induce the expression of numerous gene products such as induced pluripotency-associated transcription factors (Oct-3/4, Nanog and Sox-2), glycolysis- and epithelial-mesenchymal transition (EMT) programme-associated molecules, including CXC chemokine receptor 4 (CXCR4), snail and twist, microRNAs and angiogenic factors such as vascular endothelial growth factor (VEGF). These gene products in turn can play critical roles for high self-renewal ability, survival, altered energy metabolism, invasion and metastases of cancer cells, angiogenic switch and treatment resistance. Consequently, the targeting of HIF signalling network and altered metabolic pathways represents new promising strategies to eradicate the total mass of cancer cells and improve the efficacy of current therapies against aggressive and metastatic cancers and prevent disease relapse. PMID:23301832

[Effect of Yiguan Decoction on differentiation of bone marrow mesenchymal stem cells into hepatocyte-like cells: an experimental research].

PubMed

Ping, Jian; Chen, Hong-Yun; Yang, Zhou; Yang, Cheng; Xu, Lie-Ming

2014-03-01

To observe the effect of Yiguan Decoction (YGD) on differentiation of bone marrow mesenchymal stem cells (BMSCs) into hepatocyte-like cells in vitro. Rat BMSCs were isolated using whole bone marrow adherent method. The properties of BMSCs were identified by analyzing the expression of surface cytokines by flow cytometry. The third passage cells were differentiated into fat cells to identify their features. BMSCs were incubated with hepatocyte growth factor (HGF) plus fibroblast growth factor 4 (FGF4) or YGD containing serum YGD for 21 days. The mRNA expression of alpha-fetoprotein (alphaAFP), albumin (Alb), and hepatocyte nuclear factor 4alpha (HNF4alpha) were detected by real time PCR. Expression of AFP and cytokeratin 18 (CK18) protein was detected by cell immunofluorescence. Glycogen synthesis was observed using periodic acid-Schiff stain (PAS). CK18, Wnt 3alpha, and alphacatenin protein expressions were detected by Western blot. High expression of CD90, CD29, and CD44, and low expression of CD34 and CD11b were observed in BMSCs isolated by whole bone mar- row adherent method, and numerous lipid droplets were observed in BMSCs using oil red O staining. Both YGD containing serum and growth factor stimulated the expression levels of Alb, AFP, HNF4alpha mRNA and CK18 protein. The down-regulated expression of Wnt 3alpha and beta-catenin could be detected at 21 days after induction. The synthesized glycogen granule could be seen. Down-regulated Wnt 3alpha and beta-catenin expression could also be observed. YGD could induce the differentiation of rat BMSCs into hepatocyte-like cells, which was related to down-regulating Wnt/beta-catenin signal pathway.

SAUR Proteins as Effectors of Hormonal and Environmental Signals in Plant Growth

PubMed Central

Ren, Hong; Gray, William M.

2016-01-01

The plant hormone auxin regulates numerous aspects of plant growth and development. Early auxin response genes mediate its genomic effects on plant growth and development. Discovered in 1987, SMALL AUXIN UP RNAs (SAURs) are the largest family of early auxin response genes. SAUR functions have remained elusive, however, presumably due to extensive genetic redundancy. However, recent molecular, genetic, biochemical, and genomic studies have implicated SAURs in the regulation of a wide range of cellular, physiological, and developmental processes. Recently, crucial mechanistic insight into SAUR function was provided by the demonstration that SAURs inhibit PP2C.D phosphatases to activate plasma membrane (PM) H+-ATPases and promote cell expansion. In addition to auxin, several other hormones and environmental factors also regulate SAUR gene expression. We propose that SAURs are key effector outputs of hormonal and environmental signals that regulate plant growth and development. PMID:25983207

Beta-lactam antibiotics modulate T-cell functions and gene expression via covalent binding to cellular albumin.

PubMed

Mor, Felix; Cohen, Irun R

2013-02-19

Recent work has suggested that beta-lactam antibiotics might directly affect eukaryotic cellular functions. Here, we studied the effects of commonly used beta-lactam antibiotics on rodent and human T cells in vitro and in vivo on T-cell-mediated experimental autoimmune diseases. We now report that experimental autoimmune encephalomyelitis and adjuvant arthritis were significantly more severe in rats treated with cefuroxime and other beta-lactams. T cells appeared to mediate the effect: an anti-myelin basic protein T-cell line treated with cefuroxime or penicillin was more encephalitogenic in adoptive transfer experiments. The beta-lactam ampicillin, in contrast to cefuroxime and penicillin, did not enhance encephalomyelitis, but did inhibit the autoimmune diabetes developing spontaneously in nonobese diabetic mice. Gene expression analysis of human peripheral blood T cells showed that numerous genes associated with T helper 2 (Th2) and T regulatory (Treg) differentiation were down-regulated in T cells stimulated in the presence of cefuroxime; these genes were up-regulated in the presence of ampicillin. The T-cell protein that covalently bound beta-lactam antibiotics was found to be albumin. Human and rodent T cells expressed albumin mRNA and protein, and penicillin-modified albumin was taken up by rat T cells, leading to enhanced encephalitogenicity. Thus, beta-lactam antibiotics in wide clinical use have marked effects on T-cell behavior; beta-lactam antibiotics can function as immunomodulators, apparently through covalent binding to albumin.

Chronic alcohol binging injures the liver and other organs by reducing NAD⁺ levels required for sirtuin's deacetylase activity.

PubMed

French, Samuel W

2016-04-01

NAD(+) levels are markedly reduced when blood alcohol levels are high during binge drinking. This causes liver injury to occur because the enzymes that require NAD(+) as a cofactor such as the sirtuin de-acetylases cannot de-acetylate acetylated proteins such as acetylated histones. This prevents the epigenetic changes that regulate metabolic processes and which prevent organ injury such as fatty liver in response to alcohol abuse. Hyper acetylation of numerous regulatory proteins develops. Systemic multi-organ injury occurs when NAD(+) is reduced. For instance the Circadian clock is altered if NAD(+) is not available. Cell cycle arrest occurs due to up regulation of cell cycle inhibitors leading to DNA damage, mutations, apoptosis and tumorigenesis. NAD(+) is linked to aging in the regulation of telomere stability. NAD(+) is required for mitochondrial renewal. Alcohol dehydrogenase is present in every visceral organ in the body so that there is a systemic reduction of NAD(+) levels in all of these organs during binge drinking. Copyright © 2016. Published by Elsevier Inc.

Polycomb-like 2 Associates with PRC2 and Regulates Transcriptional Networks during Mouse Embryonic Stem Cell Self-Renewal and Differentiation

PubMed Central

Walker, Emily; Chang, Wing Y.; Hunkapiller, Julie; Cagney, Gerard; Garcha, Kamal; Torchia, Joseph; Krogan, Nevan J.; Reiter, Jeremy F.; Stanford, William L.

2010-01-01

Summary Polycomb group (PcG) proteins are conserved epigenetic transcriptional repressors that control numerous developmental gene expression programs and have recently been implicated in modulating embryonic stem cell (ESC) fate. We identified the PcG protein PCL2 (polycomb-like 2) in a genome-wide screen for regulators of self-renewal and pluripotency and predicted that it would play an important role in mouse ESC fate determination. Using multiple biochemical strategies, we provide evidence that PCL2 is a Polycomb Repressive Complex 2 (PRC2)-associated protein in mouse ESCs. Knockdown of Pcl2 in ESCs resulted in heightened self-renewal characteristics, defects in differentiation and altered patterns of histone methylation. Integration of global gene expression and promoter occupancy analyses allowed us to identify PCL2 and PRC2 transcriptional targets and draft regulatory networks. We describe the role of PCL2 in both modulating transcription of ESC self-renewal genes in undifferentiated ESCs as well as developmental regulators during early commitment and differentiation. PMID:20144788

Vitamin D3 supplementation alleviates rotavirus infection in pigs and IPEC-J2 cells via regulating the autophagy signaling pathway.

PubMed

Tian, Gang; Liang, Xiaofang; Chen, Daiwen; Mao, Xiangbing; Yu, Jie; Zheng, Ping; He, Jun; Huang, Zhiqing; Yu, Bing

2016-10-01

Vitamin D had an anti-infection effect and benefited to the intestinal health. Autophagy signaling pathway was regulated by vitamin D3 to inhibit the infection of human immunodeficiency virus type-1. Rotavirus (RV) was a major cause of the severe diarrheal disease in young children and young animals. Although evidence suggested that vitamin D3 attenuates the negative effects of RV infection via the retinoic acid-inducible gene I signaling pathway, little is known of its antiviral effect whether through the regulation of autophagy. The present study was performed to investigate whether vitamin D3 alleviates RV infection in pig and porcine small intestinal epithelial cell line (IPEC-J2) models via regulating the autophagy signaling pathway. RV administration increased the Beclin 1 mRNA abundance in porcine jejunum and ileum. 5000 IU/kg dietary vitamin D3 supplementation greatly up-regulated LC3-II/LC3-I ratios and PR-39 mRNA expression under the condition of RV challenged. The viability of IPEC-J2 was significantly inhibited by RV infection. Incubation with 25-hydroxyvitamin D3 significantly decreased the concentrations of RV antigen and non-structural protein 4 (NSP4), and up-regulated the mRNA expression of Beclin 1 and PR-39 in the RV-infected IPEC-J2 cells. And then, based on the 25-hydroxyvitamin D3 treatment and RV infection, LC3-II mRNA expression in cells was inhibited by an autophagy inhibitor 3-methyladenine (3-MA). Bafilomycin A1 (Baf A1, a class of inhibitors of membrane ATPases, inhibits maturation of autophagic vacuoles) treatment numerically enhanced the LC3-II mRNA abundance, but had no effect on NSP4 concentration. Furthermore, 25-hydroxyvitamin D3 decreased the p62 mRNA expression and increased porcine cathelicidins (PMAP23, PG1-5 and PR-39) mRNA expression in the RV-infected cells. Taken together, these results indicated that vitamin D3 attenuates RV infection through regulating autophagic maturation and porcine cathelicidin genes expression. Copyright © 2016 Elsevier Ltd. All rights reserved.

Regulatory Oversight of Cell and Gene Therapy Products in Canada.

PubMed

Ridgway, Anthony; Agbanyo, Francisca; Wang, Jian; Rosu-Myles, Michael

2015-01-01

Health Canada regulates gene therapy products and many cell therapy products as biological drugs under the Canadian Food and Drugs Act and its attendant regulations. Cellular products that meet certain criteria, including minimal manipulation and homologous use, may be subjected to a standards-based approach under the Safety of Human Cells, Tissues and Organs for Transplantation Regulations. The manufacture and clinical testing of cell and gene therapy products (CGTPs) presents many challenges beyond those for protein biologics. Cells cannot be subjected to pathogen removal or inactivation procedures and must frequently be administered shortly after final formulation. Viral vector design and manufacturing control are critically important to overall product quality and linked to safety and efficacy in patients through concerns such as replication competence, vector integration, and vector shedding. In addition, for many CGTPs, the value of nonclinical studies is largely limited to providing proof of concept, and the first meaningful data relating to appropriate dosing, safety parameters, and validity of surrogate or true determinants of efficacy must come from carefully designed clinical trials in patients. Addressing these numerous challenges requires application of various risk mitigation strategies and meeting regulatory expectations specifically adapted to the product types. Regulatory cooperation and harmonisation at an international level are essential for progress in the development and commercialisation of these products. However, particularly in the area of cell therapy, new regulatory paradigms may be needed to harness the benefits of clinical progress in situations where the resources and motivation to pursue a typical drug product approval pathway may be lacking.

Regulatory T cells and type 2 innate lymphoid cell-dependent asthma.

PubMed

Aron, J L; Akbari, O

2017-08-01

Group 2 innate lymphoid cells (ILC2s) are a recently identified group of cells with the potent capability to produce Th2-type cytokines such as interleukin (IL)-5 and IL-13. Several studies suggest that ILC2s play an important role in the development of allergic diseases and asthma. Activation of pulmonary ILC2s in murine models lacking T and B cells induces eosinophilia and airway hyper-reactivity (AHR), which are cardinal features of asthma. More importantly, numerous recent studies have highlighted the role of ILC2s in asthma persistence and exacerbation among human subjects, and thus, regulation of pulmonary ILC2s is a major area of investigation aimed at curbing allergic lung inflammation and exacerbation. Emerging evidence reveals that a group of regulatory T cells, induced Tregs (iTregs), effectively suppress the production of ILC2-driven, pro-inflammatory cytokines IL-5 and IL-13. The inhibitory effects of iTregs are blocked by preventing direct cellular contact or by inhibiting the ICOS-ICOS-ligand (ICOSL) pathway, suggesting that both direct contact and ICOS-ICOSL interaction are important in the regulation of ILC2 function. Also, cytokines such as IL-10 and TGF-β1 significantly reduce cytokine secretion by ILC2s. Altogether, these new findings uncover iTregs as potent regulators of ILC2 activation and implicate their utility as a therapeutic approach for the treatment of ILC2-mediated allergic asthma and respiratory disease. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

The synthetic α-bromo-2',3,4,4'-tetramethoxychalcone (α-Br-TMC) inhibits the JAK/STAT signaling pathway.

PubMed

Pinz, Sophia; Unser, Samy; Brueggemann, Susanne; Besl, Elisabeth; Al-Rifai, Nafisah; Petkes, Hermina; Amslinger, Sabine; Rascle, Anne

2014-01-01

Signal transducer and activator of transcription STAT5 and its upstream activating kinase JAK2 are essential mediators of cytokine signaling. Their activity is normally tightly regulated and transient. However, constitutive activation of STAT5 is found in numerous cancers and a driving force for malignant transformation. We describe here the identification of the synthetic chalcone α-Br-2',3,4,4'-tetramethoxychalcone (α-Br-TMC) as a novel JAK/STAT inhibitor. Using the non-transformed IL-3-dependent B cell line Ba/F3 and its oncogenic derivative Ba/F3-1*6 expressing constitutively activated STAT5, we show that α-Br-TMC targets the JAK/STAT pathway at multiple levels, inhibiting both JAK2 and STAT5 phosphorylation. Moreover, α-Br-TMC alters the mobility of STAT5A/B proteins in SDS-PAGE, indicating a change in their post-translational modification state. These alterations correlate with a decreased association of STAT5 and RNA polymerase II with STAT5 target genes in chromatin immunoprecipitation assays. Interestingly, expression of STAT5 target genes such as Cis and c-Myc was differentially regulated by α-Br-TMC in normal and cancer cells. While both genes were inhibited in IL-3-stimulated Ba/F3 cells, expression of the oncogene c-Myc was down-regulated and that of the tumor suppressor gene Cis was up-regulated in transformed Ba/F3-1*6 cells. The synthetic chalcone α-Br-TMC might therefore represent a promising novel anticancer agent for therapeutic intervention in STAT5-associated malignancies.

The Respiratory Environment Diverts the Development of Antiviral Memory CD8 T Cells.

PubMed

Shane, Hillary L; Reagin, Katie L; Klonowski, Kimberly D

2018-06-01

Our understanding of memory CD8 + T cells has been largely derived from acute, systemic infection models. However, memory CD8 + T cells generated from mucosal infection exhibit unique properties and, following respiratory infection, are not maintained in the lung long term. To better understand how infection route modifies memory differentiation, we compared murine CD8 + T cell responses to a vesicular stomatitis virus (VSV) challenge generated intranasally (i.n.) or i.v. The i.n. infection resulted in greater peak expansion of VSV-specific CD8 + T cells. However, this numerical advantage was rapidly lost during the contraction phase of the immune response, resulting in memory CD8 + T cell numerical deficiencies when compared with i.v. infection. Interestingly, the antiviral CD8 + T cells generated in response to i.n. VSV exhibited a biased and sustained proportion of early effector cells (CD127 lo KLRG1 lo ) akin to the developmental program favored after i.n. influenza infection, suggesting that respiratory infection broadly favors an incomplete memory differentiation program. Correspondingly, i.n. VSV infection resulted in lower CD122 expression and eomesodermin levels by VSV-specific CD8 + T cells, further indicative of an inferior transition to bona fide memory. These results may be due to distinct (CD103 + CD11b + ) dendritic cell subsets in the i.n. versus i.v. T cell priming environments, which express molecules that regulate T cell signaling and the balance between tolerance and immunity. Therefore, we propose that distinct immunization routes modulate both the quality and quantity of antiviral effector and memory CD8 + T cells in response to an identical pathogen and should be considered in CD8 + T cell-based vaccine design. Copyright © 2018 by The American Association of Immunologists, Inc.

Genome-Scale Transcriptome Analysis in Response to Nitric Oxide in Birch Cells: Implications of the Triterpene Biosynthetic Pathway

PubMed Central

Zeng, Fansuo; Sun, Fengkun; Li, Leilei; Liu, Kun; Zhan, Yaguang

2014-01-01

Evidence supporting nitric oxide (NO) as a mediator of plant biochemistry continues to grow, but its functions at the molecular level remains poorly understood and, in some cases, controversial. To study the role of NO at the transcriptional level in Betula platyphylla cells, we conducted a genome-scale transcriptome analysis of these cells. The transcriptome of untreated birch cells and those treated by sodium nitroprusside (SNP) were analyzed using the Solexa sequencing. Data were collected by sequencing cDNA libraries of birch cells, which had a long period to adapt to the suspension culture conditions before SNP-treated cells and untreated cells were sampled. Among the 34,100 UniGenes detected, BLASTX search revealed that 20,631 genes showed significant (E-values≤10−5) sequence similarity with proteins from the NR-database. Numerous expressed sequence tags (i.e., 1374) were identified as differentially expressed between the 12 h SNP-treated cells and control cells samples: 403 up-regulated and 971 down-regulated. From this, we specifically examined a core set of NO-related transcripts. The altered expression levels of several transcripts, as determined by transcriptome analysis, was confirmed by qRT-PCR. The results of transcriptome analysis, gene expression quantification, the content of triterpenoid and activities of defensive enzymes elucidated NO has a significant effect on many processes including triterpenoid production, carbohydrate metabolism and cell wall biosynthesis. PMID:25551661

NFκB is a central regulator of protein quality control in response to protein aggregation stresses via autophagy modulation

PubMed Central

Nivon, Mathieu; Fort, Loïc; Muller, Pascale; Richet, Emma; Simon, Stéphanie; Guey, Baptiste; Fournier, Maëlenn; Arrigo, André-Patrick; Hetz, Claudio; Atkin, Julie D.; Kretz-Remy, Carole

2016-01-01

During cell life, proteins often misfold, depending on particular mutations or environmental changes, which may lead to protein aggregates that are toxic for the cell. Such protein aggregates are the root cause of numerous diseases called “protein conformational diseases,” such as myofibrillar myopathy and familial amyotrophic lateral sclerosis. To fight against aggregates, cells are equipped with protein quality control mechanisms. Here we report that NFκB transcription factor is activated by misincorporation of amino acid analogues into proteins, inhibition of proteasomal activity, expression of the R120G mutated form of HspB5 (associated with myofibrillar myopathy), or expression of the G985R and G93A mutated forms of superoxide dismutase 1 (linked to familial amyotrophic lateral sclerosis). This noncanonical stimulation of NFκB triggers the up-regulation of BAG3 and HspB8 expression, two activators of selective autophagy, which relocalize to protein aggregates. Then NFκB-dependent autophagy allows the clearance of protein aggregates. Thus NFκB appears as a central and major regulator of protein aggregate clearance by modulating autophagic activity. In this context, the pharmacological stimulation of this quality control pathway might represent a valuable strategy for therapies against protein conformational diseases. PMID:27075172

The ever-evolving role of mTOR in translation.

PubMed

Fonseca, Bruno D; Smith, Ewan M; Yelle, Nicolas; Alain, Tommy; Bushell, Martin; Pause, Arnim

2014-12-01

Control of translation allows for the production of stoichiometric levels of each protein in the cell. Attaining such a level of fine-tuned regulation of protein production requires the coordinated temporal and spatial control of numerous cellular signalling cascades impinging on the various components of the translational machinery. Foremost among these is the mTOR signalling pathway. The mTOR pathway regulates both the initiation and elongation steps of protein synthesis through the phosphorylation of numerous translation factors, while simultaneously ensuring adequate folding of nascent polypeptides through co-translational degradation of misfolded proteins. Perhaps most remarkably, mTOR is also a key regulator of the synthesis of ribosomal proteins and translation factors themselves. Two seminal studies have recently shown in translatome analysis that the mTOR pathway preferentially regulates the translation of mRNAs encoding ribosomal proteins and translation factors. Therefore, the role of the mTOR pathway in the control of protein synthesis extends far beyond immediate translational control. By controlling ribosome production (and ultimately ribosome availability), mTOR is a master long-term controller of protein synthesis. Herein, we review the literature spanning the early discoveries of mTOR on translation to the latest advances in our understanding of how the mTOR pathway controls the synthesis of ribosomal proteins. Crown Copyright © 2014. Published by Elsevier Ltd. All rights reserved.

Identification and characterization of cis-acting elements involved in the regulation of ABA- and/or GA-mediated LuPLR1 gene expression and lignan biosynthesis in flax (Linum usitatissimum L.) cell cultures.

PubMed

Corbin, Cyrielle; Renouard, Sullivan; Lopez, Tatiana; Lamblin, Frédéric; Lainé, Eric; Hano, Christophe

2013-03-15

Pinoresinol lariciresinol reductase 1, encoded by the LuPLR1 gene in flax (Linum usitatissimum L.), is responsible for the biosynthesis of (+)-secoisolariciresinol, a cancer chemopreventive phytoestrogenic lignan accumulated in high amount in the hull of flaxseed. Our recent studies have demonstrated a key role of abscisic acid (ABA) in the regulation of LuPLR1 gene expression and thus of the (+)-secoisolariciresinol synthesis during the flax seedcoat development. It is well accepted that gibberellins (GA) and ABA play antagonistic roles in the regulation of numerous developmental processes; therefore it is of interest to clarify their respective effects on lignan biosynthesis. Herein, using flax cell suspension cultures, we demonstrate that LuPLR1 gene expression and (+)-secoisolariciresinol synthesis are up-regulated by ABA and down-regulated by GA. The LuPLR1 gene promoter analysis and mutation experiments allow us to identify and characterize two important cis-acting sequences (ABRE and MYB2) required for these regulations. These results imply that a cross-talk between ABA and GA signaling orchestrated by transcription factors is involved in the regulation of lignan biosynthesis. This is particularly evidenced in the case of the ABRE cis-regulatory sequence of LuPLR1 gene promoter that appears to be a common target sequence of GA and ABA signals. Copyright © 2012 Elsevier GmbH. All rights reserved.

Human recombinant RNASET2-induced inflammatory response and connective tissue remodeling in the medicinal leech.

PubMed

Baranzini, Nicolò; Pedrini, Edoardo; Girardello, Rossana; Tettamanti, Gianluca; de Eguileor, Magda; Taramelli, Roberto; Acquati, Francesco; Grimaldi, Annalisa

2017-05-01

In recent years, several studies have demonstrated that the RNASET2 gene is involved in the control of tumorigenicity in ovarian cancer cells. Furthermore, a role in establishing a functional cross-talk between cancer cells and the surrounding tumor microenvironment has been unveiled for this gene, based on its ability to act as an inducer of the innate immune response. Although several studies have reported on the molecular features of RNASET2, the details on the mechanisms by which this evolutionarily conserved ribonuclease regulates the immune system are still poorly defined. In the effort to clarify this aspect, we report here the effect of recombinant human RNASET2 injection and its role in regulating the innate immune response after bacterial challenge in an invertebrate model, the medicinal leech. We found that recombinant RNASET2 injection induces fibroplasias, connective tissue remodeling and the recruitment of numerous infiltrating cells expressing the specific macrophage markers CD68 and HmAIF1. The RNASET2-mediated chemotactic activity for macrophages has been further confirmed by using a consolidated experimental approach based on injection of the Matrigel biomatrice (MG) supplemented with recombinant RNASET2 in the leech body wall. One week after injection, a large number of CD68 + and HmAIF-1 + macrophages massively infiltrated MG sponges. Finally, in leeches challenged with lipopolysaccharides (LPS) or with the environmental bacteria pathogen Micrococcus nishinomiyaensis, numerous macrophages migrating to the site of inoculation expressed high levels of endogenous RNASET2. Taken together, these results suggest that RNASET2 is likely involved in the initial phase of the inflammatory response in leeches.

The ER-mitochondria interface: the social network of cell death.

PubMed

Grimm, Stefan

2012-02-01

When cellular organelles communicate bad things can happen. Recent findings uncovered that the junction between the endoplasmic reticulum (ER) and the mitochondria holds a crucial role for cell death regulation. Not only does this locale connect the two best-known organelles in apoptosis, numerous regulators of cell death are concentrated at this spot, providing a terrain for intense signal transfers. Ca2+ is the most prominent signalling factor that is released from the ER and, at high concentration, mediates the transfer of an apoptosis signal to mitochondria as the executioner organelle for cell death. An elaborate array of checks and balances is fine-tuning this process including Bcl-2 family members. Moreover, MAMs, "mitochondria-associated membranes", are distinct membrane sections at the ER that are in close contact with mitochondria and have been found to exchange lipids and lipid-derived molecules such as ceramide for apoptosis induction. Recent work has also described a reverse transfer of apoptosis signals, from mitochondria to the ER, via cytochrome c release and prolonged IP3R opening or through the mitochondrial fission factor Fis1 and Bap31 at the ER, which form the ARCosome, a novel caspase-activation complex. Copyright © 2011 Elsevier B.V. All rights reserved.

The Role of the Calcium-sensing Receptor in Cancer

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rodland, Karin D.

2004-03-01

The cell surface calcium receptor (Ca2+ receptor) is a particularly difficult receptor to study because its primary physiological ligand, Ca2+, affects numerous biological processes both within and outside of cells. Because of this, distinguishing effects of extracellular Ca2+ mediated by the Ca2+ receptor from those mediated by other mechanisms is challenging. Certain pharmacological approaches, however, when combined with appropriate experimental designs, can be used to more confidently identify cellular responses regulated by the Ca2+ receptor and select those that might be targeted therapeutically. The Ca2+ receptor on parathyroid cells, because it is the primary mechanism regulating secretion of parathyroid hormonemore » (PTH), is one such target. Calcimimetic compounds, which active this Ca2+ receptor and lower circulating levels of PTH, have been developed for treating hyperparathyroidism. The converse pharmaceutical approach, involving calcilytic compounds that block parathyroid cell Ca2+ receptors and stimulate PTH secretion thereby providing an anabolic therapy for osteoporosis, still awaits clinical validation. Although Ca2+ receptors are expressed throughout the body and in many tissues that are not intimately involved in systemic Ca2+ homeostasis, their physiological and/or pathological significance remains speculative and their value as therapeutic targets is unknown.« less

GILZ overexpression attenuates endoplasmic reticulum stress-mediated cell death via the activation of mitochondrial oxidative phosphorylation

DOE Office of Scientific and Technical Information (OSTI.GOV)

André, Fanny; Corazao-Rozas, Paola; Idziorek, Thierry

The Glucocorticoïd-induced leucine zipper (GILZ) protein has profound anti-inflammatory activities in haematopoietic cells. GILZ regulates numerous signal transduction pathways involved in proliferation and survival of normal and neoplastic cells. Here, we have demonstrated the potential of GILZ in alleviating apoptosis induced by ER stress inducers. Whereas the glucocorticoid, dexamethasone, protects from tunicamycin-induced cell death, silencing endogeneous GILZ in dexamethasone-treated cancer cells alter the capacity of glucocorticoids to protect from tunicamycin-mediated apoptosis. Under ER stress conditions, overexpression of GILZ significantly reduced activation of mitochondrial pathway of apoptosis by maintaining Bcl-xl level. GILZ protein affects the UPR signaling shifting the balance towardsmore » pro-survival signals as judged by down-regulation of CHOP, ATF4, XBP1s mRNA and increase in GRP78 protein level. Interestingly, GILZ sustains high mitochondrial OXPHOS during ER stress and cytoprotection mediated by GILZ is abolished in cells depleted of mitochondrial DNA, which are OXPHOS-deficient. These findings reveal a new role of GILZ, which acts as a cytoprotector against ER stress through a pathway involving mitochondrial OXPHOS. - Highlights: • GILZ attenuates apoptotic cell death induced by ER stress conditions. • GILZ promotes pro-survival signaling of the UPR. • GILZ overexpression sustains high mitochondrial activity under ER stress. • Mitochondrial OXPHOX is required for GILZ protective effects against ER stress-mediated apoptosis.« less

Pyrimidine tract-binding protein 1 mediates pyruvate kinase M2-dependent phosphorylation of signal transducer and activator of transcription 3 and oncogenesis in anaplastic large cell lymphoma.

PubMed

Hwang, Steven R; Murga-Zamalloa, Carlos; Brown, Noah; Basappa, Johnvesly; McDonnell, Scott Rp; Mendoza-Reinoso, Veronica; Basrur, Venkatesha; Wilcox, Ryan; Elenitoba-Johnson, Kojo; Lim, Megan S

2017-08-01

PKM2 (pyruvate kinase M2), a critical regulator of glycolysis, is phosphorylated by numerous growth factor receptors and oncogenic tyrosine kinases including NPM-ALK which is expressed in a subset of aggressive T-cell non-Hodgkin lymphomas known as anaplastic large cell lymphoma, ALK-positive. Our previous work demonstrated that phosphorylation of Y105-PKM2 by NPM-ALK regulates a major metabolic shift to promote lymphomagenesis. In addition to its role in metabolism, recent studies have shown that PKM2 promotes oncogenesis by phosphorylating nuclear STAT3 (signal transducer and activator of transcription 3) and regulating transcription of genes involved in cell survival and proliferation. We hypothesized that identification of novel PKM2 interactors could provide additional insights into its expanding functional role in cancer. To this end, immunocomplexes of FLAG-tagged PKM2 were isolated from NPM-ALK-positive ALCL (anaplastic large cell lymphoma) cells and subjected to liquid chromatography tandem mass spectrometry (LC-MS/MS) which led to the identification of polypyrimidine tract-binding protein (PTBP1) as a novel interactor of PKM2. The interaction between PTBP1 and PKM2 was restricted to the nucleus and was dependent on NPM-ALK mediated Y105 phosphorylation of PKM2. Stable shRNA-mediated silencing of PTBP1 resulted in a marked decrease in pY105-PKM2 and pY705-STAT3 which led to decreased ALCL cell proliferation and colony formation. Overall, our data demonstrate that PTBP1 interacts with PKM2 and promotes ALCL oncogenesis by facilitating PKM2-dependent activation of STAT3 within the nucleus.

Aft2, a Novel Transcription Regulator, Is Required for Iron Metabolism, Oxidative Stress, Surface Adhesion and Hyphal Development in Candida albicans

PubMed Central

Xu, Ning; Cheng, Xinxin; Yu, Qilin; Qian, Kefan; Ding, Xiaohui; Liu, Ruming; Zhang, Biao; Xing, Laijun; Li, Mingchun

2013-01-01

Morphological transition and iron metabolism are closely relevant to Candida albicans pathogenicity and virulence. In our previous study, we demonstrated that C. albicans Aft2 plays an important role in ferric reductase activity and virulence. Here, we further explored the roles of C. albicans Aft2 in numerous cellular processes. We found that C. albicans Aft2 exhibited an important role in iron metabolism through bi-directional regulation effects on iron-regulon expression. Deletion of AFT2 reduced cellular iron accumulation under iron-deficient conditions. Furthermore, both reactive oxygen species (ROS) generation and superoxide dismutase (SOD) activity were remarkably increased in the aft2Δ/Δ mutant, which were thought to be responsible for the defective responses to oxidative stress. However, we found that over-expression of C. albicans AFT2 under the regulation of the strong PGK1 promoter could not effectively rescue Saccharomyces cerevisiae aft1Δ mutant defects in some cellular processes, such as cell-wall assembly, ion homeostasis and alkaline resistance, suggesting a possibility that C. albicans Aft2 weakened its functional role of regulating some cellular metabolism during the evolutionary process. Interestingly, deletion of AFT2 in C. albicans increased cell surface hydrophobicity, cell flocculation and the ability of adhesion to polystyrene surfaces. In addition, our results also revealed that C. albicans Aft2 played a dual role in regulating hypha-specific genes under solid and liquid hyphal inducing conditions. Deletion of AFT2 caused an impaired invasive growth in solid medium, but an increased filamentous aggregation and growth in liquid conditions. Moreover, iron deficiency and environmental cues induced nuclear import of Aft2, providing additional evidence for the roles of Aft2 in transcriptional regulation. PMID:23626810

Osteoporosis and its association with non-gonadal hormones involved in hypertension, adiposity and hyperglycaemia.

PubMed

Poudyal, Hemant; Brown, Lindsay

2013-12-01

Osteoporosis is a high-prevalence disease, particularly in developed countries, and results in high costs both to the individual and to society through associated fragility fractures. There is an urgent need for identification of novel drug targets and development of new anti-osteoporotic agents. Between 30 and 80% of osteoporotic fractures cannot be prevented despite current treatments achieving relative fracture risk reduction of up to 20%, 50%, and 70% for non-vertebral, hip and spine fractures, respectively. Traditionally, the decline in gonadal hormones has been studied as the sole hormonal determinant for the loss of bone mineral density in osteoporosis. However, recent studies have identified receptors for numerous non-gonadal hormones such as PTH, angiotensin II, leptin, adiponectin, insulin and insulin-like growth factor-1 on the osteoblast lineage cells that directly regulate bone turnover. These hormones are also involved in the pathogenesis of metabolic syndrome risk factors, particularly hypertension, type-II diabetes and obesity. By activating their respective receptors on osteoblastic lineage cells, these hormones appear to act through a common mechanism by down-regulating receptors for activation of nuclear factor kappa B ligand (RANKL) and up-regulating osteoprotegerin (OPG) with inverse responses for adiponectin. Receptors for amylin, gastric inhibitory polypeptide and ghrelin and have also been identified on the osteoblast lineage cells although the roles of these receptors in bone turnover are controversial or poorly studied. Moreover, bone turnover may be independently regulated by modulation of osteoclast-osteoblast function and bone marrow adiposity. Leptin appears to be the only hormone that is a known regulator of both bone mineralisation and bone adiposity.

Myostatin-induced inhibition of the long noncoding RNA Malat1 is associated with decreased myogenesis.

