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Sample records for gene-related peptide gene

  1. Calcitonin Gene-Related Peptide (CGRP)

    PubMed Central

    Russo, Andrew F.

    2015-01-01

    Migraine is a neurological disorder that manifests as a debilitating headache associated with altered sensory perception. The neuropeptide calcitonin gene-related peptide (CGRP) is now firmly established as a key player in migraine. Clinical trials carried out during the past decade have proved that CGRP receptor antagonists are effective for treating migraine, and antibodies to the receptor and CGRP are currently under investigation. Despite this progress in the clinical arena, the mechanisms by which CGRP triggers migraine remain uncertain. This review discusses mechanisms whereby CGRP enhances sensitivity to sensory input at multiple levels in both the periphery and central nervous system. Future studies on epistatic and epigenetic regulators of CGRP actions are expected to shed further light on CGRP actions in migraine. In conclusion, targeting CGRP represents an approachable therapeutic strategy for migraine. PMID:25340934

  2. Calcitonin Gene-Related Peptide: Physiology and Pathophysiology

    PubMed Central

    Russell, F. A.; King, R.; Smillie, S.-J.; Kodji, X.; Brain, S. D.

    2014-01-01

    Calcitonin gene-related peptide (CGRP) is a 37-amino acid neuropeptide. Discovered 30 years ago, it is produced as a consequence of alternative RNA processing of the calcitonin gene. CGRP has two major forms (α and β). It belongs to a group of peptides that all act on an unusual receptor family. These receptors consist of calcitonin receptor-like receptor (CLR) linked to an essential receptor activity modifying protein (RAMP) that is necessary for full functionality. CGRP is a highly potent vasodilator and, partly as a consequence, possesses protective mechanisms that are important for physiological and pathological conditions involving the cardiovascular system and wound healing. CGRP is primarily released from sensory nerves and thus is implicated in pain pathways. The proven ability of CGRP antagonists to alleviate migraine has been of most interest in terms of drug development, and knowledge to date concerning this potential therapeutic area is discussed. Other areas covered, where there is less information known on CGRP, include arthritis, skin conditions, diabetes, and obesity. It is concluded that CGRP is an important peptide in mammalian biology, but it is too early at present to know if new medicines for disease treatment will emerge from our knowledge concerning this molecule. PMID:25287861

  3. Calcitonin Gene-Related Peptide: Key Regulator of Cutaneous Immunity

    PubMed Central

    Granstein, Richard D.; Wagner, John A.; Stohl, Lori L.; Ding, Wanhong

    2014-01-01

    Calcitonin gene-related peptide (CGRP) has been viewed as a neuropeptide and vasodilator. However, CGRP is more appropriately thought of as a pleiotropic signaling molecule. Indeed, CGRP has key regulatory functions on immune and inflammatory processes within the skin. CGRP-containing nerves are intimately associated with epidermal LCs and CGRP has profound regulatory effects on Langerhans cell antigen-presenting capability. When LCs are exposed to CGRP in vitro, their ability to present antigen for in vivo priming of naïve mice or elicitation of delayed-type hypersensitivity is inhibited in at least some situations. Administration of CGRP intradermally inhibits acquisition of immunity to Th1-dominant haptens applied to the injected site while augmenting immunity to Th2-dominant haptens, although the cellular targets of activity in these experiments remains unclear. Although CGRP can be a pro-inflammatory agent, several studies have demonstrated that administration of CGRP can inhibit the elicitation of inflammation by inflammatory stimuli in vivo. In this regard, CGRP inhibits the release of certain chemokines by stimulated endothelial cells. This is likely to be physiologically relevant since cutaneous blood vessels are innervated by sensory nerves. Exciting new studies suggest a significant role for CGRP in the pathogenesis of psoriasis and, most strikingly, that CGRP inhibit the ability of LCs to transmit the human immunodeficiency virus 1 to T lymphocytes. A more complete understanding of the role of CGRP in the skin immune system may lead to new and novel approaches for the therapy of immune mediated skin disorders. PMID:25534428

  4. Inflammatory mediators release calcitonin gene-related peptide from dorsal root ganglion neurons of the rat.

    PubMed

    Averbeck, B; Izydorczyk, I; Kress, M

    2000-01-01

    The interactions between the inflammatory mediators bradykinin, serotonin, prostaglandin E(2) and acid pH were studied in rat dorsal root ganglion neurons in culture. For this purpose, the cultures were stimulated by inflammatory mediators (bradykinin, serotonin, prostaglandin E(2), 10(-5)M each) or acid solution (pH 6.1) for 5 min and the content of calcitonin gene-related peptide was determined in the supernatant before, during and after stimulation, using an enzyme immunoassay. Acid solution resulted in a threefold increase of the basal calcitonin gene-related peptide release which was entirely dependent on the presence of extracellular calcium. The release could not be blocked by the addition of the capsaicin antagonist capsazepine (10(-5)M). Bradykinin (10(-5)M) caused a 50% increase of the basal calcitonin gene-related peptide release which was again dependent on the presence of extracellular calcium, whereas serotonin and prostaglandin E(2) were each ineffective at 10(-5)M concentration. The combination of bradykinin, serotonin and prostaglandin E(2) led to a fivefold increase of the calcitonin gene-related peptide release which could not be further enhanced by acidification. The competitive capsaicin receptor antagonist capsazepine (10(-5)M) significantly reduced the release induced by the combination of bradykinin, serotonin and prostaglandin E(2). It is suggested that the inflammatory mediators co-operate and together may act as endogenous agonists at the capsaicin receptor to cause calcium influx and consecutive neuropeptide release. PMID:10858619

  5. Calcitonin gene-related peptide immunoreactive (CGRP-IR) intradental nerves in the dog.

    PubMed

    Ngassapa, D; Narhi, M; Hirvonen, T; Markkula, I

    1998-03-01

    Calcitonin gene-related peptide (CGRP) is the most frequently occurring peptide in sensory neurons in the rat. Most of the Calcitonin Gene-Related Peptide Immunoreactive (CGRP-IP) nerves have been found to be Capsaicin-sensitive suggesting an involvement in certain types of pain. In the dental pulp CGRP-IR nerve fibres have been studied in the rat, guinea pig, cat, sheep, pig, cow and horse but not in the dog. Extensive sprouting of CGRP-IP intradental nerves has been demonstrated in the teeth with pulpal inflammation in rat molars. In the present investigation the occurrence and distribution of CGRP-IR intradental nerves both in the normal and the injured and inflamed teeth of the dog was studied. Immunolabelling was done by the Avidin-Biotin Complex (ABC) immunoperoxidase technique. CGRP-IR intradental nerves were demonstrated in the dog. The pattern of distribution was found to be similar to that in other animal species. However, the tip one third of the coronal pulp was sparsely innervated compared to that in the rat. Also much fewer fibres were seen to penetrate predentine and dentine, and this appears characteristic of the dog teeth. Sprouting phenomenon seen in the rat was not found in the dog teeth. It is suggested that there might be a species difference in the innervation pattern of CGRP-IR intradental nerves between the rat molar and the dog canine and incisor teeth. PMID:9640812

  6. Calcitonin gene-related peptide targeted immunotherapy for migraine: progress and challenges in treating headache.

    PubMed

    Peroutka, Stephen J

    2014-06-01

    A role for calcitonin gene-related peptide (CGRP) in the pathophysiology of migraine has been established over the past 25 years. There have now been at least five different small-molecule CGRP antagonists that have demonstrated statistical proof of efficacy in the acute treatment of migraine. At present, multiple clinical trials are underway that are assessing the ability of long-acting antibodies against CGRP to prevent frequent migraine attacks. This review summarizes the existing data concerning the role of CGRP in migraine and attempts to highlight some possible outcomes from the ongoing anti-CGRP antibody trials. PMID:24452707

  7. Inhibition of Calcitonin Gene-Related Peptide Function: A Promising Strategy for Treating Migraine

    PubMed Central

    Durham, Paul L.

    2011-01-01

    The neuropeptide calcitonin gene-related peptide (CGRP) is implicated in the underlying pathology of migraine. Serum levels of CGRP, which are elevated during a migraine attack, have been reported to return to normal with alleviation of pain. In addition, CGRP administration has been shown to cause a migraine-like headache in susceptible individuals. Importantly, CGRP receptors are found on many cell types within the trigeminovascular system that are thought to play important roles in controlling inflammatory and nociceptive processes. Based on these findings, it was proposed that blockage of CGRP receptor function and, hence, the physiological effects of CGRP would be effective in aborting a migraine attack. This review will summarize key preclinical data that support the therapeutic potential of using CGRP receptor antagonists or molecules that bind CGRP within the context of current neurovascular theories on migraine pathology. PMID:18808507

  8. Structure–activity relationships for α-calcitonin gene-related peptide

    PubMed Central

    Watkins, Harriet A; Rathbone, Dan L; Barwell, James; Hay, Debbie L; Poyner, David R

    2013-01-01

    Calcitonin gene-related peptide (CGRP) is a member of the calcitonin (CT) family of peptides. It is a widely distributed neuropeptide implicated in conditions such as neurogenic inflammation. With other members of the CT family, it shares an N-terminal disulphide-bonded ring which is essential for biological activity, an area of potential α-helix, and a C-terminal amide. CGRP binds to the calcitonin receptor-like receptor (CLR) in complex with receptor activity-modifying protein 1 (RAMP1), a member of the family B (or secretin-like) GPCRs. It can also activate other CLR or calcitonin-receptor/RAMP complexes. This 37 amino acid peptide comprises the N-terminal ring that is required for receptor activation (residues 1–7); an α-helix (residues 8–18), a region incorporating a β-bend (residues 19–26) and the C-terminal portion (residues 27–37), that is characterized by bends between residues 28–30 and 33–34. A few residues have been identified that seem to make major contributions to receptor binding and activation, with a larger number contributing either to minor interactions (which collectively may be significant), or to maintaining the conformation of the bound peptide. It is not clear if CGRP follows the pattern of other family B GPCRs in binding largely as an α-helix. LINKED ARTICLES This article is part of a themed section on Neuropeptides. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2013.170.issue-7 PMID:23186257

  9. Peripheral amplification of sweating – a role for calcitonin gene-related peptide

    PubMed Central

    Schlereth, Tanja; Dittmar, Jan Oliver; Seewald, Bianca; Birklein, Frank

    2006-01-01

    Neuropeptides are the mediators of neurogenic inflammation. Some pain disorders, e.g. complex regional pain syndromes, are characterized by increased neurogenic inflammation and by exaggerated sudomotor function. The aim of this study was to explore whether neuropeptides have a peripheral effect on human sweating. We investigated the effects of different concentrations of calcitonin gene-related peptide (CGRP), vasoactive intestinal peptide (VIP) and substance P (SP) on acetylcholine-induced axon reflex sweating in healthy subjects (total n = 18). All substances were applied via dermal microdialysis. The experiments were done in a parallel setting: ACh alone and ACh combined with CGRP, VIP or SP in various concentrations were applied. Acetylcholine (10−2m) always elicited a sweating response, neuropeptides alone did not. However, CGRP significantly enhanced ACh-induced sweating (P < 0.01). Post hoc tests revealed that CGRP in physiological concentrations of 10−7–10−9m was most effective. VIP at any concentration had no significant effect on axon reflex sweating. The duration of the sweating response (P < 0.01), but not the amount of sweat, was reduced by SP. ACh-induced skin blood flow was significantly increased by CGRP (P < 0.01), but unaltered by VIP and SP. The results indicate that CGRP amplifies axon reflex sweating in human skin. PMID:16931551

  10. Superior Cervical Ganglia Neurons Induce Foxp3+ Regulatory T Cells via Calcitonin Gene-Related Peptide

    PubMed Central

    Szklany, Kirsten; Ruiter, Evelyn; Mian, Firoz; Kunze, Wolfgang; Bienenstock, John; Forsythe, Paul; Karimi, Khalil

    2016-01-01

    The nervous and immune systems communicate bidirectionally, utilizing diverse molecular signals including cytokines and neurotransmitters to provide an integrated response to changes in the body’s internal and external environment. Although, neuro-immune interactions are becoming better understood under inflammatory circumstances and it has been evidenced that interaction between neurons and T cells results in the conversion of encephalitogenic T cells to T regulatory cells, relatively little is known about the communication between neurons and naïve T cells. Here, we demonstrate that following co-culture of naïve CD4+ T cells with superior cervical ganglion neurons, the percentage of Foxp3 expressing CD4+CD25+ cells significantly increased. This was mediated in part by immune-regulatory cytokines TGF-β and IL-10, as well as the neuropeptide calcitonin gene-related peptide while vasoactive intestinal peptide was shown to play no role in generation of T regulatory cells. Additionally, T cells co-cultured with neurons showed a decrease in the levels of pro-inflammatory cytokine IFN-γ released upon in vitro stimulation. These findings suggest that the generation of Tregs may be promoted by naïve CD4+ T cell: neuron interaction through the release of neuropeptide CGRP. PMID:27022966

  11. Calcitonin gene-related peptide in migraine: intersection of peripheral inflammation and central modulation

    PubMed Central

    Raddant, Ann C.; Russo, Andrew F.