PubMed

Watts, Rani; Johnsen, Virginia L; Shearer, Jane; Hittel, Dustin S

2013-05-15

Myostatin, a member of the transforming growth factor-β (TGF-β) superfamily of secreted proteins, is a potent negative regulator of myogenesis. Free myostatin induces the phosphorylation of the Smad family of transcription factors, which, in turn, regulates gene expression, via the canonical TGF-β signaling pathway. There is, however, emerging evidence that myostatin can regulate gene expression independent of Smad signaling. As such, we acquired global gene expression data from the gastrocnemius muscle of C57BL/6 mice following a 6-day treatment with recombinant myostatin compared with vehicle-treated animals. Of the many differentially expressed genes, the myostatin-associated decrease (-11.20-fold; P < 0.05) in the noncoding metastasis-associated lung adenocarcinoma transcript 1 (Malat1) was the most significant and the most intriguing because of numerous reports describing its novel role in regulating cell growth. We therefore sought to further characterize the role of Malat1 expression in skeletal muscle myogenesis. RT-PCR-based quantification of C2C12 and primary human skeletal muscle cells revealed a significant and persistent upregulation (4- to 7-fold; P < 0.05) of Malat1 mRNA during the differentiation of myoblasts into myotubes. Conversely, targeted knockdown of Malat1 using siRNA suppressed myoblast proliferation by arresting cell growth in the G(0)/G(1) phase. These results reveal Malat1 as novel downstream target of myostatin with a considerable ability to regulate myogenesis. The identification of new targets of myostatin will have important repercussions for regenerative biology through inhibition and/or reversal of muscle atrophy and wasting diseases.

Regulation of MicroRNAs, and the Correlations of MicroRNAs and Their Targeted Genes by Zinc Oxide Nanoparticles in Ovarian Granulosa Cells

PubMed Central

Zhao, Yong; Li, Lan; Min, Ling-Jiang; Zhu, Lian-Qin; Sun, Qing-Yuan; Zhang, Hong-Fu; Liu, Xin-Qi; Zhang, Wei-Dong; Ge, Wei; Wang, Jun-Jie; Liu, Jing-Cai

2016-01-01

Zinc oxide (ZnO) nanoparticles (NPs) have been applied in numerous industrial products and personal care products like sunscreens and cosmetics. The released ZnO NPs from consumer and household products into the environment might pose potential health issues for animals and humans. In this study the expression of microRNAs and the correlations of microRNAs and their targeted genes in ZnO NPs treated chicken ovarian granulosa cells were investigated. ZnSO4 was used as the sole Zn2+ provider to differentiate the effects of NPs from Zn2+. It was found that ZnO-NP-5 μg/ml specifically regulated the expression of microRNAs involved in embryonic development although ZnO-NP-5 μg/ml and ZnSO4-10 μg/ml treatments produced the same intracellular Zn concentrations and resulted in similar cell growth inhibition. And ZnO-NP-5 μg/ml also specifically regulated the correlations of microRNAs and their targeted genes. This is the first investigation that intact NPs in ZnO-NP-5 μg/ml treatment specifically regulated the expression of microRNAs, and the correlations of microRNAs and their targeted genes compared to that by Zn2+. This expands our knowledge for biological effects of ZnO NPs and at the same time it raises the health concerns that ZnO NPs might adversely affect our biological systems, even the reproductive systems through regulation of specific signaling pathways. PMID:27196542

Regulation of MicroRNAs, and the Correlations of MicroRNAs and Their Targeted Genes by Zinc Oxide Nanoparticles in Ovarian Granulosa Cells.

PubMed

Zhao, Yong; Li, Lan; Min, Ling-Jiang; Zhu, Lian-Qin; Sun, Qing-Yuan; Zhang, Hong-Fu; Liu, Xin-Qi; Zhang, Wei-Dong; Ge, Wei; Wang, Jun-Jie; Liu, Jing-Cai; Hao, Zhi-Hui

2016-01-01

Zinc oxide (ZnO) nanoparticles (NPs) have been applied in numerous industrial products and personal care products like sunscreens and cosmetics. The released ZnO NPs from consumer and household products into the environment might pose potential health issues for animals and humans. In this study the expression of microRNAs and the correlations of microRNAs and their targeted genes in ZnO NPs treated chicken ovarian granulosa cells were investigated. ZnSO4 was used as the sole Zn2+ provider to differentiate the effects of NPs from Zn2+. It was found that ZnO-NP-5 μg/ml specifically regulated the expression of microRNAs involved in embryonic development although ZnO-NP-5 μg/ml and ZnSO4-10 μg/ml treatments produced the same intracellular Zn concentrations and resulted in similar cell growth inhibition. And ZnO-NP-5 μg/ml also specifically regulated the correlations of microRNAs and their targeted genes. This is the first investigation that intact NPs in ZnO-NP-5 μg/ml treatment specifically regulated the expression of microRNAs, and the correlations of microRNAs and their targeted genes compared to that by Zn2+. This expands our knowledge for biological effects of ZnO NPs and at the same time it raises the health concerns that ZnO NPs might adversely affect our biological systems, even the reproductive systems through regulation of specific signaling pathways.

Final Technical Report

DOE Office of Scientific and Technical Information (OSTI.GOV)

Michael Laub

2008-12-29

Our team of investigators from MIT (Michael Laub) and Stanford (Harley McAdams and Lucy Shapiro) conducted a multi-faceted, systematic experimental analysis of the 106 Caulobacter two-component signal transduction system proteins (62 histidine kinases and 44 response regulators) to understand how they coordinate cell cycle progression, metabolism, and response to environmental changes. These two-component signaling proteins were characterized at the genetic, biochemical, and genomic levels. The results generated by our laboratories have provided numerous insights into how Caulobacter cells sense and respond to a myriad of signals. As nearly all bacteria use two-component signaling for cell regulation, the results from thismore » project help to deepen our general understanding of bacterial signal transduction. The tools and approaches developed can be applied to other bacteria. In particular, work from the Laub laboratory now enables the systematic, rational rewiring of two-component signaling proteins, a major advance that stands to impact synthetic biology and the development of biosensors and other designer molecular circuits. Results are summarized from our work. Each section lists publications and publicly-available resources which result from the work described.« less

Thymic function in the regulation of T cells, and molecular mechanisms underlying the modulation of cytokines and stress signaling (Review).

PubMed

Yan, Fenggen; Mo, Xiumei; Liu, Junfeng; Ye, Siqi; Zeng, Xing; Chen, Dacan

2017-11-01

The thymus is critical in establishing and maintaining the appropriate microenvironment for promoting the development and selection of T cells. The function and structure of the thymus gland has been extensively studied, particularly as the thymus serves an important physiological role in the lymphatic system. Numerous studies have investigated the morphological features of thymic involution. Recently, research attention has increasingly been focused on thymic proteins as targets for drug intervention. Omics approaches have yielded novel insights into the thymus and possible drug targets. The present review addresses the signaling and transcriptional functions of the thymus, including the molecular mechanisms underlying the regulatory functions of T cells and their role in the immune system. In addition, the levels of cytokines secreted in the thymus have a significant effect on thymic functions, including thymocyte migration and development, thymic atrophy and thymic recovery. Furthermore, the regulation and molecular mechanisms of stress‑mediated thymic atrophy and involution were investigated, with particular emphasis on thymic function as a potential target for drug development and discovery using proteomics.

Extracellular calcium acts as a “third messenger” to regulate enzyme and alkaline secretion

PubMed Central

Caroppo, Rosa; Gerbino, Andrea; Fistetto, Gregorio; Colella, Matilde; Debellis, Lucantonio; Hofer, Aldebaran M.; Curci, Silvana

2004-01-01

It is generally assumed that the functional consequences of stimulation with Ca2+-mobilizing agonists are derived exclusively from the second messenger action of intracellular Ca2+, acting on targets inside the cells. However, during Ca2+ signaling events, Ca2+ moves in and out of the cell, causing changes not only in intracellular Ca2+, but also in local extracellular Ca2+. The fact that numerous cell types possess an extracellular Ca2+ “sensor” raises the question of whether these dynamic changes in external [Ca2+] may serve some sort of messenger function. We found that in intact gastric mucosa, the changes in extracellular [Ca2+] secondary to carbachol-induced increases in intracellular [Ca2+] were sufficient and necessary to elicit alkaline secretion and pepsinogen secretion, independent of intracellular [Ca2+] changes. These findings suggest that extracellular Ca2+ can act as a “third messenger” via Ca2+ sensor(s) to regulate specific subsets of tissue function previously assumed to be under the direct control of intracellular Ca2+. PMID:15240573

Inhibition of Curcumin on ZAKα Activity Resultant in Apoptosis and Anchorage-Independent Growth in Cancer Cells.

PubMed

Lee, Jin-Sun; Wang, Tsu-Shing; Lin, Ming Cheng; Lin, Wei-Wen; Yang, Jaw-Ji

2017-10-31

Curcumin, a popular yellow pigment of the dietary spice turmeric, has been reported to inhibit cell growth and to induce apoptosis in a wide variety of cancer cells. Although numerous studies have investigated anticancer effects of curcumin, the precise molecular mechanism of action remains unidentified. Whereas curcumin mediates cell survival and apoptosis through mitogen-activated protein kinase (MAPK) and nuclear factor-kappa B (NF-κB) signaling cascades, its impact on the upstream regulation of MAPK is unclear. The leucine-zipper and sterile-α motif kinase alpha (ZAKα), a mitogen-activated protein kinase kinase kinase (MAP3K), activates the c-Jun N-terminal kinase (JNK) and NF-κB pathway. This paper investigated the prospective involvement of ZAKα in curcumin-induced effects on cancer cells. Our results suggest that the antitumor activity of curcumin is mediated via a mechanism involving inhibition of ZAKα activity.

Mitochondrial Calcium Transport in Trypanosomes

PubMed Central

Docampo, Roberto; Vercesi, Anibal E.; Huang, Guozhong

2014-01-01

The biochemical peculiarities of trypanosomes were fundamental for the recent molecular identification of the long-sought channel involved in mitochondrial Ca2+ uptake, the mitochondrial Ca2+ uniporter or MCU. This discovery led to the finding of numerous regulators of the channel, which form a high molecular weight complex with MCU. Some of these regulators have been bioinformatically identified in trypanosomes, which are the first eukaryotic organisms described for which MCU is essential. In trypanosomes MCU is important for buffering cytosolic Ca2+ changes and for activation of the bioenergetics of the cells. Future work on this pathway in trypanosomes promises further insight into the biology of these fascinating eukaryotes, as well as the potential for novel target discovery. PMID:25218432

Requirement of matrix metalloproteinase-1 for intestinal homeostasis in the adult Drosophila midgut

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Shin-Hae; Park, Joung-Sun; Kim, Young-Shin

Stem cells are tightly regulated by both intrinsic and extrinsic signals as well as the extracellular matrix (ECM) for tissue homeostasis and regenerative capacity. Matrix metalloproteinases (MMPs), proteolytic enzymes, modulate the turnover of numerous substrates, including cytokine precursors, growth factors, and ECM molecules. However, the roles of MMPs in the regulation of adult stem cells are poorly understood. In the present study, we utilize the Drosophila midgut, which is an excellent model system for studying stem cell biology, to show that Mmp1 is involved in the regulation of intestinal stem cells (ISCs). The results showed that Mmp1 is expressed inmore » the adult midgut and that its expression increases with age and with exposure to oxidative stress. Mmp1 knockdown or Timp-overexpressing flies and flies heterozygous for a viable, hypomorphic Mmp1 allele increased ISC proliferation in the gut, as shown by staining with an anti-phospho-histone H3 antibody and BrdU incorporation assays. Reduced Mmp1 levels induced intestinal hyperplasia, and the Mmp1depletion-induced ISC proliferation was rescued by the suppression of the EGFR signaling pathway, suggesting that Mmp1 regulates ISC proliferation through the EGFR signaling pathway. Furthermore, adult gut-specific knockdown and whole-animal heterozygotes of Mmp1 increased additively sensitivity to paraquat-induced oxidative stress and shortened lifespan. Our data suggest that Drosophila Mmp1 is involved in the regulation of ISC proliferation for maintenance of gut homeostasis. -- Highlights: Black-Right-Pointing-Pointer Mmp1 is expressed in the adult midgut. Black-Right-Pointing-Pointer Mmp1 is involved in the regulation of ISC proliferation activity. Black-Right-Pointing-Pointer Mmp1-related ISC proliferation is associated with EGFR signaling. Black-Right-Pointing-Pointer Mmp1 in the gut is required for the intestinal homeostasis and longevity.« less

Innate defense regulator IDR-1018 activates human mast cells through G protein-, phospholipase C-, MAPK- and NF-ĸB-sensitive pathways.

PubMed

Yanashima, Kensuke; Chieosilapatham, Panjit; Yoshimoto, Eri; Okumura, Ko; Ogawa, Hideoki; Niyonsaba, François

2017-08-01

Host defense (antimicrobial) peptides not only display antimicrobial activities against numerous pathogens but also exert a broader spectrum of immune-modulating functions. Innate defense regulators (IDRs) are a class of host defense peptides synthetically developed from natural or endogenous cationic host defense peptides. Of the IDRs developed to date, IDR-1018 is more efficient not only in killing bacteria but also in regulating the various functions of macrophages and neutrophils and accelerating the wound healing process. Because mast cells intimately participate in wound healing and a number of host defense peptides involved in wound healing are also known to activate mast cells, this study aimed to investigate the effects of IDR-1018 on mast cell activation. Here, we showed that IDR-1018 induced the degranulation of LAD2 human mast cells and caused their production of leukotrienes, prostaglandins and various cytokines and chemokines, including granulocyte-macrophage colony-stimulating factor, interleukin-8, monocyte chemoattractant protein-1 and -3, macrophage-inflammatory protein-1α and -1β, and tumor necrosis factor-α. Furthermore, IDR-1018 increased intracellular calcium mobilization and induced mast cell chemotaxis. The mast cell activation was markedly suppressed by pertussis toxin, U-73122, U0126, SB203580, JNK inhibitor II, and NF-κB activation inhibitor II, suggesting the involvement of G-protein, phospholipase C, ERK, p38, JNK and NF-κB pathways, respectively, in IDR-1018-induced mast cell activation. Notably, we confirmed that IDR-1018 caused the phosphorylation of MAPKs and IκB. Altogether, the current study suggests a novel immunomodulatory role of IDR-1018 through its ability to recruit and activate human mast cells at the sites of inflammation and wounds. We report that IDR-1018 stimulates various functions of human mast cells. IDR-1018-induced mast cell activation is mediated through G protein, PLC, MAPK and NF-κB pathways. IDR-1018 will be a useful therapeutic agent for wound healing.

Coordinated regulation by two VPS9 domain-containing guanine nucleotide exchange factors in small GTPase Rab5 signaling pathways in fission yeast

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tsukamoto, Yuta; Kagiwada, Satoshi; Shimazu, Sayuri

The small GTPase Rab5 is reported to regulate various cellular functions, such as vesicular transport and endocytosis. VPS9 domain-containing proteins are thought to activate Rab5(s) by their guanine-nucleotide exchange activities. Numerous VPS9 proteins have been identified and are structurally conserved from yeast to mammalian cells. However, the functional relationships among VPS9 proteins in cells remain unclear. Only one Rab5 and two VPS9 proteins were identified in the Schizosaccharomyces pombe genome. Here, we examined the cellular function of two VPS9 proteins and the relationship between these proteins in cellular functions. Vps901-GFP and Vps902-GFP exhibited dotted signals in vegetative and differentiated cells.more » vps901 deletion mutant (Δvps901) cells exhibited a phenotype deficient in the mating process and responses to high concentrations of ions, such as calcium and metals, and Δvps901Δvps902 double mutant cells exhibited round cell shapes similar to ypt5-909 (Rab5 mutant allele) cells. Deletion of both vps901 and vps902 genes completely abolished the mating process and responses to various stresses. A lack of vacuole formation and aberrant inner cell membrane structures were also observed in Δvps901Δvps902 cells by electron microscopy. These data strongly suggest that Vps901 and Vps902 are cooperatively involved in the regulation of cellular functions, such as cell morphology, sexual development, response to ion stresses, and vacuole formation, via Rab5 signaling pathways in fission yeast cells. - Highlights: • Roles of Rab5 activator VPS9 proteins in cellular functions. • Cooperation between VPS9 proteins in Rab5 signaling pathway. • Roles of each VPS9 protein in Rab5 signaling pathway are discussed.« less

Usp7 promotes medulloblastoma cell survival and metastasis by activating Shh pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhan, Meixiao; Zhuhai Precision Medicine Center, Zhuhai People's Hospital, Jinan University, Zhuhai; Sun, Xiaohan

The ubiquitin-specific protease Usp7 plays roles in multiple cellular processes through deubiquitinating and stabilizing numerous substrates, including P53, Pten and Gli. Aberrant Usp7 activity has been implicated in many disorders and tumorigenesis, making it as a potential target for therapeutic intervention. Although it is clear that Usp7 is involved in many types of cancer, its role in regulating medulloblastoma (MB) is still unknown. In this study, we show that knockdown of Usp7 inhibits the proliferation and migration of MB cells, while Usp7 overexpression exerts an opposite effect. Furthermore, we establish Usp7 knockout MB cell line using the CRISPR/Cas9 system andmore » further confirm that Usp7 knockout also blocks MB cell proliferation and metastasis. In addition, we reveal that knockdown of Usp7 compromises Shh pathway activity and decrease Gli protein levels, while P53 level and P53 target gene expression have no obvious changes. Finally, we find that Usp7 inhibitors apparently inhibit MB cell viability and migration. Taken together, our findings suggest that Usp7 is important for MB cell proliferation and metastasis by activating Shh pathway, and is a putative therapeutic target for MBs. - Highlights: • Loss of usp7 blocks the proliferation and metastasis of MB cells. • Usp7 regulates MB cell growth and migration through stimulating Shh pathway. • Usp7 inhibitors hamper MB cell proliferation and migration. • Usp7 inhibitors could attenuate Shh pathway activity.« less

Determinants of cell-to-cell variability in protein kinase signaling.

PubMed

Jeschke, Matthias; Baumgärtner, Stephan; Legewie, Stefan

2013-01-01

Cells reliably sense environmental changes despite internal and external fluctuations, but the mechanisms underlying robustness remain unclear. We analyzed how fluctuations in signaling protein concentrations give rise to cell-to-cell variability in protein kinase signaling using analytical theory and numerical simulations. We characterized the dose-response behavior of signaling cascades by calculating the stimulus level at which a pathway responds ('pathway sensitivity') and the maximal activation level upon strong stimulation. Minimal kinase cascades with gradual dose-response behavior show strong variability, because the pathway sensitivity and the maximal activation level cannot be simultaneously invariant. Negative feedback regulation resolves this trade-off and coordinately reduces fluctuations in the pathway sensitivity and maximal activation. Feedbacks acting at different levels in the cascade control different aspects of the dose-response curve, thereby synergistically reducing the variability. We also investigated more complex, ultrasensitive signaling cascades capable of switch-like decision making, and found that these can be inherently robust to protein concentration fluctuations. We describe how the cell-to-cell variability of ultrasensitive signaling systems can be actively regulated, e.g., by altering the expression of phosphatase(s) or by feedback/feedforward loops. Our calculations reveal that slow transcriptional negative feedback loops allow for variability suppression while maintaining switch-like decision making. Taken together, we describe design principles of signaling cascades that promote robustness. Our results may explain why certain signaling cascades like the yeast pheromone pathway show switch-like decision making with little cell-to-cell variability.

Eosinophil-derived CCL-6 impairs hematopoietic stem cell homeostasis

PubMed Central

Zhang, Chao; Yi, Weiwei; Li, Fei; Du, Xufei; Wang, Hu; Wu, Ping; Peng, Chao; Luo, Man; Hua, Wen; Wong, Catherine CL; Lee, James J; Li, Wen; Chen, Zhihua; Ying, Songmin; Ju, Zhenyu; Shen, Huahao

2018-01-01

Eosinophils (Eos) have been long considered as end-stage effector cells in the hierarchical hematopoietic system. Numerous lines of evidence have suggested that Eos are multifunctional leukocytes with respect to the initiation, propagation and regulation of various inflammatory or immune reactions, especially in allergic diseases. Recent studies have shown that Eos are also required for maintenance of bone marrow plasma cells and differentiation of B cells. However, it remains unclear whether Eos contributes to regulation of hematopoietic stem cell (HSC) homeostasis. Here, we demonstrate that Eos disrupt HSC homeostasis by impairing HSC quiescence and reconstitution ability in wild-type mice following ovalbumin (OVA) challenge and even by causing bone marrow HSC failure and exhaustion in Cd3δ-Il-5 transgenic mice. The impaired maintenance and function of HSCs were associated with Eos-induced redox imbalance (increased oxidative phosphorylation and decreased anti-oxidants levels). More importantly, using mass spectrometry, we determined that CCL-6 is expressed at a high level under eosinophilia. We demonstrate that CCL-6 is Eos-derived and responsible for the impaired HSC homeostasis. Interestingly, blockage of CCL-6 with a specific neutralizing antibody, restored the reconstitution ability of HSCs while exacerbating eosinophilia airway inflammation in OVA-challenged mice. Thus, our study reveals an unexpected function of Eos/CCL-6 in HSC homeostasis. PMID:29327730

Eosinophil-derived CCL-6 impairs hematopoietic stem cell homeostasis.

PubMed

Zhang, Chao; Yi, Weiwei; Li, Fei; Du, Xufei; Wang, Hu; Wu, Ping; Peng, Chao; Luo, Man; Hua, Wen; Wong, Catherine Cl; Lee, James J; Li, Wen; Chen, Zhihua; Ying, Songmin; Ju, Zhenyu; Shen, Huahao

2018-03-01

Eosinophils (Eos) have been long considered as end-stage effector cells in the hierarchical hematopoietic system. Numerous lines of evidence have suggested that Eos are multifunctional leukocytes with respect to the initiation, propagation and regulation of various inflammatory or immune reactions, especially in allergic diseases. Recent studies have shown that Eos are also required for maintenance of bone marrow plasma cells and differentiation of B cells. However, it remains unclear whether Eos contributes to regulation of hematopoietic stem cell (HSC) homeostasis. Here, we demonstrate that Eos disrupt HSC homeostasis by impairing HSC quiescence and reconstitution ability in wild-type mice following ovalbumin (OVA) challenge and even by causing bone marrow HSC failure and exhaustion in Cd3δ-Il-5 transgenic mice. The impaired maintenance and function of HSCs were associated with Eos-induced redox imbalance (increased oxidative phosphorylation and decreased anti-oxidants levels). More importantly, using mass spectrometry, we determined that CCL-6 is expressed at a high level under eosinophilia. We demonstrate that CCL-6 is Eos-derived and responsible for the impaired HSC homeostasis. Interestingly, blockage of CCL-6 with a specific neutralizing antibody, restored the reconstitution ability of HSCs while exacerbating eosinophilia airway inflammation in OVA-challenged mice. Thus, our study reveals an unexpected function of Eos/CCL-6 in HSC homeostasis.

Metabolic Enzymes Enjoying New Partnerships as RNA-Binding Proteins.

PubMed

Castello, Alfredo; Hentze, Matthias W; Preiss, Thomas

2015-12-01

In the past century, few areas of biology advanced as much as our understanding of the pathways of intermediary metabolism. Initially considered unimportant in terms of gene regulation, crucial cellular fate changes, cell differentiation, or malignant transformation are now known to involve 'metabolic remodeling' with profound changes in the expression of many metabolic enzyme genes. This review focuses on the recent identification of RNA-binding activity of numerous metabolic enzymes. We discuss possible roles of this unexpected second activity in feedback gene regulation ('moonlighting') and/or in the control of enzymatic function. We also consider how metabolism-driven post-translational modifications could regulate enzyme-RNA interactions. Thus, RNA emerges as a new partner of metabolic enzymes with far-reaching possible consequences to be unraveled in the future. Copyright © 2015 Elsevier Ltd. All rights reserved.

Regulation of Endoplasmic Reticulum-Mitochondria Ca2+ Transfer and Its Importance for Anti-Cancer Therapies.

PubMed

Pedriali, Gaia; Rimessi, Alessandro; Sbano, Luigi; Giorgi, Carlotta; Wieckowski, Mariusz R; Previati, Maurizio; Pinton, Paolo

2017-01-01

Inter-organelle membrane contact sites are emerging as major sites for the regulation of intracellular Ca 2+ concentration and distribution. Here, extracellular stimuli operate on a wide array of channels, pumps, and ion exchangers to redistribute intracellular Ca 2+ among several compartments. The resulting highly defined spatial and temporal patterns of Ca 2+ movement can be used to elicit specific cellular responses, including cell proliferation, migration, or death. Plasma membrane (PM) also can directly contact mitochondria and endoplasmic reticulum (ER) through caveolae, small invaginations of the PM that ensure inter-organelle contacts, and can contribute to the regulation of numerous cellular functions through scaffolding proteins such as caveolins. PM and ER organize specialized junctions. Here, many components of the receptor-dependent Ca 2+ signals are clustered, including the ORAI1-stromal interaction molecule 1 complex. This complex constitutes a primary mechanism for Ca 2+ entry into non-excitable cells, modulated by intracellular Ca 2+ . Several contact sites between the ER and mitochondria, termed mitochondria-associated membranes, show a very complex and specialized structure and host a wide number of proteins that regulate Ca 2+ transfer. In this review, we summarize current knowledge of the particular action of several oncogenes and tumor suppressors at these specialized check points and analyze anti-cancer therapies that specifically target Ca 2+ flow at the inter-organelle contacts to alter the metabolism and fate of the cancer cell.

Antagonistic Basic Helix-Loop-Helix/bZIP Transcription Factors Form Transcriptional Modules That Integrate Light and Reactive Oxygen Species Signaling in Arabidopsis[W

PubMed Central

Chen, Dongqin; Xu, Gang; Tang, Weijiang; Jing, Yanjun; Ji, Qiang; Fei, Zhangjun; Lin, Rongcheng

2013-01-01

The critical developmental switch from heterotrophic to autotrophic growth of plants involves light signaling transduction and the production of reactive oxygen species (ROS). ROS function as signaling molecules that regulate multiple developmental processes, including cell death. However, the relationship between light and ROS signaling remains unclear. Here, we identify transcriptional modules composed of the basic helix-loop-helix and bZIP transcription factors PHYTOCHROME-INTERACTING FACTOR1 (PIF1), PIF3, ELONGATED HYPOCOTYL5 (HY5), and HY5 HOMOLOGY (HYH) that bridge light and ROS signaling to regulate cell death and photooxidative response. We show that pif mutants release more singlet oxygen and exhibit more extensive cell death than the wild type during Arabidopsis thaliana deetiolation. Genome-wide expression profiling indicates that PIF1 represses numerous ROS and stress-related genes. Molecular and biochemical analyses reveal that PIF1/PIF3 and HY5/HYH physically interact and coordinately regulate the expression of five ROS-responsive genes by directly binding to their promoters. Furthermore, PIF1/PIF3 and HY5/HYH function antagonistically during the seedling greening process. In addition, phytochromes, cryptochromes, and CONSTITUTIVE PHOTOMORPHOGENIC1 act upstream to regulate ROS signaling. Together, this study reveals that the PIF1/PIF3-HY5/HYH transcriptional modules mediate crosstalk between light and ROS signaling and sheds light on a new mechanism by which plants adapt to the light environments. PMID:23645630

Identification of Genes Potentially Regulated by Human Polynucleotide Phosphorylase (hPNPaseold-35) Using Melanoma as a Model

PubMed Central

Sokhi, Upneet K.; Bacolod, Manny D.; Dasgupta, Santanu; Emdad, Luni; Das, Swadesh K.; Dumur, Catherine I.; Miles, Michael F.; Sarkar, Devanand; Fisher, Paul B.

2013-01-01

Human Polynucleotide Phosphorylase (hPNPaseold-35 or PNPT1) is an evolutionarily conserved 3′→5′ exoribonuclease implicated in the regulation of numerous physiological processes including maintenance of mitochondrial homeostasis, mtRNA import and aging-associated inflammation. From an RNase perspective, little is known about the RNA or miRNA species it targets for degradation or whose expression it regulates; except for c-myc and miR-221. To further elucidate the functional implications of hPNPaseold-35 in cellular physiology, we knocked-down and overexpressed hPNPaseold-35 in human melanoma cells and performed gene expression analyses to identify differentially expressed transcripts. Ingenuity Pathway Analysis indicated that knockdown of hPNPaseold-35 resulted in significant gene expression changes associated with mitochondrial dysfunction and cholesterol biosynthesis; whereas overexpression of hPNPaseold-35 caused global changes in cell-cycle related functions. Additionally, comparative gene expression analyses between our hPNPaseold-35 knockdown and overexpression datasets allowed us to identify 77 potential “direct” and 61 potential “indirect” targets of hPNPaseold-35 which formed correlated networks enriched for cell-cycle and wound healing functional association, respectively. These results provide a comprehensive database of genes responsive to hPNPaseold-35 expression levels; along with the identification new potential candidate genes offering fresh insight into cellular pathways regulated by PNPT1 and which may be used in the future for possible therapeutic intervention in mitochondrial- or inflammation-associated disease phenotypes. PMID:24143183

Lipopolysaccharide induces autotaxin expression in human monocytic THP-1 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li Song; Zhang Junjie

2009-01-09

Autotaxin (ATX) is a secreted enzyme with lysophospholipase D (lysoPLD) activity, which converts lysophosphatidylcholine (LPC) into lysophosphatidic acid (LPA), a bioactive phospholipid involved in numerous biological activities, including cell proliferation, differentiation, and migration. In the present study, we found that bacterial lipopolysaccharide (LPS), a well-known initiator of the inflammatory response, induced ATX expression in monocytic THP-1 cells. The activation of PKR, JNK, and p38 MAPK was required for the ATX induction. The LPS-induced ATX in THP-1 cells was characterized as the {beta} isoform. In the presence of LPC, ATX could promote the migrations of THP-1 and Jurkat cells, which wasmore » inhibited by pertussis toxin (PTX), an inhibitor of Gi-mediated LPA receptor signaling. In summary, LPS induces ATX expression in THP-1 cells via a PKR, JNK and p38 MAPK-mediated mechanism, and the ATX induction is likely to enhance immune cell migration in proinflammatory response by regulating LPA levels in the microenvironment.« less

Neural stem/progenitor cell properties of glial cells in the adult mouse auditory nerve

PubMed Central

Lang, Hainan; Xing, Yazhi; Brown, LaShardai N.; Samuvel, Devadoss J.; Panganiban, Clarisse H.; Havens, Luke T.; Balasubramanian, Sundaravadivel; Wegner, Michael; Krug, Edward L.; Barth, Jeremy L.

2015-01-01

The auditory nerve is the primary conveyor of hearing information from sensory hair cells to the brain. It has been believed that loss of the auditory nerve is irreversible in the adult mammalian ear, resulting in sensorineural hearing loss. We examined the regenerative potential of the auditory nerve in a mouse model of auditory neuropathy. Following neuronal degeneration, quiescent glial cells converted to an activated state showing a decrease in nuclear chromatin condensation, altered histone deacetylase expression and up-regulation of numerous genes associated with neurogenesis or development. Neurosphere formation assays showed that adult auditory nerves contain neural stem/progenitor cells (NSPs) that were within a Sox2-positive glial population. Production of neurospheres from auditory nerve cells was stimulated by acute neuronal injury and hypoxic conditioning. These results demonstrate that a subset of glial cells in the adult auditory nerve exhibit several characteristics of NSPs and are therefore potential targets for promoting auditory nerve regeneration. PMID:26307538

Modelling cell wall growth using a fibre-reinforced hyperelastic-viscoplastic constitutive law

NASA Astrophysics Data System (ADS)

Huang, R.; Becker, A. A.; Jones, I. A.

2012-04-01

A fibre-reinforced hyperelastic-viscoplastic model using a finite strain Finite Element (FE) analysis is presented to study the expansive growth of cell walls. Based on the connections between biological concepts and plasticity theory, e.g. wall-loosening and plastic yield, wall-stiffening and plastic hardening, the modelling of cell wall growth is established within a framework of anisotropic viscoplasticity aiming to represent the corresponding biology-controlled behaviour of a cell wall. In order to model in vivo growth, special attention is paid to the differences between a living cell and an isolated wall. The proposed hyperelastic-viscoplastic theory provides a unique framework to clarify the interplay between cellulose microfibrils and cell wall matrix and how this interplay regulates sustainable growth in a particular direction while maintaining the mechanical strength of the cell walls by new material deposition. Moreover, the effect of temperature is taken into account. A numerical scheme is suggested and FE case studies are presented and compared with experimental data.

A combined numerical and experimental framework for determining permeability properties of the arterial media.

PubMed

Comerford, A; Chooi, K Y; Nowak, M; Weinberg, P D; Sherwin, S J

2015-04-01

The medial layer of the arterial wall may play an important role in the regulation of water and solute transport across the wall. In particular, a high medial resistance to transport could cause accumulation of lipid-carrying molecules in the inner wall. In this study, the water transport properties of medial tissue were characterised in a numerical model, utilising experimentally obtained data for the medial microstructure and the relative permeability of different constituents. For the model, a new solver for flow in porous materials, based on a high-order splitting scheme, was implemented in the spectral/hp element library nektar++ and validated. The data were obtained by immersing excised aortic bifurcations in a solution of fluorescent protein tracer and subsequently imaging them with a confocal microscope. Cuboidal regions of interest were selected in which the microstructure and relative permeability of different structures were transformed to a computational mesh. Impermeable objects were treated fictitiously in the numerical scheme. On this cube, a pressure drop was applied in the three coordinate directions and the principal components of the permeability tensor were determined. The reconstructed images demonstrated the arrangement of elastic lamellae and interspersed smooth muscle cells in rat aortic media; the distribution and alignment of the smooth muscle cells varied spatially within the extracellular matrix. The numerical simulations highlighted that the heterogeneity of the medial structure is important in determining local water transport properties of the tissue, resulting in regional and directional variation of the permeability tensor. A major factor in this variation is the alignment and density of smooth muscle cells in the media, particularly adjacent to the adventitial layer.

Copper Trafficking in Plants and Its Implication on Cell Wall Dynamics

PubMed Central

Printz, Bruno; Lutts, Stanley; Hausman, Jean-Francois; Sergeant, Kjell

2016-01-01

In plants, copper (Cu) acts as essential cofactor of numerous proteins. While the definitive number of these so-called cuproproteins is unknown, they perform central functions in plant cells. As micronutrient, a minimal amount of Cu is needed to ensure cellular functions. However, Cu excess may exert in contrast detrimental effects on plant primary production and even survival. Therefore it is essential for a plant to have a strictly controlled Cu homeostasis, an equilibrium that is both tissue and developmentally influenced. In the current review an overview is presented on the different stages of Cu transport from the soil into the plant and throughout the different plant tissues. Special emphasis is on the Cu-dependent responses mediated by the SPL7 transcription factor, and the crosstalk between this transcriptional regulation and microRNA-mediated suppression of translation of seemingly non-essential cuproproteins. Since Cu is an essential player in electron transport, we also review the recent insights into the molecular mechanisms controlling chloroplastic and mitochondrial Cu transport and homeostasis. We finally highlight the involvement of numerous Cu-proteins and Cu-dependent activities in the properties of one of the major Cu-accumulation sites in plants: the cell wall. PMID:27200069

Proanthocyanidins-Will they effectively restrain conspicuous bacterial strains devolving on urinary tract infection?