    2012-01-01

    Over the past two decades, a convergence of basic and clinical evidence has established the neuropeptide calcitonin-gene-related peptide (CGRP) as a key player in migraine. Although CGRP is a recognised neuromodulator of nociception, its mechanism of action in migraine remains elusive. In this review, we present evidence that led us to propose that CGRP is well poised to enhance neurotransmission in migraine by both peripheral and central mechanisms. In the periphery, it is thought that local release of CGRP from the nerve endings of meningeal nociceptors following their initial activation by cortical spreading depression is critical for the induction of vasodilation, plasma protein extravasation, neurogenic inflammation and the consequential sensitisation of meningeal nociceptors. Mechanistically, we propose that CGRP release can give rise to a positive-feedback loop involved in localised increased synthesis and release of CGRP from neurons and a CGRP-like peptide called procalcitonin from trigeminal ganglion glia. Within the brain, the wide distribution of CGRP and CGRP receptors provides numerous possible targets for CGRP to act as a neuromodulator. PMID:22123247

  12. A novel function of calcitonin gene-related peptide in body fluid Cl- homeostasis.

    PubMed

    Wang, Yi-Fang; Lafont, Anne-Gaëlle; Lee, Yi-Chun; Hwang, Pung-Pung

    2016-06-15

    Vertebrates need to maintain extracellular chloride (Cl(-)) concentrations to ensure the normal operation of physiological processes; the transition from aquatic to terrestrial environments necessitated the development of sophisticated mechanisms to ensure Cl(-) homeostasis in the face of fluctuating Cl(-) levels. Zebrafish calcitonin gene-related peptide (CGRP), unlike its splice variant calcitonin, does not respond to environmental Ca(2+) levels. This study aimed to test the hypothesis that CGRP is involved in the control of body fluid Cl(-) homeostasis. Acclimation to high-Cl(-) artificial water stimulated the mRNA expression of cgrp and the receptor (crlr1) when compared with low-Cl(-) CGRP knockdown induced upregulation of the Na(+)-Cl(-) co-transporter (ncc2b), while overexpression of CGRP resulted in the downregulation of ncc2b mRNA synthesis and a simultaneous decrease in Cl(-) uptake in embryos. Consistent with these findings, knockdown of either cgrp or crlr1 was found to increase the density of NCC2b-expressing cells in embryos. This is the first demonstration that CGRP acts as a hypochloremic hormone through suppressing NCC2b expression and the differentiation of NCC-expressing ionocytes. Elucidation of this novel function of CGRP in fish body fluid Cl(-) homeostasis promises to enhance our understanding of the related physiology in vertebrates. PMID:27306053

  13. Calcitonin gene-related peptide increases acetylcholine quantal size in neuromuscular junctions of mice.

    PubMed

    Gaydukov, Alexander E; Bogacheva, Polina O; Balezina, Olga P

    2016-08-15

    We used an intracellular microelectrode technique to study the mechanisms of action of two isoforms (human and rat) of calcitonin gene-related peptide (CGRP) on the evoked and spontaneous quantal secretion of acetylcholine (ACh) in mouse diaphragm motor synapses. Recordings of miniature endplate potentials (MEPPs) and evoked multiquantal endplate potentials (EPPs) in a cut neuromuscular preparation showed that CGRP increased the amplitude of EPPs without influencing their quantal content. Both isoforms of CGRP in a wide range of concentrations (1nM-1μM) provoked a similar considerable increase in MEPPs amplitude in a dose-dependent manner (up to 150-160% compared to control) without changing their frequency, rise-time, and decay. Inhibition of CGRP-receptors by truncated CGRP (CGRP8-37) completely prevented the potentiating effect of CGRP on the MEPPs amplitude. The effect of CGRP was not accompanied by changes in input resistance of muscle fiber membrane but was fully prevented by inhibition of vesicular ACh transport by vesamicol. Inhibition of protein kinase A (PKA) by H-89 also prevented CGRP action on the MEPPs amplitude. It is concluded that, in mammalian neuromuscular junctions, different isoforms of exogenously applied CGRP uniformly potentiate amplitudes of evoked and spontaneous postsynaptic potentials acting presynaptically via an increase in ACh quantal size. PMID:27288020

  14. Calcitonin gene-related peptide is a key neurotransmitter in the neuro-immune axis

    PubMed Central

    Assas, Bakri M.; Pennock, Joanne I.; Miyan, Jaleel A.

    2014-01-01

    The question of how the neural and immune systems interact in host defense is important, integrating a system that senses the whole body with one that protects. Understanding the mechanisms and routes of control could produce novel and powerful ways of promoting and enhancing normal functions as well as preventing or treating abnormal functions. Fragmentation of biological research into specialities has resulted in some failures in recognizing and understanding interactions across different systems and this is most striking across immunology, hematology, and neuroscience. This reductionist approach does not allow understanding of the in vivo orchestrated response generated through integration of all systems. However, many factors make the understanding of multisystem cross-talk in response to a threat difficult, for instance the nervous and immune systems share communication molecules and receptors for a wide range of physiological signals. But, it is clear that physical, hard-wired connections exist between the two systems, with the key link involving sensory, unmyelinated nerve fibers (c fibers) containing the neuropeptide calcitonin gene-related peptide (CGRP), and modified macrophages, mast cells and other immune and host defense cells in various locations throughout the body. In this review we will therefore focus on the induction of CGRP and its key role in the neuroimmune axis. PMID:24592205

  15. Involvement of calcitonin gene-related peptide and receptor component protein in experimental autoimmune encephalomyelitis

    PubMed Central

    Sardi, Claudia; Zambusi, Laura; Finardi, Annamaria; Ruffini, Francesca; Tolun, Adviye A.; Dickerson, Ian M.; Righi, Marco; Zacchetti, Daniele; Grohovaz, Fabio; Provini, Luciano; Furlan, Roberto; Morara, Stefano

    2015-01-01

    Calcitonin Gene-Related Peptide (CGRP) inhibits microglia inflammatory activation in vitro. We here analyzed the involvement of CGRP and Receptor Component Protein (RCP) in experimental autoimmune encephalomyelitis (EAE). Alpha-CGRP deficiency increased EAE scores which followed the scale alpha-CGRP null > heterozygote > wild type. In wild type mice, CGRP delivery into the cerebrospinal fluid (CSF) 1) reduced chronic EAE (C-EAE) signs, 2) inhibited microglia activation (revealed by quantitative shape analysis), and 3) did not alter GFAP expression, cell density, lymphocyte infiltration, and peripheral lymphocyte production of IFN-gamma, TNF-alpha, IL-17, IL-2, and IL-4. RCP (probe for receptor involvement) was expressed in white matter microglia, astrocytes, oligodendrocytes, and vascular-endothelial cells: in EAE, also in infiltrating lymphocytes. In relapsing–remitting EAE (R-EAE) RCP increased during relapse, without correlation with lymphocyte density. RCP nuclear localization (stimulated by CGRP in vitro) was I) increased in microglia and decreased in astrocytes (R-EAE), and II) increased in microglia by CGRP CSF delivery (C-EAE). Calcitonin like receptor was rarely localized in nuclei of control and relapse mice. CGRP increased in motoneurons. In conclusion, CGRP can inhibit microglia activation in vivo in EAE. CGRP and its receptor may represent novel protective factors in EAE, apparently acting through the differential cell-specific intracellular translocationof RCP. PMID:24746422

  16. Distribution of calcitonin gene-related peptide in vertebrate neuromuscular junctions: relationship to the acetylcholine receptor.

    PubMed

    Csillik, B; Tajti, L; Kovács, T; Kukla, E; Rakic, P; Knyihár-Csillik, E

    1993-10-01

    Calcitonin gene-related peptide (CGRP), regarded by several authors to be involved in maintenance of the acetylcholine receptor, is present in the motor axons of various striated rat muscles. It is present, however, only in motor endplates of several selected striated muscles, where it is located in presynaptic axon terminals of neuromuscular junctions. No immunoreactivity could be seen within synaptic vesicles themselves. In the non-human primate Macaca fasciculata, neuromuscular junctions, including those in the diaphragm, display an intense CGRP reaction. The structure of the simian motor endplates is more elaborate than that of the rat. Amphibian motor nerve endings, both in tetanic and tonic muscles, display CGRP immunoreactivity. In tetanic muscles the CGRP reaction outlines "terminaisons en placque" (true motor end plates) and weakly reacting "terminaisons en grappe" (grape-like endings) in tonic muscles. On supramaximal stimulation of the motor nerve, CGRP is depleted from the affected neuromuscular junctions. Wallerian degeneration of the motor axon results in complete disappearance of CGRP. In most rat muscles in which motor endplates do not normally exhibit CGRP immunoreactivity, e.g., the diaphragm and buccinator muscles, the pre-terminal motor axons are CGRP-positive. After immobilization of such muscles by local bupivacaine injection to rats under brief chloral hydrate anesthesia, CGRP immunoreactivity of the neuromuscular junctions can be elicited because blockade of neuromuscular transmission results in accumulation of CGRP in the endplates. Even more striking is the appearance of CGRP immunoreactivity in normally non-reactive motor endplates during axon regeneration after an experimentally induced Wallerian degeneration of the motor axons. We conclude that CGRP is a regular, genotypically determined component of neuromuscular junctions, present either in a manifest or in a latent form. The latter can be elicited by various experimental approaches

  17. Adrenal responses to calcitonin gene-related peptide in conscious hypophysectomized calves.

    PubMed Central

    Bloom, S R; Edwards, A V; Jones, C T

    1989-01-01

    1. Right adrenal and various cardiovascular responses to an intra-aortic infusion of calcitonin gene-related peptide (CGRP; 4 micrograms min-1) have been investigated in the presence and absence of exogenous adrenocorticotrophin ACTH1-24 (2 or 5 ng min-1 kg-1, I.V.). The adrenal clamp technique was employed in conscious calves in which the pituitary stalk had been cauterized 3-7 days previously. 2. At the higher dose (5 ng min-1 kg-1) the I.V. infusion of ACTH raised mean plasma ACTH concentration by about 1000 pg ml-1 and mean right adrenal cortisol output by about 750 ng min-1 kg-1. Under these conditions the intra-aortic infusion of CGRP had no apparent effect on adrenal cortisol output by about 750 ng min-1 kg-1. Under these conditions the intra-aortic infusion of CGRP had no apparent effect on adrenal function, other than to produce moderate adrenal vasodilatation. In contrast, in the absence of exogenous ACTH, the same dose of CGRP produced a substantial rise in cortisol output, which rose steadily to a peak mean value of 409 +/- 31 pg min-1 kg-1 at 10 min. It also significantly inhibited the release of free, but not of total, met5-enkephalin-like immunoreactivity from the gland (P less than 0.001) together with a significantly greater fall in adrenal vascular resistance (P less than 0.001). 3. At the lower dose of ACTH (2 ng min-1 kg-1, I.V.) CGRP raised mean plasma cortisol output from 314 +/- 31 to 486 +/- 44 ng min-1 kg-1 (P less than 0.01) and this effect was not attributable to an increase in the adrenal presentation rate of ACTH. 4. It is concluded that this peptide exerts a steroidogenic action on the adrenal cortex which is manifest in the absence of exogenous ACTH in the functionally hypophysectomized calf. PMID:2555477

  18. Inotropic and lusitropic effects of calcitonin gene-related peptide in the heart

    PubMed Central

    Al-Rubaiee, Mustafa; Gangula, Pandu R.; Millis, Richard M.; Walker, Robin K.; Umoh, Nsini A.; Cousins, Valerie M.; Jeffress, Miara A.

    2013-01-01

    Previous studies have demonstrated positive-inotropic effects of calcitonin gene-related peptide (CGRP), but the mechanisms remain unclear. Therefore, two experiments were performed to determine the physiological correlates of the positive-inotropic effects of CGRP. Treatments designed to antagonize the effects of physiologically active CGRP1–37 included posttreatment with CGRP8–37 and pretreatment with LY-294002 (LY, an inhibitor of phosphatidylinositol 3-kinase), 17β-estradiol (E), and progesterone (P) were also used to modulate the effects of CGRP1–37. Experiment 1 was in vitro studies on sarcomeres and cells of isolated adult rat cardiac myocytes. CGRP1–37, alone and in combination with E and P, decreased sarcomere shortening velocities and increased shortening percentages, effects that were antagonized by CGRP8–37, but not by LY. CGRP1–37 increased resting intracellular calcium ion concentrations and Ca2+ influxes, effects that were also antagonized by both CGRP8–37 and LY. Experiment 2 was in vivo studies on left ventricular pressure-volume (PV) loops. CGRP1–37 increased end-systolic pressure, ejection fraction, and velocities of contraction and relaxation while decreasing stroke volume, cardiac output, stroke work, PV area, and compliance. After partial occlusion of the vena cava, CGRP1–37 increased the slope of the end-systolic PV relationship. CGRP8–37 and LY attenuated most of the CGRP-induced changes. These findings suggest that CGRP-induced positive-inotropic effects may be increased by treatments with estradiol and progesterone and inhibited by LY. The physiological correlates of CGRP-induced positive inotropy observed in rat sarcomeres, cells, and intact hearts are likely to reveal novel mechanisms of heart failure in humans. PMID:23585136

  19. Endosomal proteolysis regulates calcitonin gene-related peptide responses in mesenteric arteries

    PubMed Central

    McNeish, AJ; Roux, BT; Aylett, S-B; Van Den Brink, AM; Cottrell, GS

    2012-01-01

    Background and Purpose Calcitonin gene-related peptide (CGRP) is a potent vasodilator, implicated in the pathogenesis of migraine. CGRP activates a receptor complex comprising, calcitonin receptor-like receptor (CLR) and receptor activity-modifying protein 1 (RAMP1). In vitro studies indicate recycling of CLR•RAMP1 is regulated by degradation of CGRP in early endosomes by endothelin-converting enzyme-1 (ECE-1). However, it is not known if ECE-1 regulates the resensitization of CGRP-induced responses in functional arterial tissue. Experimental Approach CLR, ECE-1a-d and RAMP1 expression in rat mesenteric artery smooth muscle cells (RMA-SMCs) and mesenteric arteries was analysed by RT-PCR and by immunofluorescence and confocal microscopy. CGRP-induced signalling in cells was examined by measuring cAMP production and ERK activation. CGRP-induced relaxation of arteries was measured by isometric wire myography. ECE-1 was inhibited using the specific inhibitor, SM-19712. Key Results RMA-SMCs and arteries contained mRNA for CLR, ECE-1a-d and RAMP1. ECE-1 was present in early endosomes of RMA-SMCs and in the smooth muscle layer of arteries. CGRP induced endothelium-independent relaxation of arteries. ECE-1 inhibition had no effect on initial CGRP-induced responses but reduced cAMP generation in RMA-SMCs and vasodilation in mesenteric arteries responses to subsequent CGRP challenges. Conclusions And Implications ECE-1 regulated the resensitization of responses to CGRP in RMA-SMCs and mesenteric arteries. CGRP-induced relaxation did not involve endothelium-derived pathways. This is the first report of ECE-1 regulating CGRP responses in SMCs and arteries. ECE-1 inhibitors may attenuate an important vasodilatory pathway, implicated in primary headaches and may represent a new therapeutic approach for the treatment of migraine. PMID:22881710

  20. Calcitonin Gene-related Peptide Inhibits Chemokine Production by Human Dermal Microvascular Endothelial Cells

    PubMed Central

    Huang, Jing; Stohl, Lori L.; Zhou, Xi; Ding, Wanhong; Granstein, Richard D.