PubMed

Jagannathan, Venkataseshan; Viswanathan, Pragasam

2018-05-18

Struvite or infection stones are one of the major clinical burdens among urinary tract infection, which occur due to the interaction between microbes and urine mineral components. Numerous urinary tract infection (UTI) causing microbes regulate through biofilm formation for survival from host defense, it is often found difficult in its eradication with simple anti-microbial agents and also the chance of recurrence and resistance development is significantly high. Cranberry consumption and maintenance of urinary tract health have been supported by clinical, epidemiological, and mechanistic studies. It predominantly contains proanthocyanidins that belong to the class of polyphenols with repeating catechin and epicatechin monomeric units. Numerous studies have correlated proanthocyanidin consumption and prevention of bacterial adhesion to uroepithelial cells. Quorum sensing (QS) is the prime mechanism that drives bacteria to coordinate biofilm development and virulence expression. Reports have shown that proanthocyanidins are effective in disrupting cell-cell communication by quenching signal molecules. Overall, this review assesses the merits of proanthocyanidins and its effective oppression on adherence, motility, QS, and biofilm formation of major UTI strains such as Escherichia coli, Pseudomonas aeruginosa, and Proteus mirabilis by comparing and evaluating results from many significant findings. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The Role of Dynamic m6 A RNA Methylation in Photobiology.

PubMed

Robinson, Myles; Shah, Palak; Cui, Yan-Hong; He, Yu-Ying

2018-05-04

N 6 -methyladenosine (m 6 A) is the most abundant internal RNA modification among numerous post-transcriptional modifications identified in eukaryotic mRNA. m 6 A modification of RNA is catalyzed by the "writer" m 6 A methyltransferase enzyme complex, consisting of METTL3, METTL14, WTAP and KIAA1429. The m 6 A modification is reversible and can be removed by "eraser" m 6 A demethylase enzymes, namely, FTO and ALKBH5. The biological function of m 6 A modification on RNA is carried out by RNA-binding effector proteins called "readers." Varied functions of the reader proteins regulate mRNA metabolism by affecting stability, translation, splicing or nuclear export. The epitranscriptomic gene regulation by m 6 A RNA methylation regulates various pathways, which contribute to basic cellular processes essential for cell maintenance, development and cell fate, and affect response to external stimuli and stressors. In this review, we summarize the recent advances in the regulation and function of m 6 A RNA methylation, with a focus on UV-induced DNA damage response and the circadian clock machinery. Insights into the mechanisms of m 6 A RNA regulation and post-transcriptional regulatory function in these biological processes may facilitate the development of new preventive and therapeutic strategies for various diseases related to dysregulation of UV damage response and circadian rhythm. © 2018 The American Society of Photobiology.

The Integrated Role of Wnt/β-Catenin, N-Glycosylation, and E-Cadherin-Mediated Adhesion in Network Dynamics

PubMed Central

Vargas, Diego A.; Sun, Meng; Sadykov, Khikmet; Kukuruzinska, Maria A.; Zaman, Muhammad H.

2016-01-01

The cellular network composed of the evolutionarily conserved metabolic pathways of protein N-glycosylation, Wnt/β-catenin signaling pathway, and E-cadherin-mediated cell-cell adhesion plays pivotal roles in determining the balance between cell proliferation and intercellular adhesion during development and in maintaining homeostasis in differentiated tissues. These pathways share a highly conserved regulatory molecule, β-catenin, which functions as both a structural component of E-cadherin junctions and as a co-transcriptional activator of the Wnt/β-catenin signaling pathway, whose target is the N-glycosylation-regulating gene, DPAGT1. Whereas these pathways have been studied independently, little is known about the dynamics of their interaction. Here we present the first numerical model of this network in MDCK cells. Since the network comprises a large number of molecules with varying cell context and time-dependent levels of expression, it can give rise to a wide range of plausible cellular states that are difficult to track. Using known kinetic parameters for individual reactions in the component pathways, we have developed a theoretical framework and gained new insights into cellular regulation of the network. Specifically, we developed a mathematical model to quantify the fold-change in concentration of any molecule included in the mathematical representation of the network in response to a simulated activation of the Wnt/ β-catenin pathway with Wnt3a under different conditions. We quantified the importance of protein N-glycosylation and synthesis of the DPAGT1 encoded enzyme, GPT, in determining the abundance of cytoplasmic β-catenin. We confirmed the role of axin in β-catenin degradation. Finally, our data suggest that cell-cell adhesion is insensitive to E-cadherin recycling in the cell. We validate the model by inhibiting β-catenin-mediated activation of DPAGT1 expression and predicting changes in cytoplasmic β-catenin concentration and stability of E-cadherin junctions in response to DPAGT1 inhibition. We show the impact of pathway dysregulation through measurements of cell migration in scratch-wound assays. Collectively, our results highlight the importance of numerical analyses of cellular networks dynamics to gain insights into physiological processes and potential design of therapeutic strategies to prevent epithelial cell invasion in cancer. PMID:27427963

The structural basis of arrestin-mediated regulation of G-protein-coupled receptors

PubMed Central

Gurevich, Vsevolod V.; Gurevich, Eugenia V.

2008-01-01

The 4 mammalian arrestins serve as almost universal regulators of the largest known family of signaling proteins, G-protein-coupled receptors (GPCRs). Arrestins terminate receptor interactions with G proteins, redirect the signaling to a variety of alternative pathways, and orchestrate receptor internalization and subsequent intracellular trafficking. The elucidation of the structural basis and fine molecular mechanisms of the arrestin–receptor interaction paved the way to the targeted manipulation of this interaction from both sides to produce very stable or extremely transient complexes that helped to understand the regulation of many biologically important processes initiated by active GPCRs. The elucidation of the structural basis of arrestin interactions with numerous non-receptor-binding partners is long overdue. It will allow the construction of fully functional arrestins in which the ability to interact with individual partners is specifically disrupted or enhanced by targeted mutagenesis. These “custom-designed” arrestin mutants will be valuable tools in defining the role of various interactions in the intricate interplay of multiple signaling pathways in the living cell. The identification of arrestin-binding sites for various signaling molecules will also set the stage for designing molecular tools for therapeutic intervention that may prove useful in numerous disorders associated with congenital or acquired disregulation of GPCR signaling. PMID:16460808

RNA Expression Profiling Reveals Differentially Regulated Growth Factor and Receptor Expression in Redirected Cancer Cells.

PubMed

Schmucker, Hannah S; Park, Jang Pyo; Coissieux, Marie-May; Bentires-Alj, Mohamed; Feltus, F Alex; Booth, Brian W

2017-05-01

Tumorigenic cells can be redirected to adopt a normal phenotype when transplanted into cleared mammary fat pads of juvenile female mice in specific ratios with normal epithelial cells. The redirected tumorigenic cells enter stem cell niches and provide progeny that differentiate into all mammary epithelial subtypes. We have developed an in vitro model that mimics the in vivo phenomenon. The shift in phenotype to redirection should be accomplished through a return to a normal gene expression state. To measure this shift, we interrogated the transcriptome of various in vitro model states in search for casual genes. For this study, expression of growth factors, cytokines, and their associated receptors was examined. In all, we queried 251 growth factor and cytokine-related genes. We found numerous growth factor and cytokine genes whose expression levels switched from expression levels seen in cancer cells to expression levels observed in normal cells. The comparisons of gene expression between normal mammary epithelial cells, tumor-derived cells, and redirected cancer cells have revealed insight into active and inactive growth factors and cytokines in cancer cell redirection.

Tks5-dependent formation of circumferential podosomes/invadopodia mediates cell-cell fusion.

PubMed

Oikawa, Tsukasa; Oyama, Masaaki; Kozuka-Hata, Hiroko; Uehara, Shunsuke; Udagawa, Nobuyuki; Saya, Hideyuki; Matsuo, Koichi

2012-05-14

Osteoclasts fuse to form multinucleated cells during osteoclastogenesis. This process is mediated by dynamic rearrangement of the plasma membrane and cytoskeleton, and it requires numerous factors, many of which have been identified. The underlying mechanism remains obscure, however. In this paper, we show that Tks5, a master regulator of invadopodia in cancer cells, is crucial for osteoclast fusion downstream of phosphoinositide 3-kinase and Src. Expression of Tks5 was induced during osteoclastogenesis, and prevention of this induction impaired both the formation of circumferential podosomes and osteoclast fusion without affecting cell differentiation. Tyrosine phosphorylation of Tks5 was attenuated in Src-/- osteoclasts, likely accounting for defects in podosome organization and multinucleation in these cells. Circumferential invadopodia formation in B16F0 melanoma cells was also accompanied by Tks5 phosphorylation. Co-culture of B16F0 cells with osteoclasts in an inflammatory milieu promoted the formation of melanoma-osteoclast hybrid cells. Our results thus reveal an unexpected link between circumferential podosome/invadopodium formation and cell-cell fusion in and beyond osteoclasts.

Role of nuclear bodies in apoptosis signalling.

PubMed

Krieghoff-Henning, Eva; Hofmann, Thomas G

2008-11-01

Promyelocytic leukemia nuclear bodies (PML NBs) are dynamic macromolecular multiprotein complexes that recruit and release a plethora of proteins. A considerable number of PML NB components play vital roles in apoptosis, senescence regulation and tumour suppression. The molecular basis by which PML NBs control these cellular responses is still just beginning to be understood. In addition to PML itself, numerous further tumour suppressors including transcriptional regulator p53, acetyl transferase CBP (CREB binding protein) and protein kinase HIPK2 (homeodomain interacting protein kinase 2) are recruited to PML NBs in response to genotoxic stress or oncogenic transformation and drive the senescence and apoptosis response by regulating p53 activity. Moreover, in response to death-receptor activation, PML NBs may act as nuclear depots that release apoptotic factors, such as the FLASH (FLICE-associated huge) protein, to amplify the death signal. PML NBs are also associated with other nuclear domains including Cajal bodies and nucleoli and share apoptotic regulators with these domains, implying crosstalk between NBs in apoptosis regulation. In conclusion, PML NBs appear to regulate cell death decisions through different, pathway-specific molecular mechanisms.

Thiol peroxidases mediate specific genome-wide regulation of gene expression in response to hydrogen peroxide

PubMed Central

Fomenko, Dmitri E.; Koc, Ahmet; Agisheva, Natalia; Jacobsen, Michael; Kaya, Alaattin; Malinouski, Mikalai; Rutherford, Julian C.; Siu, Kam-Leung; Jin, Dong-Yan; Winge, Dennis R.; Gladyshev, Vadim N.

2011-01-01

Hydrogen peroxide is thought to regulate cellular processes by direct oxidation of numerous cellular proteins, whereas antioxidants, most notably thiol peroxidases, are thought to reduce peroxides and inhibit H2O2 response. However, thiol peroxidases have also been implicated in activation of transcription factors and signaling. It remains unclear if these enzymes stimulate or inhibit redox regulation and whether this regulation is widespread or limited to a few cellular components. Herein, we found that Saccharomyces cerevisiae cells lacking all eight thiol peroxidases were viable and withstood redox stresses. They transcriptionally responded to various redox treatments, but were unable to activate and repress gene expression in response to H2O2. Further studies involving redox transcription factors suggested that thiol peroxidases are major regulators of global gene expression in response to H2O2. The data suggest that thiol peroxidases sense and transfer oxidative signals to the signaling proteins and regulate transcription, whereas a direct interaction between H2O2 and other cellular proteins plays a secondary role. PMID:21282621

CSL protein regulates transcription of genes required to prevent catastrophic mitosis in fission yeast.

PubMed

Převorovský, Martin; Oravcová, Martina; Zach, Róbert; Jordáková, Anna; Bähler, Jürg; Půta, František; Folk, Petr

2016-11-16

For every eukaryotic cell to grow and divide, intricately coordinated action of numerous proteins is required to ensure proper cell-cycle progression. The fission yeast Schizosaccharomyces pombe has been instrumental in elucidating the fundamental principles of cell-cycle control. Mutations in S. pombe 'cut' (cell untimely torn) genes cause failed coordination between cell and nuclear division, resulting in catastrophic mitosis. Deletion of cbf11, a fission yeast CSL transcription factor gene, triggers a 'cut' phenotype, but the precise role of Cbf11 in promoting mitotic fidelity is not known. We report that Cbf11 directly activates the transcription of the acetyl-coenzyme A carboxylase gene cut6, and the biotin uptake/biosynthesis genes vht1 and bio2, with the former 2 implicated in mitotic fidelity. Cbf11 binds to a canonical, metazoan-like CSL response element (GTGGGAA) in the cut6 promoter. Expression of Cbf11 target genes shows apparent oscillations during the cell cycle using temperature-sensitive cdc25-22 and cdc10-M17 block-release experiments, but not with other synchronization methods. The penetrance of catastrophic mitosis in cbf11 and cut6 mutants is nutrient-dependent. We also show that drastic decrease in biotin availability arrests cell proliferation but does not cause mitotic defects. Taken together, our results raise the possibility that CSL proteins play conserved roles in regulating cell-cycle progression, and they could guide experiments into mitotic CSL functions in mammals.

Assembly and Functional Analysis of an S/MAR Based Episome with the Cystic Fibrosis Transmembrane Conductance Regulator Gene.

PubMed

De Rocco, Davide; Pompili, Barbara; Castellani, Stefano; Morini, Elena; Cavinato, Luca; Cimino, Giuseppe; Mariggiò, Maria A; Guarnieri, Simone; Conese, Massimo; Del Porto, Paola; Ascenzioni, Fiorentina

2018-04-17

Improving the efficacy of gene therapy vectors is still an important goal toward the development of safe and efficient gene therapy treatments. S/MAR (scaffold/matrix attached region)-based vectors are maintained extra-chromosomally in numerous cell types, which is similar to viral-based vectors. Additionally, when established as an episome, they show a very high mitotic stability. In the present study we tested the idea that addition of an S/MAR element to a CFTR (cystic fibrosis transmembrane conductance regulator) expression vector, may allow the establishment of a CFTR episome in bronchial epithelial cells. Starting from the observation that the S/MAR vector pEPI-EGFP (enhanced green fluorescence protein) is maintained as an episome in human bronchial epithelial cells, we assembled the CFTR vector pBQ-S/MAR. This vector, transfected in bronchial epithelial cells with mutated CFTR , supported long term wt CFTR expression and activity, which in turn positively impacted on the assembly of tight junctions in polarized epithelial cells. Additionally, the recovery of intact pBQ-S/MAR, but not the parental vector lacking the S/MAR element, from transfected cells after extensive proliferation, strongly suggested that pBQ-S/MAR was established as an episome. These results add a new element, the S/MAR, that can be considered to improve the persistence and safety of gene therapy vectors for cystic fibrosis pulmonary disease.

The human cumulus--oocyte complex gene-expression profile

PubMed Central

Assou, Said; Anahory, Tal; Pantesco, Véronique; Le Carrour, Tanguy; Pellestor, Franck; Klein, Bernard; Reyftmann, Lionel; Dechaud, Hervé; De Vos, John; Hamamah, Samir

2006-01-01

BACKGROUND The understanding of the mechanisms regulating human oocyte maturation is still rudimentary. We have identified transcripts differentially expressed between immature and mature oocytes, and cumulus cells. METHODS Using oligonucleotides microarrays, genome wide gene expression was studied in pooled immature and mature oocytes or cumulus cells from patients who underwent IVF. RESULTS In addition to known genes such as DAZL, BMP15 or GDF9, oocytes upregulated 1514 genes. We show that PTTG3 and AURKC are respectively the securin and the Aurora kinase preferentially expressed during oocyte meiosis. Strikingly, oocytes overexpressed previously unreported growth factors such as TNFSF13/APRIL, FGF9, FGF14, and IL4, and transcription factors including OTX2, SOX15 and SOX30. Conversely, cumulus cells, in addition to known genes such as LHCGR or BMPR2, overexpressed cell-tocell signaling genes including TNFSF11/RANKL, numerous complement components, semaphorins (SEMA3A, SEMA6A, SEMA6D) and CD genes such as CD200. We also identified 52 genes progressively increasing during oocyte maturation, comprising CDC25A and SOCS7. CONCLUSION The identification of genes up and down regulated during oocyte maturation greatly improves our understanding of oocyte biology and will provide new markers that signal viable and competent oocytes. Furthermore, genes found expressed in cumulus cells are potential markers of granulosa cell tumors. PMID:16571642

Mapping Human Pluripotent-to-Cardiomyocyte Differentiation: Methylomes, Transcriptomes, and Exon DNA Methylation “Memories”

PubMed Central

Tompkins, Joshua D.; Jung, Marc; Chen, Chang-yi; Lin, Ziguang; Ye, Jingjing; Godatha, Swetha; Lizhar, Elizabeth; Wu, Xiwei; Hsu, David; Couture, Larry A.; Riggs, Arthur D.

2016-01-01

The directed differentiation of human cardiomyocytes (CMs) from pluripotent cells provides an invaluable model for understanding mechanisms of cell fate determination and offers considerable promise in cardiac regenerative medicine. Here, we utilize a human embryonic stem cell suspension bank, produced according to a good manufacturing practice, to generate CMs using a fully defined and small molecule-based differentiation strategy. Primitive and cardiac mesoderm purification was used to remove non-committing and multi-lineage populations and this significantly aided the identification of key transcription factors, lncRNAs, and essential signaling pathways that define cardiomyogenesis. Global methylation profiles reflect CM development and we report on CM exon DNA methylation “memories” persisting beyond transcription repression and marking the expression history of numerous developmentally regulated genes, especially transcription factors. PMID:26981572

Transcriptional regulation of cranial sensory placode development

PubMed Central

Moody, Sally A.; LaMantia, Anthony-Samuel

2015-01-01

Cranial sensory placodes derive from discrete patches of the head ectoderm, and give rise to numerous sensory structures. During gastrulation, a specialized “neural border zone” forms around the neural plate in response to interactions between the neural and non-neural ectoderm and signals from adjacent mesodermal and/or endodermal tissues. This zone subsequently gives rise to two distinct precursor populations of the peripheral nervous system: the neural crest and the pre-placodal ectoderm (PPE). The PPE is a common field from which all cranial sensory placodes arise (adenohypophyseal, olfactory, lens, trigeminal, epibranchial, otic). Members of the Six family of transcription factors are major regulators of PPE specification, in partnership with co-factor proteins such as Eya. Six gene activity also maintains tissue boundaries between the PPE, neural crest and epidermis by repressing genes that specify the fates of those adjacent ectodermally-derived domains. As the embryo acquires anterior-posterior identity, the PPE becomes transcriptionally regionalized, and it subsequently subdivides into specific placodes with distinct developmental fates in response to signaling from adjacent tissues. Each placode is characterized by a unique transcriptional program that leads to the differentiation of highly specialized cells, such as neurosecretory cells, somatic sensory receptor cells, chemosensory neurons, peripheral glia and supporting cells. In this review, we summarize the transcriptional and signaling factors that regulate key steps of placode development, influence subsequent sensory neuron specification, and discuss what is known about mutations in some of the essential PPE genes that underlie human congenital syndromes. PMID:25662264

Targeted overexpression of EZH2 in the mammary gland disrupts ductal morphogenesis and causes epithelial hyperplasia.

PubMed

Li, Xin; Gonzalez, Maria E; Toy, Katherine; Filzen, Tracey; Merajver, Sofia D; Kleer, Celina G

2009-09-01

The Polycomb group protein enhancer of zeste homolog 2 (EZH2), which has roles during development of numerous tissues, is a critical regulator of cell type identity. Overexpression of EZH2 has been detected in invasive breast carcinoma tissue samples and is observed in human breast tissue samples of morphologically normal lobules up to 12 years before the development of breast cancer. The function of EZH2 during preneoplastic progression in the mammary gland is unknown. To investigate the role of EZH2 in the mammary gland, we targeted the expression of EZH2 to mammary epithelial cells using the mouse mammary tumor virus long terminal repeat. EZH2 overexpression resulted in aberrant terminal end bud architecture. By the age of 4 months, 100% of female mouse mammary tumor virus-EZH2 virgin mice developed intraductal epithelial hyperplasia resembling the human counterpart accompanied by premature differentiation of ductal epithelial cells and up-regulation of the luminal marker GATA-3. In addition, remodeling of the mammary gland after parturition was impaired and EZH2 overexpression caused delayed involution. Mechanistically, we found that EZH2 physically interacts with beta-catenin, inducing beta-catenin nuclear accumulation in mammary epithelial cells and activating Wnt/beta-catenin signaling. The biological significance of these data to human hyperplasias is demonstrated by EZH2 up-regulation and colocalization with beta-catenin in human intraductal epithelial hyperplasia, the earliest histologically identifiable precursor of breast carcinoma.

Targeted Overexpression of EZH2 in the Mammary Gland Disrupts Ductal Morphogenesis and Causes Epithelial Hyperplasia

PubMed Central

Li, Xin; Gonzalez, Maria E.; Toy, Katherine; Filzen, Tracey; Merajver, Sofia D.; Kleer, Celina G.

2009-01-01

The Polycomb group protein enhancer of zeste homolog 2 (EZH2), which has roles during development of numerous tissues, is a critical regulator of cell type identity. Overexpression of EZH2 has been detected in invasive breast carcinoma tissue samples and is observed in human breast tissue samples of morphologically normal lobules up to 12 years before the development of breast cancer. The function of EZH2 during preneoplastic progression in the mammary gland is unknown. To investigate the role of EZH2 in the mammary gland, we targeted the expression of EZH2 to mammary epithelial cells using the mouse mammary tumor virus long terminal repeat. EZH2 overexpression resulted in aberrant terminal end bud architecture. By the age of 4 months, 100% of female mouse mammary tumor virus-EZH2 virgin mice developed intraductal epithelial hyperplasia resembling the human counterpart accompanied by premature differentiation of ductal epithelial cells and up-regulation of the luminal marker GATA-3. In addition, remodeling of the mammary gland after parturition was impaired and EZH2 overexpression caused delayed involution. Mechanistically, we found that EZH2 physically interacts with β-catenin, inducing β-catenin nuclear accumulation in mammary epithelial cells and activating Wnt/β-catenin signaling. The biological significance of these data to human hyperplasias is demonstrated by EZH2 up-regulation and colocalization with β-catenin in human intraductal epithelial hyperplasia, the earliest histologically identifiable precursor of breast carcinoma. PMID:19661437

Ankrd6 is a mammalian functional homolog of Drosophila planar cell polarity gene diego and regulates coordinated cellular orientation in the mouse inner ear.

PubMed

Jones, Chonnettia; Qian, Dong; Kim, Sun Myoung; Li, Shuangding; Ren, Dongdong; Knapp, Lindsey; Sprinzak, David; Avraham, Karen B; Matsuzaki, Fumio; Chi, Fanglu; Chen, Ping

2014-11-01

The coordinated polarization of neighboring cells within the plane of the tissue, known as planar cell polarity (PCP), is a recurring theme in biology. It is required for numerous developmental processes for the form and function of many tissues and organs across species. The genetic pathway regulating PCP was first discovered in Drosophila, and an analogous but distinct pathway is emerging in vertebrates. It consists of membrane protein complexes known as core PCP proteins that are conserved across species. Here we report that the over-expression of the murine Ankrd6 (mAnkrd6) gene that shares homology with Drosophila core PCP gene diego causes a typical PCP phenotype in Drosophila, and mAnkrd6 can rescue the loss of function of diego in Drosophila. In mice, mAnkrd6 protein is asymmetrically localized in cells of the inner ear sensory organs, characteristic of components of conserved core PCP complexes. The loss of mAnkrd6 causes PCP defects in the inner ear sensory organs. Moreover, canonical Wnt signaling is significantly increased in mouse embryonic fibroblasts from mAnkrd6 knockout mice in comparison to wild type controls. Together, these results indicated that mAnkrd6 is a functional homolog of the Drosophila diego gene for mammalian PCP regulation and act to suppress canonical Wnt signaling. Copyright © 2014 Elsevier Inc. All rights reserved.

Microfluidic co-culture platform for investigating osteocyte-osteoclast signalling during fluid shear stress mechanostimulation.

PubMed

Middleton, K; Al-Dujaili, S; Mei, X; Günther, A; You, L

2017-07-05

Bone cells exist in a complex environment where they are constantly exposed to numerous dynamic biochemical and mechanical stimuli. These stimuli regulate bone cells that are involved in various bone disorders, such as osteoporosis. Knowledge of how these stimuli affect bone cells have been utilised to develop various treatments, such as pharmaceuticals, hormone therapy, and exercise. To investigate the role that bone loading has on these disorders in vitro, bone cell mechanotransduction studies are typically performed using parallel plate flow chambers (PPFC). However, these chambers do not allow for dynamic cellular interactions among different cell populations to be investigated. We present a microfluidic approach that exposes different cell populations, which are located at physiologically relevant distances within adjacent channels, to different levels of fluid shear stress, and promotes cell-cell communication between the different channels. We employed this microfluidic system to assess mechanically regulated osteocyte-osteoclast communication. Osteoclast precursors (RAW264.7 cells) responded to cytokine gradients (e.g., RANKL, OPG, PGE-2) developed by both mechanically stimulated (fOCY) and unstimulated (nOCY) osteocyte-like MLO-Y4 cells simultaneously. Specifically, we observed increased osteoclast precursor cell densities and osteoclast differentiation towards nOCY. We also used this system to show an increased mechanoresponse of osteocytes when in co-culture with osteoclasts. We envision broad applicability of the presented approach for microfluidic perfusion co-culture of multiple cell types in the presence of fluid flow stimulation, and as a tool to investigate osteocyte mechanotransduction, as well as bone metastasis extravasation. This system could also be applied to any multi-cell population cross-talk studies that are typically performed using PPFCs (e.g. endothelial cells, smooth muscle cells, and fibroblasts). Copyright © 2017 Elsevier Ltd. All rights reserved.

Suppression of microRNA-629 enhances sensitivity of cervical cancer cells to 1'S-1'-acetoxychavicol acetate via regulating RSU1.

PubMed

Phuah, Neoh Hun; Azmi, Mohamad Nurul; Awang, Khalijah; Nagoor, Noor Hasima

2017-01-01

Cervical cancer is the fourth most frequent malignancy affecting women worldwide, but drug resistance and toxicities remain a major challenge in chemotherapy. The use of natural compounds is promising because they are less toxic and able to target multiple signaling pathways. The 1'S-1'-acetoxychavicol acetate (ACA), a natural compound isolated from wild ginger Alpinia conchigera , induced cytotoxicity on various cancer cells including cervical cancer. MicroRNAs (miRNAs) are short noncoding RNAs that regulate numerous biological processes, such as apoptosis and chemosensitivity. Past studies reported that miR-629 is upregulated in many cancers, and its expression was altered in ACA-treated cervical cancer cells. However, the role of miR-629 in regulating sensitivity toward ACA or other anticancer agents has not been reported. Hence, this study aims to investigate the role of miR-629 in regulating response toward ACA on cervical cancer cells. The miR-629 expression following transfection with miR-629 hairpin inhibitor and hairpin inhibitor negative control was measured using quantitative real-time polymerase chain reaction (RT-qPCR). The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to investigate sensitivity toward ACA. Apoptosis was detected using Annexin V/propidium iodide and Caspase 3/7 assays. The gene target for miR-629 was identified using miRNA target prediction programs, luciferase reporter assay and Western blots. Gene overexpression studies were performed to evaluate its role in regulating response toward ACA. Transfection with miR-629 hairpin inhibitor downregulated its expression in both cervical cancer cell lines. Suppression of miR-629 increased sensitivity toward ACA by reducing cell proliferation and inducing apoptosis. Luciferase reporter assay confirmed RSU1 as a direct target of miR-629. Overexpression of miR-629 decreased RSU1 protein expression, while inhibition of miR-629 increased RSU1 protein expression. Overexpression of RSU1 augmented antiproliferative and apoptosis-inducing effects of ACA. Our findings showed that combination of ACA with miR-629 and RSU1 may provide a potential strategy in treating cervical cancer.

Functional roles of fibroblast growth factor receptors (FGFRs) signaling in human cancers.

PubMed

Tiong, Kai Hung; Mah, Li Yen; Leong, Chee-Onn

2013-12-01

The fibroblast growth factor receptors (FGFRs) regulate important biological processes including cell proliferation and differentiation during development and tissue repair. Over the past decades, numerous pathological conditions and developmental syndromes have emerged as a consequence of deregulation in the FGFRs signaling network. This review aims to provide an overview of FGFR family, their complex signaling pathways in tumorigenesis, and the current development and application of therapeutics targeting the FGFRs signaling for treatment of refractory human cancers.

Differential chemokine, chemokine receptor and cytokine expression in Epstein-Barr virus-associated lymphoproliferative diseases.

PubMed

Ohshima, Koichi; Karube, Kennosuke; Hamasaki, Makoto; Tutiya, Takeshi; Yamaguchi, Takahiro; Suefuji, Hiroaki; Suzuki, Keiko; Suzumiya, Junji; Ohga, Shouichi; Kikuchi, Masahiro

2003-08-01

T cell immunity plays an important role in the clinicopathology of Epstein-Barr virus (EBV)-associated diseases. Acute EBV-induced infectious mononucleosis (IM) is a common self-limiting disease, however, other EBV-associated diseases, including chronic active EBV infection (CAEBV), NK cell lymphoma (NKL), and Hodgkin's lymphoma (HL), exhibit distinct clinical features. Chemokines are members of a family of small-secreted proteins. The relationships between chemokines and the chemokine receptor (R) are thought to be important for selectivity of local immunity. Some chemokines, chemokine R and cytokines closely associate with the T cell subtypes, Th1 and Th2 T cells and cytotoxic cells. To clarify the role of T cell immunity in EBV-associated diseases, we conducted gene expression profiling, using chemokine, chemokine R and cytokine DNA chips. Compared to EBV negative non-specific lymphadenitis, CAEBV and NKL exhibited diffuse down- and up-regulation, respectively, of these gene profiles. IM had a predominantly Th1-type profile, whereas HL had a mixed Th1/Th2-type profile. Reduction of the Th1-type cytokine interferon gamma (IFN-gamma) in CAEBV was confirmed by Reverse transcriptase-polymerase chain reaction, whereas IFN-gamma expression was markedly enhanced in NKL, and moderately enhanced in IM. Compared to IM, CAEBV showed slight elevation of "regulated upon activation, normal T expressed and secreted" (RANTES), but almost all other genes assayed were down-regulated. NKL exhibited elevated expression of numerous genes, particularly IFN-gamma-inducible-10 (IP-10) and monokine induced by IFN-gamma (MIG). HL showed variable elevated and reduced expression of various genes, with increased expression of IL-13 receptor and MIG. Our study demonstrated the enormous potential of gene expression profiling for clarifying the pathogenesis of EBV-associated diseases.

Host responses in tissue repair and fibrosis.

PubMed

Duffield, Jeremy S; Lupher, Mark; Thannickal, Victor J; Wynn, Thomas A

2013-01-24

Myofibroblasts accumulate in the spaces between organ structures and produce extracellular matrix (ECM) proteins, including collagen I. They are the primary "effector" cells in tissue remodeling and fibrosis. Previously, leukocyte progenitors termed fibrocytes and myofibroblasts generated from epithelial cells through epithelial-to-mesenchymal transition (EMT) were considered the primary sources of ECM-producing myofibroblasts in injured tissues. However, genetic fate mapping experiments suggest that mesenchyme-derived cells, known as resident fibroblasts, and pericytes are the primary precursors of scar-forming myofibroblasts, whereas epithelial cells, endothelial cells, and myeloid leukocytes contribute to fibrogenesis predominantly by producing key fibrogenic cytokines and by promoting cell-to-cell communication. Numerous cytokines derived from T cells, macrophages, and other myeloid cell populations are important drivers of myofibroblast differentiation. Monocyte-derived cell populations are key regulators of the fibrotic process: They act as a brake on the processes driving fibrogenesis, and they dismantle and degrade established fibrosis. We discuss the origins, modes of activation, and fate of myofibroblasts in various important fibrotic diseases and describe how manipulation of macrophage activation could help ameliorate fibrosis.

Peripheral Serotonin Regulates Maternal Calcium Trafficking in Mammary Epithelial Cells during Lactation in Mice

PubMed Central

Laporta, Jimena; Keil, Kimberly P.; Vezina, Chad M.; Hernandez, Laura L.

2014-01-01

Lactation is characterized by massive transcellular flux of calcium, from the basolateral side of the mammary alveolar epithelium (blood) into the ductal lumen (milk). Regulation of calcium transport during lactation is critical for maternal and neonatal health. The monoamine serotonin (5-HT) is synthesized by the mammary gland and functions as a homeostatic regulation of lactation. Genetic ablation of tryptophan hydroxylase 1 (Tph1), which encodes the rate-limiting enzyme in non-neuronal serotonin synthesis, causes a deficiency in circulating serotonin. As a consequence maternal calcium concentrations decrease, mammary epithelial cell morphology is altered, and cell proliferation is decreased during lactation. Here we demonstrate that serotonin deficiency decreases the expression and disrupts the normal localization of calcium transporters located in the apical (PMCA2) and basolateral (CaSR, ORAI-1) membranes of the lactating mammary gland. In addition, serotonin deficiency decreases the mRNA expression of calcium transporters located in intracellular compartments (SERCA2, SPCA1 and 2). Mammary expression of serotonin receptor isoform 2b and its downstream pathways (PLCβ3, PKC and MAP-ERK1/2) are also decreased by serotonin deficiency, which might explain the numerous phenotypic alterations described above. In most cases, addition of exogenous 5-hydroxy-L-tryptophan to the Tph1 deficient mice rescued the phenotype. Our data supports the hypothesis that serotonin is necessary for proper mammary gland structure and function, to regulate blood and mammary epithelial cell transport of calcium during lactation. These findings can be applicable to the treatment of lactation-induced hypocalcemia in dairy cows and can have profound implications in humans, given the wide-spread use of selective serotonin reuptake inhibitors as antidepressants during pregnancy and lactation. PMID:25299122

The Synthetic α-Bromo-2′,3,4,4′-Tetramethoxychalcone (α-Br-TMC) Inhibits the JAK/STAT Signaling Pathway

PubMed Central

Brueggemann, Susanne; Besl, Elisabeth; Al-Rifai, Nafisah; Petkes, Hermina; Amslinger, Sabine; Rascle, Anne

2014-01-01

Signal transducer and activator of transcription STAT5 and its upstream activating kinase JAK2 are essential mediators of cytokine signaling. Their activity is normally tightly regulated and transient. However, constitutive activation of STAT5 is found in numerous cancers and a driving force for malignant transformation. We describe here the identification of the synthetic chalcone α-Br-2′,3,4,4′-tetramethoxychalcone (α-Br-TMC) as a novel JAK/STAT inhibitor. Using the non-transformed IL-3-dependent B cell line Ba/F3 and its oncogenic derivative Ba/F3-1*6 expressing constitutively activated STAT5, we show that α-Br-TMC targets the JAK/STAT pathway at multiple levels, inhibiting both JAK2 and STAT5 phosphorylation. Moreover, α-Br-TMC alters the mobility of STAT5A/B proteins in SDS-PAGE, indicating a change in their post-translational modification state. These alterations correlate with a decreased association of STAT5 and RNA polymerase II with STAT5 target genes in chromatin immunoprecipitation assays. Interestingly, expression of STAT5 target genes such as Cis and c-Myc was differentially regulated by α-Br-TMC in normal and cancer cells. While both genes were inhibited in IL-3-stimulated Ba/F3 cells, expression of the oncogene c-Myc was down-regulated and that of the tumor suppressor gene Cis was up-regulated in transformed Ba/F3-1*6 cells. The synthetic chalcone α-Br-TMC might therefore represent a promising novel anticancer agent for therapeutic intervention in STAT5-associated malignancies. PMID:24595334

Guava Leaf Extract Inhibits Quorum-Sensing and Chromobacterium violaceum Induced Lysis of Human Hepatoma Cells: Whole Transcriptome Analysis Reveals Differential Gene Expression

PubMed Central

Tiwary, Bipransh Kumar; Kumar, Anoop

2014-01-01

Quorum sensing (QS) is a process mediated via small molecules termed autoinducers (AI) that allow bacteria to respond and adjust according to the cell population density by altering the expression of multitudinous genes. Since QS governs numerous bioprocesses in bacteria, including virulence, its inhibition promises to be an ideal target for the development of novel therapeutics. We found that the aqueous leaf extract of Psidium guajava (GLE) exhibited anti-QS properties as evidenced by inhibition of violacein production in Chromobacterium violaceum and swarming motility of Pseudomonas aeruginosa. The gram-negative bacterium, C. violaceum is a rare pathogen with high mortality rate. In this study, perhaps for the first time, we identified the target genes of GLE in C. violaceum MTCC 2656 by whole transcriptome analysis on Ion Torrent. Our data revealed that GLE significantly down-regulated 816 genes at least three fold, with p value≤0.01, which comprises 19% of the C. violaceum MTCC 2656 genome. These genes were distributed throughout the genome and were associated with virulence, motility and other cellular processes, many of which have been described as quorum regulated in C. violaceum and other gram negative bacteria. Interestingly, GLE did not affect the growth of the bacteria. However, consistent with the gene expression pattern, GLE treated C. violaceum cells were restrained from causing lysis of human hepatoma cell line, HepG2, indicating a positive relationship between the QS-regulated genes and pathogenicity. Overall, our study proposes GLE as a QS inhibitor (QSI) with the ability to attenuate virulence without affecting growth. To the best of our knowledge, this is the first report which provides with a plausible set of candidate genes regulated by the QS system in the neglected pathogen C. violaceum. PMID:25229331

Guava leaf extract inhibits quorum-sensing and Chromobacterium violaceum induced lysis of human hepatoma cells: whole transcriptome analysis reveals differential gene expression.

PubMed

Ghosh, Runu; Tiwary, Bipransh Kumar; Kumar, Anoop; Chakraborty, Ranadhir

2014-01-01

Quorum sensing (QS) is a process mediated via small molecules termed autoinducers (AI) that allow bacteria to respond and adjust according to the cell population density by altering the expression of multitudinous genes. Since QS governs numerous bioprocesses in bacteria, including virulence, its inhibition promises to be an ideal target for the development of novel therapeutics. We found that the aqueous leaf extract of Psidium guajava (GLE) exhibited anti-QS properties as evidenced by inhibition of violacein production in Chromobacterium violaceum and swarming motility of Pseudomonas aeruginosa. The gram-negative bacterium, C. violaceum is a rare pathogen with high mortality rate. In this study, perhaps for the first time, we identified the target genes of GLE in C. violaceum MTCC 2656 by whole transcriptome analysis on Ion Torrent. Our data revealed that GLE significantly down-regulated 816 genes at least three fold, with p value ≤ 0.01, which comprises 19% of the C. violaceum MTCC 2656 genome. These genes were distributed throughout the genome and were associated with virulence, motility and other cellular processes, many of which have been described as quorum regulated in C. violaceum and other gram negative bacteria. Interestingly, GLE did not affect the growth of the bacteria. However, consistent with the gene expression pattern, GLE treated C. violaceum cells were restrained from causing lysis of human hepatoma cell line, HepG2, indicating a positive relationship between the QS-regulated genes and pathogenicity. Overall, our study proposes GLE as a QS inhibitor (QSI) with the ability to attenuate virulence without affecting growth. To the best of our knowledge, this is the first report which provides with a plausible set of candidate genes regulated by the QS system in the neglected pathogen C. violaceum.

Tcf19 is a novel islet factor necessary for proliferation and survival in the INS-1 β-cell line

PubMed Central

Krautkramer, Kimberly A.; Linnemann, Amelia K.; Fontaine, Danielle A.; Whillock, Amy L.; Harris, Ted W.; Schleis, Gregory J.; Truchan, Nathan A.; Marty-Santos, Leilani; Lavine, Jeremy A.; Cleaver, Ondine; Kimple, Michelle E.

2013-01-01

Recently, a novel type 1 diabetes association locus was identified at human chromosome 6p31.3, and transcription factor 19 (TCF19) is a likely causal gene. Little is known about Tcf19, and we now show that it plays a role in both proliferation and apoptosis in insulinoma cells. Tcf19 is expressed in mouse and human islets, with increasing mRNA expression in nondiabetic obesity. The expression of Tcf19 is correlated with β-cell mass expansion, suggesting that it may be a transcriptional regulator of β-cell mass. Increasing proliferation and decreasing apoptotic cell death are two strategies to increase pancreatic β-cell mass and prevent or delay diabetes. siRNA-mediated knockdown of Tcf19 in the INS-1 insulinoma cell line, a β-cell model, results in a decrease in proliferation and an increase in apoptosis. There was a significant reduction in the expression of numerous cell cycle genes from the late G1 phase through the M phase, and cells were arrested at the G1/S checkpoint. We also observed increased apoptosis and susceptibility to endoplasmic reticulum (ER) stress after Tcf19 knockdown. There was a reduction in expression of genes important for the maintenance of ER homeostasis (Bip, p58IPK, Edem1, and calreticulin) and an increase in proapoptotic genes (Bim, Bid, Nix, Gadd34, and Pdia2). Therefore, Tcf19 is necessary for both proliferation and survival and is a novel regulator of these pathways. PMID:23860123

Noncanonical Hedgehog Signaling

PubMed Central

Brennan, Donna; Chen, Xiaole; Cheng, Lan; Mahoney, My; Riobo, Natalia A.

2012-01-01

The notion of noncanonical hedgehog (Hh) signaling in mammals has started to receive support from numerous observations. By noncanonical, we refer to all those cellular and tissue responses to any of the Hh isoforms that are independent of transcriptional changes mediated by the Gli family of transcription factors. In this chapter, we discuss the most recent findings that suggest that Patched1 can regulate cell proliferation and apoptosis independently of Smoothened (Smo) and Gli and the reports that Smo modulates actin cytoskeleton-dependent processes such as fibroblast migration, endothelial cell tubulogenesis, axonal extension, and neurite formation by diverse mechanisms that exclude any involvement of Gli-dependent transcription. We also acknowledge the existence of less stronger evidence of noncanonical signaling in Drosophila. PMID:22391299

S-nitrosylation in the regulation of gene transcription☆

PubMed Central

Sha, Yonggang; Marshall, Harvey E.

2015-01-01

Background Post-translational modification of proteins by S-nitrosylation serves as a major mode of signaling in mammalian cells and a growing body of evidence has shown that transcription factors and their activating pathways are primary targets. S-nitrosylation directly modifies a number of transcription factors, including NF-κB, HIF-1, and AP-1. In addition, S-nitrosylation can indirectly regulate gene transcription by modulating other cell signaling pathways, in particular JNK kinase and ras. Scope of review The evolution of S-nitrosylation as a signaling mechanism in the regulation of gene transcription, physiological advantages of protein S-nitrosylation in the control of gene transcription, and discussion of the many transcriptional proteins modulated by S-nitrosylation is summarized. Major conclusions S-nitrosylation plays a crucial role in the control of mammalian gene transcription with numerous transcription factors regulated by this modification. Many of these proteins serve as immunomodulators, and inducible nitric oxide synthase (iNOS) is regarded as a principal mediatiator of NO-dependent S-nitrosylation. However, additional targets within the nucleus (e.g. histone deacetylases) and alternative mechanisms of S-nitrosylation (e.g. GAPDH-mediated trans-nitrosylation) are thought to play a role in NOS-dependent transcriptional regulation. General significance Derangement of SNO-regulated gene transcription is an important factor in a variety of pathological conditions including neoplasia and sepsis. A better understanding of protein S-nitrosylation as it relates to gene transcription and the physiological mechanisms behind this process is likely to lead to novel therapies for these disorders. This article is part of a Special Issue entitled Regulation of Cellular Processes by S-nitrosylation. PMID:21640163

Axon guidance molecules in vascular patterning.

PubMed

Adams, Ralf H; Eichmann, Anne

2010-05-01

Endothelial cells (ECs) form extensive, highly branched and hierarchically organized tubular networks in vertebrates to ensure the proper distribution of molecular and cellular cargo in the vertebrate body. The growth of this vascular system during development, tissue repair or in disease conditions involves the sprouting, migration and proliferation of endothelial cells in a process termed angiogenesis. Surprisingly, specialized ECs, so-called tip cells, which lead and guide endothelial sprouts, share many feature with another guidance structure, the axonal growth cone. Tip cells are motile, invasive and extend numerous filopodial protrusions sensing growth factors, extracellular matrix and other attractive or repulsive cues in their tissue environment. Axonal growth cones and endothelial tip cells also respond to signals belonging to the same molecular families, such as Slits and Roundabouts, Netrins and UNC5 receptors, Semaphorins, Plexins and Neuropilins, and Eph receptors and ephrin ligands. Here we summarize fundamental principles of angiogenic growth, the selection and function of tip cells and the underlying regulation by guidance cues, the Notch pathway and vascular endothelial growth factor signaling.

F-box protein interactions with the hallmark pathways in cancer.

PubMed

Randle, Suzanne J; Laman, Heike

2016-02-01

F-box proteins (FBP) are the substrate specifying subunit of Skp1-Cul1-FBP (SCF)-type E3 ubiquitin ligases and are responsible for directing the ubiquitination of numerous proteins essential for cellular function. Due to their ability to regulate the expression and activity of oncogenes and tumour suppressor genes, FBPs themselves play important roles in cancer development and progression. In this review, we provide a comprehensive overview of FBPs and their targets in relation to their interaction with the hallmarks of cancer cell biology, including the regulation of proliferation, epigenetics, migration and invasion, metabolism, angiogenesis, cell death and DNA damage responses. Each cancer hallmark is revealed to have multiple FBPs which converge on common signalling hubs or response pathways. We also highlight the complex regulatory interplay between SCF-type ligases and other ubiquitin ligases. We suggest six highly interconnected FBPs affecting multiple cancer hallmarks, which may prove sensible candidates for therapeutic intervention. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

Identification of the miRNA targetome in hippocampal neurons using RIP-seq.

PubMed

Malmevik, Josephine; Petri, Rebecca; Klussendorf, Thies; Knauff, Pina; Åkerblom, Malin; Johansson, Jenny; Soneji, Shamit; Jakobsson, Johan

2015-07-28

MicroRNAs (miRNAs) are key players in the regulation of neuronal processes by targeting a large network of target messenger RNAs (mRNAs). However, the identity and function of mRNAs targeted by miRNAs in specific cells of the brain are largely unknown. Here, we established an adeno-associated viral vector (AAV)-based neuron-specific Argonaute2:GFP-RNA immunoprecipitation followed by high-throughput sequencing to analyse the regulatory role of miRNAs in mouse hippocampal neurons. Using this approach, we identified more than two thousand miRNA targets in hippocampal neurons, regulating essential neuronal features such as cell signalling, transcription and axon guidance. Furthermore, we found that stable inhibition of the highly expressed miR-124 and miR-125 in hippocampal neurons led to significant but distinct changes in the AGO2 binding of target mRNAs, resulting in subsequent upregulation of numerous miRNA target genes. These findings greatly enhance our understanding of the miRNA targetome in hippocampal neurons.

Hey Factors at the Crossroad of Tumorigenesis and Clinical Therapeutic Modulation of Hey for Anticancer Treatment.

PubMed

Liu, Zihao; Sanders, Andrew J; Liang, Gehao; Song, Erwei; Jiang, Wen G; Gong, Chang

2017-05-01

Hairy and Enhancer-of-split related with YRPW motif (Hey) transcription factors are important regulators of stem cell embryogenesis. Clinical relevance shows that they are also highly expressed in malignant carcinoma. Recent studies have highlighted functions for the Hey factors in tumor metastasis, the maintenance of cancer cell self-renewal, as well as proliferation and the promotion of tumor angiogenesis. Pathways that regulate Hey gene expression, such as Notch and TGFβ signaling, are frequently aberrant in numerous cancers. In addition, Hey factors control downstream targets via recruitment of histone deacetylases (HDAC). Targeting these signaling pathways or HDACs may reverse tumor progression and provide clinical benefit for cancer patients. Thus, some small molecular inhibitors or monoclonal antibodies of each of these signaling pathways have been studied in clinical trials. This review focuses on the involvement of Hey proteins in malignant carcinoma progression and provides valuable therapeutic information for anticancer treatment. Mol Cancer Ther; 16(5); 775-86. ©2017 AACR . ©2017 American Association for Cancer Research.

CREB at the Crossroads of Activity-Dependent Regulation of Nervous System Development and Function.

PubMed

Belgacem, Yesser H; Borodinsky, Laura N

2017-01-01

The central nervous system is a highly plastic network of cells that constantly adjusts its functions to environmental stimuli throughout life. Transcription-dependent mechanisms modify neuronal properties to respond to external stimuli regulating numerous developmental functions, such as cell survival and differentiation, and physiological functions such as learning, memory, and circadian rhythmicity. The discovery and cloning of the cyclic adenosine monophosphate (cAMP) responsive element binding protein (CREB) constituted a big step toward deciphering the molecular mechanisms underlying neuronal plasticity. CREB was first discovered in learning and memory studies as a crucial mediator of activity-dependent changes in target gene expression that in turn impose long-lasting modifications of the structure and function of neurons. In this chapter, we review the molecular and signaling mechanisms of neural activity-dependent recruitment of CREB and its cofactors. We discuss the crosstalk between signaling pathways that imprints diverse spatiotemporal patterns of CREB activation allowing for the integration of a wide variety of stimuli.

Multifunctional Role of ATM/Tel1 Kinase in Genome Stability: From the DNA Damage Response to Telomere Maintenance

PubMed Central

2014-01-01

The mammalian protein kinase ataxia telangiectasia mutated (ATM) is a key regulator of the DNA double-strand-break response and belongs to the evolutionary conserved phosphatidylinositol-3-kinase-related protein kinases. ATM deficiency causes ataxia telangiectasia (AT), a genetic disorder that is characterized by premature aging, cerebellar neuropathy, immunodeficiency, and predisposition to cancer. AT cells show defects in the DNA damage-response pathway, cell-cycle control, and telomere maintenance and length regulation. Likewise, in Saccharomyces cerevisiae, haploid strains defective in the TEL1 gene, the ATM ortholog, show chromosomal aberrations and short telomeres. In this review, we outline the complex role of ATM/Tel1 in maintaining genomic stability through its control of numerous aspects of cellular survival. In particular, we describe how ATM/Tel1 participates in the signal transduction pathways elicited by DNA damage and in telomere homeostasis and its importance as a barrier to cancer development. PMID:25247188

Gene Architectures that Minimize Cost of Gene Expression.

PubMed

Frumkin, Idan; Schirman, Dvir; Rotman, Aviv; Li, Fangfei; Zahavi, Liron; Mordret, Ernest; Asraf, Omer; Wu, Song; Levy, Sasha F; Pilpel, Yitzhak

2017-01-05

Gene expression burdens cells by consuming resources and energy. While numerous studies have investigated regulation of expression level, little is known about gene design elements that govern expression costs. Here, we ask how cells minimize production costs while maintaining a given protein expression level and whether there are gene architectures that optimize this process. We measured fitness of ∼14,000 E. coli strains, each expressing a reporter gene with a unique 5' architecture. By comparing cost-effective and ineffective architectures, we found that cost per protein molecule could be minimized by lowering transcription levels, regulating translation speeds, and utilizing amino acids that are cheap to synthesize and that are less hydrophobic. We then examined natural E. coli genes and found that highly expressed genes have evolved more forcefully to minimize costs associated with their expression. Our study thus elucidates gene design elements that improve the economy of protein expression in natural and heterologous systems. Copyright © 2017 Elsevier Inc. All rights reserved.

SREBP-regulated lipid metabolism: convergent physiology - divergent pathophysiology.

PubMed

Shimano, Hitoshi; Sato, Ryuichiro

2017-12-01

Cellular lipid metabolism and homeostasis are controlled by sterol regulatory-element binding proteins (SREBPs). In addition to performing canonical functions in the transcriptional regulation of genes involved in the biosynthesis and uptake of lipids, genome-wide system analyses have revealed that these versatile transcription factors act as important nodes of convergence and divergence within biological signalling networks. Thus, they are involved in myriad physiological and pathophysiological processes, highlighting the importance of lipid metabolism in biology. Changes in cell metabolism and growth are reciprocally linked through SREBPs. Anabolic and growth signalling pathways branch off and connect to multiple steps of SREBP activation and form complex regulatory networks. In addition, SREBPs are implicated in numerous pathogenic processes such as endoplasmic reticulum stress, inflammation, autophagy and apoptosis, and in this way, they contribute to obesity, dyslipidaemia, diabetes mellitus, nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, chronic kidney disease, neurodegenerative diseases and cancers. This Review aims to provide a comprehensive understanding of the role of SREBPs in physiology and pathophysiology at the cell, organ and organism levels.

Cysteine-rich peptides (CRPs) mediate diverse aspects of cell-cell communication in plant reproduction and development.

PubMed

Marshall, Eleanor; Costa, Liliana M; Gutierrez-Marcos, Jose

2011-03-01

Cell-cell communication in plants is essential for the correct co-ordination of reproduction, growth, and development. Studies to dissect this mode of communication have previously focussed primarily on the action of plant hormones as mediators of intercellular signalling. In animals, peptide signalling is a well-documented intercellular communication system, however, relatively little is known about this system in plants. In recent years, numerous reports have emerged about small, secreted peptides controlling different aspects of plant reproduction. Interestingly, most of these peptides are cysteine-rich, and there is convincing evidence suggesting multiple roles for related cysteine-rich peptides (CRPs) as signalling factors in developmental patterning as well as during plant pathogen responses and symbiosis. In this review, we discuss how CRPs are emerging as key signalling factors in regulating multiple aspects of vegetative growth and reproductive development in plants.

Transcriptional Differences between Rhesus Embryonic Stem Cells Generated from In Vitro and In Vivo Derived Embryos

PubMed Central

Harvey, Alexandra J.; Mao, Shihong; Lalancette, Claudia; Krawetz, Stephen A.; Brenner, Carol A.

2012-01-01

Numerous studies have focused on the transcriptional signatures that underlie the maintenance of embryonic stem cell (ESC) pluripotency. However, it remains unclear whether ESC retain transcriptional aberrations seen in in vitro cultured embryos. Here we report the first global transcriptional profile comparison between ESC generated from either in vitro cultured or in vivo derived primate embryos by microarray analysis. Genes involved in pluripotency, oxygen regulation and the cell cycle were downregulated in rhesus ESC generated from in vitro cultured embryos (in vitro ESC). Significantly, several gene differences are similarly downregulated in preimplantation embryos cultured in vitro, which have been associated with long term developmental consequences and disease predisposition. This data indicates that prior to derivation, embryo quality may influence the molecular signature of ESC lines, and may differentially impact the physiology of cells prior to or following differentiation. PMID:23028448

The protein expression landscape of the Arabidopsis root

PubMed Central

Petricka, Jalean J.; Schauer, Monica A.; Megraw, Molly; Breakfield, Natalie W.; Thompson, J. Will; Georgiev, Stoyan; Soderblom, Erik J.; Ohler, Uwe; Moseley, Martin Arthur; Grossniklaus, Ueli; Benfey, Philip N.

2012-01-01

Because proteins are the major functional components of cells, knowledge of their cellular localization is crucial to gaining an understanding of the biology of multicellular organisms. We have generated a protein expression map of the Arabidopsis root providing the identity and cell type-specific localization of nearly 2,000 proteins. Grouping proteins into functional categories revealed unique cellular functions and identified cell type-specific biomarkers. Cellular colocalization provided support for numerous protein–protein interactions. With a binary comparison, we found that RNA and protein expression profiles are weakly correlated. We then performed peak integration at cell type-specific resolution and found an improved correlation with transcriptome data using continuous values. We performed GeLC-MS/MS (in-gel tryptic digestion followed by liquid chromatography-tandem mass spectrometry) proteomic experiments on mutants with ectopic and no root hairs, providing complementary proteomic data. Finally, among our root hair-specific proteins we identified two unique regulators of root hair development. PMID:22447775

Epigenetic and Epitranscriptomic Factors Make a Mark on Hematopoietic Stem Cell Development.

PubMed

Kasper, Dionna M; Nicoli, Stefania

2018-03-01

Blood specification is a highly dynamic process, whereby committed hemogenic endothelial cells (ECs) progressively transdifferentiate into multipotent, self-renewing hematopoietic stem cells (HSCs). Massive changes in gene expression must occur to switch cell identity, however the factors that mediate such an effect were a mystery until recently. This review summarizes the higher-order mechanisms involved in endothelial to hematopoietic reprogramming identified thus far. Accumulating evidence from mouse and zebrafish studies reveal that numerous chromatin-modifying (epigenetic) and RNA-modifying (epitranscriptomic) factors are required for the formation of HSCs from hemogenic endothelium. These genes function throughout the endothelial-hematopoietic transition, suggesting a dynamic interplay between 'epi'-machineries. Epigenetic and epitranscriptomic regulation are key mechanisms for reshaping global EC gene expression patterns to those that support HSC production. Future studies that capture modification dynamics should bring us closer to a complete understanding of how HSCs transition from hemogenic endothelium at the molecular level.

Tks5-dependent formation of circumferential podosomes/invadopodia mediates cell–cell fusion

PubMed Central

Oyama, Masaaki; Kozuka-Hata, Hiroko; Uehara, Shunsuke; Udagawa, Nobuyuki; Saya, Hideyuki; Matsuo, Koichi

2012-01-01

Osteoclasts fuse to form multinucleated cells during osteoclastogenesis. This process is mediated by dynamic rearrangement of the plasma membrane and cytoskeleton, and it requires numerous factors, many of which have been identified. The underlying mechanism remains obscure, however. In this paper, we show that Tks5, a master regulator of invadopodia in cancer cells, is crucial for osteoclast fusion downstream of phosphoinositide 3-kinase and Src. Expression of Tks5 was induced during osteoclastogenesis, and prevention of this induction impaired both the formation of circumferential podosomes and osteoclast fusion without affecting cell differentiation. Tyrosine phosphorylation of Tks5 was attenuated in Src−/− osteoclasts, likely accounting for defects in podosome organization and multinucleation in these cells. Circumferential invadopodia formation in B16F0 melanoma cells was also accompanied by Tks5 phosphorylation. Co-culture of B16F0 cells with osteoclasts in an inflammatory milieu promoted the formation of melanoma–osteoclast hybrid cells. Our results thus reveal an unexpected link between circumferential podosome/invadopodium formation and cell–cell fusion in and beyond osteoclasts. PMID:22584907

Protein profiling of single epidermal cell types from Arabidopsis thaliana using surface-enhanced laser desorption and ionization technology.

PubMed

Ebert, Berit; Melle, Christian; Lieckfeldt, Elke; Zöller, Daniela; von Eggeling, Ferdinand; Fisahn, Joachim

2008-08-25

Here, we describe a novel approach for investigating differential protein expression within three epidermal cell types. In particular, 3000 single pavement, basal, and trichome cells from leaves of Arabidopsis thaliana were harvested by glass micro-capillaries. Subsequently, these single cell samples were joined to form pools of 100 individual cells and analyzed using the ProteinChip technology; SELDI: surface-enhanced laser desorption and ionization. As a result, numerous protein signals that were differentially expressed in the three epidermal cell types could be detected. One of these proteins was characterized by tryptical digestion and subsequent identification via tandem quadrupole-time of flight (Q-TOF) mass spectrometry. Down regulation of this sequenced small subunit precursor of ribulose-1,5 bisphosphate carboxylase(C) oxygenase(O) (RuBisCo) in trichome and basal cells indicates the sink status of these cell types that are located on the surface of A. thaliana source leaves. Based on the obtained protein profiles, we suggest a close functional relationship between basal and trichome cells at the protein level.

Targeting neuronal activity-regulated neuroligin-3 dependency in high-grade glioma

PubMed Central

Venkatesh, Humsa S.; Tam, Lydia T.; Woo, Pamelyn J.; Lennon, James; Nagaraja, Surya; Gillespie, Shawn M.; Ni, Jing; Duveau, Damien Y.; Morris, Patrick J.; Zhao, Jean J.; Thomas, Craig J.; Monje, Michelle

2017-01-01

Summary High-grade gliomas (HGG) are a devastating group of cancers, representing the leading cause of brain tumor-related death in both children and adults. Therapies aimed at mechanisms intrinsic to the glioma cell have translated to only limited success; effective therapeutic strategies will need to also target elements of the tumor microenvironment that promote glioma progression. We recently demonstrated that neuronal activity robustly promotes the growth of a range of molecularly and clinically distinct HGG types, including adult glioblastoma (GBM), anaplastic oligodendroglioma, pediatric GBM, and diffuse intrinsic pontine glioma (DIPG)1. An important mechanism mediating this neural regulation of brain cancer is activity-dependent cleavage and secretion of the synaptic molecule neuroligin-3 (NLGN3), which promotes glioma proliferation through the PI3K-mTOR pathway1. However, neuroligin-3 necessity to glioma growth, proteolytic mechanism of secretion and further molecular consequences in glioma remain to be clarified. Here, we demonstrate a striking dependence of HGG growth on microenvironmental neuroligin-3, elucidate signaling cascades downstream of neuroligin-3 binding in glioma and determine a therapeutically targetable mechanism of secretion. Patient-derived orthotopic xenografts of pediatric GBM, DIPG and adult GBM fail to grow in Nlgn3 knockout mice. Neuroligin-3 stimulates numerous oncogenic pathways, including early focal adhesion kinase activation upstream of PI3K-mTOR, and induces transcriptional changes including upregulation of numerous synapse-related genes in glioma cells. Neuroligin-3 is cleaved from both neurons and oligodendrocyte precursor cells via the ADAM10 sheddase. ADAM10 inhibitors prevent release of neuroligin-3 into the tumor microenvironment and robustly block HGG xenograft growth. This work defines a promising strategy for targeting neuroligin-3 secretion, which could prove transformative for HGG therapy. PMID:28959975

Human nutrigenomics of gene regulation by dietary fatty acids.

PubMed

Afman, Lydia A; Müller, Michael

2012-01-01

Nutrigenomics employs high-throughput genomics technologies to unravel how nutrients modulate gene and protein expression and ultimately influence cellular and organism metabolism. The most often-applied genomics technique so far is transcriptomics, which allows quantifying genome-wide changes in gene expression of thousands of genes at the same time in one sample. The performance of gene expression quantification requires sufficient high-quality homogenous cellular material, therefore research in healthy volunteers is restricted to biopsies from easy accessible tissues such as subcutaneous adipose tissue, skeletal muscle and intestinal biopsies or even more easily accessible cells such as peripheral blood mononuclear cells from blood. There is now significant evidence that fatty acids, in particular unsaturated fatty acids, exert many of their effects through modulation of gene transcription by regulating the activity of numerous transcription factors, including nuclear receptors such as peroxisome proliferator activated receptors, liver X receptor and sterol regulatory binding proteins. This review evaluates the human nutrigenomics studies performed on dietary fat since the initiation of nutrigenomics research around 10 years ago. Although the number of studies is still limited, all studies clearly suggest that changes in dietary fatty acids intake and composition can have a significant impact on cellular adaptive response capacity by gene transcription changes in humans. This adds important knowledge to our understanding of the strong effects that various fatty acids can have on numerous metabolic and inflammatory pathways, signaling routes and homeostatic control in the cell and ultimately on whole body health. It is important to use and integrate nutrigenomics in all future nutrition studies to build up the necessary framework for evidence-based nutrition in near future. Copyright © 2011 Elsevier Ltd. All rights reserved.

cAMP-specific PDE4 Phosphodiesterases and AIP in the Pathogenesis of Pituitary Tumors

PubMed Central

Bolger, Graeme B.; Bizzi, Mariana Ferreira; Brant Pinheiro, Sergio Veloso; Trivellin, Giampaolo; Smoot, Lisa; Accavitti, Mary-Ann; Korbonits, Márta; Ribeiro-Oliveira, Antonio

2016-01-01

PDE4 cyclic nucleotide phosphodiesterases regulate cAMP abundance in cells and thereby regulate numerous processes, including cell growth and differentiation. The rat PDE4A5 isoform (human homologue PDE4A4) interacts with the AIP protein (also called XAP2 or ARA-9). Germline mutations in AIP occur in approximately 20% of patients with Familial Isolated Pituitary Adenoma (FIPA) and 20% of childhood-onset simplex somatotroph adenomas. We therefore examined the protein expression of PDE4A4 and the closely-related isoform PDE4A8 in normal human pituitary tissue and in pituitary adenomas. PDE4A4 had low expression in normal pituitary, but was significantly over-expressed in somatotroph, lactotroph, corticotroph and clinically non-functioning gonadotroph adenomas (P<0.0001 for all subtypes). Likewise, PDE4A8 was expressed in normal pituitary and was also significantly over-expressed in the adenoma subtypes (P<0.0001 for all). Among the different adenoma subtypes, corticotroph and lactotroph adenomas were the highest and lowest expressed for PDE4A4, respectively, whereas the opposite was observed for PDE4A8. Naturally occurring oncogenic variants in AIP were shown by a two-hybrid assay to disrupt the ability of AIP to interact with PDE4A5. A reverse-two-hybrid screen identified numerous additional variants in the TPR region of AIP that also disrupted its ability to interact with PDE4A5. The expression of PDE4A4 and PDE4A8 in normal pituitary, their increased expression in adenomatous pituitary cells where AIP is meant to participate, and the disruption of the PDE4A4-AIP interaction by AIP mutants may play a role in pituitary tumorigenesis. PMID:27267386

Mammary Gland Development

PubMed Central

Macias, Hector

2012-01-01

The mammary gland develops through several distinct stages. The first transpires in the embryo as the ectoderm forms a mammary line that resolves into placodes. Regulated by epithelial/mesenchymal interactions, the placodes descend into the underlying mesenchyme and produce the rudimentary ductal structure of the gland present at birth. Subsequent stages of development – pubertal growth, pregnancy, lactation and involution – occur postnatally under the regulation of hormones. Puberty initiates branching morphogenesis, which requires growth hormone and estrogen, as well as IGF1, to create a ductal tree that fills the fat pad. Upon pregnancy the combined actions of progesterone and prolactin generate alveoli, which secrete milk during lactation. Lack of demand for milk at weaning initiates the process of involution whereby the gland is remodeled back to its pre-pregnancy state. These processes require numerous signaling pathways that have distinct regulatory functions at different stages of gland development. Signaling pathways also regulate a specialized subpopulation of mammary stem cells that fuel the dramatic changes in the gland occurring with each pregnancy. Our knowledge of mammary gland development and mammary stem cell biology has significantly contributed to our understanding of breast cancer and has advanced the discovery of therapies to treat this disease. PMID:22844349

The Hippo Pathway as Drug Targets in Cancer Therapy and Regenerative Medicine.

PubMed

Nagashima, Shunta; Bao, Yijun; Hata, Yutaka

2017-01-01

Yes-associated protein 1 (YAP1) and transcriptional co-activator with PDZ-binding motif (TAZ) co-operate with numerous transcription factors to regulate gene transcriptions. YAP1 and TAZ are negatively regulated by the tumor suppressive Hippo pathway. In human cancers, the Hippo pathway is frequently deregulated and YAP1 and TAZ escape the inhibition by the Hippo pathway. The upregulation of YAP1 and TAZ induces epithelial-mesenchymal transition and increases drug resistance in cancer cells. TAZ is implicated in cancer stemness. In consequence cancers with hyperactive YAP1 and TAZ are associated with poor clinical prognosis. Inhibitors of YAP1 and TAZ are reasoned to be beneficial in cancer therapy. On the other hand, since YAP1 and TAZ play important roles in the regulation of various tissue stem cells and in tissue repair, activators of YAP1 and TAZ are useful in the regenerative medicine. We discuss the potential application of inhibitors and activators of YAP1 and TAZ in human diseases and review the progress of drug screenings to search for them. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Identification of antisense long noncoding RNAs that function as SINEUPs in human cells.

PubMed

Schein, Aleks; Zucchelli, Silvia; Kauppinen, Sakari; Gustincich, Stefano; Carninci, Piero

2016-09-20

Mammalian genomes encode numerous natural antisense long noncoding RNAs (lncRNAs) that regulate gene expression. Recently, an antisense lncRNA to mouse Ubiquitin carboxyl-terminal hydrolase L1 (Uchl1) was reported to increase UCHL1 protein synthesis, representing a new functional class of lncRNAs, designated as SINEUPs, for SINE element-containing translation UP-regulators. Here, we show that an antisense lncRNA to the human protein phosphatase 1 regulatory subunit 12A (PPP1R12A), named as R12A-AS1, which overlaps with the 5' UTR and first coding exon of the PPP1R12A mRNA, functions as a SINEUP, increasing PPP1R12A protein translation in human cells. The SINEUP activity depends on the aforementioned sense-antisense interaction and a free right Alu monomer repeat element at the 3' end of R12A-AS1. In addition, we identify another human antisense lncRNA with SINEUP activity. Our results demonstrate for the first time that human natural antisense lncRNAs can up-regulate protein translation, suggesting that endogenous SINEUPs may be widespread and present in many mammalian species.