    2011-01-01

    This study examined whether the sensory neuropeptide calcitonin gene-related peptide (CGRP) inhibits release of chemokines by dermal microvascular endothelial cells. Dermal blood vessels are associated with nerves containing CGRP, suggesting that CGRP-containing nerves may regulate cutaneous inflammation through effects on vessels. We examined CGRP effects on stimulated chemokine production by a human dermal microvascular endothelial cell line (HMEC-1) and primary human dermal microvascular endothelial cells (pHDMECs). HMEC-1 cells and pHDMECs expressed mRNA for components of the CGRP and adrenomedullin receptors and CGRP inhibited LPS-induced production of the chemokines CXCL8, CCL2, and CXCL1 by both HMEC-1 cells and pHDMECs. The receptor activity-modifying protein (RAMP)1/calcitonin receptor-like receptor (CL)-specific antagonists CGRP8-37 and BIBN4096BS, blocked this effect of CGRP in a dose-dependent manner. CGRP prevented LPS-induced IκBα degradation and NF-κB binding to the promoters of CXCL1, CXCL8 and CCL2 in HMEC-1 cells and Bay 11-7085, an inhibitor of NF-κB activation, suppressed LPS-induced production of CXCL1, CXCL8 and CCL2. Thus, the NF-κB pathway appears to be involved in CGRP-mediated suppression of chemokine production. Accordingly, CGRP treatment of LPS-stimulated HMEC-1 cells inhibited their ability to chemoattract human neutrophils and mononuclear cells. Elucidation of this pathway may suggest new avenues for therapeutic manipulation of cutaneous inflammation. PMID:21334428

  1. Skeletal muscle microcirculatory response to rat alpha-calcitonin gene-related peptide.

    PubMed

    Arden, W A; Fiscus, R R; Beihn, L D; Derbin, M; Oremus, R; Gross, D R

    1994-07-01

    We used in vivo video microscopy to determine the effect of increasing doses of rat alpha-calcitonin gene-related peptide (rCGRP) on rat cremaster muscle arterioles in the presence or absence of the nitric oxide synthase inhibitor N-omega-nitro-L-arginine (L-NNA). Male Sprague-Dawley rats (118-148 g) were anaesthetized with pentobarbital, and neurovascularly intact cremaster muscles were imaged. Changes in the diameter, erythrocyte velocity and volume flow in second-(A2), third-(A3), and fourth-(A4) order arterioles were determined. To produce uniform arteriolar tone, the cremaster preparation was challenged with norepinephrine (NE: 10(-7) M). L-NNA (10(-4) M), which was shown to inhibit acetylcholine-(ACh: 10(-6) M) induced arteriolar dilations, was added to 16 of the preparations. Preparations were then challenged by adding cumulative log concentrations of rCGRP (10(-12)-10-7) M; n = 16) or an equivalent volume of vehicle (n = 19) to the bath. Following rCGRP challenge, arterioles were maximally dilated with 10(-5) M nitroprusside (NP). rCGRP caused significant dose-dependent increases in erythrocyte velocity and volume flow in A2 arterioles, and in diameter, velocity, and volume flow in A3 and A4 arterioles, by 10(-8) M, when compared with vehicle-treated controls. L-NNA had no significant effect on rCGRP-induced responses. These data indicate that rCGRP causes dose-dependent dilation of skeletal muscle resistance arterioles at a concentration similar to that observed in larger vessels. This dilation does not appear to be dependent on the vascular production of nitric oxide from L-arginine. PMID:7526261

  2. Calcitonin/calcitonin gene-related peptide transcription unit: tissue-specific expression involves selective use of alternative polyadenylation sites.

    PubMed Central

    Amara, S G; Evans, R M; Rosenfeld, M G

    1984-01-01

    Different 3' coding exons in the rat calcitonin gene are used to generate distinct mRNAs encoding either the hormone calcitonin in thyroidal C-cells or a new neuropeptide referred to as calcitonin gene-related peptide in neuronal tissue, indicating the RNA processing regulation is one strategy used in tissue-specific regulation of gene expression in the brain. Although the two mRNAs use the same transcriptional initiation site and have identical 5' terminal sequences, their 3' termini are distinct. The polyadenylation sites for calcitonin and calcitonin gene-related peptide mRNAs are located at the end of the exons 4 and 6, respectively. Termination of transcription after the calcitonin exon does not dictate the production of calcitonin mRNA, because transcription proceeds through both calcitonin and calcitonin gene-related peptide exons irrespective of which mRNA is ultimately produced. In isolated nuclei, both polyadenylation sites appear to be utilized; however, the proximal (calcitonin) site is preferentially used in nuclei from tissues producing calcitonin mRNA. These data suggest that the mechanism dictating production of each mRNA involves the selective use of alternative polyadenylation sites. Images PMID:6334229

  3. Effect of vasoactive peptides in Tetrahymena: chemotactic activities of adrenomedullin, proadrenomedullin N-terminal 20 peptide (PAMP) and calcitonin gene-related peptide (CGRP).

    PubMed

    Kőhidai, László; Tóth, Katalin; Samotik, Paul; Ranganathan, Kiran; Láng, Orsolya; Tóth, Miklós; Ruskoaho, Heikki

    2016-01-01

    Adrenomedullin (AMD), proadrenomedullin N-terminal 20 peptide (PAMP) and calcitonin gene-related peptide (CGRP) were studied for chemotaxis, chemotactic selection and G-actin/F-actin transition in Tetrahymena. The aim of the experiments was to study the effects of two different peptides encoded by the same gene compared to a peptide related to one of the two, but encoded by a different gene, at a low level of phylogeny. The positive, chemotactic effect of ADM and the strong negative, chemorepellent effect of PAMP suggest that in Tetrahymena, the two peptides elicit their chemotactic effects via different signalling mechanisms. The complexity of swimming behaviour modulated by the three peptides underlines that chemotaxis, chemokinesis and some characteristics of migratory behaviour (velocity, tortuosity) are working as a sub-population level complex functional unit. Chemotactic responsiveness to ADM and CGRP is short-term, in contrast to PAMP, which as a chemorepellent ligand, has the ability to select sub-populations with negative chemotactic responsiveness. The different effects of ADM and PAMP on the polymerization of actin networks show that the microtubular structure of cilia is more essential to chemotactic response than are transitions of the actin network. The results draw attention to the characteristic effects of vasoactive peptides at this low level of phylogeny. PMID:26481478

  4. [The pharmacological mechanism of gastrodin on calcitonin gene-related peptide of cultured rat trigeminal ganglion].

    PubMed

    Luo, Guo-Gang; Fan, Wen-Jing; Yuan, Xing-Yun; Yuan, Bo-Bo; Lü, She-Min; Cao, Yong-Xiao; Xu, Cang-Bao

    2011-12-01

    The Chinese herbal medicine Tianma (Gastrodia elata) has been used for treating and preventing primary headache over thousands of years, but the exact pharmacological mechanism of the main bioactive ingredient gastrodin remains unclear. In present study, the effects of gastrodin on calcitonin gene-related peptide (CGRP) and phosphorylated extracellular signal-regulated kinase1/2 (pERK1/2) expression were observed in rat trigeminal ganglion (TG) after in vitro organ culture to explore the underlying intracellular mechanism of gastrodin on primary vascular-associated headache. CGRP-immunoreactivity (CGRP-ir) positive neurons count, positive area, mean optical density and integrated optical density by means of immunohistochemistry stain were compared at different concentrations of gastrodin, which was separately co-incubated with DMEM in SD rat TG for 24 hours. Only at 5 or 10 mmol L(-1) concentration, gastrodin demonstrated significantly concentration-dependent reduction of CGRP-ir (+) expression and its action closed to 1.2 mmol L(-1) sumatriptan succinate. While at 2.5, 20, and 40 mmol L(-1) concentration, gastrodin did not show remarkable effects on CGRP-ir (+) expression. The optimal concentration of gastrodin (5 and 10 mmol L(-1)) similarly inhibited CGRP-mRNA expression level separately compared with 1.2 mmol L(-1) sumatriptan succinate and 10 micromol L(-1) flunarizine hydrochloride, which was quantitatively analyzed by real-time PCR (RT-PCR). pERK1/2 level was examined by Western blotting after co-cultured with optimal concentration of gastrodin and effective specific ERK1/2 pathway inhibitors PD98059, U0126. The result indicated that gastrodin significantly reduced pERK1/2 protein actions similarly to ERK1/2 pathway specific blockade. It suggests ERK1/2 signaling transduction pathway may be involved in gastrodin intracellular mechanism. This study indicates gastrodin (5 and 10 mmol L(-1)) can remarkably reduce CGRP-ir (+) neuron, CGRP-mRNA and pERK1

  5. Structure of the mouse calcitonin/calcitonin gene-related peptide alpha and beta genes.

    PubMed

    Thomas, P M; Nasonkin, I; Zhang, H; Gagel, R F; Cote, G J

    2001-01-01

    We report the cloning, genomic organization and sequence of the mouse alpha-CALC and beta-CALC genes. The two genes share extensive sequence homology. The transcription units of both genes contain 6 exons. Transcripts of the alpha-CALC gene were found to alternatively include exon 4 or exons 5 and 6. For the beta-CALC gene exon 4 was not detected in transcripts derived from this gene. The predicted mouse alpha-CGRP was found to be identical to rat alpha-CGRP, however, beta-CGRP predicted amino acid sequences revealed three amino acid differences suggesting these residues are not critical to CGRP function. PMID:11761712

  6. Phospholipase C gamma mediates endogenous brain-derived neurotrophic factor-regulated calcitonin gene-related peptide expression in colitis-induced visceral pain

    PubMed Central

    Hashmi, Fiza; Liu, Miao; Shen, Shanwei

    2016-01-01

    Background Visceral hypersensitivity is a complex pathophysiological paradigm with unclear mechanisms. Primary afferent neuronal plasticity marked by alterations in neuroactive compounds such as calcitonin gene-related peptide is suggested to underlie the heightened sensory responses. Signal transduction that leads to calcitonin gene-related peptide expression thereby sensory neuroplasticity during colitis remains to be elucidated. Results In a rat model with colitis induced by 2,4,6-trinitrobenzene sulfonic acid, we found that endogenously elevated brain-derived neurotrophic factor elicited an up-regulation of calcitonin gene-related peptide in the lumbar L1 dorsal root ganglia. At seven days of colitis, neutralization of brain-derived neurotrophic factor with a specific brain-derived neurotrophic factor antibody reversed calcitonin gene-related peptide up-regulation in the dorsal root ganglia. Colitis-induced calcitonin gene-related peptide transcription was also inhibited by brain-derived neurotrophic factor antibody treatment. Signal transduction studies with dorsal root ganglia explants showed that brain-derived neurotrophic factor-induced calcitonin gene-related peptide expression was mediated by the phospholipase C gamma, but not the phosphatidylinositol 3-kinase/Akt or the mitogen-activated protein kinase/extracellular signal-regulated protein kinase pathway. Application of PLC inhibitor U73122 in vivo confirmed that colitis-induced and brain-derived neurotrophic factor-mediated calcitonin gene-related peptide up-regulation in the dorsal root ganglia was regulated by the phospholipase C gamma pathway. In contrast, suppression of the phosphatidylinositol 3-kinase activity in vivo had no effect on colitis-induced calcitonin gene-related peptide expression. During colitis, calcitonin gene-related peptide also co-expressed with phospholipase C gamma but not with p-Akt. Calcitonin gene-related peptide up-regulation during colitis correlated to the activation

  7. Calcitonin gene-related peptide (CGRP), peptide YY (PYY) gastrin releasing peptide (GRP) and others in hamster lung and plasma

    SciTech Connect

    Ekman, R.; Keith, I.M.

    1986-03-05

    Rabbit antisera to CGRP, PYY, neuropeptide Y (NPY) and GRP were used for immunocytochemical localization of these peptides in lungs of neonate hamsters at birth and 6 d of age and young (70 gm) and adult (107 gm) hamsters. The peroxidase-antiperoxidase method was applied to paraffin sections of tissue fixed in Bouin's or Zamboni's solution. Furthermore, radioimmunoassay (RIA) was used to quantify these peptides in lung tissue and plasma from the young hamsters (n=13). Distinct CGRP-like immunoreactivity (IR) was noted in grouped (NEB) and individual (NEC) neuroendocrine cells at all ages including all airways from trachea (NECs only) to alveoli. In some NEBs this IR coexisted with 5-HT-like IR. PYY- and NPY-like Ir was mainly noted in NEBs and NECs at the level of bronchioles and alveoli, and weak GRP-like IR was present in neuroendocrine-like cells of small airways. Measurable quantities of all peptides were recorded by RIA. Females had higher lung and plasma levels of CGRP and plasma levels of PYY than males and tended to have higher lung levels of GRP. The neuropeptides CGRP, PYY and the analog NPY are putative regulators of local pulmonary blood flow by vasodilation (CGRP) and constriction (PYY, NPY), and GRP is known to regulate peptide release.

  8. Microglial content-dependent inhibitory effects of calcitonin gene-related peptide (CGRP) on murine retroviral infection of glial cells.

    PubMed

    Malon, Jennifer T; Grlickova-Duzevik, Eliza; Vaughn, James; Beaulac, Holly; Vunk, Tyler R; Cao, Ling

    2015-02-15

    C57BL/6 (B6) mice develop peripheral neuropathy post-LP-BM5 infection, a murine model of HIV-1 infection, along with the up-regulation of select spinal cord cytokines. We investigated if calcitonin gene-related peptide (CGRP) contributed to the development of peripheral neuropathy by stimulating glial responses. An increased expression of lumbar spinal cord CGRP was observed in vivo, post-LP-BM5 infection. Consequently, in vitro CGRP co-treatments led to a microglial content-dependent attenuation of viral loads in spinal cord mixed glia infected with selected doses of LP-BM5. This inhibition was neither caused by the loss of glia nor induced via the direct inhibition of LP-BM5 by CGRP. PMID:25670002

  9. Effects of Voluntary Locomotion and Calcitonin Gene-Related Peptide on the Dynamics of Single Dural Vessels in Awake Mice

    PubMed Central

    Gao, Yu-Rong

    2016-01-01

    The dura mater is a vascularized membrane surrounding the brain and is heavily innervated by sensory nerves. Our knowledge of the dural vasculature has been limited to pathological conditions, such as headaches, but little is known about the dural blood flow regulation during behavior. To better understand the dynamics of dural vessels during behavior, we used two-photon laser scanning microscopy (2PLSM) to measure the diameter changes of single dural and pial vessels in the awake mouse during voluntary locomotion. Surprisingly, we found that voluntary locomotion drove the constriction of dural vessels, and the dynamics of these constrictions could be captured with a linear convolution model. Dural vessel constrictions did not mirror the large increases in intracranial pressure (ICP) during locomotion, indicating that dural vessel constriction was not caused passively by compression. To study how behaviorally driven dynamics of dural vessels might be altered in pathological states, we injected the vasodilator calcitonin gene-related peptide (CGRP), which induces headache in humans. CGRP dilated dural, but not pial, vessels and significantly reduced spontaneous locomotion but did not block locomotion-induced constrictions in dural vessels. Sumatriptan, a drug commonly used to treat headaches, blocked the vascular and behavioral the effects of CGRP. These findings suggest that, in the awake animal, the diameters of dural vessels are regulated dynamically during behavior and during drug-induced pathological states. SIGNIFICANT STATEMENT The vasculature of the dura has been implicated in the pathophysiology of headaches, but how individual dural vessels respond during behavior, both under normal conditions and after treatment with the headache-inducing peptide calcitonin gene-related peptide (CGRP), is poorly understood. To address these issues, we imaged individual dural vessels in awake mice and found that dural vessels constricted during voluntary locomotion, and

  10. Studies on rat and human thymus to demonstrate immunoreactivity of calcitonin gene-related peptide, tyrosine hydroxylase and neuropeptide Y

    PubMed Central

    KRANZ, ANDREA; KENDALL, MARION D.; VON GAUDECKER, BRITA

    1997-01-01

    The peptidergic and noradrenergic innervation of rat and human thymus was investigated by immunohistochemistry at the light and electron microscopical level (avidin-biotin-complex, sucrose-phosphate-glyoxylic-acid, and immunogold techniques). The distribution of noradrenergic neural profiles, and positive immunoreactivity for calcitonin gene-related peptide (CGRP), tyrosine hydroxylase (TH) and neuropeptide Y (NPY) is described in female rats during ageing, and in human children. In the neonatal rat thymus, the arteries and septa are well supplied by fine varicose nerves. In older animals (2 wk–1 y) the number of septa and blood vessels increase and consequently also the innervation. No nerves were found in the cortex. Apart from the innervation of the septal areas, immunoreactivity for CGRP and TH was present in thymic cells. Except for the young rats (neonatal–14 d), all rats showed CGRP positivity in subcapsular/perivascular epithelial cells (type 1 cells). All rat thymuses also contained a few TH positive cells in the medulla, which could only be confirmed as epithelial cells (type 6 cells) in children. Type 1 cells in the human thymus were not CGRP positive, but as in the rat, there were similar TH positive cells in the medulla. It was concluded that in addition to nerves containing CGRP, noradrenaline or dopamine, epithelial cells also contain these transmitters. They could therefore act on different cells (compared with neural targets) in a paracrine manner. PMID:9419001

  11. Control of calcitonin/calcitonin gene-related peptide pre-mRNA processing by constitutive intron and exon elements.

    PubMed Central

    Yeakley, J M; Hedjran, F; Morfin, J P; Merillat, N; Rosenfeld, M G; Emeson, R B

    1993-01-01

    The calcitonin/calcitonin gene-related peptide (CGRP) primary transcript is alternatively spliced in thyroid C cells and neurons, resulting in the tissue-specific production of calcitonin and CGRP mRNAs. Analyses of mutated calcitonin/CGRP transcription units in permanently transfected cell lines have indicated that alternative splicing is regulated by a differential capacity to utilize the calcitonin-specific splice acceptor. The analysis of an extensive series of mutations suggests that tissue-specific regulation of calcitonin mRNA production does not depend on the presence of a single, unique cis-active element but instead appears to be a consequence of suboptimal constitutive splicing signals. While only those mutations that altered constitutive splicing signals affected splice choices, the action of multiple regulatory sequences cannot be formally excluded. Further, we have identified a 13-nucleotide purine-rich element from a constitutive exon that, when placed in exon 4, entirely switches splice site usage in CGRP-producing cells. These data suggest that specific exon recruitment sequences, in combination with other constitutive elements, serve an important function in exon recognition. These results are consistent with the hypothesis that tissue-specific alternative splicing of the calcitonin/CGRP primary transcript is mediated by cell-specific differences in components of the constitutive splicing machinery. Images PMID:8413203

  12. Role of calcitonin gene-related peptide in cerebral vasospasm, and as a therapeutic approach to subarachnoid hemorrhage

    PubMed Central

    Kokkoris, Stelios; Andrews, Peter; Webb, David J.

    2012-01-01

    Calcitonin gene-related peptide (CGRP) is one of the most potent microvascular vasodilators identified to date. Vascular relaxation and vasodilation is mediated via activation of the CGRP receptor. This atypical receptor is made up of a G protein-coupled receptor called calcitonin receptor-like receptor (CLR), a single transmembrane protein called receptor activity-modifying protein (RAMP), and an additional protein that is required for Gas coupling, known as receptor component protein (RCP). Several mechanisms involved in CGRP-mediated relaxation have been identified. These include nitric oxide (NO)-dependent endothelium-dependent mechanisms or cAMP-mediated endothelium-independent pathways; the latter being more common. Subarachnoid hemorrhage (SAH) is associated with cerebral vasoconstriction that occurs several days after the hemorrhage and is often fatal. The vasospasm occurs in 30–40% of patients and is the major cause of death from this condition. The vasoconstriction is associated with a decrease in CGRP levels in nerves and an increase in CGRP levels in draining blood, suggesting that CGRP is released from nerves to oppose the vasoconstriction. This evidence has led to the concept that exogenous CGRP may be beneficial in a condition that has proven hard to treat. The present article reviews: (a) the pathophysiology of delayed ischemic neurologic deficit after SAH (b) the basics of the CGRP receptor structure, signal transduction, and vasodilatation mechanisms and (c) the studies that have been conducted so far using CGRP in both animals and humans with SAH. PMID:23162536

  13. Calcitonin Gene-Related Peptide Enhances Release of Native Brain-Derived Neurotrophic Factor from Trigeminal Ganglion Neurons

    PubMed Central

    Buldyrev, Ilya; Tanner, Nathan M.; Hsieh, Hui-ya; Dodd, Emily G.; Nguyen, Loi T.; Balkowiec, Agnieszka

    2008-01-01

    Activity-dependent plasticity in nociceptive pathways has been implicated in pathomechanisms of chronic pain syndromes. Calcitonin gene-related peptide (CGRP), which is expressed by trigeminal nociceptors, has recently been identified as a key player in the mechanism of migraine headaches. Here we show that CGRP is co-expressed with brain-derived neurotrophic factor (BDNF) in a large subset of adult rat trigeminal ganglion neurons in vivo. Using ELISA in situ, we show that CGRP (1–1000 nM) potently enhances BDNF release from cultured trigeminal neurons. The effect of CGRP is dose–dependent and abolished by pretreatment with CGRP receptor antagonist, CGRP(8–37). Intriguingly, CGRP-mediated BDNF release, unlike BDNF release evoked by physiological patterns of electrical stimulation, is independent of extracellular calcium. Depletion of intracellular calcium stores with thapsigargin blocks the CGRP-mediated BDNF release. Using transmission electron microscopy, our study also shows that BDNF-immunoreactivity is present in dense core vesicles of unmyelinated axons and axon terminals in the subnucleus caudalis of the spinal trigeminal nucleus, the primary central target of trigeminal nociceptors. Together, these results reveal a previously unknown role for CGRP in regulating BDNF availability, and point to BDNF as a candidate mediator of trigeminal nociceptive plasticity. PMID:17064360

  14. Calcitonin gene-related peptide enhances release of native brain-derived neurotrophic factor from trigeminal ganglion neurons.

    PubMed

    Buldyrev, Ilya; Tanner, Nathan M; Hsieh, Hui-ya; Dodd, Emily G; Nguyen, Loi T; Balkowiec, Agnieszka

    2006-12-01

    Activity-dependent plasticity in nociceptive pathways has been implicated in pathomechanisms of chronic pain syndromes. Calcitonin gene-related peptide (CGRP), which is expressed by trigeminal nociceptors, has recently been identified as a key player in the mechanism of migraine headaches. Here we show that CGRP is coexpressed with brain-derived neurotrophic factor (BDNF) in a large subset of adult rat trigeminal ganglion neurons in vivo. Using ELISA in situ, we show that CGRP (1-1000 nM) potently enhances BDNF release from cultured trigeminal neurons. The effect of CGRP is dose-dependent and abolished by pretreatment with CGRP receptor antagonist, CGRP(8-37). Intriguingly, CGRP-mediated BDNF release, unlike BDNF release evoked by physiological patterns of electrical stimulation, is independent of extracellular calcium. Depletion of intracellular calcium stores with thapsigargin blocks the CGRP-mediated BDNF release. Using transmission electron microscopy, our study also shows that BDNF-immunoreactivity is present in dense core vesicles of unmyelinated axons and axon terminals in the subnucleus caudalis of the spinal trigeminal nucleus, the primary central target of trigeminal nociceptors. Together, these results reveal a previously unknown role for CGRP in regulating BDNF availability, and point to BDNF as a candidate mediator of trigeminal nociceptive plasticity. PMID:17064360

  15. Calcitonin Gene-Related Peptide Modulates Heat Nociception in the Human Brain - An fMRI Study in Healthy Volunteers

    PubMed Central

    Asghar, Mohammad Sohail; Becerra, Lino; Larsson, Henrik B. W.; Borsook, David; Ashina, Messoud

    2016-01-01

    Background Intravenous infusion of calcitonin-gene-related-peptide (CGRP) provokes headache and migraine in humans. Mechanisms underlying CGRP-induced headache are not fully clarified and it is unknown to what extent CGRP modulates nociceptive processing in the brain. To elucidate this we recorded blood-oxygenation-level-dependent (BOLD) signals in the brain by functional MRI after infusion of CGRP in a double-blind placebo-controlled crossover study of 27 healthy volunteers. BOLD-signals were recorded in response to noxious heat stimuli in the V1-area of the trigeminal nerve. In addition, we measured BOLD-signals after injection of sumatriptan (5-HT1B/1D antagonist). Results Brain activation to noxious heat stimuli following CGRP infusion compared to baseline resulted in increased BOLD-signal in insula and brainstem, and decreased BOLD-signal in the caudate nuclei, thalamus and cingulate cortex. Sumatriptan injection reversed these changes. Conclusion The changes in BOLD-signals in the brain after CGRP infusion suggests that systemic CGRP modulates nociceptive transmission in the trigeminal pain pathways in response to noxious heat stimuli. PMID:26990646

  16. Cells showing immunoreactivity for calcitonin or calcitonin gene-related peptide (CGRP) in the central nervous system of some invertebrates.

    PubMed

    Sasayama, Y; Katoh, A; Oguro, C; Kambegawa, A; Yoshizawa, H

    1991-09-01

    In the central nervous system of some species of several invertebrate phyla, including land planarians (Platyhelminthes), ribbon worms (Nemertina), slugs (Mollusca), polychaetes, earthworms and leeches (Annelida), pill bugs (Arthropoda), and beard worms (Pogonophora), salmon calcitonin-immunoreactive cells and rat calcitonin gene-related peptide (CGRP)-immunoreactive cells were found by immunohistochemistry. These immunoreactive cells were located in the region surrounding the neuropile, although the sizes of the cells varied according to species. Some of them were round or polygonal and regarded as apolar nerve cells because of their lack of cytoplasmic processes, whereas others were spindle-shaped or elongated, being comparable with unipolar nerve cells because of extension of their cytoplasmic processes in the direction of the neuropile. In some cases, it was noted that the cytoplasmic processes had complicated branches or formed loop-like structures at their ends. These observations suggest that a calcitonin-like or CGRP-like substance is extensively present in invertebrates as well as vertebrates. PMID:1936921

  17. Reductions in calcitonin gene-related peptide may be associated with the impairment of the contralateral testis in unilateral cryptorchidism

    PubMed Central

    ZHU, BAOPING; LIU, QING; LIN, LI; ZHENG, XINMIN

    2015-01-01

    The aim of the present study was to investigate the mechanism underlying the impairment of the contralateral testis in unilateral cryptorchidism in experimental rats using a molecular neurophysiological approach. Thirty-six male rats (21 days old) were divided into a cryptorchidism group, a cryptorchidism with division of the genitofemoral nerve (GFN) group and a control group (n=12/group). The distribution of the calcitonin gene-related peptide (CGRP) immunoreactive nerve fibers in the testes was studied using an immunohistochemistry technique. Germ cell apoptosis was detected using the terminal deoxynucleotidyl-transferase-mediated dUTP nick end labeling method. The concentration of malondialdehyde (MDA) in the testis tissue was evaluated using a spectrophotometric determination method, and the ultrastructure of Sertoli cells was observed using transmission electron microscopy. It was found that, 100 days after the surgery, the concentration of CGRP in the cryptorchidism group was decreased significantly, whereas the levels of MDA and the number of apoptotic germ cells were increased significantly compared with the control group (P<0.01). Following the division of the GFN, the damaging effects were decreased (P<0.01). The impairment mechanism may therefore be associated with a reduction in the level of CGRP in the contralateral testis. The reflex decrease in CGRP may be caused by germ cell apoptosis, decreased blood flow and oxygen levels, and the increase in reactive oxygen free radicals and lipid peroxidation. PMID:26136895

  18. Interaction of histamine and calcitonin gene-related peptide in the formalin induced pain perception in rats.

    PubMed

    Mobarakeh, Jalal Izadi; Torkaman-Boutorabi, Anahita; Rahimi, Amir Abbas; Ghasri, Shahrooz; Nezhad, Reza Mohammad Ali; Hamzely, Arash; Khoshkholgh Sima, Baharak; Takahashi, Kazuhiro; Nunoki, Kazuo; Yanai, Kazuhiko

    2011-06-01

    Histamine and calcitonin gene-related peptide (CGRP) contribute to the pain perception. The aim of the present study is to clarify the interaction of histamine and CGRP in the perception of inflammatory pain. The effects of a histamine H1 receptor antagonist (pyrilamine, i.p.), an H2 receptor antagonist (ranitidine, i.p.) and a CGRP antagonist (CGRP 8-37, i.t.) on the formalininduced pain was studied in rats. Pyrilamine and ranitidine produced a dose-dependent antinociceptive response in the first and the second phases of the formalin test. A single administration of pyrilamine (1 mg/kg, i.p.), ranitidine (10 mg/kg, i.p.) or CGRP 8-37 (10 µg/µL, i.t.) had no significant effects on the pain perception in the second phase. A combination of CGRP 8-37 and pyrilamine or ranitidine at these sub-effective doses, however, showed nociceptive response in the second phase. Moreover, a histamine (i.t.)-induced hyperalgesia was completely prevented by treatment with GGRP 8-37 at this dose. Our findings have raised the possibility that the CGRP system has interaction with histamine in the perception of inflammatory pain. PMID:21673449

  19. Calcitonin gene-related peptide activated ATP-sensitive K+ currents in rabbit arterial smooth muscle via protein kinase A.