Comparing anterior and posterior Hox complex formation reveals guidelines for predicting cis-regulatory elements

PubMed Central

Uhl, Juli D.; Cook, Tiffany A.; Gebelein, Brian

2010-01-01

Hox transcription factors specify numerous cell fates along the anterior-posterior axis by regulating the expression of downstream target genes. While expression analysis has uncovered large numbers of de-regulated genes in cells with altered Hox activity, determining which are direct versus indirect targets has remained a significant challenge. Here, we characterize the DNA binding activity of Hox transcription factor complexes on eight experimentally verified cis-regulatory elements. Hox factors regulate the activity of each element by forming protein complexes with two cofactor proteins, Extradenticle (Exd) and Homothorax (Hth). Using comparative DNA binding assays, we found that a number of flexible arrangements of Hox, Exd, and Hth binding sites mediate cooperative transcription factor complexes. Moreover, analysis of a Distal-less regulatory element (DMXR) that is repressed by abdominal Hox factors revealed that suboptimal binding sites can be combined to form high affinity transcription complexes. Lastly, we determined that the anterior Hox factors are more dependent upon Exd and Hth for complex formation than posterior Hox factors. Based upon these findings, we suggest a general set of guidelines to serve as a basis for designing bioinformatics algorithms aimed at identifying Hox regulatory elements using the wealth of recently sequenced genomes. PMID:20398649

Regulation of skeletal growth and mineral acquisition by the GH/IGF-1 axis: Lessons from mouse models.

PubMed

Yakar, Shoshana; Isaksson, Olle

2016-06-01

The growth hormone (GH) and its downstream mediator, the insulin-like growth factor-1 (IGF-1), construct a pleotropic axis affecting growth, metabolism, and organ function. Serum levels of GH/IGF-1 rise during pubertal growth and associate with peak bone acquisition, while during aging their levels decline and associate with bone loss. The GH/IGF-1 axis was extensively studied in numerous biological systems including rodent models and cell cultures. Both hormones act in an endocrine and autocrine/paracrine fashion and understanding their distinct and overlapping contributions to skeletal acquisition is still a matter of debate. GH and IGF-1 exert their effects on osteogenic cells via binding to their cognate receptor, leading to activation of an array of genes that mediate cellular differentiation and function. Both hormones interact with other skeletal regulators, such as sex-steroids, thyroid hormone, and parathyroid hormone, to facilitate skeletal growth and metabolism. In this review we summarized several rodent models of the GH/IGF-1 axis and described key experiments that shed new light on the regulation of skeletal growth by the GH/IGF-1 axis. Copyright © 2015 Elsevier Ltd. All rights reserved.

Regulation of skeletal growth and mineral acquisition by the GH/IGF-1 axis: Lessons from mouse models

PubMed Central

Yakar, Shoshana; Isaksson, Olle

2015-01-01

The growth hormone (GH) and its downstream mediator, the insulin-like growth factor-1 (IGF-1), construct a pleotropic axis affecting growth, metabolism, and organ function. Serum levels of GH/IGF-1 rise during pubertal growth and associate with peak bone acquisition, while during aging their levels decline and associate with bone loss. The GH/IGF-1 axis was extensively studied in numerous biological systems including rodent models and cell cultures. Both hormones act in an endocrine and autocrine/paracrine fashion and understanding their distinct and overlapping contributions to skeletal acquisition is still a matter of debate. GH and IGF-1 exert their effects on osteogenic cells via binding to their cognate receptor, leading to activation of an array of genes that mediate cellular differentiation and function. Both hormones interact with other skeletal regulators, such as sex-steroids, thyroid hormone, and parathyroid hormone, to facilitate skeletal growth and metabolism. In this review we summarized several rodent models of the GH/IGF-1 axis and described key experiments that shed new light on the regulation of skeletal growth by the GH/IGF-1 axis. PMID:26432542

Does stress remove the HDAC brakes for the formation and persistence of long-term memory?

PubMed

White, André O; Wood, Marcelo A

2014-07-01

It has been known for numerous decades that gene expression is required for long-lasting forms of memory. In the past decade, the study of epigenetic mechanisms in memory processes has revealed yet another layer of complexity in the regulation of gene expression. Epigenetic mechanisms do not only provide complexity in the protein regulatory complexes that control coordinate transcription for specific cell function, but the epigenome encodes critical information that integrates experience and cellular history for specific cell functions as well. Thus, epigenetic mechanisms provide a unique mechanism of gene expression regulation for memory processes. This may be why critical negative regulators of gene expression, such as histone deacetylases (HDACs), have powerful effects on the formation and persistence of memory. For example, HDAC inhibition has been shown to transform a subthreshold learning event into robust long-term memory and also generate a form of long-term memory that persists beyond the point at which normal long-term memory fails. A key question that is explored in this review, from a learning and memory perspective, is whether stress-dependent signaling drives the formation and persistence of long-term memory via HDAC-dependent mechanisms. Copyright © 2013 Elsevier Inc. All rights reserved.

Does stress remove the HDAC brakes for the formation and persistence of long-term memory?

PubMed Central

White, André O.; Wood, Marcelo A.

2013-01-01

It has been known for numerous decades that gene expression is required for long-lasting forms of memory. In the past decade, the study of epigenetic mechanisms in memory processes has revealed yet another layer of complexity in the regulation of gene expression. Epigenetic mechanisms do not only provide complexity in the protein regulatory complexes that control coordinate transcription for specific cell function, but the epigenome encodes critical information that integrates experience and cellular history for specific cell functions as well. Thus, epigenetic mechanisms provide a unique mechanism of gene expression regulation for memory processes. This may be why critical negative regulators of gene expression, such as histone deacetylases (HDACs), have powerful effects on the formation and persistence of memory. For example, HDAC inhibition has been shown to transform a subthreshold learning event into robust long-term memory and also generate a form of long-term memory that persists beyond the point at which normal long-term memory fails. A key question that is explored in this review, from a learning and memory perspective, is whether stress-dependent signaling drives the formation and persistence of long-term memory via HDAC-dependent mechanisms. PMID:24149059

State of the APC/C: Organization, function, and structure

PubMed Central

McLean, Janel R.; Chaix, Denis; Ohi, Melanie D.; Gould, Kathleen L.

2016-01-01

The ubiquitin-proteasome protein degradation system is involved in many essential cellular processes including cell cycle regulation, cell differentiation, and the unfolded protein response.The anaphase-promoting complex/cyclosome (APC/C), an evolutionary conserved E3 ubiquitin ligase, was discovered 15 years ago because of its pivotal role in cyclin degradation and mitotic progression. Since then, we have learned that the APC/C is a very large, complex E3 ligase composed of 13 subunits, yielding a molecular machine of approximately 1 MDa. The intricate regulation of the APC/C is mediated by the Cdc20 family of activators, pseudosubstrate inhibitors, protein kinases and phosphatases and the spindle assembly checkpoint. The large size, complexity, and dynamic nature of the APC/C represent significant obstacles toward high-resolution structural techniques; however, over the last decade, there have been a number of lower resolution APC/C structures determined using single particle electron microscopy. These structures, when combined with data generated from numerous genetic and biochemical studies, have begun to shed light on how APC/C activity is regulated. Here, we discuss the most recent developments in the APC/C field concerning structure, substrate recognition, and catalysis. PMID:21261459

Electroporation of the Testis

NASA Astrophysics Data System (ADS)

Yomogida, Kentaro

The mature mammalian testis is a marvelous organ that produces numerous sperm cells during its reproductive phase. This biologically significant process consists of three steps: stem cell self-renewal and differentiation, meiosis and genetic recombination, and haploid cell morphogenesis into sperm (Russell et al., 1990). The first step provides a good model for investigating the molecular mechanism of stem cell regulation. Currently, the mechanism underlying sperm cell production is a very exciting topic in regenerative medicine (Lensch et al. 2007; Okita et al., 2007). The spermatogonial stem cell system has several advantages, including the easy histological identification of stem cells (Russell et al., 1990), a clear relationship between stem cells and the supporting Sertoli cells, which provide a stem cell niche (Tadokoro et al., 2002; Yomogida et al., 2003), and a transplantation assay for stem cell activity (Oatley & Brinster, 2006). Although germline stem (GS) cells derived from the gonocytes in newborn testis constitute a suitable in vitro system for investigating the properties of spermatogonial stem cells (Kanatsu-Shinohara et al., 2003, 2004), studies using living mammalian testes continue to provide information regarding the roles of the stem cell niche. In vivo electroporation of the supporting cells in the testis will expand our ability to study it.

Role of Melatonin in Cell-Wall Disassembly and Chilling Tolerance in Cold-Stored Peach Fruit.

PubMed

Cao, Shifeng; Bian, Kun; Shi, Liyu; Chung, Hsiao-Hang; Chen, Wei; Yang, Zhenfeng

2018-06-06

Melatonin reportedly increases chilling tolerance in postharvest peach fruit during cold storage, but information on its effects on cell-wall disassembly in chilling-injured peaches is limited. In this study, we investigated the role of cell-wall depolymerization in chilling-tolerance induction in melatonin-treated peaches. Treatment with 100 μM melatonin alleviated chilling symptoms (mealiness) characterized by a decrease in fruit firmness and increase in juice extractability in treated peaches during storage. The loss of neutral sugars, such as arabinose and galactose, in both the 1,2-cyclohexylenedinitrilotetraacetic acid (CDTA)- and Na 2 CO 3 -soluble fractions was observed at 7 days in treated peaches, but the contents increased after 28 days of storage. Atomic-force-microscopy (AFM) analysis revealed that the polysaccharide widths in the CDTA- and Na 2 CO 3 -soluble fractions in the treated fruit were mainly distributed in a shorter range, as compared with those in the control fruit. In addition, the expression profiles of a series of cell-wall-related genes showed that melatonin treatment maintained the balance between transcripts of PpPME and PpPG, which accompany the up-regulation of several other genes involved in cell-wall disassembly. Taken together, our results suggested that the reduced mealiness by melatonin was probably associated with its positive regulation of numerous cell-wall-modifying enzymes and proteins; thus, the depolymerization of the cell-wall polysaccharides in the peaches treated with melatonin was maintained, and the treated fruit could soften gradually during cold storage.

The Mediator Complex and Lipid Metabolism.

PubMed

Zhang, Yi; Xiaoli; Zhao, Xiaoping; Yang, Fajun

2013-03-01

The precise control of gene expression is essential for all biological processes. In addition to DNA-binding transcription factors, numerous transcription cofactors contribute another layer of regulation of gene transcription in eukaryotic cells. One of such transcription cofactors is the highly conserved Mediator complex, which has multiple subunits and is involved in various biological processes through directly interacting with relevant transcription factors. Although the current understanding on the biological functions of Mediator remains incomplete, research in the past decade has revealed an important role of Mediator in regulating lipid metabolism. Such function of Mediator is dependent on specific transcription factors, including peroxisome proliferator-activated receptor-gamma (PPARγ) and sterol regulatory element-binding proteins (SREBPs), which represent the master regulators of lipid metabolism. The medical significance of these findings is apparent, as aberrant lipid metabolism is intimately linked to major human diseases, such as type 2 diabetes and cardiovascular disease. Here, we briefly review the functions and molecular mechanisms of Mediator in regulation of lipid metabolism.

Genome-Wide RNAi Ionomics Screen Reveals New Genes and Regulation of Human Trace Element Metabolism

PubMed Central

Malinouski, Mikalai; Hasan, Nesrin M.; Zhang, Yan; Seravalli, Javier; Lin, Jie; Avanesov, Andrei; Lutsenko, Svetlana; Gladyshev, Vadim N.

2017-01-01

Trace elements are essential for human metabolism and dysregulation of their homeostasis is associated with numerous disorders. Here we characterize mechanisms that regulate trace elements in human cells by designing and performing a genome-wide high-throughput siRNA/ionomics screen, and examining top hits in cellular and biochemical assays. The screen reveals high stability of the ionomes, especially the zinc ionome, and yields known regulators and novel candidates. We further uncover fundamental differences in the regulation of different trace elements. Specifically, selenium levels are controlled through the selenocysteine machinery and expression of abundant selenoproteins; copper balance is affected by lipid metabolism and requires machinery involved in protein trafficking and posttranslational modifications; and the iron levels are influenced by iron import and expression of the iron/heme-containing enzymes. Our approach can be applied to a variety of disease models and/or nutritional conditions, and the generated dataset opens new directions for studies of human trace element metabolism. PMID:24522796

Regulation of autophagy by amino acid availability in S. cerevisiae and mammalian cells.

PubMed

Abeliovich, Hagai

2015-10-01

Autophagy is a catabolic membrane-trafficking process that occurs in all eukaryotic organisms analyzed to date. The study of autophagy has exploded over the last decade or so, branching into numerous aspects of cellular and organismal physiology. From basic functions in starvation and quality control, autophagy has expanded into innate immunity, aging, neurological diseases, redox regulation, and ciliogenesis, to name a few roles. In the present review, I would like to narrow the discussion to the more classical roles of autophagy in supporting viability under nutrient limitation. My aim is to provide a semblance of a historical overview, together with a concise, and perhaps subjective, mechanistic and functional analysis of the central questions in the autophagy field.

Biochemical regulation of pigment motility in vertebrate chromatophores: a review of physiological color change mechanisms

PubMed Central

McCartney, Kristen L.

2016-01-01

Abstract The fundamental unit of rapid, physiological color change in vertebrates is the dermal chromatophore unit. This unit, comprised of cellular associations between different chromatophore types, is relatively conserved across the fish, amphibian, and reptilian species capable of physiological color change and numerous attempts have been made to understand the nature of the four major chromatophore types (melanophores, erythrophores, xanthophores, and iridophores) and their biochemical regulation. In this review, we attempt to describe the current state of knowledge regarding what classifies a pigment cell as a dynamic chromatophore, the unique characteristics of each chromatophore type, and how different hormones, neurotransmitters, or other signals direct pigment reorganization in a variety of vertebrate taxa. PMID:29491911

Canonical Wnt signaling in megakaryocytes regulates proplatelet formation

PubMed Central

Macaulay, Iain C.; Thon, Jonathan N.; Tijssen, Marloes R.; Steele, Brian M.; MacDonald, Bryan T.; Meade, Gerardene; Burns, Philippa; Rendon, Augusto; Salunkhe, Vishal; Murphy, Ronan P.; Bennett, Cavan; Watkins, Nicholas A.; He, Xi; Fitzgerald, Desmond J.; Italiano, Joseph E.

2013-01-01

Wnt signaling is involved in numerous aspects of vertebrate development and homeostasis, including the formation and function of blood cells. Here, we show that canonical and noncanonical Wnt signaling pathways are present and functional in megakaryocytes (MKs), with several Wnt effectors displaying MK-restricted expression. Using the CHRF288-11 cell line as a model for human MKs, the canonical Wnt3a signal was found to induce a time and dose-dependent increase in β-catenin expression. β-catenin accumulation was inhibited by the canonical antagonist dickkopf-1 (DKK1) and by the noncanonical agonist Wnt5a. Whole genome expression analysis demonstrated that Wnt3a and Wnt5a regulated distinct patterns of gene expression in MKs, and revealed a further interplay between canonical and noncanonical Wnt pathways. Fetal liver cells derived from low-density-lipoprotein receptor-related protein 6-deficient mice (LRP6−/−), generated dramatically reduced numbers of MKs in culture of lower ploidy (2N and 4N) than wild-type controls, implicating LRP6-dependent Wnt signaling in MK proliferation and maturation. Finally, in wild-type mature murine fetal liver-derived MKs, Wnt3a potently induced proplatelet formation, an effect that could be completely abrogated by DKK1. These data identify novel extrinsic regulators of proplatelet formation, and reveal a profound role for Wnt signaling in platelet production. PMID:23160460

Ankyrin-G Inhibits Endocytosis of Cadherin Dimers.

PubMed

Cadwell, Chantel M; Jenkins, Paul M; Bennett, Vann; Kowalczyk, Andrew P

2016-01-08

Dynamic regulation of endothelial cell adhesion is central to vascular development and maintenance. Furthermore, altered endothelial adhesion is implicated in numerous diseases. Therefore, normal vascular patterning and maintenance require tight regulation of endothelial cell adhesion dynamics. However, the mechanisms that control junctional plasticity are not fully understood. Vascular endothelial cadherin (VE-cadherin) is an adhesive protein found in adherens junctions of endothelial cells. VE-cadherin mediates adhesion through trans interactions formed by its extracellular domain. Trans binding is followed by cis interactions that laterally cluster the cadherin in junctions. VE-cadherin is linked to the actin cytoskeleton through cytoplasmic interactions with β- and α-catenin, which serve to increase adhesive strength. Furthermore, p120-catenin binds to the cytoplasmic tail of cadherin and stabilizes it at the plasma membrane. Here we report that induced cis dimerization of VE-cadherin inhibits endocytosis independent of both p120 binding and trans interactions. However, we find that ankyrin-G, a protein that links membrane proteins to the spectrin-actin cytoskeleton, associates with VE-cadherin and inhibits its endocytosis. Ankyrin-G inhibits VE-cadherin endocytosis independent of p120 binding. We propose a model in which ankyrin-G associates with and inhibits the endocytosis of VE-cadherin cis dimers. Our findings support a novel mechanism for regulation of VE-cadherin endocytosis through ankyrin association with cadherin engaged in lateral interactions. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Membrane transporters mediating root signalling and adaptive responses to oxygen deprivation and soil flooding.

PubMed

Shabala, Sergey; Shabala, Lana; Barcelo, Juan; Poschenrieder, Charlotte

2014-10-01

This review provides a comprehensive assessment of a previously unexplored topic: elucidating the role that plasma- and organelle-based membrane transporters play in plant-adaptive responses to flooding. We show that energy availability and metabolic shifts under hypoxia and anoxia are critical in regulating membrane-transport activity. We illustrate the high tissue and time dependence of this regulation, reveal the molecular identity of transporters involved and discuss the modes of their regulation. We show that both reduced oxygen availability and accumulation of transition metals in flooded roots result in a reduction in the cytosolic K(+) pool, ultimately determining the cell's fate and transition to programmed cell death (PCD). This process can be strongly affected by hypoxia-induced changes in the amino acid pool profile and, specifically, ϒ-amino butyric acid (GABA) accumulation. It is suggested that GABA plays an important regulatory role, allowing plants to proceed with H2 O2 signalling to activate a cascade of genes that mediate plant adaptation to flooding while at the same time, preventing the cell from entering a 'suicide program'. We conclude that progress in crop breeding for flooding tolerance can only be achieved by pyramiding the numerous physiological traits that confer efficient energy maintenance, cytosolic ion homeostasis, and reactive oxygen species (ROS) control and detoxification. © 2014 John Wiley & Sons Ltd.

Dynamics in the tomato root transcriptome on infection with the potato cyst nematode Globodera rostochiensis.

PubMed

Swiecicka, Magdalena; Filipecki, Marcin; Lont, Dieuwertje; Van Vliet, Joke; Qin, Ling; Goverse, Aska; Bakker, Jaap; Helder, Johannes

2009-07-01

Plant parasitic nematodes infect roots and trigger the formation of specialized feeding sites by substantial reprogramming of the developmental process of root cells. In this article, we describe the dynamic changes in the tomato root transcriptome during early interactions with the potato cyst nematode Globodera rostochiensis. Using amplified fragment length polymorphism-based mRNA fingerprinting (cDNA-AFLP), we monitored 17 600 transcript-derived fragments (TDFs) in infected and uninfected tomato roots, 1-14 days after inoculation with nematode larvae. Six hundred and twenty-four TDFs (3.5%) showed significant differential expression on nematode infection. We employed GenEST, a computer program which links gene expression profiles generated by cDNA-AFLP and databases of cDNA sequences, to identify 135 tomato sequences. These sequences were grouped into eight functional categories based on the presence of genes involved in hormone regulation, plant pathogen defence response, cell cycle and cytoskeleton regulation, cell wall modification, cellular signalling, transcriptional regulation, primary metabolism and allocation. The presence of unclassified genes was also taken into consideration. This article describes the responsiveness of numerous tomato genes hitherto uncharacterized during infection with endoparasitic cyst nematodes. The analysis of transcriptome profiles allowed the sequential order of expression to be dissected for many groups of genes and the genes to be connected with the biological processes involved in compatible interactions between the plant and nematode.

Coordinated action of histone modification and microRNA regulations in human genome.

PubMed

Wang, Xuan; Zheng, Guantao; Dong, Dong

2015-10-10

Both histone modifications and microRNAs (miRNAs) play pivotal role in gene expression regulation. Although numerous studies have been devoted to explore the gene regulation by miRNA and epigenetic regulations, their coordinated actions have not been comprehensively examined. In this work, we systematically investigated the combinatorial relationship between miRNA and epigenetic regulation by taking advantage of recently published whole genome-wide histone modification data and high quality miRNA targeting data. The results showed that miRNA targets have distinct histone modification patterns compared with non-targets in their promoter regions. Based on this finding, we proposed a machine learning approach to fit predictive models on the task to discern whether a gene is targeted by a specific miRNA. We found a considerable advantage in both sensitivity and specificity in diverse human cell lines. Finally, we found that our predicted miRNA targets are consistently annotated with Gene Ontology terms. Our work is the first genome-wide investigation of the coordinated action of miRNA and histone modification regulations, which provide a guide to deeply understand the complexity of transcriptional regulation. Copyright © 2015 Elsevier B.V. All rights reserved.

A whole organism screen identifies novel regulators of fat storage

PubMed Central

Lemieux, George A.; Liu, Jason; Mayer, Nasima; Bainton, Roland J.; Ashrafi, Kaveh; Werb, Zena

2011-01-01

The regulation of energy homeostasis integrates diverse biological processes ranging from behavior to metabolism and is linked fundamentally to numerous disease states. To identify new molecules that can bypass homeostatic compensatory mechanisms of energy balance in intact animals, we screened for small molecule modulators of C. elegans fat content. We report on several molecules that modulate fat storage without obvious deleterious effects on feeding, growth, and reproduction. A subset of these compounds also altered fat storage in mammalian and insect cell culture. We found that one of the newly identified compounds exerts its effects in C. elegans through a pathway that requires novel functions of an AMP-activated kinase catalytic subunit and a transcription factor previously unassociated with fat regulation. Thus, our strategy identifies small molecules that are effective within the context of intact animals and reveals relationships between new pathways that operate across phyla to influence energy homeostasis. PMID:21390037

STK33 potentiates the malignancy of hypopharyngeal squamous carcinoma: Possible relation to calcium

PubMed Central

Chen, Chen; Huang, Lingyan; Zhang, Guodong; Li, Yang; Li, Li; Bai, Xiaohui; Liu, Wenwen; Wang, Haibo; Li, Jianfeng

2016-01-01

ABSTRACT Background: The present study aims to further explore the role of STK33 in hypopharyngeal squamous cell carcinoma (HSCC), with special attention given to the possible relationship between STK33 alteration and calcium. Methods: An in vivo experiment and microarray analysis were performed to investigate the impact of STK33 knockdown (STK33-RNAi) on the biological behaviors and the gene profile alterations of a HSCC cell line (Fadu). Cell viability and morphological change of Fadu cells in response to Ionomycin were measured by MTT assay and acridine orange staining. The concentration of intracellular calcium ([Ca2+]i) was detected by laser scanning confocal microscope with fluo-3/AM. The mRNA and protein expressions of relevant genes were examined by real-time PCR and Western blot. Results: STK33-RNAi retarded the Fadu cell proliferation and the metastasis in nude mice and led to up- and down-regulation of the expressions of abundance of genes, especially, the downregulation of the CAPN1 gene. Ionomycin increased the [Ca2+]i and decreased the survival rates of Fadu cells in a time-dependent manner. Moreover, Ionomycin resulted in the elevation of CAPN1 mRNA expression in normal Fadu cells and, conversely, had almost no effect on CAPN1 expression in STK33-RNAi cells. Conclusions: Findings from this work further validate that STK33 is a potential oncogene and plays an important role in tumorigenesis of HSCC via regulation of numerous genes. In addition, there exists the reciprocal influence between STK33 and [Ca2+]i in Fadu cells. PMID:27414193

Factors that regulate embryonic gustatory development

PubMed Central

Krimm, Robin F

2007-01-01

Numerous molecular factors orchestrate the development of the peripheral taste system. The unique anatomy/function of the taste system makes this system ideal for understanding the mechanisms by which these factors function; yet the taste system is underutilized for this role. This review focuses on some of the many factors that are known to regulate gustatory development, and discusses a few topics where more work is needed. Some attention is given to factors that regulate epibranchial placode formation, since gustatory neurons are thought to be primarily derived from this region. Epibranchial placodes appear to arise from a pan-placodal region and a number of regulatory factors control the differentiation of individual placodes. Gustatory neuron differentiation is regulated by a series of transcription factors and perhaps bone morphongenic proteins (BMP). As neurons differentiate, they also proliferate such that their numbers exceed those in the adult, and this is followed by developmental death. Some of these cell-cycling events are regulated by neurotrophins. After gustatory neurons become post-mitotic, axon outgrowth occurs. Axons are guided by multiple chemoattractive and chemorepulsive factors, including semaphorins, to the tongue epithelium. Brain derived neurotrophic factor (BDNF), functions as a targeting factor in the final stages of axon guidance and is required for gustatory axons to find and innervate taste epithelium. Numerous factors are involved in the development of gustatory papillae including Sox-2, Sonic hedge hog and Wnt-β-catenin signaling. It is likely that just as many factors regulate taste bud differentiation; however, these factors have not yet been identified. Studies examining the molecular factors that regulate terminal field formation in the nucleus of the solitary tract are also lacking. However, it is possible that some of the factors that regulate geniculate ganglion development, outgrowth, guidance and targeting of peripheral axons may have the same functions in the gustatory CNS. PMID:17903280

Stabilization of diastolic calcium signal via calcium pump regulation of complex local calcium releases and transient decay in a computational model of cardiac pacemaker cell with individual release channels

PubMed Central

Maltsev, Alexander V.; Maltsev, Victor A.; Stern, Michael D.

2017-01-01

Intracellular Local Ca releases (LCRs) from sarcoplasmic reticulum (SR) regulate cardiac pacemaker cell function by activation of electrogenic Na/Ca exchanger (NCX) during diastole. Prior studies demonstrated the existence of powerful compensatory mechanisms of LCR regulation via a complex local cross-talk of Ca pump, release and NCX. One major obstacle to study these mechanisms is that LCR exhibit complex Ca release propagation patterns (including merges and separations) that have not been characterized. Here we developed new terminology, classification, and computer algorithms for automatic detection of numerically simulated LCRs and examined LCR regulation by SR Ca pumping rate (Pup) that provides a major contribution to fight-or-flight response. In our simulations the faster SR Ca pumping accelerates action potential-induced Ca transient decay and quickly clears Ca under the cell membrane in diastole, preventing premature releases. Then the SR generates an earlier, more synchronized, and stronger diastolic LCR signal activating an earlier and larger inward NCX current. LCRs at higher Pup exhibit larger amplitudes and faster propagation with more collisions to each other. The LCRs overlap with Ca transient decay, causing an elevation of the average diastolic [Ca] nadir to ~200 nM (at Pup = 24 mM/s). Background Ca (in locations lacking LCRs) quickly decays to resting Ca levels (<100 nM) at high Pup, but remained elevated during slower decay at low Pup. Release propagation is facilitated at higher Pup by a larger LCR amplitude, whereas at low Pup by higher background Ca. While at low Pup LCRs show smaller amplitudes, their larger durations and sizes combined with longer transient decay stabilize integrals of diastolic Ca and NCX current signals. Thus, the local interplay of SR Ca pump and release channels regulates LCRs and Ca transient decay to insure fail-safe pacemaker cell operation within a wide range of rates. PMID:28792496

Atg7-Mediated Autophagy Is Involved in the Neural Crest Cell Generation in Chick Embryo.

PubMed

Wang, Guang; Chen, En-Ni; Liang, Chang; Liang, Jianxin; Gao, Lin-Rui; Chuai, Manli; Münsterberg, Andrea; Bao, Yongping; Cao, Liu; Yang, Xuesong

2018-04-01

Autophagy plays a very important role in numerous physiological and pathological events. However, it still remains unclear whether Atg7-induced autophagy is involved in the regulation of neural crest cell production. In this study, we found the co-location of Atg7 and Pax7 + neural crest cells in early chick embryo development. Upregulation of Atg7 with unilateral transfection of full-length Atg7 increased Pax7 + and HNK-1 + cephalic and trunk neural crest cell numbers compared to either Control-GFP transfection or opposite neural tubes, suggesting that Atg7 over-expression in neural tubes could enhance the production of neural crest cells. BMP4 in situ hybridization and p-Smad1/5/8 immunofluorescent staining demonstrated that upregulation of Atg7 in neural tubes suppressed the BMP4/Smad signaling, which is considered to promote the delamination of neural crest cells. Interestingly, upregulation of Atg7 in neural tubes could significantly accelerate cell progression into the S phase, implying that Atg7 modulates cell cycle progression. However, β-catenin expression was not significantly altered. Finally, we demonstrated that upregulation of the Atg7 gene could activate autophagy as did Atg8. We have also observed that similar phenotypes, such as more HNK-1 + neural crest cells in the unilateral Atg8 transfection side of neural tubes, and the transfection with full-length Atg8-GFP certainly promote the numbers of BrdU + neural crest cells in comparison to the GFP control. Taken together, we reveal that Atg7-induced autophagy is involved in regulating the production of neural crest cells in early chick embryos through the modification of the cell cycle.

Molecular Pathways: Gene-environment interactions regulating dietary fiber induction of proliferation and apoptosis via butyrate for cancer prevention

PubMed Central

Bultman, Scott J.

2013-01-01

Gene-environment interactions are so numerous and biologically complicated that it can be challenging to understand their role in cancer. However, dietary fiber and colorectal cancer prevention may represent a tractable model system. Fiber is fermented by colonic bacteria into short-chain fatty acids such as butyrate. One molecular pathway that has emerged involves butyrate having differential effects depending on its concentration and the metabolic state of the cell. Low-moderate concentrations, which are present near the base of colonic crypts, are readily metabolized in the mitochondria to stimulate cell proliferation via energetics. Higher concentrations, which are present near the lumen, exceed the metabolic capacity of the colonocyte. Unmetabolized butyrate enters the nucleus and functions as a histone deacetylase (HDAC) inhibitor that epigenetically regulates gene expression to inhibit cell proliferation and induce apoptosis as the colonocytes exfoliate into the lumen. Butyrate may therefore play a role in normal homeostasis by promoting turnover of the colonic epithelium. Because cancerous colonocytes undergo the Warburg effect, their preferred energy source is glucose instead of butyrate. Consequently, even moderate concentrations of butyrate accumulate in cancerous colonocytes and function as HDAC inhibitors to inhibit cell proliferation and induce apoptosis. These findings implicate a bacterial metabolite with metaboloepigenetic properties in tumor suppression. PMID:24270685

Turtle Functions Downstream of Cut in Differentially Regulating Class Specific Dendrite Morphogenesis in Drosophila

PubMed Central

Sulkowski, Mikolaj J.; Iyer, Srividya Chandramouli; Kurosawa, Mathieu S.; Iyer, Eswar Prasad R.; Cox, Daniel N.

2011-01-01

Background Dendritic morphology largely determines patterns of synaptic connectivity and electrochemical properties of a neuron. Neurons display a myriad diversity of dendritic geometries which serve as a basis for functional classification. Several types of molecules have recently been identified which regulate dendrite morphology by acting at the levels of transcriptional regulation, direct interactions with the cytoskeleton and organelles, and cell surface interactions. Although there has been substantial progress in understanding the molecular mechanisms of dendrite morphogenesis, the specification of class-specific dendritic arbors remains largely unexplained. Furthermore, the presence of numerous regulators suggests that they must work in concert. However, presently, few genetic pathways regulating dendrite development have been defined. Methodology/Principal Findings The Drosophila gene turtle belongs to an evolutionarily conserved class of immunoglobulin superfamily members found in the nervous systems of diverse organisms. We demonstrate that Turtle is differentially expressed in Drosophila da neurons. Moreover, MARCM analyses reveal Turtle acts cell autonomously to exert class specific effects on dendritic growth and/or branching in da neuron subclasses. Using transgenic overexpression of different Turtle isoforms, we find context-dependent, isoform-specific effects on mediating dendritic branching in class II, III and IV da neurons. Finally, we demonstrate via chromatin immunoprecipitation, qPCR, and immunohistochemistry analyses that Turtle expression is positively regulated by the Cut homeodomain transcription factor and via genetic interaction studies that Turtle is downstream effector of Cut-mediated regulation of da neuron dendrite morphology. Conclusions/Significance Our findings reveal that Turtle proteins differentially regulate the acquisition of class-specific dendrite morphologies. In addition, we have established a transcriptional regulatory interaction between Cut and Turtle, representing a novel pathway for mediating class specific dendrite development. PMID:21811639

Oncogene miR-187-5p is associated with cellular proliferation, migration, invasion, apoptosis and an increased risk of recurrence in bladder cancer.

PubMed

Li, Zuwei; Lin, Canbin; Zhao, Liwen; Zhou, Liang; Pan, Xiang; Quan, Jing; Peng, Xiqi; Li, Weiqing; Li, Hang; Xu, Jinling; Xu, Weijie; Guan, Xin; Chen, Yun; Lai, Yongqing

2018-06-05

Bladder cancer, the ninth-most-common malignancy worldwide with an estimated 356,000 new cases and 145,000 deaths annually, has a propensity to relapse, requiring lifelong monitoring after diagnosis. 75% patients diagnosed with BC are non-muscle invasive BC and over 50% of them experience recurrences within 6-12 years of initial diagnosis. miRNA are small, noncoding RNA and shown to be oncogenes or anti-oncogenes in bladder cancer, contributing to numerous BC cell processes, including cell proliferation, differentiation, migration and apoptosis. RT-qPCR were performed to detect the expression of miR-187-5p in tissues and cell lines, After which, clinicopathological variables and the prognostic value of altered miR-187-5p expression in BC was analyzed with the 48 formalin-fixed paraffin-embedded BC samples. Moreover, Cell functional assays (wound healing assay, CCK-8 assay, transwell assay and flow cytometry assay) were performed to explore the relationship between miR-187-5p expression and cell proliferation, migration, invasion and apoptosis in BC. Up-regulation of miR-187-5p was observed in BC tissues and BC cell lines. Cox proportional hazard regression analysis demonstrated that the patients with low expression of miR-187-5p experience lower risks of recurrence in the univariate and multivariate analysis. The Kaplan-Meier recurrence-free curves suggested that the patients with low expression of miR-187-5p experience lower risks of recurrence. Up-regulation of miR-187-5p promotes cell proliferation and mobility and inhibits the apoptosis of 5637 and UM-UC-3 cell, while down-regulation of miR-187-5p reverses these effects. The results of our study demonstrated that oncogene miR-187-5p is associated with cellular proliferation, migration, invasion, apoptosis and an increased risk of recurrence in bladder cancer. Copyright © 2018 The Authors. Published by Elsevier Masson SAS.. All rights reserved.