    PubMed Central

    Quayle, J M; Bonev, A D; Brayden, J E; Nelson, M T

    1994-01-01

    1. Whole-cell K+ currents activated by calcitonin gene-related peptide (CGRP) in smooth muscle cells enzymatically isolated from rabbit mesenteric arteries were measured in the conventional and perforated configurations of the patch clamp technique. The signal transduction pathway from CGRP receptors to activation of potassium currents was investigated. 2. CGRP (10 nM) activated a whole-cell current that was blocked by glibenclamide (10 microM), an inhibitor of ATP-sensitive K+ channels. Elevating intracellular ATP reduced glibenclamide-sensitive currents. CGRP increased the glibenclamide-sensitive currents by 3- to 6-fold in cells dialysed with 0.1 mM ATP, 3.0 mM ATP or in intact cells. The reversal potential of the glibenclamide-sensitive current in the presence of CGRP shifted with the potassium equilibrium potential, while its current-voltage relationship exhibited little voltage dependence. 3. Forskolin (10 microM), an adenylyl cyclase activator, Sp-cAMPS (500 microM) and the catalytic subunit of protein kinase A increased glibenclamide-sensitive K+ currents 2.1-, 3.3- and 8.2-fold, respectively. 4. Nitric oxide and nitroprusside did not activate glibenclamide-sensitive K+ currents. 5. Dialysis of the cell's interior with inhibitors of protein kinase A (synthetic peptide inhibitor, 4.6 microM or H-8, 100 microM) completely blocked activation of K+ currents by CGRP. 6. Our results suggest the following signal transduction scheme for activation of K+ currents by CGRP in arterial smooth muscle: (1) CGRP stimulates adenylyl cyclase, which leads to an elevation of cAMP; (2) cAMP activates protein kinase A, which opens ATP-sensitive K+ channels. PMID:8189394

  20. Pharmacologic Characterization of AMG 334, a Potent and Selective Human Monoclonal Antibody against the Calcitonin Gene-Related Peptide Receptor.

    PubMed

    Shi, Licheng; Lehto, Sonya G; Zhu, Dawn X D; Sun, Hong; Zhang, Jianhua; Smith, Brian P; Immke, David C; Wild, Kenneth D; Xu, Cen

    2016-01-01

    Therapeutic agents that block the calcitonin gene-related peptide (CGRP) signaling pathway are a highly anticipated and promising new drug class for migraine therapy, especially after reports that small-molecule CGRP-receptor antagonists are efficacious for both acute migraine treatment and migraine prevention. Using XenoMouse technology, we successfully generated AMG 334, a fully human monoclonal antibody against the CGRP receptor. Here we show that AMG 334 competes with [(125)I]-CGRP binding to the human CGRP receptor, with a Ki of 0.02 nM. AMG 334 fully inhibited CGRP-stimulated cAMP production with an IC50 of 2.3 nM in cell-based functional assays (human CGRP receptor) and was 5000-fold more selective for the CGRP receptor than other human calcitonin family receptors, including adrenomedullin, calcitonin, and amylin receptors. The potency of AMG 334 at the cynomolgus monkey (cyno) CGRP receptor was similar to that at the human receptor, with an IC50 of 5.7 nM, but its potency at dog, rabbit, and rat receptors was significantly reduced (>5000-fold). Therefore, in vivo target coverage of AMG 334 was assessed in cynos using the capsaicin-induced increase in dermal blood flow model. AMG 334 dose-dependently prevented capsaicin-induced increases in dermal blood flow on days 2 and 4 postdosing. These results indicate AMG 334 is a potent, selective, full antagonist of the CGRP receptor and show in vivo dose-dependent target coverage in cynos. AMG 334 is currently in clinical development for the prevention of migraine. PMID:26559125

  1. Immunohistochemical localization of the calcitonin gene-related peptide binding site in the primate trigeminovascular system using functional antagonist antibodies.

    PubMed

    Miller, Silke; Liu, Hantao; Warfvinge, Karin; Shi, Licheng; Dovlatyan, Mary; Xu, Cen; Edvinsson, Lars

    2016-07-22

    Calcitonin gene-related peptide (CGRP) is a potent vasodilator and a neuromodulator implicated in the pathophysiology of migraine. It binds to the extracellular domains of calcitonin receptor-like receptor (CLR) and receptor activity-modifying protein (RAMP) 1 that together form the CGRP receptor. Antagonist antibodies against CGRP and its binding site at the receptor are clinically effective in preventing migraine attacks. The blood-brain barrier penetration of these antagonist antibodies is limited, suggesting that a potential peripheral site of action is sufficient to prevent migraine attacks. To further understand the sites of CGRP-mediated signaling in migraine, we used immunohistochemical staining with recently developed antagonist antibodies specifically recognizing a fusion protein of the extracellular domains of RAMP1 and CLR that comprise the CGRP binding pocket at the CGRP receptor in monkey and man. We confirmed binding of the antagonist antibodies to human vascular smooth muscle cells (VSMCs) of dural meningeal arteries and neurons in the trigeminal ganglion, both of which are likely sites of action for therapeutic antibodies in migraine patients. We further used one of these antibodies for detailed mapping on cynomolgus monkey tissue and found antagonist antibody binding sites at multiple levels in the trigeminovascular system: in the dura mater VSMCs, in neurons and satellite glial cells in the trigeminal ganglion, and in neurons in the spinal trigeminal nucleus caudalis. These data reinforce and clarify our understanding of CGRP receptor localization in a pattern consistent with a role for CGRP receptors in trigeminal sensitization and migraine pathology. PMID:27155150

  2. Calcitonin gene-related peptide inhibits autophagic-lysosomal proteolysis through cAMP/PKA signaling in rat skeletal muscles.

    PubMed

    Machado, Juliano; Manfredi, Leandro H; Silveira, Wilian A; Gonçalves, Dawit A P; Lustrino, Danilo; Zanon, Neusa M; Kettelhut, Isis C; Navegantes, Luiz C

    2016-03-01

    Calcitonin gene-related peptide (CGRP) is a neuropeptide released by motor neuron in skeletal muscle and modulates the neuromuscular transmission by induction of synthesis and insertion of acetylcholine receptor on postsynaptic muscle membrane; however, its role in skeletal muscle protein metabolism remains unclear. We examined the in vitro and in vivo effects of CGRP on protein breakdown and signaling pathways in control skeletal muscles and muscles following denervation (DEN) in rats. In isolated muscles, CGRP (10(-10) to 10(-6)M) reduced basal and DEN-induced activation of overall proteolysis in a concentration-dependent manner. The in vitro anti-proteolytic effect of CGRP was completely abolished by CGRP8-37, a CGRP receptor antagonist. CGRP down-regulated the lysosomal proteolysis, the mRNA levels of LC3b, Gabarapl1 and cathepsin L and the protein content of LC3-II in control and denervated muscles. In parallel, CGRP elevated cAMP levels, stimulated PKA/CREB signaling and increased Foxo1 phosphorylation in both conditions. In denervated muscles and starved C2C12 cells, Rp-8-Br-cAMPs or PKI, two PKA inhibitors, completely abolished the inhibitory effect of CGRP on Foxo1, 3 and 4 and LC3 lipidation. A single injection of CGRP (100 μg kg(-1)) in denervated rats increased the phosphorylation levels of CREB and Akt, inhibited Foxo transcriptional activity, the LC3 lipidation as well as the mRNA levels of LC3b and cathepsin L, two bona fide targets of Foxo. This study shows for the first time that CGRP exerts a direct inhibitory action on autophagic-lysosomal proteolysis in control and denervated skeletal muscle by recruiting cAMP/PKA signaling, effects that are related to inhibition of Foxo activity and LC3 lipidation. PMID:26718975

  3. Demonstration of the neurotransmitter role of calcitonin gene-related peptides (CGRP) by immunoblockade with anti-CGRP monoclonal antibodies.

    PubMed Central

    Tan, K. K.; Brown, M. J.; Longmore, J.; Plumpton, C.; Hill, R. G.

    1994-01-01

    1. Monoclonal antibodies (MAbs) against rat alpha-calcitonin gene-related peptide (alpha CGRP) were produced. Those which bound CGRP in a radioimmunoassay and inhibited the binding of 2-[125I]-iodohistidyl10-CGRP in a receptor binding assay were selected for immunoblockade experiments. 2. The effect of MAbs on CGRP inhibition of electrically stimulated contractions of the rat isolated vas deferens was characterized. Four out of 11 MAbs tested shifted the concentration-response curve of CGRP to the right compared with vehicle or irrelevant MAb control. MAb C4.19 produced equipotent blockade of rat alpha CGRP and rat beta CGRP and was chosen for further studies. MAb C4.19 had no pharmacologically significant effect on the concentration-response relationship of isoprenaline, rat beta-endorphin or somatostatin. 3. We demonstrated that the pharmacological response to CGRP in the presence of MAb C4.19 could be predicted when the dissociation constant and concentration of binding sites of the antibody were known. Comparison of experimental and computer simulated data showed good agreement for EC50 and maximum effect of CGRP in the presence of MAb C4.19. 4. Capsaicin at 1 microM inhibited the electrically stimulated contractions by 60.8% (95% confidence interval 51.8% to 69.9%). This effect was significantly attenuated by MAb C4.19 to 26.0% (95% confidence interval 15.2% to 36.8%; P < 0.003). 5. The immunoblockade of exogenous and endogenous CGRP described here, together with complementary evidence from other studies, strongly suggest that CGRP has a major neurotransmitter role at the neuroeffector junction of the rat vas deferens. PMID:7912623

  4. Nitric oxide synthase inhibitors can antagonize neurogenic and calcitonin gene-related peptide induced dilation of dural meningeal vessels

    PubMed Central

    Akerman, S; Williamson, D J; Kaube, H; Goadsby, P J

    2002-01-01

    The detailed pathophysiology of migraine is beginning to be understood and is likely to involve activation of trigeminovascular afferents. Clinically effective anti-migraine compounds are believed to have actions that include peripheral inhibition of calcitonin gene-related peptide (CGRP) release from trigeminal neurones, or preventing dural vessel dilation, or both. CGRP antagonists can block both neurogenic and CGRP-induced dural vessel dilation. Nitric oxide (NO) can induce headache in migraine patients and often triggers a delayed migraine. The initial headache is thought to be caused via a direct action of the NO–cGMP pathway that causes vasodilation by vascular smooth muscle relaxation, while the delayed headache is likely to be a result of triggering trigeminovascular activation. Nitric oxide synthase (NOS) inhibitors are effective in the treatment of acute migraine. The present studies used intravital microscopy to examine the effects of specific NOS inhibitors on neurogenic dural vasodilation (NDV) and CGRP-induced dilation. The non-specific and neuronal NOS (nNOS) inhibitors were able to partially inhibit NDV, while the non-specific and endothelial NOS (eNOS) inhibitors were able to partially inhibit the CGRP induced dilation. There was no effect of the inducible NOS (iNOS) inhibitor. The data suggest that the delayed headache response triggered by NO donors in humans may be due, in part, to increased nNOS activity in the trigeminal system that causes CGRP release and dural vessel dilation. Further, eNOS activity in the endothelium causes NO production and smooth muscle relaxation by direct activation of the NO–cGMP pathway, and may be involved in the initial headache response. PMID:12183331

  5. Identification of specific calcitonin-like receptor residues important for calcitonin gene-related peptide high affinity binding

    PubMed Central

    Banerjee, Sugato; Evanson, Janel; Harris, Erik; Lowe, Stephen L; Thomasson, Kathryn A; Porter, James E

    2006-01-01

    Background Calcitonin gene-related peptide (CGRP) is a vasoactive neuropeptide whose biological activity has potential therapeutic value for many vascular related diseases. CGRP is a 37 amino acid neuropeptide that signals through a G protein-coupled receptor belonging to the secretin receptor family. Previous studies on the calcitonin-like receptor (CLR), which requires co-expression of the receptor-activity-modifying protein-1 (RAMP1) to function as a CGRP receptor, have shown an 18 amino acid N-terminus sequence important for binding CGRP. Moreover, several investigations have recognized the C-terminal amidated phenylalanine (F37) of CGRP as essential for docking to the mature receptor. Therefore, we hypothesize that hydrophobic amino acids within the previously characterized 18 amino acid CLR N-terminus domain are important binding contacts for the C-terminal phenylalaninamide of CGRP. Results Two leucine residues within this previously characterized CLR N-terminus domain, when mutated to alanine and expressed on HEK293T cells stably transfected with RAMP1, demonstrated a significantly decreased binding affinity for CGRP compared to wild type receptor. Additional decreases in binding affinity for CGRP were not found when both leucine mutations were expressed in the same CLR construct. Decreased binding characteristic of these leucine mutant receptors was observed for all CGRP ligands tested that contained the necessary amidated phenylalanine at their C-terminus. However, there was no difference in the potency of CGRP to increase cAMP production by these leucine mutant receptors when compared to wild type CLR, consistent with the notion that the neuropeptide C-terminal F37 is important for docking but not activation of the receptor. This observation was conserved when modified CGRP ligands lacking the amidated F37 demonstrated similar potencies to generate cAMP at both wild type and mutant CLRs. Furthermore, these modified CGRP ligands displayed a significant

  6. The peptide semax affects the expression of genes related to the immune and vascular systems in rat brain focal ischemia: genome-wide transcriptional analysis

    PubMed Central

    2014-01-01

    Background The nootropic neuroprotective peptide Semax (Met-Glu-His-Phe-Pro-Gly-Pro) has proved efficient in the therapy of brain stroke; however, the molecular mechanisms underlying its action remain obscure. Our genome-wide study was designed to investigate the response of the transcriptome of ischemized rat brain cortex tissues to the action of Semax in vivo. Results The gene-expression alteration caused by the action of the peptide Semax was compared with the gene expression of the “ischemia” group animals at 3 and 24 h after permanent middle cerebral artery occlusion (pMCAO). The peptide predominantly enhanced the expression of genes related to the immune system. Three hours after pMCAO, Semax influenced the expression of some genes that affect the activity of immune cells, and, 24 h after pMCAO, the action of Semax on the immune response increased considerably. The genes implicated in this response represented over 50% of the total number of genes that exhibited Semax-induced altered expression. Among the immune-response genes, the expression of which was modulated by Semax, genes that encode immunoglobulins and chemokines formed the most notable groups. In response to Semax administration, 24 genes related to the vascular system exhibited altered expression 3 h after pMCAO, whereas 12 genes were changed 24 h after pMCAO. These genes are associated with such processes as the development and migration of endothelial tissue, the migration of smooth muscle cells, hematopoiesis, and vasculogenesis. Conclusions Semax affects several biological processes involved in the function of various systems. The immune response is the process most markedly affected by the drug. Semax altered the expression of genes that modulate the amount and mobility of immune cells and enhanced the expression of genes that encode chemokines and immunoglobulins. In conditions of rat brain focal ischemia, Semax influenced the expression of genes that promote the formation and

  7. Calcitonin gene-related peptide promotes the expression of osteoblastic genes and activates the WNT signal transduction pathway in bone marrow stromal stem cells

    PubMed Central

    ZHOU, RI; YUAN, ZHI; LIU, JIERONG; LIU, JIAN

    2016-01-01

    Calcitonin gene-related peptide (CGRP) is known to induce osteoblastic differentiation and alkaline phosphatase activity in bone marrow stromal stem cells (BMSCs). However, it has remained elusive whether this effect is mediated by CGRP receptors directly or whether other signaling pathways are involved. The present study assessed the possible involvement of the Wnt/β-catenin signaling pathway in the activation of CGRP signaling during the differentiation of BMSCs. First, the differentiation of BMSCs was induced in vitro and the expression of CGRP receptors was examined by western blot analysis. The effects of exogenous CGRP and LiCl, a stimulator of the Wnt/β-catenin signaling pathway, on the osteoblastic differentiation of BMSCs were assessed; furthermore, the expression of mRNA and proteins involved in the Wnt/β-catenin signaling pathway was assessed using quantitative PCR and western blot analyses. The results revealed that CGRP receptors were expressed throughout the differentiation of BMSCs, at days 7 and 14. Incubation with CGRP and LiCl led to the upregulation of the expression of osteoblastic genes associated with the Wnt/β-catenin pathway, including the mRNA of c-myc, cyclin D1, Lef1, Tcf7 and β-catenin as well as β-catenin protein. However, the upregulation of these genes and β-catenin protein was inhibited by CGRP receptor antagonist or secreted frizzled-related protein, an antagonist of the Wnt/β-catenin pathway. The results of the present study therefore suggested that the Wnt/β-catenin signaling pathway may be involved in CGRP- and LiCl-promoted osteoblastic differentiation of BMSCs. PMID:27082317

  8. Calcitonin gene-related peptide promotes the expression of osteoblastic genes and activates the WNT signal transduction pathway in bone marrow stromal stem cells.

    PubMed

    Zhou, Ri; Yuan, Zhi; Liu, Jierong; Liu, Jian

    2016-06-01

    Calcitonin gene-related peptide (CGRP) is known to induce osteoblastic differentiation and alkaline phosphatase activity in bone marrow stromal stem cells (BMSCs). However, it has remained elusive whether this effect is mediated by CGRP receptors directly or whether other signaling pathways are involved. The present study assessed the possible involvement of the Wnt/β‑catenin signaling pathway in the activation of CGRP signaling during the differentiation of BMSCs. First, the differentiation of BMSCs was induced in vitro and the expression of CGRP receptors was examined by western blot analysis. The effects of exogenous CGRP and LiCl, a stimulator of the Wnt/β‑catenin signaling pathway, on the osteoblastic differentiation of BMSCs were assessed; furthermore, the expression of mRNA and proteins involved in the Wnt/β‑catenin signaling pathway was assessed using quantitative PCR and western blot analyses. The results revealed that CGRP receptors were expressed throughout the differentiation of BMSCs, at days 7 and 14. Incubation with CGRP and LiCl led to the upregulation of the expression of osteoblastic genes associated with the Wnt/β‑catenin pathway, including the mRNA of c‑myc, cyclin D1, Lef1, Tcf7 and β‑catenin as well as β‑catenin protein. However, the upregulation of these genes and β‑catenin protein was inhibited by CGRP receptor antagonist or secreted frizzled‑related protein, an antagonist of the Wnt/β‑catenin pathway. The results of the present study therefore suggested that the Wnt/β-catenin signaling pathway may be involved in CGRP‑ and LiCl-promoted osteoblastic differentiation of BMSCs. PMID:27082317

  9. Diverse Physiological Roles of Calcitonin Gene-Related Peptide in Migraine Pathology: Modulation of Neuronal-Glial-Immune Cells to Promote Peripheral and Central Sensitization.

    PubMed

    Durham, Paul L

    2016-08-01

    The neuropeptide calcitonin gene-related peptide (CGRP) is implicated in the underlying pathology of migraine by promoting the development of a sensitized state of primary and secondary nociceptive neurons. The ability of CGRP to initiate and maintain peripheral and central sensitization is mediated by modulation of neuronal, glial, and immune cells in the trigeminal nociceptive signaling pathway. There is accumulating evidence to support a key role of CGRP in promoting cross excitation within the trigeminal ganglion that may help to explain the high co-morbidity of migraine with rhinosinusitis and temporomandibular joint disorder. In addition, there is emerging evidence that CGRP facilitates and sustains a hyperresponsive neuronal state in migraineurs mediated by reported risk factors such as stress and anxiety. In this review, the significant role of CGRP as a modulator of the trigeminal system will be discussed to provide a better understanding of the underlying pathology associated with the migraine phenotype. PMID:27334137

  10. Calcitonin gene-related peptide contributes to peripheral nerve injury-induced mechanical hypersensitivity through CCL5 and p38 pathways.

    PubMed

    Malon, Jennifer T; Cao, Ling

    2016-08-15

    The role of calcitonin gene related peptide (CGRP) in neuropathic pain was investigated in a mouse model of neuropathic pain, spinal nerve L5 transection (L5Tx). Intrathecal injection (i.t.) of CGRP8-37, a CGRP antagonist, significantly reduced L5Tx-induced mechanical hypersensitivity and lumbar spinal cord CCL5 expression. i.t. injection of a CCL5 neutralizing antibody significantly inhibited L5Tx-induced mechanical hypersensitivity. Further, pre-treatment with a p38-inhibitor, SB203580, was able to reduce CGRP-induced mechanical hypersensitivity, but not CGRP-induced CCL5 production. Our data indicate that CGRP can play its pro-nociceptive role through both a spinal cord CCL5-dependent, p38-independent pathway, and a p38-depenented, CCL5-independent pathway. PMID:27397078

  11. Effects of rizatriptan on the expression of calcitonin gene-related peptide and cholecystokinin in the periaqueductal gray of a rat migraine model.

    PubMed

    Yao, Gang; Han, Ximei; Hao, Tingting; Huang, Qian; Yu, Tingmin

    2015-02-01

    Triptans are serotonin 5-hydroxytryptamine receptor 1B/D agonists that are highly effective in the treatment of migraine. We previously found that rizatriptan can reduce the expression of proenkephalin and P substance in the rat midbrain, suggesting that rizatriptan may exert its analgesic effects by influencing the endogenous pain modulatory system. Calcitonin gene-related peptide (CGRP) and cholecystokinin (CCK) are mainly responsible for antagonizing the analgesic effects of opioid peptides in the endogenous pain modulatory system. In this study, we investigated the effects of rizatriptan on the expression of CGRP and CCK in the periaqueductal gray (PAG), a key structure of the endogenous pain modulatory system, in a rat migraine model induced by nitroglycerin. We found that the mRNA and protein levels of CGRP and CCK in the PAG of migraine rats were significantly increased compared to those in control rats, and these levels were significantly reduced upon treatment with rizatriptan in migraine rats (P<0.05). Our results suggest that the expression of CGRP and CCK in the endogenous pain modulatory system may be increased during migraine attacks, which further antagonizes the analgesic effects of endogenous opioid peptides and induces sustained migraine. Rizatriptan, however, significantly reduces the levels of CGRP and CCK to enhance the inhibition of pain signals via the endogenous pain modulatory system, resulting in effective treatment of migraine. PMID:25524408

  12. Calcitonin gene-related peptide is a key factor in the homing of transplanted human MSCs to sites of spinal cord injury.

    PubMed

    Zhang, Yu; Yang, Jinhua; Zhang, Peng; Liu, Tao; Xu, Jianwei; Fan, Zhihai; Shen, Yixin; Li, Wenjie; Zhang, Huanxiang

    2016-01-01

    Mesenchymal stem cells (MSCs) can be used to treat many diseases, including spinal cord injury (SCI). Treatment relies mostly on the precise navigation of cells to the injury site for rebuilding the damaged spinal cord. However, the key factors guiding MSCs to the epicenter of SCI remain unknown. Here, we demonstrated that calcitonin gene-related peptide (CGRP), a neural peptide synthesized in spinal cord, can dramatically aid the homing of human umbilical cord mesenchymal stem cells (HUMSCs) in spinal cord-transected SCI rats. First, HUMSCs exhibited chemotactic responses in vitro to CGRP. By time-lapse video analysis, increased chemotactic index (CMI), forward migration index (FMI) and speed contributed to this observed migration. Then, through enzyme immunoassay, higher CGRP concentrations at the lesion site were observed after injury. The release of CGRP directed HUMSCs to the injury site, which was suppressed by CGRP 8-37, a CGRP antagonist. We also verified that the PI3K/Akt and p38MAPK signaling pathways played a critical role in the CGRP-induced chemotactic migration of HUMSCs. Collectively, our data reveal that CGRP is a key chemokine that helps HUMSCs migrate to the lesion site and thereby can be used as a model molecule to study MSCs homing after SCI. PMID:27296555

  13. Calcitonin gene-related peptide is a key factor in the homing of transplanted human MSCs to sites of spinal cord injury

    PubMed Central

    Zhang, Yu; Yang, Jinhua; Zhang, Peng; Liu, Tao; Xu, Jianwei; Fan, Zhihai; Shen, Yixin; Li, Wenjie; Zhang, Huanxiang

    2016-01-01

    Mesenchymal stem cells (MSCs) can be used to treat many diseases, including spinal cord injury (SCI). Treatment relies mostly on the precise navigation of cells to the injury site for rebuilding the damaged spinal cord. However, the key factors guiding MSCs to the epicenter of SCI remain unknown. Here, we demonstrated that calcitonin gene-related peptide (CGRP), a neural peptide synthesized in spinal cord, can dramatically aid the homing of human umbilical cord mesenchymal stem cells (HUMSCs) in spinal cord-transected SCI rats. First, HUMSCs exhibited chemotactic responses in vitro to CGRP. By time-lapse video analysis, increased chemotactic index (CMI), forward migration index (FMI) and speed contributed to this observed migration. Then, through enzyme immunoassay, higher CGRP concentrations at the lesion site were observed after injury. The release of CGRP directed HUMSCs to the injury site, which was suppressed by CGRP 8–37, a CGRP antagonist. We also verified that the PI3K/Akt and p38MAPK signaling pathways played a critical role in the CGRP-induced chemotactic migration of HUMSCs. Collectively, our data reveal that CGRP is a key chemokine that helps HUMSCs migrate to the lesion site and thereby can be used as a model molecule to study MSCs homing after SCI. PMID:27296555

  14. Hydrogen sulfide inhibits opioid withdrawal-induced pain sensitization in rats by down-regulation of spinal calcitonin gene-related peptide expression in the spine.

    PubMed

    Yang, Hai-Yu; Wu, Zhi-Yuan; Bian, Jin-Song

    2014-09-01

    Hyperalgesia often occurs in opioid-induced withdrawal syndrome. In the present study, we found that three hourly injections of DAMGO (a μ-opioid receptor agonist) followed by naloxone administration at the fourth hour significantly decreased rat paw nociceptive threshold, indicating the induction of withdrawal hyperalgesia. Application of NaHS (a hydrogen sulfide donor) together with each injection of DAMGO attenuated naloxone-precipitated withdrawal hyperalgesia. RT-PCR and Western blot analysis showed that NaHS significantly reversed the gene and protein expression of up-regulated spinal calcitonin gene-related peptide (CGRP) in naloxone-treated animals. NaHS also inhibited naloxone-induced cAMP rebound and cAMP response element-binding protein (CREB) phosphorylation in rat spinal cord. In SH-SY5Y neuronal cells, NaHS inhibited forskolin-stimulated cAMP production and adenylate cyclase (AC) activity. Moreover, NaHS pre-treatment suppressed naloxone-stimulated activation of protein kinase C (PKC) α, Raf-1, and extracellular signal-regulated kinase (ERK) 1/2 in rat spinal cord. Our data suggest that H2S prevents the development of opioid withdrawal-induced hyperalgesia via suppression of synthesis of CGRP in spine through inhibition of AC/cAMP and PKC/Raf-1/ERK pathways. PMID:24824948

  15. Calcitonin gene-related peptide erases the fear memory and facilitates long-term potentiation in the central nucleus of the amygdala in rats.