Learning a Markov Logic network for supervised gene regulatory network inference.

PubMed

Brouard, Céline; Vrain, Christel; Dubois, Julie; Castel, David; Debily, Marie-Anne; d'Alché-Buc, Florence

2013-09-12

Gene regulatory network inference remains a challenging problem in systems biology despite the numerous approaches that have been proposed. When substantial knowledge on a gene regulatory network is already available, supervised network inference is appropriate. Such a method builds a binary classifier able to assign a class (Regulation/No regulation) to an ordered pair of genes. Once learnt, the pairwise classifier can be used to predict new regulations. In this work, we explore the framework of Markov Logic Networks (MLN) that combine features of probabilistic graphical models with the expressivity of first-order logic rules. We propose to learn a Markov Logic network, e.g. a set of weighted rules that conclude on the predicate "regulates", starting from a known gene regulatory network involved in the switch proliferation/differentiation of keratinocyte cells, a set of experimental transcriptomic data and various descriptions of genes all encoded into first-order logic. As training data are unbalanced, we use asymmetric bagging to learn a set of MLNs. The prediction of a new regulation can then be obtained by averaging predictions of individual MLNs. As a side contribution, we propose three in silico tests to assess the performance of any pairwise classifier in various network inference tasks on real datasets. A first test consists of measuring the average performance on balanced edge prediction problem; a second one deals with the ability of the classifier, once enhanced by asymmetric bagging, to update a given network. Finally our main result concerns a third test that measures the ability of the method to predict regulations with a new set of genes. As expected, MLN, when provided with only numerical discretized gene expression data, does not perform as well as a pairwise SVM in terms of AUPR. However, when a more complete description of gene properties is provided by heterogeneous sources, MLN achieves the same performance as a black-box model such as a pairwise SVM while providing relevant insights on the predictions. The numerical studies show that MLN achieves very good predictive performance while opening the door to some interpretability of the decisions. Besides the ability to suggest new regulations, such an approach allows to cross-validate experimental data with existing knowledge.

The expression of cancer stem cell markers in human colorectal carcinoma cells in a microenvironment dependent manner.

PubMed

Stankevicius, Vaidotas; Kunigenas, Linas; Stankunas, Edvinas; Kuodyte, Karolina; Strainiene, Egle; Cicenas, Jonas; Samalavicius, Narimantas E; Suziedelis, Kestutis

2017-03-18

Numerous lines of evidence support the hierarchical model of cancer development and tumor initiation. According to the theory, cancer stem cells play a crucial role in the formation of the tumor and should be targeted for more effective anticancer treatment. However, cancer stem cells quickly loose their characteristics when propagated as 2D cell culture, indicating that the 2D cell culture does not provide the appropriate settings to maintain an in vivo environment. In this study we have investigated the expression of self-renewal, cancer stem cell and epithelial to mesenchymal transition markers after the transfer of human colorectal carcinoma cell DLD1 and HT29 lines from 2D cell cultures to scaffold-attached laminin rich extracellular matrix and scaffold-free multicellular spheroid 3D culture models. Based on the up-regulated expression of multipotency, CSC and EMT markers, our data suggests that human colorectal carcinoma cells grown in 3D exhibit enhanced cancer stem cell characteristics. Therefore, in order to design more efficient targeted therapies, we suggest that 3D cell culture models should be employed in cancer stem cell research. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Segregated Excitatory–Inhibitory Recurrent Subnetworks in Layer 5 of the Rat Frontal Cortex

PubMed Central

Morishima, Mieko; Kobayashi, Kenta; Kato, Shigeki; Kobayashi, Kazuto; Kawaguchi, Yasuo

2017-01-01

Abstract A prominent feature of neocortical pyramidal cells (PCs) is their numerous projections to diverse brain areas. In layer 5 (L5) of the rat frontal cortex, there are 2 major subtypes of PCs that differ in their long-range axonal projections, corticopontine (CPn) cells and crossed corticostriatal (CCS) cells. The outputs of these L5 PCs can be regulated by feedback inhibition from neighboring cortical GABAergic cells. Two major subtypes of GABAergic cells are parvalbumin (PV)-positive and somatostatin (SOM)-positive cells. PV cells have a fast-spiking (FS) firing pattern, while SOM cells have a low threshold spike (LTS) and regular spiking. In this study, we found that the 2 PC subtypes in L5 selectively make recurrent connections with LTS cells. The connection patterns correlated with the morphological and physiological diversity of LTS cells. LTS cells with high input resistance (Ri) exhibited more compact dendrites and more rebound spikes than LTS cells with low Ri, which had vertically elongated dendrites. LTS subgroups differently inhibited the PC subtypes, although FS cells made nonselective connections with both projection subtypes. These results demonstrate a novel recurrent network of inhibitory and projection-specific excitatory neurons within the neocortex. PMID:29045559

miRNA let-7b modulates macrophage polarization and enhances tumor-associated macrophages to promote angiogenesis and mobility in prostate cancer.

PubMed

Wang, Zhigang; Xu, Lu; Hu, Yinying; Huang, Yanqin; Zhang, Yujuan; Zheng, Xiufen; Wang, Shanshan; Wang, Yifan; Yu, Yanrong; Zhang, Meng; Yuan, Keng; Min, Weiping

2016-05-09

Macrophage polarization is a highly plastic physiological process that responds to a variety of environmental factors by changing macrophage phenotype and function. Tumor-associated macrophages (TAMs) are generally recognized as promoting tumor progression. As universal regulators, microRNAs (miRNAs) are functionally involved in numerous critical cellular processes including macrophage polarization. Let-7b, a miRNA, has differential expression patterns in inflamed tissues compared with healthy controls. However, whether and how miRNA let-7b regulates macrophage phenotype and function is unclear. In this report, we find that up-regulation of let-7b is characteristic of prostatic TAMs, and down-regulation of let-7b in TAMs leads to changes in expression profiles of inflammatory cytokines, such as IL-12, IL-23, IL-10 and TNF-α. As a result, TAMs treated with let-7b inhibitors reduce angiogenesis and prostate carcinoma (PCa) cell mobility. Let-7b may play a vital role in regulating macrophage polarization, thus modulating the prognosis of prostate cancer.

Glucose regulates enzymatic sources of mitochondrial NADPH in skeletal muscle cells; a novel role for glucose-6-phosphate dehydrogenase.

PubMed

Mailloux, Ryan J; Harper, Mary-Ellen

2010-07-01

Reduced nicotinamide adenine dinucleotide (NADPH) is a functionally important metabolite required to support numerous cellular processes. However, despite the identification of numerous NADPH-producing enzymes, the mechanisms underlying how the organellar pools of NADPH are maintained remain elusive. Here, we have identified glucose-6-phosphate dehydrogenase (G6PDH) as an important source of NADPH in mitochondria. Activity analysis, submitochondrial fractionation, fluorescence microscopy, and protease sensitivity assays revealed that G6PDH is localized to the mitochondrial matrix. 6-ANAM, a specific G6PDH inhibitor, depleted mitochondrial NADPH pools and increased oxidative stress revealing the importance of G6PDH in NADPH maintenance. We also show that glucose availability and differences in metabolic state modulate the enzymatic sources of NADPH in mitochondria. Indeed, cells cultured in high glucose (HG) not only adopted a glycolytic phenotype but also relied heavily on matrix-associated G6PDH as a source of NADPH. In contrast, cells exposed to low-glucose (LG) concentrations, which displayed increased oxygen consumption, mitochondrial metabolic efficiency, and decreased glycolysis, relied predominantly on isocitrate dehydrogenase (ICDH) as the principal NADPH-producing enzyme in the mitochondria. Culturing glycolytic cells in LG for 48 h decreased G6PDH and increased ICDH protein levels in the mitochondria, further pointing to the regulatory role of glucose. 2-Deoxyglucose treatment also prevented the increase of mitochondrial G6PDH in response to HG. The role of glucose in regulating enzymatic sources of mitochondrial NADPH pool maintenance was confirmed using human myotubes from obese adults with a history of type 2 diabetes mellitus (post-T2DM). Myotubes from post-T2DM participants failed to increase mitochondrial G6PDH in response to HG in contrast to mitochondria in myotubes from control participants (non-T2DM). Hence, we not only identified a matrix-associated G6PDH but also provide evidence that metabolic state/glucose availability modulate enzymatic sources of NADPH.

Apolipoprotein D Internalization Is a Basigin-dependent Mechanism.

PubMed

Najyb, Ouafa; Brissette, Louise; Rassart, Eric

2015-06-26

Apolipoprotein D (apoD), a member of the lipocalin family, is a 29-kDa secreted glycoprotein that binds and transports small lipophilic molecules. Expressed in several tissues, apoD is up-regulated under different stress stimuli and in a variety of pathologies. Numerous studies have revealed that overexpression of apoD led to neuroprotection in various mouse models of acute stress and neurodegeneration. This multifunctional protein is internalized in several cells types, but the specific internalization mechanism remains unknown. In this study, we demonstrate that the internalization of apoD involves a specific cell surface receptor in 293T cells, identified as the transmembrane glycoprotein basigin (BSG, CD147); more particularly, its low glycosylated form. Our results show that internalized apoD colocalizes with BSG into vesicular compartments. Down-regulation of BSG disrupted the internalization of apoD in cells. In contrast, overexpression of basigin in SH-5YSY cells, which poorly express BSG, restored the uptake of apoD. Cyclophilin A, a known ligand of BSG, competitively reduced apoD internalization, confirming that BSG is a key player in the apoD internalization process. In summary, our results demonstrate that basigin is very likely the apoD receptor and provide additional clues on the mechanisms involved in apoD-mediated functions, including neuroprotection. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Human Milk Cells Contain Numerous miRNAs that May Change with Milk Removal and Regulate Multiple Physiological Processes

PubMed Central

Alsaweed, Mohammed; Lai, Ching Tat; Hartmann, Peter E.; Geddes, Donna T.; Kakulas, Foteini

2016-01-01

Human milk (HM) is a complex biofluid conferring nutritional, protective and developmental components for optimal infant growth. Amongst these are maternal cells, which change in response to feeding and were recently shown to be a rich source of miRNAs. We used next generation sequencing to characterize the cellular miRNA profile of HM collected before and after feeding. HM cells conserved higher miRNA content than the lipid and skim HM fractions or other body fluids, in accordance with previous studies. In total, 1467 known mature and 1996 novel miRNAs were identified, with 89 high-confidence novel miRNAs. HM cell content was higher post-feeding (p < 0.05), and was positively associated with total miRNA content (p = 0.014) and species number (p < 0.001). This coincided with upregulation of 29 known and 2 novel miRNAs, and downregulation of 4 known and 1 novel miRNAs post-feeding, but no statistically significant change in expression was found for the remaining miRNAs. These findings suggest that feeding may influence the miRNA content of HM cells. The most highly and differentially expressed miRNAs were key regulators of milk components, with potential diagnostic value in lactation performance. They are also involved in the control of body fluid balance, thirst, appetite, immune response, and development, implicating their functional significance for the infant. PMID:27322254

AKT inhibition mitigates GRP78 (glucose-regulated protein) expression and contribution to chemoresistance in endometrial cancers.

PubMed

Gray, Michael J; Mhawech-Fauceglia, Paulette; Yoo, Eunjeong; Yang, Wangrong; Wu, Eijean; Lee, Amy S; Lin, Yvonne G

2013-07-01

Overexpression of the unfolded protein response master regulator GRP78 is associated with poor prognosis and therapeutic resistance in numerous human cancers, yet its role in endometrial cancers (EC) is undefined. To better understand the contribution of GRP78 to EC, we examined its expression levels in EC patient samples and EC cell lines. We demonstrate that GRP78 overexpression occurs more frequently in EC tissues compared with that found in normal endometrium, and that GRP78 expression occurs in most EC cell lines examined. Functional analysis demonstrated that GRP78 is inducible by cisplatin in EC cells, and siRNA knockdown of GRP78 augments chemotherapy-mediated cell death. Examination of AKT and GRP78 expression demonstrated that inhibition of AKT activity by MK2206 blocks GRP78 expression in EC cells. SiRNA studies also revealed that knockdown of GRP78 reduces but does not abrogate AKT activity, demonstrating that GRP78 is required for optimal AKT activity. In the presence of MK2206, siRNA knockdown of GRP78 does not augment AKT mediated survival in response to cisplatin treatment, suggesting that GRP78's antiapoptosis functions are part of the AKT survival pathway. Targeted therapies that reduce GRP78 expression or activity in cancers may serve to increase the effectiveness of current therapies for EC patients. Copyright © 2012 UICC.

Regulation of vacuolar H{sup +}-ATPase in microglia by RANKL

DOE Office of Scientific and Technical Information (OSTI.GOV)

Serrano, Eric M.; Ricofort, Ryan D.; Zuo, Jian

2009-11-06

Vacuolar H{sup +}-ATPases (V-ATPases) are large electrogenic proton pumps composed of numerous subunits that play vital housekeeping roles in the acidification of compartments of the endocytic pathway. Additionally, V-ATPases play specialized roles in certain cell types, a capacity that is linked to cell type selective expression of isoforms of some of the subunits. We detected low levels of the a3 isoform of the a-subunit in mouse brain extracts. Examination of various brain-derived cell types by immunoblotting showed a3 was expressed in the N9 microglia cell line and in primary microglia, but not in other cell types. The expression of a3more » in osteoclasts requires stimulation by Receptor Activator of Nuclear Factor {kappa}B-ligand (RANKL). We found that Receptor Activator of Nuclear Factor {kappa}B (RANK) was expressed by microglia. Stimulation of microglia with RANKL triggered increased expression of a3. V-ATPases in microglia were shown to bind microfilaments, and stimulation with RANKL increased the proportion of V-ATPase associated with the detergent-insoluble cytoskeletal fraction and with actin. In summary, microglia express the a3-subunit of V-ATPase. The expression of a3 and the interaction between V-ATPases and microfilaments was modulated by RANKL. These data suggest a novel molecular pathway for regulating microglia.« less

Evolution and development of the vertebrate ear

NASA Technical Reports Server (NTRS)

Fritzsch, B.; Beisel, K. W.

2001-01-01

This review outlines major aspects of development and evolution of the ear, specifically addressing issues of cell fate commitment and the emerging molecular governance of these decisions. Available data support the notion of homology of subsets of mechanosensors across phyla (proprioreceptive mechanosensory neurons in insects, hair cells in vertebrates). It is argued that this conservation is primarily related to the specific transducing environment needed to achieve mechanosensation. Achieving this requires highly conserved transcription factors that regulate the expression of the relevant structural genes for mechanosensory transduction. While conserved at the level of some cell fate assignment genes (atonal and its mammalian homologue), the ear has also radically reorganized its development by implementing genes used for cell fate assignment in other parts of the developing nervous systems (e.g., neurogenin 1) and by evolving novel sets of genes specifically associated with the novel formation of sensory neurons that contact hair cells (neurotrophins and their receptors). Numerous genes have been identified that regulate morphogenesis, but there is only one common feature that emerges at the moment: the ear appears to have co-opted genes from a large variety of other parts of the developing body (forebrain, limbs, kidneys) and establishes, in combination with existing transcription factors, an environment in which those genes govern novel, ear-related morphogenetic aspects. The ear thus represents a unique mix of highly conserved developmental elements combined with co-opted and newly evolved developmental elements.

The skeletal cell-derived molecule sclerostin drives bone marrow adipogenesis.

PubMed

Fairfield, Heather; Falank, Carolyne; Harris, Elizabeth; Demambro, Victoria; McDonald, Michelle; Pettitt, Jessica A; Mohanty, Sindhu T; Croucher, Peter; Kramer, Ina; Kneissel, Michaela; Rosen, Clifford J; Reagan, Michaela R

2018-02-01

The bone marrow niche is a dynamic and complex microenvironment that can both regulate, and be regulated by the bone matrix. Within the bone marrow (BM), mesenchymal stromal cell (MSC) precursors reside in a multi-potent state and retain the capacity to differentiate down osteoblastic, adipogenic, or chondrogenic lineages in response to numerous biochemical cues. These signals can be altered in various pathological states including, but not limited to, osteoporotic-induced fracture, systemic adiposity, and the presence of bone-homing cancers. Herein we provide evidence that signals from the bone matrix (osteocytes) determine marrow adiposity by regulating adipogenesis in the bone marrow. Specifically, we found that physiologically relevant levels of Sclerostin (SOST), which is a Wnt-inhibitory molecule secreted from bone matrix-embedded osteocytes, can induce adipogenesis in 3T3-L1 cells, mouse ear- and BM-derived MSCs, and human BM-derived MSCs. We demonstrate that the mechanism of SOST induction of adipogenesis is through inhibition of Wnt signaling in pre-adipocytes. We also demonstrate that a decrease of sclerostin in vivo, via both genetic and pharmaceutical methods, significantly decreases bone marrow adipose tissue (BMAT) formation. Overall, this work demonstrates a direct role for SOST in regulating fate determination of BM-adipocyte progenitors. This provides a novel mechanism for which BMAT is governed by the local bone microenvironment, which may prove relevant in the pathogenesis of certain diseases involving marrow adipose. Importantly, with anti-sclerostin therapy at the forefront of osteoporosis treatment and a greater recognition of the role of BMAT in disease, these data are likely to have important clinical implications. © 2017 Wiley Periodicals, Inc.

Prostate cancer epigenetics and its clinical implications

PubMed Central

Yegnasubramanian, Srinivasan

2016-01-01

Normal cells have a level of epigenetic programming that is superimposed on the genetic code to establish and maintain their cell identity and phenotypes. This epigenetic programming can be thought as the architecture, a sort of cityscape, that is built upon the underlying genetic landscape. The epigenetic programming is encoded by a complex set of chemical marks on DNA, on histone proteins in nucleosomes, and by numerous context-specific DNA, RNA, protein interactions that all regulate the structure, organization, and function of the genome in a given cell. It is becoming increasingly evident that abnormalities in both the genetic landscape and epigenetic cityscape can cooperate to drive carcinogenesis and disease progression. Large-scale cancer genome sequencing studies have revealed that mutations in genes encoding the enzymatic machinery for shaping the epigenetic cityscape are among the most common mutations observed in human cancers, including prostate cancer. Interestingly, although the constellation of genetic mutations in a given cancer can be quite heterogeneous from person to person, there are numerous epigenetic alterations that appear to be highly recurrent, and nearly universal in a given cancer type, including in prostate cancer. The highly recurrent nature of these alterations can be exploited for development of biomarkers for cancer detection and risk stratification and as targets for therapeutic intervention. Here, we explore the basic principles of epigenetic processes in normal cells and prostate cancer cells and discuss the potential clinical implications with regards to prostate cancer biomarker development and therapy. PMID:27212125

Prostate cancer epigenetics and its clinical implications.

PubMed

Yegnasubramanian, Srinivasan

2016-01-01

Normal cells have a level of epigenetic programming that is superimposed on the genetic code to establish and maintain their cell identity and phenotypes. This epigenetic programming can be thought as the architecture, a sort of cityscape, that is built upon the underlying genetic landscape. The epigenetic programming is encoded by a complex set of chemical marks on DNA, on histone proteins in nucleosomes, and by numerous context-specific DNA, RNA, protein interactions that all regulate the structure, organization, and function of the genome in a given cell. It is becoming increasingly evident that abnormalities in both the genetic landscape and epigenetic cityscape can cooperate to drive carcinogenesis and disease progression. Large-scale cancer genome sequencing studies have revealed that mutations in genes encoding the enzymatic machinery for shaping the epigenetic cityscape are among the most common mutations observed in human cancers, including prostate cancer. Interestingly, although the constellation of genetic mutations in a given cancer can be quite heterogeneous from person to person, there are numerous epigenetic alterations that appear to be highly recurrent, and nearly universal in a given cancer type, including in prostate cancer. The highly recurrent nature of these alterations can be exploited for development of biomarkers for cancer detection and risk stratification and as targets for therapeutic intervention. Here, we explore the basic principles of epigenetic processes in normal cells and prostate cancer cells and discuss the potential clinical implications with regards to prostate cancer biomarker development and therapy.

Exploding a myth: the capsule dehiscence mechanism and the function of pseudostomata in Sphagnum.

PubMed

Duckett, Jeffrey G; Pressel, Silvia; P'ng, Ken M Y; Renzaglia, Karen S

2009-01-01

The nineteenth century air-gun explanation for explosive spore discharge in Sphagnum has never been tested experimentally. Similarly, the function of the numerous stomata ubiquitous in the capsule walls has never been investigated. Both intact and pricked Sphagnum capsules, that were allowed to dry out, all dehisced over an 8-12 h period during which time the stomatal guard cells gradually collapsed and their potassium content, measured by X-ray microanalysis in a cryoscanning electron microscope, gradually increased. By contrast, guard cell potassium fell in water-stressed Arabidopsis. The pricking experiments demonstrate that the air-gun notion for explosive spore discharge in Sphagnum is inaccurate; differential shrinkage of the capsule walls causes popping off the rigid operculum. The absence of evidence for a potassium-regulating mechanism in the stomatal guard cells and their gradual collapse before spore discharge indicates that their sole role is facilitation of sporophyte desiccation that ultimately leads to capsule dehiscence. Our novel functional data on Sphagnum, when considered in relation to bryophyte phylogeny, suggest the possibility that stomata first appeared in land plants as structures that facilitated sporophyte drying out before spore discharge and only subsequently acquired their role in the regulation of gaseous exchange.

Taurine activates delayed rectifier KV channels via a metabotropic pathway in retinal neurons

PubMed Central

Bulley, Simon; Liu, Yufei; Ripps, Harris; Shen, Wen

2013-01-01

Taurine is one of the most abundant amino acids in the retina, throughout the CNS, and in heart and muscle cells. In keeping with its broad tissue distribution, taurine serves as a modulator of numerous basic processes, such as enzyme activity, cell development, myocardial function and cytoprotection. Despite this multitude of functional roles, the precise mechanism underlying taurine's actions has not yet been identified. In this study we report findings that indicate a novel role for taurine in the regulation of voltage-gated delayed rectifier potassium (KV) channels in retinal neurons by means of a metabotropic receptor pathway. The metabotropic taurine response was insensitive to the Cl− channel blockers, picrotoxin and strychnine, but it was inhibited by a specific serotonin 5-HT2A receptor antagonist, MDL11939. Moreover, we found that taurine enhanced KV channels via intracellular protein kinase C-mediated pathways. When 5-HT2A receptors were expressed in human embryonic kidney cells, taurine and AL34662, a non-specific 5-HT2 receptor activator, produced a similar regulation of KIR channels. In sum, this study provides new evidence that taurine activates a serotonin system, apparently via 5-HT2A receptors and related intracellular pathways. PMID:23045337

Calcium and mitochondrial metabolism in ceramide-induced cardiomyocyte death.

PubMed

Parra, Valentina; Moraga, Francisco; Kuzmicic, Jovan; López-Crisosto, Camila; Troncoso, Rodrigo; Torrealba, Natalia; Criollo, Alfredo; Díaz-Elizondo, Jessica; Rothermel, Beverly A; Quest, Andrew F G; Lavandero, Sergio

2013-08-01

Ceramides are important intermediates in the biosynthesis and degradation of sphingolipids that regulate numerous cellular processes, including cell cycle progression, cell growth, differentiation and death. In cardiomyocytes, ceramides induce apoptosis by decreasing mitochondrial membrane potential and promoting cytochrome-c release. Ca(2+) overload is a common feature of all types of cell death. The aim of this study was to determine the effect of ceramides on cytoplasmic Ca(2+) levels, mitochondrial function and cardiomyocyte death. Our data show that C2-ceramide induces apoptosis and necrosis in cultured cardiomyocytes by a mechanism involving increased Ca(2+) influx, mitochondrial network fragmentation and loss of the mitochondrial Ca(2+) buffer capacity. These biochemical events increase cytosolic Ca(2+) levels and trigger cardiomyocyte death via the activation of calpains. Copyright © 2013 Elsevier B.V. All rights reserved.

Differential KrasV12 protein levels control a switch regulating lung cancer cell morphology and motility

PubMed Central

Schäfer, C.; Mohan, A.; Burford, W.; Driscoll, M. K.; Ludlow, A. T.; Wright, W. E.; Shay, J. W.; Danuser, G.

2016-01-01

Introduction Oncogenic Kras mutations are important drivers of lung cancer development and metastasis. They are known to activate numerous cellular signaling pathways implicated in enhanced proliferation, survival, tumorigenicity and motility during malignant progression. Objectives Most previous studies of Kras in cancer have focused on the comparison of cell states in the absence or presence of oncogenic Kras mutations. Here we show that differential expression of the constitutively active mutation KrasV12 has profound effects on cell morphology and motility that drive metastatic processes. Methods The study relies on lung cancer cell transformation models, patient-derived lung cancer cell lines, and human lung tumor sections combined with molecular biology techniques, live-cell imaging and staining methods. Results Our analysis shows two cell functional states driven by KrasV12 protein levels: a non-motile state associated with high KrasV12 levels and tumorigenicity, and a motile state associated with low KrasV12 levels and cell dissemination. Conversion between the states is conferred by differential activation of a mechano-sensitive double-negative feedback between KrasV12/ERK/Myosin II and matrix-adhesion signaling. KrasV12 expression levels change upon cues such as hypoxia and integrin-mediated cell-matrix adhesion, rendering KrasV12 levels an integrator of micro-environmental signals that translate into cellular function. By live cell imaging of tumor models we observe shedding of mixed high and low KrasV12 expressers forming multi-functional collectives with potentially optimal metastatic properties composed of a highly mobile and a highly tumorigenic unit. Discussion Together these data highlight previously unappreciated roles for the quantitative effects of expression level variation of oncogenic signaling molecules in conferring fundamental alterations in cell function regulation required for cancer progression. PMID:29057096

Feedback regulation in a stem cell model with acute myeloid leukaemia.

PubMed

Jiao, Jianfeng; Luo, Min; Wang, Ruiqi

2018-04-24

The haematopoietic lineages with leukaemia lineages are considered in this paper. In particular, we mainly consider that haematopoietic lineages are tightly controlled by negative feedback inhibition of end-product. Actually, leukemia has been found 100 years ago. Up to now, the exact mechanism is still unknown, and many factors are thought to be associated with the pathogenesis of leukemia. Nevertheless, it is very necessary to continue the profound study of the pathogenesis of leukemia. Here, we propose a new mathematical model which include some negative feedback inhibition from the terminally differentiated cells of haematopoietic lineages to the haematopoietic stem cells and haematopoietic progenitor cells in order to describe the regulatory mechanisms mentioned above by a set of ordinary differential equations. Afterwards, we carried out detailed dynamical bifurcation analysis of the model, and obtained some meaningful results. In this work, we mainly perform the analysis of the mathematic model by bifurcation theory and numerical simulations. We have not only incorporated some new negative feedback mechanisms to the existing model, but also constructed our own model by using the modeling method of stem cell theory with probability method. Through a series of qualitative analysis and numerical simulations, we obtain that the weak negative feedback for differentiation probability is conducive to the cure of leukemia. However, with the strengthening of negative feedback, leukemia will be more difficult to be cured, and even induce death. In contrast, strong negative feedback for differentiation rate of progenitor cells can promote healthy haematopoiesis and suppress leukaemia. These results demonstrate that healthy progenitor cells are bestowed a competitive advantage over leukaemia stem cells. Weak g 1 , g 2 , and h 1 enable the system stays in the healthy state. However, strong h 2 can promote healthy haematopoiesis and suppress leukaemia.

Effects of Tobacco Smoking on the Degeneration of the Intervertebral Disc: A Finite Element Study

PubMed Central

Elmasry, Shady; Asfour, Shihab; de Rivero Vaccari, Juan Pablo; Travascio, Francesco

2015-01-01

Tobacco smoking is associated with numerous pathological conditions. Compelling experimental evidence associates smoking to the degeneration of the intervertebral disc (IVD). In particular, it has been shown that nicotine down-regulates both the proliferation rate and glycosaminoglycan (GAG) biosynthesis of disc cells. Moreover, tobacco smoking causes the constriction of the vascular network surrounding the IVD, thus reducing the exchange of nutrients and anabolic agents from the blood vessels to the disc. It has been hypothesized that both nicotine presence in the IVD and the reduced solute exchange are responsible for the degeneration of the disc due to tobacco smoking, but their effects on tissue homeostasis have never been quantified. In this study, a previously presented computational model describing the homeostasis of the IVD was deployed to investigate the effects of impaired solute supply and nicotine-mediated down-regulation of cell proliferation and biosynthetic activity on the health of the disc. We found that the nicotine-mediated down-regulation of cell anabolism mostly affected the GAG concentration at the cartilage endplate, reducing it up to 65% of the value attained in normal physiological conditions. In contrast, the reduction of solutes exchange between blood vessels and disc tissue mostly affected the nucleus pulposus, whose cell density and GAG levels were reduced up to 50% of their normal physiological levels. The effectiveness of quitting smoking on the regeneration of a degenerated IVD was also investigated, and showed to have limited benefit on the health of the disc. A cell-based therapy in conjunction with smoke cessation provided significant improvements in disc health, suggesting that, besides quitting smoking, additional treatments should be implemented in the attempt to recover the health of an IVD degenerated by tobacco smoking. PMID:26301590

Effects of histone acetylation and DNA methylation on p21( WAF1) regulation.

PubMed

Fang, Jing-Yuan; Lu, You-Yong

2002-06-01

Cell cycle progression is regulated by interactions between cyclins and cyclin-dependent kinases (CDKs). p21(WAF1) is one of the CIP/KIP family which inhibits CDKs activity. Increased expression of p21(WAF1) may play an important role in the growth arrest induced in transformed cells. Although the stability of the p21( WAF1) mRNA could be altered by different signals, cell differentiation and numerous influencing factors. However, recent studies suggest that two known mechanisms of epigenesis, i.e.gene inactivation by methylation in promoter region and changes to an inactive chromatin by histone deacetylation, seem to be the best candidate mechanisms for inactivation of p21( WAF1). To date, almost no coding region p21(WAF1) mutations have been found in tumor cells, despite extensive screening of hundreds of various tumors. Hypermethylation of the p21(WAF1) promoter region may represent an alternative mechanism by which the p21(WAF1/CIP1) gene can be inactivated. The reduction of cellular DNMT protein levels also induces a corresponding rapid increase in the cell cycle regulator p21(WAF1) protein demonstrating a regulatory link between DNMT and p21(WAF1) which is independent of methylation of DNA. Both histone hyperacetylation and hypoacetylation appear to be important in the carcinoma process, and induction of the p21(WAF1) gene by histone hyperacetylation may be a mechanism by which dietary fiber prevents carcinogenesis. Here, we review the influence of histone acetylation and DNA methylation on p21(WAF1) transcription, and affection of pathways or factors associated such as p 53, E2A, Sp1 as well as several histone deacetylation inhibitors.

TOR signaling pathway and autophagy are involved in the regulation of circadian rhythms in behavior and plasticity of L2 interneurons in the brain of Drosophila melanogaster.

PubMed

Kijak, Ewelina; Pyza, Elżbieta

2017-01-01

Drosophila melanogaster is a common model used to study circadian rhythms in behavior and circadian clocks. However, numerous circadian rhythms have also been detected in non-clock neurons, especially in the first optic neuropil (lamina) of the fly's visual system. Such rhythms have been observed in the number of synapses and in the structure of interneurons, which exhibit changes in size and shape in a circadian manner. Although the patterns of these changes are known, the mechanism remains unclear. In the present study, we investigated the role of the TOR signaling pathway and autophagy in regulating circadian rhythms based on the behavior and structural plasticity of the lamina L2 monopolar cell dendritic trees. In addition, we examined the cyclic expression of the TOR signaling pathway (Tor, Pi3K class 1, Akt1) and autophagy (Atg5 and Atg7) genes in the fly's brain. We observed that Tor, Atg5 and Atg7 exhibit rhythmic expressions in the brain of wild-type flies in day/night conditions (LD 12:12) that are abolished in per01 clock mutants. The silencing of Tor in per expressing cells shortens a period of the locomotor activity rhythm of flies. In addition, silencing of the Tor and Atg5 genes in L2 cells disrupts the circadian plasticity of the L2 cell dendritic trees measured in the distal lamina. In turn, silencing of the Atg7 gene in L2 cells changes the pattern of this rhythm. Our results indicate that the TOR signaling pathway and autophagy are involved in the regulation of circadian rhythms in the behavior and plasticity of neurons in the brain of adult flies.

Characterisation and expression of myogenesis regulatory factors during in vitro myoblast development and in vivo fasting in the gilthead sea bream (Sparus aurata).

PubMed

García de la serrana, Daniel; Codina, Marta; Capilla, Encarnación; Jiménez-Amilburu, Vanesa; Navarro, Isabel; Du, Shao-Jun; Johnston, Ian A; Gutiérrez, Joaquim

2014-01-01

The aim of this study was to characterise a primary cell culture isolated from fast skeletal muscle of the gilthead sea bream. Gene expression profiles during culture maturation were compared with those obtained from a fasting-refeeding model which is widely used to modulate myogenesis in vivo. Myogenesis is controlled by numerous extracellular signals together with intracellular transcriptional factors whose coordinated expression is critical for the appropriate development of muscle fibres. Full-length cDNAs for the transcription factors Myf5, Mrf4, Pax7 and Sox8 were cloned and sequenced for gilthead sea bream. Pax7, sox8, myod2 and myf5 levels were up-regulated during the proliferating phase of the myogenic cultures coincident with the highest expression of proliferating cell nuclear antigen (PCNA). In contrast, myogenin and mrf4 transcript abundance was highest during the differentiation phase of the culture when myotubes were present, and was correlated with increased myosin heavy chain (mhc) and desmin expression. In vivo, 30days of fasting resulted in muscle fibre atrophy, a reduction in myod2, myf5 and igf1 expression, lower number of Myod-positive cells, and decreased PCNA protein expression, whereas myogenin expression was not significantly affected. Myostatin1 (mstn1) and pax7 expression were up-regulated in fasted relative to well-fed individuals, consistent with a role for Pax7 in the reduction of myogenic cell activity with fasting. The primary cell cultures and fasting-feeding experiments described provide a foundation for the future investigations on the regulation of muscle growth in gilthead sea bream. © 2013.