    PubMed

    Wu, Xin; Zhang, Jie-Ting; Liu, Jue; Yang, Si; Chen, Tao; Chen, Jian-Guo; Wang, Fang

    2015-11-01

    Calcitonin gene-related peptide (CGRP) is a 37 amino acid neuropeptide, which plays a critical role in the central nervous system. CGRP binds to G protein-coupled receptors, including CGRP1, which couples positively to adenylyl cyclase (AC) and protein kinase A (PKA) activation. CGRP and CGRP1 receptors are enriched in central nucleus of the amygdala (CeA), the main part of the amygdala, which regulates conditioned fear memories. Here, we reported the importance of CGRP and CGRP1 receptor for synaptic plasticity in the CeA and the extinction of fear memory in rats. Our electrophysiological and behavioral in vitro and in vivo results showed exogenous application of CGRP induced an immediate and lasting long-term potentiation in the basolateral nucleus of amygdala-CeA pathway, but not in the lateral nucleus of amygdala-CeA pathway, while bilateral intra-CeA infusion CGRP (0, 5, 13 and 21 μM/side) dose dependently enhanced fear memory extinction. The effects were blocked by CGRP1 receptor antagonist (CGRP8-37 ), N-methyl-d-aspartate receptors antagonist MK801 and PKA inhibitor H89. These results demonstrate that CGRP can lead to long-term potentiation of basolateral nucleus of amygdala-CeA pathway through a PKA-dependent postsynaptic mechanism that involved N-methyl-d-aspartate receptors and enhance the extinction of fear memory in rats. Together, the results strongly support a pivotal role of CGRP in the synaptic plasticity of CeA and extinction of fear memory. Calcitonin gene-related peptide (CGRP) plays an essential role in synaptic plasticity in the amygdala and fear memory. We found that CGRP-induced chemical long-term potentiation (LTP) in a dose-dependent way in the BLA-CeA (basolateral and central nucleus of amygdala, respectively) pathway and enhanced fear memory extinction in rats through a protein kinase A (PKA)-dependent postsynaptic mechanism that involved NMDA receptors. These results support a pivotal role of CGRP in amygdala. PMID:26179152

  16. High arterial compliance in cirrhosis is related to low adrenaline and elevated circulating calcitonin gene related peptide but not to activated vasoconstrictor systems

    PubMed Central

    Henriksen, J; Moller, S; Schifter, S; Abrahamsen, J; Becker, U

    2001-01-01

    BACKGROUND AND AIMS—Static and dynamic functions of the wall of large arteries are largely unknown in cirrhosis in vivo. The present study was undertaken to determine arterial compliance (COMPart) in relation to vasodilator and vasoconstrictor systems in patients with cirrhosis. In addition, vasoactivity was manipulated by inhalation of oxygen.
STUDY POPULATION AND METHODS—In 20 patients with alcoholic cirrhosis and 12 controls we determined COMPart (stroke volume relative to pulse pressure), cardiac output, plasma volume, systemic vascular resistance, central circulation time, plasma catecholamines, renin activity, endothelin-1, and calcitonin gene related peptide (CGRP) at baseline and during oxygen inhalation.
RESULTS—COMPart was significantly increased in cirrhotic patients compared with controls (1.32 v 1.06 ml/mm Hg; p< 0.05) and inversely related to plasma adrenaline levels (r=−0.53; p<0.02) but positively related to circulating levels of CGRP (r=0.58; p<0.01). No significant relation was found for plasma noradrenaline, renin activity, or endothelin-1. COMPart was positively related to plasma volume (r=0.50; p<0.02) and inversely to systemic vascular resistance (r=−0.69; p<0.001) and central circulation time (r=−0.49; p<0.02). During oxygen inhalation, COMPart decreased (−13%; p<0.005) and systemic vascular resistance increased (+10%; p<0.001) towards normal values without significant changes in mean arterial pressure. Plasma adrenaline (−16%; p<0.01) decreased and the relation to COMPart disappeared. The relation of COMPart to CGRP and circulatory variables remained unchanged.
CONCLUSION—Elevated arterial compliance in cirrhosis is related to low adrenaline, high CGRP, and systemic hyperdynamics but not to indicators of the activated vasoconstrictor systems (noradrenaline, renin, endothelin-1). Thus the altered static and dynamic characteristics of the wall of large arteries are intimately associated with circulatory and

  17. Involvement of calcitonin gene-related peptide and CCL2 production in CD40-mediated behavioral hypersensitivity in a model of neuropathic pain

    PubMed Central

    MALON, JENNIFER T.; MADDULA, SWATHI; BELL, HARMONY; CAO, LING

    2014-01-01

    The neuropeptide calcitonin gene-related peptide (CGRP) is known to play a pro-nociceptive role after peripheral nerve injury upon its release from primary afferent neurons in preclinical models of neuropathic pain. We previously demonstrated a critical role for spinal cord microglial CD40 in the development of spinal nerve L5 transection (L5Tx)-induced mechanical hypersensitivity. Herein, we investigated whether CGRP is involved in the CD40-mediated behavioral hypersensitivity. First, L5Tx was found to significantly induce CGRP expression in wild-type (WT) mice up to 14 days post-L5Tx. This increase in CGRP expression was reduced in CD40 knockout (KO) mice at day 14 post-L5Tx. Intrathecal injection of the CGRP antagonist CGRP8–37 significantly blocked L5Tx-induced mechanical hypersensitivity. In vitro, CGRP induced glial IL-6 and CCL2 production, and CD40 stimulation added to the effects of CGRP in neonatal glia. Further, there was decreased CCL2 production in CD40 KO mice compared to WT mice 21 days post-L5Tx. However, CGRP8–37 did not significantly affect spinal cord CCL2 production following L5Tx in WT mice. Altogether, these data suggest that CD40 contributes to the maintenance of behavioral hypersensitivity following peripheral nerve injury in part through two distinct pathways, the enhancement of CGRP expression and spinal cord CCL2 production. PMID:22377050

  18. [Suppressing effect of the serotonin 5HT1B/D receptor agonist rizatriptan on calcitonin gene-related peptide (CGRP) concentration in migraine attacks].

    PubMed

    Stepień, Adam; Jagustyn, Piotr; Trafny, Elzbieta Anna; Widerkiewicz, Krzysztof

    2003-01-01

    Calcitonin gene-related peptide (CGRP) is one of the neuropeptides most abundant in the nervous tissue. Recent studies indicate that local cranial release of CGRP from the trigeminal nerve perivascular endings within arachnoidea plays an important role in the pathophysiology of migraine attacks and cluster headaches. Elevated CGRP levels in cranial venous blood (in the jugular vein) during an acute spontaneous migraine attack have been reported in rather few studies so far. Sumatriptan--a selective serotonin 5HT1B/D receptor agonist, highly effective in terminating migraine attacks, decreases the elevated CGRP level back to normal. The aim of our study was to determine the effect of rizatriptan (a drug from a new generation of triptans) on CGRP release in migraine attacks. In 45 patients suffering from migraine attacks with and without aura, plasma CGRP levels were assessed during an attack twice: before treatment and two hours after rizatriptan administration. In the group under study the plasma CGRP level before treatment was significantly higher than that measured two hours after rizatriptan administration. The decrease in CGRP levels was associated with subsidence of the migraine attack. There was no difference between migraine patients with and without aura. These results suggest that triptans as serotonin 5HT1B/D receptor agonists decrease CGRP plasma concentration in migraine attacks. PMID:15174248

  19. Lafutidine, a novel histamine H2-receptor antagonist, increases serum calcitonin gene-related peptide in rats after water immersion-restraint stress.

    PubMed

    Sato, Hiroshi; Kawashima, Kousaku; Yuki, Mika; Kazumori, Hideaki; Rumi, Mohammad Azharul Karim; Ortega-Cava, Cesar Francisco; Ishihara, Shunji; Kinoshita, Yoshikazu

    2003-02-01

    Lafutidine is a novel histamine H(2)-receptor antagonist with a potent and long-lasting anti-acid secretory effect that has also been found to have a potent gastroprotective effect. We investigated the effect of lafutidine on gastric mucosal injury induced in rats with the use of water-immersion restraint stress (WRS) by examining serum calcitonin gene-related peptide (CGRP) concentrations, which we measured with the use of an enzyme immunometric assay. WRS-induced mucosal erosive injury in the stomach was reduced significantly by both lafutidine and famotidine pretreatment (from 7.79 +/- 2.02 mm(2) to 3.09 +/- 0.74 mm(2) and 4.05 +/- 1.18 mm(2), respectively). A single administration of lafutidine or famotidine did not change the serum CGRP concentration from the control value when these drugs were administered without WRS. Lafutidine pretreatment before WRS caused a significant increase in serum CGRP concentration compared with famotidine (lafutidine, 86.64 +/- 9.52 pg/mL; famotidine, 47.55 +/- 4.35 pg/mL; control, 58.43 +/- 6.07 pg/mL). Our results suggest that lafutidine augments CGRP release from the rat stomach when administered before the induction of WRS. PMID:12577045

  20. Rapid nitric oxide- and prostaglandin-dependent release of calcitonin gene-related peptide (CGRP) triggered by endotoxin in rat mesenteric arterial bed.

    PubMed Central

    Wang, X.; Wu, Z.; Tang, Y.; Fiscus, R. R.; Han, C.

    1996-01-01

    1. Our objective was to determine whether endotoxin (ETX) could directly trigger the release of calcitonin gene-related peptide (CGRP) from perivascular sensory nerves in the isolated mesenteric arterial bed (MAB) of the rat and to determine whether nitric oxide (NO) and prostaglandins (PGs) are involved. 2. ETX caused time- and concentration-dependent release of CGRP, and as much as a 17 fold increase in CGRP levels in the perfusate at 10-15 min after the administration of ETX (50 micrograms ml-1). 3. CGRP-like immunoreactivity in the perfusate was shown to co-elute with synthetic rat CGRP by reverse-phase h.p.l.c. 4. Pretreatment of MAB with capsaicin or ruthenium red inhibited ETX-induced CGRP release by 90% and 71%, respectively. ETX-evoked CGRP release was decreased by 84% during Ca2(+)-free perfusion. 5. The release of CGRP evoked by ETX was enhanced by L-arginine by 43% and inhibited by N omega-nitro-L-arginine (L-NOARG) and methylene blue by 37% and 38%, respectively. L-Arginine reversed the effect of L-NOARG. 6. Indomethacin and ibuprofen also inhibited the ETX-induced CGRP release by 34% and 44%, respectively. No additive inhibition could be found when L-NOARG and indomethacin were concomitantly incubated. 7. The data suggest that ETX triggers the release of CGRP from capsaicin-sensitive sensory nerves innervating blood vessels. The ETX-induced CGRP release is dependent on extracellular Ca2+ influx and involves a ruthenium red-sensitive mechanism. Both NO and PGs appear to be involved in the ETX-induced release of CGRP in the rat mesenteric arterial bed. PMID:8864557

  1. Identification of N-terminal receptor activity-modifying protein residues important for calcitonin gene-related peptide, adrenomedullin, and amylin receptor function.

    PubMed

    Qi, Tao; Christopoulos, George; Bailey, Richard J; Christopoulos, Arthur; Sexton, Patrick M; Hay, Debbie L

    2008-10-01

    Calcitonin-family receptors comprise calcitonin receptor-like receptor (CL) or calcitonin receptor and receptor activity-modifying protein (RAMP) pairings. Calcitonin gene-related peptide (CGRP) receptors are CL/RAMP1, whereas adrenomedullin (AM) receptors are CL/RAMP2 (AM1 receptor) or CL/RAMP3 (AM2 receptor). Amylin (Amy) receptors are RAMP hetero-oligomers with the calcitonin receptor (AMY1, AMY2, and AMY3, respectively). How RAMPs change G protein-coupled receptor pharmacology is not fully understood. We exploited sequence differences between RAMP1 and RAMP3 to identify individual residues capable of altering receptor pharmacology. Alignment of human RAMPs revealed eight residues that are conserved in RAMP2 and RAMP3 but are different in RAMP1. We hypothesized that residues in RAMP2 and RAMP3, but not RAMP1, are responsible for making CL/RAMP2 and CL/RAMP3 AM receptors. Using site-directed mutagenesis, we introduced individual RAMP3 residues into RAMP1 and vice versa in these eight positions. Mutant or wild-type RAMPs were transfected into Cos7 cells with CL or the insert-negative form of the calcitonin receptor [CT(a)]. Agonist-stimulated cAMP production and cell-surface expression of constructs were measured. Position 74 in RAMP1 and RAMP3 was critical for determining AM potency and affinity, and Phe93 in RAMP1 was an important contributor to alphaCGRP potency at CGRP receptors. Mutant RAMP/CT(a) receptor complexes displayed different phenotypes. It is noteworthy that RAMP1 S103N and W74E mutations led to enhanced rAmy potency, probably related to increased cell-surface expression of these complexes. This differs from the effect on CL-based receptors where expression was unchanged. Targeted substitution has emphasized the importance of position 74 in RAMP1/RAMP3 as a key determinant of AM pharmacology. PMID:18593822

  2. Structure–activity relationships of the N-terminus of calcitonin gene-related peptide: key roles of alanine-5 and threonine-6 in receptor activation

    PubMed Central

    Hay, Debbie L; Harris, Paul WR; Kowalczyk, Renata; Brimble, Margaret A; Rathbone, Dan L; Barwell, James; Conner, Alex C; Poyner, David R

    2014-01-01

    Background and Purpose: The N-terminus of calcitonin gene-related peptide (CGRP) is important for receptor activation, especially the disulphide-bonded ring (residues 1–7). However, the roles of individual amino acids within this region have not been examined and so the molecular determinants of agonism are unknown. This study has examined the role of residues 1, 3–6 and 8–9, excluding Cys-2 and Cys-7. Experimental Approach: CGRP derivatives were substituted with either cysteine or alanine; further residues were introduced at position 6. Their affinity was measured by radioligand binding and their efficacy by measuring cAMP production in SK-N-MC cells and β-arrestin 2 translocation in CHO-K1 cells at the CGRP receptor. Key Results: Substitution of Ala-5 by cysteine reduced affinity 270-fold and reduced efficacy for production of cAMP in SK-N-MCs. Potency at β-arrestin translocation was reduced by ninefold. Substitution of Thr-6 by cysteine destroyed all measurable efficacy of both cAMP and β-arrestin responses; substitution with either alanine or serine impaired potency. Substitutions at positions 1, 4, 8 and 9 resulted in approximately 10-fold reductions in potency at both responses. Similar observations were made at a second CGRP-activated receptor, the AMY1(a) receptor. Conclusions and Implications: Ala-5 and Thr-6 are key determinants of agonist activity for CGRP. Ala-5 is also very important for receptor binding. Residues outside of the 1–7 ring also contribute to agonist activity. PMID:24125506

  3. Expression and function of calcitonin gene-related peptide (CGRP) receptors in trigeminal ganglia of R192Q Cacna1a knock-in mice.