TOR signaling pathway and autophagy are involved in the regulation of circadian rhythms in behavior and plasticity of L2 interneurons in the brain of Drosophila melanogaster

PubMed Central

Kijak, Ewelina; Pyza, Elżbieta

2017-01-01

Drosophila melanogaster is a common model used to study circadian rhythms in behavior and circadian clocks. However, numerous circadian rhythms have also been detected in non-clock neurons, especially in the first optic neuropil (lamina) of the fly’s visual system. Such rhythms have been observed in the number of synapses and in the structure of interneurons, which exhibit changes in size and shape in a circadian manner. Although the patterns of these changes are known, the mechanism remains unclear. In the present study, we investigated the role of the TOR signaling pathway and autophagy in regulating circadian rhythms based on the behavior and structural plasticity of the lamina L2 monopolar cell dendritic trees. In addition, we examined the cyclic expression of the TOR signaling pathway (Tor, Pi3K class 1, Akt1) and autophagy (Atg5 and Atg7) genes in the fly’s brain. We observed that Tor, Atg5 and Atg7 exhibit rhythmic expressions in the brain of wild-type flies in day/night conditions (LD 12:12) that are abolished in per01 clock mutants. The silencing of Tor in per expressing cells shortens a period of the locomotor activity rhythm of flies. In addition, silencing of the Tor and Atg5 genes in L2 cells disrupts the circadian plasticity of the L2 cell dendritic trees measured in the distal lamina. In turn, silencing of the Atg7 gene in L2 cells changes the pattern of this rhythm. Our results indicate that the TOR signaling pathway and autophagy are involved in the regulation of circadian rhythms in the behavior and plasticity of neurons in the brain of adult flies. PMID:28196106

SIRT1 inhibition restores apoptotic sensitivity in p53-mutated human keratinocytes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Herbert, Katharine J.; Cook, Anthony L., E-mail: Anthony.Cook@utas.edu.au; Snow, Elizabeth T., E-mail: elizabeth.snow@utas.edu.au

2014-06-15

Mutations to the p53 gene are common in UV-exposed keratinocytes and contribute to apoptotic resistance in skin cancer. P53-dependent activity is modulated, in part, by a complex, self-limiting feedback loop imposed by miR-34a-mediated regulation of the lysine deacetylase, SIRT1. Expression of numerous microRNAs is dysregulated in squamous and basal cell carcinomas; however the contribution of specific microRNAs to the pathogenesis of skin cancer remains untested. Through use of RNAi, miRNA target site blocking oligonucleotides and small molecule inhibitors, this study explored the influence of p53 mutational status, SIRT1 activity and miR-34a levels on apoptotic sensitivity in primary (NHEK) and p53-mutatedmore » (HaCaT) keratinocyte cell lines. SIRT1 and p53 are overexpressed in p53-mutated keratinocytes, whilst miR-34a levels are 90% less in HaCaT cells. HaCaTs have impaired responses to p53/SIRT1/miR-34a axis manipulation which enhanced survival during exposure to the chemotherapeutic agent, camptothecin. Inhibition of SIRT1 activity in this cell line increased p53 acetylation and doubled camptothecin-induced cell death. Our results demonstrate that p53 mutations increase apoptotic resistance in keratinocytes by interfering with miR-34a-mediated regulation of SIRT1 expression. Thus, SIRT1 inhibitors may have a therapeutic potential for overcoming apoptotic resistance during skin cancer treatment. - Highlights: • Impaired microRNA biogenesis promotes apoptotic resistance in HaCaT keratinocytes. • TP53 mutations suppress miR-34a-mediated regulation of SIRT1 expression. • SIRT1 inhibition increases p53 acetylation in HaCaTs, restoring apoptosis.« less

Redox Signaling Mechanisms in Nervous System Development.

PubMed

Olguín-Albuerne, Mauricio; Morán, Julio

2018-06-20

Numerous studies have demonstrated the actions of reactive oxygen species (ROS) as regulators of several physiological processes. In this study, we discuss how redox signaling mechanisms operate to control different processes such as neuronal differentiation, oligodendrocyte differentiation, dendritic growth, and axonal growth. Recent Advances: Redox homeostasis regulates the physiology of neural stem cells (NSCs). Notably, the neuronal differentiation process of NSCs is determined by a change toward oxidative metabolism, increased levels of mitochondrial ROS, increased activity of NADPH oxidase (NOX) enzymes, decreased levels of Nrf2, and differential regulation of different redoxins. Furthermore, during the neuronal maturation processes, NOX and MICAL produce ROS to regulate cytoskeletal dynamics, which control the dendritic and axonal growth, as well as the axonal guidance. The redox homeostasis changes are, in part, attributed to cell metabolism and compartmentalized production of ROS, which is regulated, sensed, and transduced by different molecules such as thioredoxins, glutaredoxins, peroxiredoxins, and nucleoredoxin to control different signaling pathways in different subcellular regions. The study of how these elements cooperatively act is essential for the understanding of nervous system development, as well as the application of regenerative therapies that recapitulate these processes. The information about these topics in the last two decades leads us to the conclusion that the role of ROS signaling in development of the nervous system is more important than it was previously believed and makes clear the importance of exploring in more detail the mechanisms of redox signaling. Antioxid. Redox Signal. 28, 1603-1625.

The Plasma Membrane Calcium Pump: New Ways to Look at an Old Enzyme

PubMed Central

Lopreiato, Raffaele; Giacomello, Marta; Carafoli, Ernesto

2014-01-01

The three-dimensional structure of the PMCA pump has not been solved, but its basic mechanistic properties are known to repeat those of the other Ca2+ pumps. However, the pump also has unique properties. They concern essentially its numerous regulatory mechanisms, the most important of which is the autoinhibition by its C-terminal tail. Other regulatory mechanisms involve protein kinases and the phospholipids of the membrane in which the pump is embedded. Permanent activation of the pump, e.g. by calmodulin, is physiologically as harmful to cells as its absence. The concept is now emerging that the global control of cell Ca2+ may not be the main function of the pump; in some cell types, it could even be irrelevant. The main pump role would be the regulation of Ca2+ in cell microdomains in which the pump co-segregates with partners that modulate the Ca2+ message and transduce it to important cell functions. PMID:24570005

Uncovering Novel Roles of Nonneuronal Cells in Body Weight Homeostasis and Obesity

PubMed Central

Argente, Jesús

2013-01-01

Glial cells, which constitute more than 50% of the mass of the central nervous system and greatly outnumber neurons, are at the vanguard of neuroendocrine research in metabolic control and obesity. Historically relegated to roles of structural support and protection, diverse functions have been gradually attributed to this heterogeneous class of cells with their protagonism in crescendo in all areas of neuroscience during the past decade. However, this dramatic increase in attention bestowed upon glial cells has also emphasized our vast lack of knowledge concerning many aspects of their physiological functions, let alone their participation in numerous pathologies. This minireview focuses on the recent advances in our understanding of how glial cells participate in the physiological regulation of appetite and systemic metabolism as well as their role in the pathophysiological response to poor nutrition and secondary complications associated with obesity. Moreover, we highlight some of the existing lagoons of knowledge in this increasingly important area of investigation. PMID:23798599

Modelling Spatially Regulated β-Catenin Dynamics and Invasion in Intestinal Crypts

PubMed Central

Murray, Philip J.; Kang, Jun-Won; Mirams, Gary R.; Shin, Sung-Young; Byrne, Helen M.; Maini, Philip K.; Cho, Kwang-Hyun

2010-01-01

Experimental data (e.g., genetic lineage and cell population studies) on intestinal crypts reveal that regulatory features of crypt behavior, such as control via morphogen gradients, are remarkably well conserved among numerous organisms (e.g., from mouse and rat to human) and throughout the different regions of the small and large intestines. In this article, we construct a partial differential equation model of a single colonic crypt that describes the spatial distribution of Wnt pathway proteins along the crypt axis. The novelty of our continuum model is that it is based upon assumptions that can be directly related to processes at the cellular and subcellular scales. We use the model to predict how the distributions of Wnt pathway proteins are affected by mutations. The model is then extended to investigate how mutant cell populations can invade neighboring crypts. The model simulations suggest that cell crowding caused by increased proliferation and decreased cell loss may be sufficient for a mutant cell population to colonize a neighboring healthy crypt. PMID:20682248

Modeling the formation of cell-matrix adhesions on a single 3D matrix fiber.

PubMed

Escribano, J; Sánchez, M T; García-Aznar, J M

2015-11-07

Cell-matrix adhesions are crucial in different biological processes like tissue morphogenesis, cell motility, and extracellular matrix remodeling. These interactions that link cell cytoskeleton and matrix fibers are built through protein clutches, generally known as adhesion complexes. The adhesion formation process has been deeply studied in two-dimensional (2D) cases; however, the knowledge is limited for three-dimensional (3D) cases. In this work, we simulate different local extracellular matrix properties in order to unravel the fundamental mechanisms that regulate the formation of cell-matrix adhesions in 3D. We aim to study the mechanical interaction of these biological structures through a three dimensional discrete approach, reproducing the transmission pattern force between the cytoskeleton and a single extracellular matrix fiber. This numerical model provides a discrete analysis of the proteins involved including spatial distribution, interaction between them, and study of the different phenomena, such as protein clutches unbinding or protein unfolding. Copyright © 2015 Elsevier Ltd. All rights reserved.

A Microfluidic-Based Multi-Shear Device for Investigating the Effects of Low Fluid-Induced Stresses on Osteoblasts

PubMed Central

Yu, Weiliang; Qu, Hong; Hu, Guoqing; Zhang, Qian; Song, Kui; Guan, Haijie; Liu, Tingjiao; Qin, Jianhua

2014-01-01

Interstitial fluid flow (IFF) within the extracellular matrix (ECM) produces low magnitude shear stresses on cells. Fluid flow-induced stress (FSS) plays an important role during tissue morphogenesis. To investigate the effect of low FSS generated by IFF on cells, we developed a microfluidic-based cell culture device that can generate multiple low shear stresses. By changing the length and width of the flow-in channels, different continuous low level shear stresses could be generated in individual cell culture chambers. Numerical calculations demonstrate uniform shear stress distributions of the major cell culture area of each chamber. This calculation is further confirmed by the wall shear stress curves. The effects of low FSS on MC3T3-E1 proliferation and differentiation were studied using this device. It was found that FSS ranging from 1.5 to 52.6 µPa promoted MC3T3-E1 proliferation and differentiation, but FSS over 412 µPa inhibited the proliferation and differentiation of MC3T3-E1 cells. FSS ranging from 1.5 to 52.6 µPa also increased the expression of Runx2, a key transcription factor regulating osteoblast differentiation. It is suggested that Runx2 might be an important regulator in low FSS-induced MC3T3-E1 differentiation. This device allows for detailed study of the effect of low FSS on the behaviors of cells; thus, it would be a useful tool for analysis of the effects of IFF-induced shear stresses on cells. PMID:24587156

Up-regulation of Paxillin and Focal Adhesion Signaling follows Dystroglycan Complex deletions and promotes a Hypertensive State of Differentiation

PubMed Central

Sen, Shamik; Tewari, Manorama; Zajac, Allison; Barton, Elisabeth; Sweeney, H. Lee; Discher, Dennis E.

2010-01-01

Anchorage to matrix is mediated for many cells not only by integrin-based focal adhesions but also by a parallel assembly of integral and peripheral membrane proteins known as the Dystroglycan Complex. Deficiencies in either dystrophin (mdx mice) or γ-sarcoglycan (γSG−/− mice) components of the Dystroglycan Complex lead to upregulation of numerous focal adhesion proteins, and the phosphoprotein paxillin proves to be among the most prominent. In mdx muscle, paxillin-Y31 and Y118 are both hyper-phosphorylated as are key sites in focal adhesion kinase (FAK) and the stretch-stimulatable pro-survival MAPK pathway, whereas γSG−/− muscle exhibits more erratic hyper-phosphorylation. In cultured myotubes, cell tension generated by myosin-II appears required for localization of paxillin to adhesions while vinculin appears more stably integrated. Over-expression of wild-type (WT) paxillin has no obvious effect on focal adhesion density or the physical strength of adhesion, but WT and a Y118F mutant promote contractile sarcomere formation whereas a Y31F mutant shows no effect, implicating Y31 in striation. Self-peeling of cells as well as Atomic Force Microscopy (AFM) probing of cells with or without myosin II inhibition indicate an increase in cell tension within paxillin-overexpressing cells. However, prednisolone, a first-line glucocorticoid for muscular dystrophies, decreases cell tension without affecting paxillin at adhesions, suggesting a non-linear relationship between paxillin and cell tension. Hypertension that results from upregulation of integrin adhesions is thus a natural and treatable outcome of dystroglycan complex down-regulation. PMID:20663583

Novel strategies to enforce an epithelial phenotype in mesenchymal cells

PubMed Central

Dragoi, Ana-Maria; Swiss, Rachel; Gao, Beile; Agaisse, Hervé

2014-01-01

E-cadherin downregulation in cancer cells is associated with epithelial-to-mesenchymal transition (EMT) and metastatic prowess, but the underlying mechanisms are incompletely characterized. In this study, we probed E-cadherin expression at the plasma membrane as a functional assay to identify genes involved in E-cadherin downregulation. The assay was based on the E-cadherin-dependent invasion properties of the intracellular pathogen Listeria monocytogenes. On the basis of a functional readout, automated microscopy and computer-assisted image analysis were used to screen siRNAs targeting 7,000 human genes. The validity of the screen was supported by its definion of several known regulators of E-cadherin expression, including ZEB1, HDAC1 and MMP14. We identified three new regulators (FLASH, CASP7 and PCGF1), the silencing of which was sufficient to restore high levels of E-cadherin transcription. Additionally, we identified two new regulators (FBXL5 and CAV2), the silencing of which was sufficient to increase E-cadherin expression at a post-transcriptional level. FLASH silencing regulated the expression of E-cadherin and other ZEB1-dependent genes, through post-transcriptional regulation of ZEB1, but it also regulated the expression of numerous ZEB1-independent genes with functions predicted to contribute to a restoration of the epithelial phenotype. Finally, we also report the identification of siRNA duplexes that potently restored the epithelial phenotype by mimicking the activity of known and putative microRNAs. Our findings suggest new ways to enforce epithelial phenotypes as a general strategy to treat cancer by blocking invasive and metastatic phenotypes associated with EMT. PMID:24845104

Transgenesis of the Wolffian duct visualizes dynamic behavior of cells undergoing tubulogenesis in vivo.

PubMed

Atsuta, Yuji; Tadokoro, Ryosuke; Saito, Daisuke; Takahashi, Yoshiko

2013-05-01

Deciphering how the tubulogenesis is regulated is an essential but unsolved issue in developmental biology. Here, using Wolffian duct (WD) formation in chicken embryos, we have developed a novel method that enables gene manipulation during tubulogenesis in vivo. Exploiting that WD arises from a defined site located anteriorly in the embryo (pronephric region), we targeted this region with the enhanced green fluorescent protein (EGFP) gene by the in ovo electroporation technique. EGFP-positive signals were detected in a wide area of elongating WD, where transgenic cells formed an epithelial component in a mosaic manner. Time-lapse live imaging analyses further revealed dynamic behavior of cells during WD elongation: some cells possessed numerous filopodia, and others exhibited cellular tails that repeated elongation and retraction. The retraction of the tail was precisely regulated by Rho activity via actin dynamics. When electroporated with the C3 gene, encoding Rho inhibitor, WD cells failed to contract their tails, resulting in an aberrantly elongated process. We further combined with the Tol2 transposon-mediated gene transfer technique, and could trace EGFP-positive cells at later stages in the ureteric bud sprouting from WD. This is the first demonstration that exogenous gene(s) can directly be introduced into elongating tubular structures in living amniote embryos. This method has opened a way to investigate how a complex tubulogenesis proceeds in higher vertebrates. © 2013 The Authors Development, Growth & Differentiation © 2013 Japanese Society of Developmental Biologists.

Tetracycline-regulated expression of OLIG2 gene in human dental pulp stem cells lead to mouse sciatic nerve regeneration upon transplantation.

PubMed

Askari, N; Yaghoobi, M M; Shamsara, M; Esmaeili-Mahani, S

2015-10-01

Numerous studies have indicated dental pulp stem cells (DPSCs) potency to differentiate into several types of cell lineages. Oligodendrocyte lineage transcription factor 2 (OLIG2) plays an important role in the oligodendrogenic pathway. In this study, a tetracycline (Tet)-inducible system expressing OLIG2 gene was transfected into human DPSCs to direct their differentiation toward oligodendrocyte progenitor cells (OPCs). Following induction, the expression of stage-specific markers was studied by Reverse Transcription quantitative Polymerase Chain Reaction (RT-qPCR), immunocytochemistry and western blotting. In the following, the cells were transplanted into the mouse model of local sciatic demyelination damage by lysolecithin. Recovery of lysolecithin-induced lesions in sciatic nerve was studied by treadmill exercise, von Frey filament test and hind paw withdrawal in response to a thermal stimulus. Improvement of behavioral symptoms was efficiently observed from the second week to the sixth week post-transplantation. Our findings showed that exogenous expression of the OLIG2 gene by a Tet-regulated system could be used as an efficient way to induce the differentiation of DPSCs into functional oligodendrocytes. Meanwhile, the DPSC-derived OPCs have relevant therapeutic potential in the animal model of sciatic nerve injury and therefore might represent a valuable tool for stem cell-based therapy in inflammatory and degenerative diseases of the peripheral and central nervous systems (CNSs). Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.

Transcriptome Profiling of Caco-2 Cancer Cell Line following Treatment with Extracts from Iodine-Biofortified Lettuce (Lactuca sativa L.)

PubMed Central

Koronowicz, Aneta A.; Kopeć, Aneta; Master, Adam; Smoleń, Sylwester; Piątkowska, Ewa; Bieżanowska-Kopeć, Renata; Ledwożyw-Smoleń, Iwona; Skoczylas, Łukasz; Rakoczy, Roksana; Leszczyńska, Teresa; Kapusta-Duch, Joanna; Pysz, Mirosław

2016-01-01

Although iodization of salt is the most common method used to obtain iodine-enriched food, iodine deficiency disorders are still a global health problem and profoundly affect the quality of human life. Iodine is required for the synthesis of thyroid hormones, which are crucial regulators of human metabolism, cell growth, proliferation, apoptosis and have been reported to be involved in carcinogenesis. In this study, for the first time, we evaluated the effect of iodine-biofortified lettuce on transcriptomic profile of Caco-2 cancer cell line by applying the Whole Human Genome Microarray assay. We showed 1326 differentially expressed Caco-2 transcripts after treatment with iodine-biofortified (BFL) and non-fortified (NFL) lettuce extracts. We analysed pathways, molecular functions, biological processes and protein classes based on comparison between BFL and NFL specific genes. Iodine, which was expected to act as a free ion (KI-NFL) or at least in part to be incorporated into lettuce macromolecules (BFL), differently regulated pathways of numerous transcription factors leading to different cellular effects. In this study we showed the inhibition of Caco-2 cells proliferation after treatment with BFL, but not potassium iodide (KI), and BFL-mediated induction of mitochondrial apoptosis and/or cell differentiation. Our results showed that iodine-biofortified plants can be effectively used by cells as an alternative source of this trace element. Moreover, the observed differences in action of both iodine sources may suggest a potential of BFL in cancer treatment. PMID:26799209

Megakaryocyte and polyploidization.

PubMed

Mazzi, Stefania; Lordier, Larissa; Debili, Najet; Raslova, Hana; Vainchenker, William

2018-01-01

In mammals, platelets are produced in the blood by cytoplasmic fragmentation of megakaryocytes (MKs). Platelet production is thus dependent on both the MK number and size. During differentiation, MKs switch from a division by mitosis to polyploidization by endomitosis to increase their size. The endomitotic process includes several successive rounds of DNA replication with an entry in mitosis with a failure in late cytokinesis and a defect in karyokinesis. This leads to a giant cell with a modal ploidy at 16N and one multilobulated nucleus. The entire genome is duplicated several times and all alleles remain functional producing a hypermetabolic cell. A defect in abscission explains the cytokinesis failure and is related to an altered accumulation of actomyosin at the cleavage furrow as a consequence of both a low local RhoA activity and silencing of the MYH10 gene. This mechanism is regulated by transcription factors that govern differentiation explaining the intricacies of both processes. However, the endomitotic cell cycle regulation is still incompletely understood, particularly mitosis entry, escape to the tetraploid checkpoint, and defect in karyokinesis. Polyploidization is regulated during ontogeny, the first embryonic MKs being 2N. The molecular mechanism of this embryo-fetal/adult transition is beginning to be understood. In physiological conditions, MK ploidy is increased by an enhanced platelet demand through the thrombopoietin/myeloproliferative leukemia axis. In numerous hematologic malignancies, MK ploidy decreases, but it is always associated with a defect in MK differentiation. It has been proposed that polyploidization induction could be a treatment for some malignant MK disorders. Copyright © 2018 ISEH – Society for Hematology and Stem Cells. Published by Elsevier Inc. All rights reserved.

TMEM16A regulates portal vein smooth muscle cell proliferation in portal hypertension.

PubMed

Zeng, Xi; Huang, Ping; Chen, Mingkai; Liu, Shiqian; Wu, Nannan; Wang, Fang; Zhang, Jing

2018-01-01

The aim of the present study was to elucidate the effect of transmembrane protein 16A (TMEM16A) on portal vein smooth muscle cell (PVSMC) proliferation associated with portal vein remodeling in portal hypertension (PHT). Sprague-Dawley rats were subjected to bile duct ligation to establish a rat model of liver cirrhosis and PHT. Sham-operated animals served as controls. At 8 weeks after bile duct ligation, the extent of liver fibrosis and the portal vein wall thickness were assessed using hematoxylin-eosin staining. The protein expression levels of TMEM16A, extracellular signal-regulated kinase 1 and 2 (ERK1/2) and phosphorylated ERK1/2 (p-ERK1/2) in the portal vein were detected by immunohistochemistry and western blotting. In vitro , the lentivirus vectors were constructed and transfected into PVSMCs to upregulate the expression of TMEM16A. Isolated rat primary PVSMCs were treated with a small molecule inhibitor of TMEM16A, T16A-inhA01. Cell cycle was detected by flow cytometry. The activity of TMEM16A in the portal vein isolated from bile duct ligated rats was decreased, while the expression level of p-ERK1/2 was increased. However, in vitro , upregulation of TMEM16A promoted the proliferation PVSMCs, while inhibition of TMEM16A channels inhibited the proliferation of PVSMCs. The results indicated that TMEM16A contributes to PVSMCs proliferation in vitro , but in vivo , it may be a negative regulator of cell proliferation influenced by numerous factors.

Germ cell control of testin production is inverse to that of other Sertoli cell products.

PubMed

Jégou, B; Pineau, C; Velez de la Calle, J F; Touzalin, A M; Bardin, C W; Cheng, C Y

1993-06-01

Recent studies have shown that germ cells can regulate testins, two newly identified Sertoli cell proteins that are associated with junctional complexes. To investigate this possibility, several parameters of Sertoli cell function were investigated over 2-120 days post exposure of the rat testes to x-rays (3 Grays). The irradiation-induced loss of spermatogonia resulted in a maturation-depletion process progressively affecting all germ cell classes. Testis weight began to decrease when the most numerous germ cell type (spermatids) began to decline. A complete or near complete recovery of spermatogenesis and of the testis weight had occurred by day 120 post irradiation. There was no significant change in FSH, epididymal androgen-binding protein, and tubule fluid levels during the first weeks after irradiation, when the seminiferious epithelium was depleted of spermatogonia and germ cells up to early spermatids. In contrast, when the number of the more mature forms of spermatids declined (between day 21 and 54), FSH rose and androgen-binding protein as well as fluid production declined. The subsequent recovery of these parameters was also highly correlated with the number of late spermatids. By contrast, testicular testin contents reacted to the depletion of germ cells with a biphasic increase; a doubling occurred when spermatogonia, spermatocytes, and early spermatids were absent (days 4-28), and a 7-fold rise occurred by day 37 when the number of late spermatids had decreased by 50%. By day 54, when the sperm counts had reached a nadir, testin contents had returned to levels corresponding to about four times the control levels; they progressively recovered thereafter. These observations support the postulate that germ cells negatively regulate testins. This possibility was investigated with in vitro experiments showing that addition of germ cell-conditioned medium to Sertoli cell monolayers inhibited testin secretion in a dose-dependent manner. In conclusion this study; 1) highlights the complex interplay between the various germ cell classes in the control of the Sertoli cell function in the adult testis; 2) establishes that germ cell effects may be opposite on different Sertoli cell products; 3) demonstrates that several classes of germ cells negatively control testicular testin contents; and 4) emphasizes the particular role of late spermatids in Sertoli cell regulation.

Extracellular vesicles released by mesenchymal-like prostate carcinoma cells modulate EMT state of recipient epithelial-like carcinoma cells through regulation of AR signaling.

PubMed

El-Sayed, Ihsan Y; Daher, Ahmad; Destouches, Damien; Firlej, Virginie; Kostallari, Enis; Maillé, Pascale; Huet, Eric; Haidar-Ahmad, Nathaline; Jenster, Guido; de la Taille, Alexandre; Abou Merhi, Raghida; Terry, Stéphane; Vacherot, Francis

2017-12-01

Extracellular vesicles released from cancer cells may play an important role in cancer progression by shuttling oncogenic information into recipient cells. However, our knowledge is still fragmentary and there remain numerous questions regarding the mechanisms at play and the functional consequences of these interactions. We have recently established a mesenchymal-like prostate cancer cell line (22Rv1/CR-1; Mes-PCa). In this study, we assessed the effects of the extracellular vesicles released by these cells on recipient androgen-dependent epithelial VCaP prostate cancer cells. Mes-PCa derived vesicles were found to promote mesenchymal features in the recipient epithelial-like prostate cancer cells. This transformation was accompanied by a modulation of androgen receptor signaling and activation of TGFβ signaling pathway. Moreover, recipient cells acquiring mesenchymal traits displayed enhanced migratory and invasive features as well as increased resistance to the androgen receptor antagonist, enzalutamide. Our results suggest a previously unappreciated role for Mes-PCa secreted vesicles in cancer promotion by transferring cell-mediated signals and promoting phenotypic changes in recipient prostate cancer cells. Copyright © 2017 Elsevier B.V. All rights reserved.

Implications for bidirectional signaling between afferent nerves and urothelial cells-ICI-RS 2014.

PubMed

Kanai, Anthony; Fry, Christopher; Ikeda, Youko; Kullmann, Florenta Aura; Parsons, Brian; Birder, Lori

2016-02-01

To present a synopsis of the presentations and discussions from Think Tank I, "Implications for afferent-urothelial bidirectional communication" of the 2014 International Consultation on Incontinence-Research Society (ICI-RS) meeting in Bristol, UK. The participants presented what is new, currently understood or still unknown on afferent-urothelial signaling mechanisms. New avenues of research and experimental methodologies that are or could be employed were presented and discussed. It is clear that afferent-urothelial interactions are integral to the regulation of normal bladder function and that its disruption can have detrimental consequences. The urothelium is capable of releasing numerous signaling factors that can affect sensory neurons innervating the suburothelium. However, the understanding of how factors released from urothelial cells and afferent nerve terminals regulate one another is incomplete. Utilization of techniques such as viruses that genetically encode Ca(2+) sensors, based on calmodulin and green fluorescent protein, has helped to address the cellular mechanisms involved. Additionally, the epithelial-neuronal interactions in the urethra may also play a significant role in lower urinary tract regulation and merit further investigation. The signaling capabilities of the urothelium and afferent nerves are well documented, yet how these signals are integrated to regulate bladder function is unclear. There is unquestionably a need for expanded methodologies to further our understanding of lower urinary tract sensory mechanisms and their contribution to various pathologies. © 2016 Wiley Periodicals, Inc.

The Caenorhabditis elegans Iodotyrosine Deiodinase Ortholog SUP-18 Functions through a Conserved Channel SC-Box to Regulate the Muscle Two-Pore Domain Potassium Channel SUP-9

PubMed Central

de la Cruz, Ignacio Perez; Ma, Long; Horvitz, H. Robert

2014-01-01

Loss-of-function mutations in the Caenorhabditis elegans gene sup-18 suppress the defects in muscle contraction conferred by a gain-of-function mutation in SUP-10, a presumptive regulatory subunit of the SUP-9 two-pore domain K+ channel associated with muscle membranes. We cloned sup-18 and found that it encodes the C. elegans ortholog of mammalian iodotyrosine deiodinase (IYD), an NADH oxidase/flavin reductase that functions in iodine recycling and is important for the biosynthesis of thyroid hormones that regulate metabolism. The FMN-binding site of mammalian IYD is conserved in SUP-18, which appears to require catalytic activity to function. Genetic analyses suggest that SUP-10 can function with SUP-18 to activate SUP-9 through a pathway that is independent of the presumptive SUP-9 regulatory subunit UNC-93. We identified a novel evolutionarily conserved serine-cysteine-rich region in the C-terminal cytoplasmic domain of SUP-9 required for its specific activation by SUP-10 and SUP-18 but not by UNC-93. Since two-pore domain K+ channels regulate the resting membrane potentials of numerous cell types, we suggest that the SUP-18 IYD regulates the activity of the SUP-9 channel using NADH as a coenzyme and thus couples the metabolic state of muscle cells to muscle membrane excitability. PMID:24586202

Membrane cofactor protein (CD46) is a keratinocyte receptor for the M protein of the group A streptococcus.

PubMed

Okada, N; Liszewski, M K; Atkinson, J P; Caparon, M

1995-03-28

The pathogenic Gram-positive bacterium Streptococcus pyogenes (group A streptococcus) is the causative agent of numerous suppurative diseases of human skin. The M protein of S. pyogenes mediates the adherence of the bacterium to keratinocytes, the most numerous cell type in the epidermis. In this study, we have constructed and analyzed a series of mutant M proteins and have shown that the C repeat domain of the M molecule is responsible for cell recognition. The binding of factor H, a serum regulator of complement activation, to the C repeat region of M protein blocked bacterial adherence. Factor H is a member of a large family of complement regulatory proteins that share a homologous structural motif termed the short consensus repeat. Membrane cofactor protein (MCP), or CD46, is a short consensus repeat-containing protein found on the surface of keratinocytes, and purified MCP could competitively inhibit the adherence of S. pyogenes to these cells. Furthermore, the M protein was found to bind directly to MCP, whereas mutant M proteins that lacked the C repeat domain did not bind MCP, suggesting that recognition of MCP plays an important role in the ability of the streptococcus to adhere to keratinocytes.

Negative regulation of quorum-sensing systems in Pseudomonas aeruginosa by ATP-dependent Lon protease.

PubMed

Takaya, Akiko; Tabuchi, Fumiaki; Tsuchiya, Hiroko; Isogai, Emiko; Yamamoto, Tomoko

2008-06-01

Lon protease, a member of the ATP-dependent protease family, regulates numerous cellular systems by degrading specific substrates. Here, we demonstrate that Lon is involved in the regulation of quorum-sensing (QS) signaling systems in Pseudomonas aeruginosa, an opportunistic human pathogen. The organism has two acyl-homoserine lactone (HSL)-mediated QS systems, LasR/LasI and RhlR/RhlI. Many reports have demonstrated that these two systems are regulated and interconnected by global regulators. We found that lon-disrupted cells overproduce pyocyanin, the biosynthesis of which depends on the RhlR/RhlI system, and show increased levels of a transcriptional regulator, RhlR. The QS systems are organized hierarchically: the RhlR/RhlI system is subordinate to LasR/LasI. To elucidate the mechanism by which Lon negatively regulates RhlR/RhlI, we examined the effect of lon disruption on the LasR/LasI system. We found that Lon represses the expression of LasR/LasI by degrading LasI, an HSL synthase, leading to negative regulation of the RhlR/RhlI system. RhlR/RhlI was also shown to be regulated by Lon independently of LasR/LasI via regulation of RhlI, an HSL synthase. In view of these findings, it is suggested that Lon protease is a powerful negative regulator of both HSL-mediated QS systems in P. aeruginosa.

Human CD30+ B cells represent a unique subset related to Hodgkin lymphoma cells.

PubMed

Weniger, Marc A; Tiacci, Enrico; Schneider, Stefanie; Arnolds, Judith; Rüschenbaum, Sabrina; Duppach, Janine; Seifert, Marc; Döring, Claudia; Hansmann, Martin-Leo; Küppers, Ralf

2018-06-11

Very few B cells in germinal centers (GCs) and extrafollicular (EF) regions of lymph nodes express CD30. Their specific features and relationship to CD30-expressing Hodgkin and Reed/Sternberg (HRS) cells of Hodgkin lymphoma are unclear but highly relevant, because numerous patients with lymphoma are currently treated with an anti-CD30 immunotoxin. We performed a comprehensive analysis of human CD30+ B cells. Phenotypic and IgV gene analyses indicated that CD30+ GC B lymphocytes represent typical GC B cells, and that CD30+ EF B cells are mostly post-GC B cells. The transcriptomes of CD30+ GC and EF B cells largely overlapped, sharing a strong MYC signature, but were strikingly different from conventional GC B cells and memory B and plasma cells, respectively. CD30+ GC B cells represent MYC+ centrocytes redifferentiating into centroblasts; CD30+ EF B cells represent active, proliferating memory B cells. HRS cells shared typical transcriptome patterns with CD30+ B cells, suggesting that they originate from these lymphocytes or acquire their characteristic features during lymphomagenesis. By comparing HRS to normal CD30+ B cells we redefined aberrant and disease-specific features of HRS cells. A remarkable downregulation of genes regulating genomic stability and cytokinesis in HRS cells may explain their genomic instability and multinuclearity.

Regulation of spinogenesis in mature Purkinje cells via mGluR/PKC-mediated phosphorylation of CaMKIIβ

PubMed Central

Sugawara, Takeyuki; Hisatsune, Chihiro; Miyamoto, Hiroyuki; Ogawa, Naoko; Mikoshiba, Katsuhiko

2017-01-01

Dendritic spines of Purkinje cells form excitatory synapses with parallel fiber terminals, which are the primary sites for cerebellar synaptic plasticity. Nevertheless, how density and morphology of these spines are properly maintained in mature Purkinje cells is not well understood. Here we show an activity-dependent mechanism that represses excessive spine development in mature Purkinje cells. We found that CaMKIIβ promotes spine formation and elongation in Purkinje cells through its F-actin bundling activity. Importantly, activation of group I mGluR, but not AMPAR, triggers PKC-mediated phosphorylation of CaMKIIβ, which results in dissociation of the CaMKIIβ/F-actin complex. Defective function of the PKC-mediated CaMKIIβ phosphorylation promotes excess F-actin bundling and leads to abnormally numerous and elongated spines in mature IP3R1-deficient Purkinje cells. Thus, our data suggest that phosphorylation of CaMKIIβ through the mGluR/IP3R1/PKC signaling pathway represses excessive spine formation and elongation in mature Purkinje cells. PMID:28607044

Discovery of thalicthuberine as a novel antimitotic agent from nature that disrupts microtubule dynamics and induces apoptosis in prostate cancer cells

PubMed Central

2018-01-01

ABSTRACT We report for the first time the mechanism of action of the natural product thalicthuberine (TH) in prostate and cervical cancer cells. TH induced a strong accumulation of LNCaP cells in mitosis, severe mitotic spindle defects, and asymmetric cell divisions, ultimately leading to mitotic catastrophe accompanied by cell death through apoptosis. However, unlike microtubule-binding drugs (vinblastine and paclitaxel), TH did not directly inhibit tubulin polymerization when tested in a cell-free system, whereas it reduced cellular microtubule polymer mass in LNCaP cells. This suggests that TH indirectly targets microtubule dynamics through inhibition of a critical regulator or tubulin-associated protein. Furthermore, TH is not a major substrate for P-glycoprotein (Pgp), which is responsible for multidrug resistance in numerous cancers, providing a rationale to further study TH in cancers with Pgp-mediated treatment resistance. The identification of TH's molecular target in future studies will be of great value to the development of TH as potential treatment of multidrug-resistant tumors. PMID:28749250

The DNA Inflammasome in Human Myeloid Cells Is Initiated by a STING-Cell Death Program Upstream of NLRP3

PubMed Central

Gaidt, Moritz M.; Ebert, Thomas S.; Chauhan, Dhruv; Ramshorn, Katharina; Pinci, Francesca; Zuber, Sarah; O’Duill, Fionan; Schmid-Burgk, Jonathan L.; Hoss, Florian; Buhmann, Raymund; Wittmann, Georg; Latz, Eicke; Subklewe, Marion; Hornung, Veit

2018-01-01

Summary Detection of cytosolic DNA constitutes a central event in the context of numerous infectious and sterile inflammatory conditions. Recent studies have uncovered a bipartite mode of cytosolic DNA recognition, in which the cGAS-STING axis triggers antiviral immunity, whereas AIM2 triggers inflammasome activation. Here, we show that AIM2 is dispensable for DNA-mediated inflammasome activation in human myeloid cells. Instead, detection of cytosolic DNA by the cGAS-STING axis induces a cell death program initiating potassium efflux upstream of NLRP3. Forward genetics identified regulators of lysosomal trafficking to modulate this cell death program, and subsequent studies revealed that activated STING traffics to the lysosome, where it triggers membrane permeabilization and thus lysosomal cell death (LCD). Importantly, the cGAS-STING-NLRP3 pathway constitutes the default inflammasome response during viral and bacterial infections in human myeloid cells. We conclude that targeting the cGAS-STING-LCD-NLRP3 pathway will ameliorate pathology in inflammatory conditions that are associated with cytosolic DNA sensing. PMID:29033128

Allosteric inhibition of a stem cell RNA-binding protein by an intermediary metabolite

PubMed Central

Clingman, Carina C; Deveau, Laura M; Hay, Samantha A; Genga, Ryan M; Shandilya, Shivender MD; Massi, Francesca; Ryder, Sean P

2014-01-01

Gene expression and metabolism are coupled at numerous levels. Cells must sense and respond to nutrients in their environment, and specialized cells must synthesize metabolic products required for their function. Pluripotent stem cells have the ability to differentiate into a wide variety of specialized cells. How metabolic state contributes to stem cell differentiation is not understood. In this study, we show that RNA-binding by the stem cell translation regulator Musashi-1 (MSI1) is allosterically inhibited by 18–22 carbon ω-9 monounsaturated fatty acids. The fatty acid binds to the N-terminal RNA Recognition Motif (RRM) and induces a conformational change that prevents RNA association. Musashi proteins are critical for development of the brain, blood, and epithelium. We identify stearoyl-CoA desaturase-1 as a MSI1 target, revealing a feedback loop between ω-9 fatty acid biosynthesis and MSI1 activity. We propose that other RRM proteins could act as metabolite sensors to couple gene expression changes to physiological state. DOI: http://dx.doi.org/10.7554/eLife.02848.001 PMID:24935936

Oxygen-Dependent Cell-to-Cell Variability in the Output of the Escherichia coli Tor Phosphorelay.

PubMed

Roggiani, Manuela; Goulian, Mark

2015-06-15

Escherichia coli senses and responds to trimethylamine-N-oxide (TMAO) in the environment through the TorT-TorS-TorR signal transduction system. The periplasmic protein TorT binds TMAO and stimulates the hybrid kinase TorS to phosphorylate the response regulator TorR through a phosphorelay. Phosphorylated TorR, in turn, activates transcription of the torCAD operon, which encodes the proteins required for anaerobic respiration via reduction of TMAO to trimethylamine. Interestingly, E. coli respires TMAO in both the presence and absence of oxygen, a behavior that is markedly different from the utilization of other alternative electron acceptors by this bacterium. Here we describe an unusual form of regulation by oxygen for this system. While the average level of torCAD transcription is the same for aerobic and anaerobic cultures containing TMAO, the behavior across the population of cells is strikingly different under the two growth conditions. Cellular levels of torCAD transcription in aerobic cultures are highly heterogeneous, in contrast to the relatively homogeneous distribution in anaerobic cultures. Thus, oxygen regulates the variance of the output but not the mean for the Tor system. We further show that this oxygen-dependent variability stems from the phosphorelay. Trimethylamine-N-oxide (TMAO) is utilized by numerous bacteria as an electron acceptor for anaerobic respiration. In E. coli, expression of the proteins required for TMAO respiration is tightly regulated by a signal transduction system that is activated by TMAO. Curiously, although oxygen is the energetically preferred electron acceptor, TMAO is respired even in the presence of oxygen. Here we describe an interesting and unexpected form of regulation for this system in which oxygen produces highly variable expression of the TMAO utilization proteins across a population of cells without affecting the mean expression of these proteins. To our knowledge, this is the first reported example of a stimulus regulating the variance but not the mean output of a signaling system. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Genome-wide analysis of human constitutive androstane receptor (CAR) transcriptome in wild-type and CAR-knockout HepaRG cells.

PubMed

Li, Daochuan; Mackowiak, Bryan; Brayman, Timothy G; Mitchell, Michael; Zhang, Lei; Huang, Shiew-Mei; Wang, Hongbing

2015-11-01

The constitutive androstane receptor (CAR) modulates the transcription of numerous genes involving drug metabolism, energy homeostasis, and cell proliferation. Most functions of CAR however were defined from animal studies. Given the known species difference of CAR and the significant cross-talk between CAR and the pregnane X receptor (PXR), it is extremely difficult to decipher the exact role of human CAR (hCAR) in gene regulation, relying predominantly on pharmacological manipulations. Here, utilizing a newly generated hCAR-knockout (KO) HepaRG cell line, we carried out RNA-seq analysis of the global transcriptomes in wild-type (WT) and hCAR-KO HepaRG cells treated with CITCO, a selective hCAR agonist, phenobarbital (PB), a dual activator of hCAR and hPXR, or vehicle control. Real-time PCR assays in separate experiments were used to validate RNA-seq findings. Our results indicate that genes encoding drug-metabolizing enzymes are among the main clusters altered by both CITCO and PB. Specifically, CITCO significantly changed the expression of 135 genes in an hCAR-dependent manner, while PB altered the expression of 227 genes in WT cells of which 94 were simultaneously modulated in both cell lines reflecting dual effects of PB on hCAR/PXR. Notably, we found that many genes promoting cell proliferation and tumorigenesis were up-regulated in hCAR-KO cells, suggesting that hCAR may play an important role in cell growth that differs from mouse CAR. Together, our results reveal both novel and known targets of hCAR and support the role of hCAR in maintaining the homeostasis of metabolism and cell proliferation in the liver. Copyright © 2015 Elsevier Inc. All rights reserved.

The pluripotency factor Nanog is directly upregulated by the androgen receptor in prostate cancer cells.

PubMed

Kregel, Steven; Szmulewitz, Russell Z; Vander Griend, Donald J

2014-11-01

The Androgen Receptor (AR) is a nuclear hormone receptor that functions as a critical oncogene in all stages of prostate cancer progression, including progression to castration-resistance following androgen-deprivation therapy. Thus, identifying and targeting critical AR-regulated genes is one potential method to block castration-resistant cancer proliferation. Of particular importance are transcription factors that regulate stem cell pluripotency; many of these genes are emerging as critical oncogenes in numerous tumor cell types. Of these, Nanog has been previously shown to increase the self-renewal and stem-like properties of prostate cancer cells. Thus, we hypothesized that Nanog is a candidate AR target gene that may impart castration-resistance. We modulated AR signaling in LNCaP prostate cancer cells and assayed for Nanog expression. Direct AR binding to the NANOG promoter was tested using AR Chromatin Immunoprecipation (ChIP) and analyses of publically available AR ChIP-sequencing data-sets. Nanog over-expressing cells were analyzed for cell growth and cytotoxicity in response to the AR antagonist enzalutamide and the microtubule stabilizing agent docetaxel. AR signaling upregulates Nanog mRNA and protein. AR binds directly to the NANOG promoter, and was not identified within 75 kb of the NANOGP8 pseudogene, suggesting the NANOG gene locus was preferentially activated. Nanog overexpression in LNCaP cells increases overall growth, but does not increase resistance to enzalutamide or docetaxel. Nanog is a novel oncogenic AR target gene in prostate cancer cells, and stable expression of Nanog increases proliferation and growth of prostate cancer cells, but not resistance to enzalutamide or docetaxel. © 2014 Wiley Periodicals, Inc.

Inhibitory Interneurons That Express GFP in the PrP-GFP Mouse Spinal Cord Are Morphologically Heterogeneous, Innervated by Several Classes of Primary Afferent and Include Lamina I Projection Neurons among Their Postsynaptic Targets

PubMed Central

Ganley, Robert P.; Iwagaki, Noboru; del Rio, Patricia; Baseer, Najma; Dickie, Allen C.; Boyle, Kieran A.; Polgár, Erika; Watanabe, Masahiko; Abraira, Victoria E; Zimmerman, Amanda

2015-01-01

The superficial dorsal horn of the spinal cord contains numerous inhibitory interneurons, which regulate the transmission of information perceived as touch, pain, or itch. Despite the importance of these cells, our understanding of their roles in the neuronal circuitry is limited by the difficulty in identifying functional populations. One group that has been identified and characterized consists of cells in the mouse that express green fluorescent protein (GFP) under control of the prion protein (PrP) promoter. Previous reports suggested that PrP-GFP cells belonged to a single morphological class (central cells), received inputs exclusively from unmyelinated primary afferents, and had axons that remained in lamina II. However, we recently reported that the PrP-GFP cells expressed neuronal nitric oxide synthase (nNOS) and/or galanin, and it has been shown that nNOS-expressing cells are more diverse in their morphology and synaptic connections. We therefore used a combined electrophysiological, pharmacological, and anatomical approach to reexamine the PrP-GFP cells. We provide evidence that they are morphologically diverse (corresponding to “unclassified” cells) and receive synaptic input from a variety of primary afferents, with convergence onto individual cells. We also show that their axons project into adjacent laminae and that they target putative projection neurons in lamina I. This indicates that the neuronal circuitry involving PrP-GFP cells is more complex than previously recognized, and suggests that they are likely to have several distinct roles in regulating the flow of somatosensory information through the dorsal horn. PMID:25972186

DNA microarray analyses reveal a post-irradiation differential time-dependent gene expression profile in yeast cells exposed to X-rays and gamma-rays.

PubMed

Kimura, Shinzo; Ishidou, Emi; Kurita, Sakiko; Suzuki, Yoshiteru; Shibato, Junko; Rakwal, Randeep; Iwahashi, Hitoshi

2006-07-21

Ionizing radiation (IR) is the most enigmatic of genotoxic stress inducers in our environment that has been around from the eons of time. IR is generally considered harmful, and has been the subject of numerous studies, mostly looking at the DNA damaging effects in cells and the repair mechanisms therein. Moreover, few studies have focused on large-scale identification of cellular responses to IR, and to this end, we describe here an initial study on the transcriptional responses of the unicellular genome model, yeast (Saccharomyces cerevisiae strain S288C), by cDNA microarray. The effect of two different IR, X-rays, and gamma (gamma)-rays, was investigated by irradiating the yeast cells cultured in YPD medium with 50 Gy doses of X- and gamma-rays, followed by resuspension of the cells in YPD for time-course experiments. The samples were collected for microarray analysis at 20, 40, and 80 min after irradiation. Microarray analysis revealed a time-course transcriptional profile of changed gene expressions. Up-regulated genes belonged to the functional categories mainly related to cell cycle and DNA processing, cell rescue defense and virulence, protein and cell fate, and metabolism (X- and gamma-rays). Similarly, for X- and gamma-rays, the down-regulated genes belonged to mostly transcription and protein synthesis, cell cycle and DNA processing, control of cellular organization, cell fate, and C-compound and carbohydrate metabolism categories, respectively. This study provides for the first time a snapshot of the genome-wide mRNA expression profiles in X- and gamma-ray post-irradiated yeast cells and comparatively interprets/discusses the changed gene functional categories as effects of these two radiations vis-à-vis their energy levels.

Non-coding RNA networks in cancer.

PubMed

Anastasiadou, Eleni; Jacob, Leni S; Slack, Frank J

2018-01-01

Thousands of unique non-coding RNA (ncRNA) sequences exist within cells. Work from the past decade has altered our perception of ncRNAs from 'junk' transcriptional products to functional regulatory molecules that mediate cellular processes including chromatin remodelling, transcription, post-transcriptional modifications and signal transduction. The networks in which ncRNAs engage can influence numerous molecular targets to drive specific cell biological responses and fates. Consequently, ncRNAs act as key regulators of physiological programmes in developmental and disease contexts. Particularly relevant in cancer, ncRNAs have been identified as oncogenic drivers and tumour suppressors in every major cancer type. Thus, a deeper understanding of the complex networks of interactions that ncRNAs coordinate would provide a unique opportunity to design better therapeutic interventions.

Long noncoding RNA Hoxb3os is dysregulated in autosomal dominant polycystic kidney disease and regulates mTOR signaling.

PubMed

Aboudehen, Karam; Farahani, Shayan; Kanchwala, Mohammed; Chan, Siu Chiu; Avdulov, Svetlana; Mickelson, Alan; Lee, Dayeon; Gearhart, Micah D; Patel, Vishal; Xing, Chao; Igarashi, Peter

2018-06-15

Autosomal dominant polycystic kidney disease (ADPKD) is a debilitating disease that is characterized by the accumulation of numerous fluid-filled cysts in the kidney. ADPKD is primarily caused by mutations in two genes, PKD1 and PKD2 Long noncoding RNAs (lncRNA), defined by a length >200 nucleotides and absence of a long ORF, have recently emerged as epigenetic regulators of development and disease; however, their involvement in PKD has not been explored previously. Here, we performed deep RNA-Seq to identify lncRNAs that are dysregulated in two orthologous mouse models of ADPKD (kidney-specific Pkd1 and Pkd2 mutant mice). We identified a kidney-specific, evolutionarily conserved lncRNA called Hoxb3os that was down-regulated in cystic kidneys from Pkd1 and Pkd2 mutant mice. The human ortholog HOXB3-AS1 was down-regulated in cystic kidneys from ADPKD patients. Hoxb3os was highly expressed in renal tubules in adult WT mice, whereas its expression was lost in the cyst epithelium of mutant mice. To investigate the function of Hoxb3os , we utilized CRISPR/Cas9 to knock out its expression in mIMCD3 cells. Deletion of Hoxb3os resulted in increased phosphorylation of mTOR and its downstream targets, including p70 S6 kinase, ribosomal protein S6, and the translation repressor 4E-BP1. Consistent with activation of mTORC1 signaling, Hoxb3os mutant cells displayed increased mitochondrial respiration. The Hoxb3os mutant phenotype was partially rescued upon re-expression of Hoxb3os in knockout cells. These findings identify Hoxb3os as a novel lncRNA that is down-regulated in ADPKD and regulates mTOR signaling and mitochondrial respiration. © 2018 Aboudehen et al.

Cannabis: old medicine with new promise for neurological disorders.

PubMed

Carter, Gregory T; Weydt, Patrick

2002-03-01

Marijuana is a complex substance containing over 60 different forms of cannabinoids, the active ingredients. Cannabinoids are now known to have the capacity for neuromodulation, via direct, receptor-based mechanisms at numerous levels within the nervous system. These have therapeutic properties that may be applicable to the treatment of neurological disorders; including anti-oxidative, neuroprotective, analgesic and anti-inflammatory actions; immunomodulation, modulation of glial cells and tumor growth regulation. This article reviews the emerging research on the physiological mechanisms of endogenous and exogenous cannabinoids in the context of neurological disease.

Diffusion model to describe osteogenesis within a porous titanium scaffold.

PubMed

Schmitt, M; Allena, R; Schouman, T; Frasca, S; Collombet, J M; Holy, X; Rouch, P

2016-01-01

In this study, we develop a two-dimensional finite element model, which is derived from an animal experiment and allows simulating osteogenesis within a porous titanium scaffold implanted in ewe's hemi-mandible during 12 weeks. The cell activity is described through diffusion equations and regulated by the stress state of the structure. We compare our model to (i) histological observations and (ii) experimental data obtained from a mechanical test done on sacrificed animal. We show that our mechano-biological approach provides consistent numerical results and constitutes a useful tool to predict osteogenesis pattern.

Effects of Secreted Mast Cell Mediators on Retinal Pigment Epithelial Cells: Focus on Mast Cell Tryptase.

PubMed

Arai, Rei; Usui-Ouchi, Ayumi; Ito, Yosuke; Mashimo, Keitaro; Murakami, Akira; Ebihara, Nobuyuki

2017-01-01

Numerous mast cells are present in the choroid, but the effects of mast cell mediators on retinal pigment epithelial (RPE) cells are not well understood. We investigated the influence of mast cell mediators on RPE cells in vitro, focusing on tryptase. Expression of receptors was examined by the reverse transcription polymerase chain reaction. We also assessed production of interleukin 8 and vascular endothelial growth factor (VEGF) after RPE cells were stimulated with mast cell mediators by using an antibody array and enzyme-linked immunosorbent assay. Furthermore, we investigated the influence of tryptase on RPE cell migration and integrity by the scratch assay and the transepithelial resistance. RPE cells expressed protease-activated receptor 2 (PAR2), histamine receptor 1, tumor necrosis factor- α (TNF- α ) receptor 1, and CCR 1, 3, 4, 8, and 11. Tryptase, PAR2 agonists, histamine, and TNF- α all enhanced interleukin 8 production by RPE cells, while only tryptase enhanced VEGF production. Tryptase also enhanced expression of phosphorylated extracellular signal-regulated kinases 1/2, resulting in increased migration of RPE cells. However, tryptase did not alter epithelial integrity or the expression of zonula occludens-1 and junctional adhesion molecule-A by RPE cells. Mast cell mediators, especially tryptase, may influence RPE cell inflammation.

Modularized TGFbeta-Smad Signaling Pathway

NASA Technical Reports Server (NTRS)

Li, Yongfeng; Wang, M.; Carra, C.; Cucinotta, F. A.

2011-01-01

The Transforming Growth Factor beta (TGFbeta) signaling pathway is a prominent regulatory signaling pathway controlling various important cellular processes. It can be induced by several factors, including ionizing radiation. It is regulated by Smads in a negative feedback loop through promoting increases in the regulatory Smads in the cell nucleus, and subsequent expression of inhibitory Smad, Smad7 to form a ubiquitin ligase with Smurf targeting active TGF receptors for degradation. In this work, we proposed a mathematical model to study the radiation-induced Smad-regulated TGF signaling pathway. By modularization, we are able to analyze each module (subsystem) and recover the nonlinear dynamics of the entire network system. Meanwhile the excitability, a common feature observed in the biological systems, along the TGF signaling pathway is discussed by mathematical analysis and numerical simulation.

Identification of ATM Protein Kinase Phosphorylation Sites by Mass Spectrometry.

PubMed

Graham, Mark E; Lavin, Martin F; Kozlov, Sergei V

2017-01-01

ATM (ataxia-telangiectasia mutated) protein kinase is a key regulator of cellular responses to DNA damage and oxidative stress. DNA damage triggers complex cascade of signaling events leading to numerous posttranslational modification on multitude of proteins. Understanding the regulation of ATM kinase is therefore critical not only for understanding the human genetic disorder ataxia-telangiectasia and potential treatment strategies, but essential for deciphering physiological responses of cells to stress. These responses play an important role in carcinogenesis, neurodegeneration, and aging. We focus here on the identification of DNA damage inducible ATM phosphorylation sites to understand the importance of autophosphorylation in the mechanism of ATM kinase activation. We demonstrate the utility of using immunoprecipitated ATM in quantitative LC-MS/MS workflow with stable isotope dimethyl labeling of ATM peptides for identification of phosphorylation sites.

Macro- and microscale variables regulate stent haemodynamics, fibrin deposition and thrombomodulin expression

PubMed Central

Jiménez, Juan M.; Prasad, Varesh; Yu, Michael D.; Kampmeyer, Christopher P.; Kaakour, Abdul-Hadi; Wang, Pei-Jiang; Maloney, Sean F.; Wright, Nathan; Johnston, Ian; Jiang, Yi-Zhou; Davies, Peter F.

2014-01-01

Drug eluting stents are associated with late stent thrombosis (LST), delayed healing and prolonged exposure of stent struts to blood flow. Using macroscale disturbed and undisturbed fluid flow waveforms, we numerically and experimentally determined the effects of microscale model strut geometries upon the generation of prothrombotic conditions that are mediated by flow perturbations. Rectangular cross-sectional stent strut geometries of varying heights and corresponding streamlined versions were studied in the presence of disturbed and undisturbed bulk fluid flow. Numerical simulations and particle flow visualization experiments demonstrated that the interaction of bulk fluid flow and stent struts regulated the generation, size and dynamics of the peristrut flow recirculation zones. In the absence of endothelial cells, deposition of thrombin-generated fibrin occurred primarily in the recirculation zones. When endothelium was present, peristrut expression of anticoagulant thrombomodulin (TM) was dependent on strut height and geometry. Thinner and streamlined strut geometries reduced peristrut flow recirculation zones decreasing prothrombotic fibrin deposition and increasing endothelial anticoagulant TM expression. The studies define physical and functional consequences of macro- and microscale variables that relate to thrombogenicity associated with the most current stent designs, and particularly to LST. PMID:24554575

Comprehensive Identification of Long Non-coding RNAs in Purified Cell Types from the Brain Reveals Functional LncRNA in OPC Fate Determination.

PubMed

Dong, Xiaomin; Chen, Kenian; Cuevas-Diaz Duran, Raquel; You, Yanan; Sloan, Steven A; Zhang, Ye; Zong, Shan; Cao, Qilin; Barres, Ben A; Wu, Jia Qian

2015-12-01

Long non-coding RNAs (lncRNAs) (> 200 bp) play crucial roles in transcriptional regulation during numerous biological processes. However, it is challenging to comprehensively identify lncRNAs, because they are often expressed at low levels and with more cell-type specificity than are protein-coding genes. In the present study, we performed ab initio transcriptome reconstruction using eight purified cell populations from mouse cortex and detected more than 5000 lncRNAs. Predicting the functions of lncRNAs using cell-type specific data revealed their potential functional roles in Central Nervous System (CNS) development. We performed motif searches in ENCODE DNase I digital footprint data and Mouse ENCODE promoters to infer transcription factor (TF) occupancy. By integrating TF binding and cell-type specific transcriptomic data, we constructed a novel framework that is useful for systematically identifying lncRNAs that are potentially essential for brain cell fate determination. Based on this integrative analysis, we identified lncRNAs that are regulated during Oligodendrocyte Precursor Cell (OPC) differentiation from Neural Stem Cells (NSCs) and that are likely to be involved in oligodendrogenesis. The top candidate, lnc-OPC, shows highly specific expression in OPCs and remarkable sequence conservation among placental mammals. Interestingly, lnc-OPC is significantly up-regulated in glial progenitors from experimental autoimmune encephalomyelitis (EAE) mouse models compared to wild-type mice. OLIG2-binding sites in the upstream regulatory region of lnc-OPC were identified by ChIP (chromatin immunoprecipitation)-Sequencing and validated by luciferase assays. Loss-of-function experiments confirmed that lnc-OPC plays a functional role in OPC genesis. Overall, our results substantiated the role of lncRNA in OPC fate determination and provided an unprecedented data source for future functional investigations in CNS cell types. We present our datasets and analysis results via the interactive genome browser at our laboratory website that is freely accessible to the research community. This is the first lncRNA expression database of collective populations of glia, vascular cells, and neurons. We anticipate that these studies will advance the knowledge of this major class of non-coding genes and their potential roles in neurological development and diseases.

The mechanics of the primary cilium: an intricate structure with complex function.

PubMed

Hoey, David A; Downs, Matthew E; Jacobs, Christopher R

2012-01-03

The primary cilium is a non-motile singular cellular structure that extends from the surface of nearly every cell in the body. The cilium has been shown to play numerous roles in maintaining tissue homeostasis, through regulating signaling pathways and sensing both biophysical and biochemical changes in the extracellular environment. The structural performance of the cilium is paramount to its function as defective cilia have been linked to numerous pathologies. In particular, the cilium has demonstrated a mechanosensory role in tissues such as the kidney, liver, endothelium and bone, where cilium deflection under mechanical loading triggers a cellular response. Understanding of how cilium structure and subsequent mechanical behavior contributes to the roles that cilium plays in regulating cellular behavior is a compelling question, yet is a relatively untouched research area. Recent advances in biophysical measurements have demonstrated the cilium to be a structurally intricate organelle containing an array of load bearing proteins. Furthermore advances in modeling of this organelle have revealed the importance of these proteins at regulating the cilium's mechanosensitivity. Remarkably, the cilium is capable of adapting its mechanical state, altering its length and possibly it's bending resistance, to regulate its mechanosensitivity demonstrating the importance of cilium mechanics in cellular responses. In this review, we introduce the cilium as a mechanosensor; discuss the advances in the mechanical modeling of cilia; explore the structural features of the cilium, which contribute to its mechanics and finish with possible mechanisms in which alteration in structure may affect ciliary mechanics, consequently affecting ciliary based mechanosensing. Copyright © 2011 Elsevier Ltd. All rights reserved.

OCT4 expression mediates partial cardiomyocyte reprogramming of mesenchymal stromal cells.

PubMed

Yannarelli, Gustavo; Pacienza, Natalia; Montanari, Sonia; Santa-Cruz, Diego; Viswanathan, Sowmya; Keating, Armand

2017-01-01

Mesenchymal stem/stromal cells (MSCs) are in numerous cell therapy clinical trials, including for injured myocardium. Acquisition of cardiomyocyte characteristics by MSCs may improve cardiac regeneration but the mechanisms regulating this process are unclear. Here, we investigated whether the pluripotency transcription factor OCT4 is involved in the activation of cardiac lineage genetic programs in MSCs. We employed our established co-culture model of MSCs with rat embryonic cardiomyocytes showing co-expression of cardiac markers on MSCs independent of cell fusion. Bone marrow-derived MSCs were isolated from transgenic mice expressing GFP under the control of the cardiac-specific α-myosin heavy chain promoter. After 5 days of co-culture, MSCs expressed cardiac specific genes, including Nkx2.5, atrial natriuretic factor and α-cardiac actin. The frequency of GFP+ cells was 7.6±1.9%, however, these cells retained the stromal cell phenotype, indicating, as expected, only partial differentiation. Global OCT4 expression increased 2.6±0.7-fold in co-cultured MSCs and of interest, 87±5% vs 79±4% of MSCs expressed OCT4 by flow cytometry in controls and after co-culture, respectively. Consistent with the latter observation, the GFP+ cells did not express nuclear OCT4 and showed a significant increase in OCT4 promoter methylation compared with undifferentiated MSCs (92% vs 45%), inferring that OCT4 is regulated by an epigenetic mechanism. We further showed that siRNA silencing of OCT4 in MSCs resulted in a reduced frequency of GFP+ cells in co-culture to less than 1%. Our data infer that OCT4 expression may have a direct effect on partial cardiomyocyte reprogramming of MSCs and suggest a new mechanism(s) associated with MSC multipotency and a requirement for crosstalk with the cardiac microenvironment.

OCT4 expression mediates partial cardiomyocyte reprogramming of mesenchymal stromal cells

PubMed Central

Montanari, Sonia; Santa-Cruz, Diego; Viswanathan, Sowmya; Keating, Armand

2017-01-01

Mesenchymal stem/stromal cells (MSCs) are in numerous cell therapy clinical trials, including for injured myocardium. Acquisition of cardiomyocyte characteristics by MSCs may improve cardiac regeneration but the mechanisms regulating this process are unclear. Here, we investigated whether the pluripotency transcription factor OCT4 is involved in the activation of cardiac lineage genetic programs in MSCs. We employed our established co-culture model of MSCs with rat embryonic cardiomyocytes showing co-expression of cardiac markers on MSCs independent of cell fusion. Bone marrow-derived MSCs were isolated from transgenic mice expressing GFP under the control of the cardiac-specific α-myosin heavy chain promoter. After 5 days of co-culture, MSCs expressed cardiac specific genes, including Nkx2.5, atrial natriuretic factor and α-cardiac actin. The frequency of GFP+ cells was 7.6±1.9%, however, these cells retained the stromal cell phenotype, indicating, as expected, only partial differentiation. Global OCT4 expression increased 2.6±0.7-fold in co-cultured MSCs and of interest, 87±5% vs 79±4% of MSCs expressed OCT4 by flow cytometry in controls and after co-culture, respectively. Consistent with the latter observation, the GFP+ cells did not express nuclear OCT4 and showed a significant increase in OCT4 promoter methylation compared with undifferentiated MSCs (92% vs 45%), inferring that OCT4 is regulated by an epigenetic mechanism. We further showed that siRNA silencing of OCT4 in MSCs resulted in a reduced frequency of GFP+ cells in co-culture to less than 1%. Our data infer that OCT4 expression may have a direct effect on partial cardiomyocyte reprogramming of MSCs and suggest a new mechanism(s) associated with MSC multipotency and a requirement for crosstalk with the cardiac microenvironment. PMID:29216265

Learning a Markov Logic network for supervised gene regulatory network inference

PubMed Central

2013-01-01

Background Gene regulatory network inference remains a challenging problem in systems biology despite the numerous approaches that have been proposed. When substantial knowledge on a gene regulatory network is already available, supervised network inference is appropriate. Such a method builds a binary classifier able to assign a class (Regulation/No regulation) to an ordered pair of genes. Once learnt, the pairwise classifier can be used to predict new regulations. In this work, we explore the framework of Markov Logic Networks (MLN) that combine features of probabilistic graphical models with the expressivity of first-order logic rules. Results We propose to learn a Markov Logic network, e.g. a set of weighted rules that conclude on the predicate “regulates”, starting from a known gene regulatory network involved in the switch proliferation/differentiation of keratinocyte cells, a set of experimental transcriptomic data and various descriptions of genes all encoded into first-order logic. As training data are unbalanced, we use asymmetric bagging to learn a set of MLNs. The prediction of a new regulation can then be obtained by averaging predictions of individual MLNs. As a side contribution, we propose three in silico tests to assess the performance of any pairwise classifier in various network inference tasks on real datasets. A first test consists of measuring the average performance on balanced edge prediction problem; a second one deals with the ability of the classifier, once enhanced by asymmetric bagging, to update a given network. Finally our main result concerns a third test that measures the ability of the method to predict regulations with a new set of genes. As expected, MLN, when provided with only numerical discretized gene expression data, does not perform as well as a pairwise SVM in terms of AUPR. However, when a more complete description of gene properties is provided by heterogeneous sources, MLN achieves the same performance as a black-box model such as a pairwise SVM while providing relevant insights on the predictions. Conclusions The numerical studies show that MLN achieves very good predictive performance while opening the door to some interpretability of the decisions. Besides the ability to suggest new regulations, such an approach allows to cross-validate experimental data with existing knowledge. PMID:24028533

Gα73B is a downstream effector of JAK/STAT signalling and a regulator of Rho1 in Drosophila haematopoiesis.

PubMed

Bausek, Nina; Zeidler, Martin P

2014-01-01

JAK/STAT signalling regulates many essential developmental processes including cell proliferation and haematopoiesis, whereas its inappropriate activation is associated with the majority of myeloproliferative neoplasias and numerous cancers. Furthermore, high levels of JAK/STAT pathway signalling have also been associated with enhanced metastatic invasion by cancerous cells. Strikingly, gain-of-function mutations in the single Drosophila JAK homologue, Hopscotch, result in haemocyte neoplasia, inappropriate differentiation and the formation of melanised haemocyte-derived 'tumour' masses; phenotypes that are partly orthologous to human gain-of-function JAK2-associated pathologies. Here we show that Gα73B, a novel JAK/STAT pathway target gene, is necessary for JAK/STAT-mediated tumour formation in flies. In addition, although Gα73B does not affect haemocyte differentiation, it does regulate haemocyte morphology and motility under non-pathological conditions. We show that Gα73B is required for constitutive, but not injury-induced, activation of Rho1 and for the localisation of Rho1 into filopodia upon haemocyte activation. Consistent with these results, we also show that Rho1 interacts genetically with JAK/STAT signalling, and that wild-type levels of Rho1 are necessary for tumour formation. Our findings link JAK/STAT transcriptional outputs, Gα73B activity and Rho1-dependent cytoskeletal rearrangements and cell motility, therefore connecting a pathway associated with cancer with a marker indicative of invasiveness. As such, we suggest a mechanism by which JAK/STAT pathway signalling may promote metastasis.

The Regulation and Function of Fibroblast Growth Factor 8 and Its Function during Gonadotropin-Releasing Hormone Neuron Development.

PubMed

Chung, Wilson C J; Linscott, Megan L; Rodriguez, Karla M; Stewart, Courtney E

2016-01-01

Over the last few years, numerous studies solidified the hypothesis that fibroblast growth factor (FGF) signaling regulates neuroendocrine progenitor cell proliferation, fate specification, and cell survival and, therefore, is critical for the regulation and maintenance of homeostasis of the body. One important example that underscores the involvement of FGF signaling during neuroendocrine cell development is gonadotropin-releasing hormone (GnRH) neuron ontogenesis. Indeed, transgenic mice with reduced olfactory placode (OP) Fgf8 expression do not have GnRH neurons. This observation indicates the requirement of FGF8 signaling for the emergence of the GnRH neuronal system in the embryonic OP, the putative birth place of GnRH neurons. Mammalian reproductive success depends on the presence of GnRH neurons to stimulate gonadotropin secretion from the anterior pituitary, which activates gonadal steroidogenesis and gametogenesis. Together, these observations are critical for understanding the function of GnRH neurons and their control of the hypothalamus-pituitary-gonadal (HPG) axis to maintain fertility. Taken together, these studies illustrate that GnRH neuron emergence and hence HPG function is vulnerable to genomic and molecular signals that abnormally modify Fgf8 expression in the developing mouse OP. In this short review, we focus on research that is aimed at unraveling how androgen, all-trans retinoic acid, and how epigenetic factors modify control mouse OP Fgf8 transcription in the context of GnRH neuronal development and mammalian reproductive success.