    PubMed

    Vilotti, Sandra; Vana, Natascha; Van den Maagdenberg, Arn M J M; Nistri, Andrea

    2016-05-01

    Migraine is a neurovascular brain disorder suggested to be due to dysfunction of the trigeminovascular system with sensitization of trigeminal ganglion (TG) nociceptors. Since the neuropeptide calcitonin gene-related peptide (CGRP) has been established as a key player in the pathogenesis of migraine, CGRP receptor antagonists have been considered useful compounds to block headache originating from hyperactivation of such TG neurons. Whereas there is some information on the expression of CGRP receptors in postmortem human tissue, data are lacking for migraineurs suffering from common or genetic migraine. To help to clarify these issues it is very useful to study a transgenic knock-in (KI) mouse model of hemiplegic migraine expressing a R192Q missense mutation in the α1 subunit of CaV2.1 calcium channels previously found in patients with familial hemiplegic migraine type-1 (FHM-1). The aim of the present study, therefore, was to compare CGRP receptor expression and function in wildtype (WT) versus KI mouse TG. The principal components of the CGRP receptor, namely the CLR and RAMP-1 proteins, were similarly expressed in WT and KI TG neurons (in situ or in culture) and responded to exogenous CGRP with a strong rise in cAMP concentration. Hence, the previously reported phenotype of sensitization of KI TG neurons is not due to up-regulation of CGRP receptors but is likely caused by a constitutively larger release of CGRP. This observation implies that, in FHM-1 TG, normal TG sensory neuron signaling can be restored once the extracellular concentration of CGRP returns to control level with targeted treatment. PMID:27021026

  4. Calcitonin gene-related peptide cooperates with substance P to inhibit melanogenesis and induces apoptosis of B16F10 cells.

    PubMed

    Zhou, Jia; Feng, Jun-Yi; Wang, Qian; Shang, Jing

    2015-07-01

    Skin is the largest organ in human body and works as biologically active barrier to provide critical preservation of body homeostasis. The skin is highly innervated by a plenitude of nerve fiber subpopulations, each carrying one or more neuronal mediators. Melanocyte itself also intimately contact with nerve fibers to form 'synaptic-like structure' and its functions may be directly regulated by the mediators contained in terminals of intra-epidermal nerve fibers. Clinical and biochemical studies have suggested that calcitonin gene-related peptide (CGRP) is involved in vitiligo skin. The present study was designed to investigate the effect of CGRP on epidermal melanocytes. After treatment with CGRP ranging from 0 to 500 ng/mL for 48 h, tyrosinase activity and melanogenesis were with little changes compared to treatment with medium only in B16F10 cells. Treatment with 500 ng/mL of CGRP cooperates with substance P (SP) (0.1-10 nM) to decrease tyrosinase activity and decrease melanin biosynthesis in B16F10 cells in a concentration-dependent manner. Furthermore, CGRP (8-37) antagonizes the synergistic effect of CGRP. The effect of CGRP on the cell apoptosis was examined. Treatments with 0-500 ng/mL of CGRP for 24 h, the expression levels of cleaved caspase-3, total caspase-3, cleaved caspase-9 and total caspase-9 were increased in a concentration-dependent manner. And 500 ng/mL of CGRP induced B16F10 cell apoptosis showed by TUNEL assay. In addition, Bax expression was up-regulated and Bcl-2 down-regulated in response to CGRP treatment. Hence, the Bax/Bcl-2 ratio was significantly increased. These in vitro observations indicate the pro-apoptotic impact of CGRP on B16F10 cell. PMID:25982845

  5. Exogenous asymmetric dimethylarginine (ADMA) in pathogenesis of ischemia-reperfusion-induced gastric lesions: interaction with protective nitric oxide (NO) and calcitonin gene-related peptide (CGRP).

    PubMed

    Magierowski, Marcin; Jasnos, Katarzyna; Sliwowski, Zbigniew; Surmiak, Marcin; Krzysiek-Maczka, Gracjana; Ptak-Belowska, Agata; Kwiecien, Slawomir; Brzozowski, Tomasz

    2014-01-01

    Asymmetric dimethylarginine (ADMA) is an endogenous nitric oxide (NO) synthesis inhibitor and pro-inflammatory factor. We investigated the role of ADMA in rat gastric mucosa compromised through 30 min of gastric ischemia (I) and 3 h of reperfusion (R). These I/R animals were pretreated with ADMA with or without the combination of L-arginine, calcitonin gene-related peptide (CGRP) or a small dose of capsaicin, all of which are known to afford protection against gastric lesions, or with a farnesoid X receptor (FXR) agonist, GW 4064, to increase the metabolism of ADMA. In the second series, ADMA was administered to capsaicin-denervated rats. The area of gastric damage was measured with planimetry, gastric blood flow (GBF) was determined by H2-gas clearance, and plasma ADMA and CGRP levels were determined using ELISA and RIA. ADMA significantly increased I/R-induced gastric injury while significantly decreasing GBF, the luminal NO content, and the plasma level of CGRP. This effect of ADMA was significantly attenuated by pretreatment with CGRP, L-arginine, capsaicin, or a PGE2 analogue. In GW4064 pretreated animals, the I/R injury was significantly reduced and this effect was abolished by co-treatment with ADMA. I/R damage potentiated by ADMA was exacerbated in capsaicin-denervated animals with a further reduction of CGRP. Plasma levels of IL-10 were significantly decreased while malonylodialdehyde (MDA) and plasma TNF-α contents were significantly increased by ADMA. In conclusion, ADMA aggravates I/R-induced gastric lesions due to a decrease of GBF, which is mediated by a fall in NO and CGRP release, and the enhancement of lipid peroxidation and its pro-inflammatory properties. PMID:24658439

  6. Impact of Food Components on in vitro Calcitonin Gene-Related Peptide Secretion-A Potential Mechanism for Dietary Influence on Migraine.

    PubMed

    Slavin, Margaret; Bourguignon, Julia; Jackson, Kyle; Orciga, Michael-Angelo

    2016-01-01

    Calcitonin gene-related peptide (CGRP) is a pivotal messenger in the inflammatory process in migraine. Limited evidence indicates that diet impacts circulating levels of CGRP, suggesting that certain elements in the diet may influence migraine outcomes. Interruption of calcium signaling, a mechanism which can trigger CGRP release, has been suggested as one potential route by which exogenous food substances may impact CGRP secretion. The objective of this study was to investigate the effects of foods and a dietary supplement on two migraine-related mechanisms in vitro: CGRP secretion from neuroendocrine CA77 cells, and calcium uptake by differentiated PC12 cells. Ginger and grape pomace extracts were selected for their anecdotal connections to reducing or promoting migraine. S-petasin was selected as a suspected active constituent of butterbur extract, the migraine prophylactic dietary supplement. Results showed a statistically significant decrease in stimulated CGRP secretion from CA77 cells following treatment with ginger (0.2 mg dry ginger equivalent/mL) and two doses of grape pomace (0.25 and 1.0 mg dry pomace equivalent/mL) extracts. Relative to vehicle control, CGRP secretion decreased by 22%, 43%, and 87%, respectively. S-petasin at 1.0 μM also decreased CGRP secretion by 24%. Meanwhile, S-petasin and ginger extract showed inhibition of calcium influx, whereas grape pomace had no effect on calcium. These results suggest that grape pomace and ginger extracts, and S-petasin may have anti-inflammatory propensity by preventing CGRP release in migraine, although potentially by different mechanisms, which future studies may elucidate further. PMID:27376323

  7. Calcitonin Gene-Related Peptide-Exposed Endothelial Cells Bias Antigen Presentation to CD4+ T Cells toward a Th17 Response.

    PubMed

    Ding, Wanhong; Stohl, Lori L; Xu, Linghui; Zhou, Xi K; Manni, Michela; Wagner, John A; Granstein, Richard D

    2016-03-01

    Calcitonin gene-related peptide (CGRP) is a neuropeptide with well-established immunomodulatory functions. CGRP-containing nerves innervate dermal blood vessels and lymph nodes. We examined whether CGRP regulates the outcome of Ag presentation by Langerhans cells (LCs) to T cells through actions on microvascular endothelial cells (ECs). Exposure of primary murine dermal microvascular ECs (pDMECs) to CGRP followed by coculture with LCs, responsive CD4(+) T cells and Ag resulted in increased production of IL-6 and IL-17A accompanied by inhibition of IFN-γ, IL-4, and IL-22 compared with wells containing pDMECs treated with medium alone. Physical contact between ECs and LCs or T cells was not required for this effect and, except for IL-4, we demonstrated that IL-6 production by CGRP-treated pDMECs was involved in these effects. CD4(+) cells expressing cytoplasmic IL-17A were increased, whereas cells expressing cytoplasmic IFN-γ or IL-4 were decreased by the presence of CGRP-treated pDMECs. In addition, the level of retinoic acid receptor-related orphan receptor γt mRNA was significantly increased, whereas T-bet and GATA3 expression was inhibited. Immunization at the site of intradermally administered CGRP led to a similar bias in CD4(+) T cells from draining lymph node cells toward IL-17A and away from IFN-γ. Actions of nerve-derived CGRP on ECs may have important regulatory effects on the outcome of Ag presentation with consequences for the expression of inflammatory skin disorders involving Th17 cells. PMID:26829986

  8. Involvement of calcitonin gene-related peptide (CGRP) receptors in insulin-induced vasodilatation in mesenteric resistance blood vessels of rats

    PubMed Central

    Mimaki, Yuichi; Kawasaki, Hiromu; Okazaki, Masatoshi; Nakatsuma, Akira; Araki, Hiroaki; Gomita, Yutaka

    1998-01-01

    The vascular effect of insulin in the mesenteric resistance blood vessel and the role of calcitonin gene-related peptide (CGRP)-receptor in insulin-induced vascular responsiveness were investigated in rats.The mesenteric vascular beds isolated from Wistar rats were perfused with Krebs solution, and perfusion pressure was measured with a pressure transducer. In preparations contracted by perfusion with Krebs solution containing methoxamine in the presence of guanethidine, the perfusion of insulin (from 0.1 to 3000 nM) caused a concentration-dependent decrease in perfusion pressure due to vasodilatation. The pD2 value and maximum relaxation (%) were 6.94±0.22 and 43.9±5.2, respectively.This vasodilator response to insulin was unaffected by 100 nM propranolol (β-adrenoceptor antagonist) plus 100 nM atropine (muscarinic cholinoceptor antagonist), 100 μM L-NG-nitroarginine (nitric oxide synthase inhibitor), 1 μM ouabain (Na+-K+ ATPase inhibitor), or 1 μM glibenclamide (ATP sensitive K+-channel inhibitor).In preparations without endothelium, perfusion of insulin produced a marked vasodilatation. The pD2 value and maximum relaxation (%) were 7.62±0.21 and 81.0±4.6, respectively, significantly greater than in preparations with intact endothelium.The vasodilator responses to insulin in the preparations without endothelium were significantly inhibited by CGRP[8–37], a CGRP receptor antagonist, whereas pretreatment with capsaisin, a toxin for CGRP-containing nerves, did not affect insulin-induced vasodilatation.These results suggest that insulin induces non-adrenergic, non-cholinergic and endothelium-independent vasodilatation, which is partially mediated by CGRP receptors. PMID:9605576

  9. Similarity between class A and class B G-protein-coupled receptors exemplified through calcitonin gene-related peptide receptor modelling and mutagenesis studies

    PubMed Central

    Vohra, Shabana; Taddese, Bruck; Conner, Alex C.; Poyner, David R.; Hay, Debbie L.; Barwell, James; Reeves, Philip J.; Upton, Graham J. G.; Reynolds, Christopher A.

    2013-01-01

    Modelling class B G-protein-coupled receptors (GPCRs) using class A GPCR structural templates is difficult due to lack of homology. The plant GPCR, GCR1, has homology to both class A and class B GPCRs. We have used this to generate a class A–class B alignment, and by incorporating maximum lagged correlation of entropy and hydrophobicity into a consensus score, we have been able to align receptor transmembrane regions. We have applied this analysis to generate active and inactive homology models of the class B calcitonin gene-related peptide (CGRP) receptor, and have supported it with site-directed mutagenesis data using 122 CGRP receptor residues and 144 published mutagenesis results on other class B GPCRs. The variation of sequence variability with structure, the analysis of polarity violations, the alignment of group-conserved residues and the mutagenesis results at 27 key positions were particularly informative in distinguishing between the proposed and plausible alternative alignments. Furthermore, we have been able to associate the key molecular features of the class B GPCR signalling machinery with their class A counterparts for the first time. These include the [K/R]KLH motif in intracellular loop 1, [I/L]xxxL and KxxK at the intracellular end of TM5 and TM6, the NPXXY/VAVLY motif on TM7 and small group-conserved residues in TM1, TM2, TM3 and TM7. The equivalent of the class A DRY motif is proposed to involve Arg2.39, His2.43 and Glu3.46, which makes a polar lock with T6.37. These alignments and models provide useful tools for understanding class B GPCR function. PMID:23235263

  10. Impact of Food Components on in vitro Calcitonin Gene-Related Peptide Secretion—A Potential Mechanism for Dietary Influence on Migraine

    PubMed Central

    Slavin, Margaret; Bourguignon, Julia; Jackson, Kyle; Orciga, Michael-Angelo

    2016-01-01

    Calcitonin gene-related peptide (CGRP) is a pivotal messenger in the inflammatory process in migraine. Limited evidence indicates that diet impacts circulating levels of CGRP, suggesting that certain elements in the diet may influence migraine outcomes. Interruption of calcium signaling, a mechanism which can trigger CGRP release, has been suggested as one potential route by which exogenous food substances may impact CGRP secretion. The objective of this study was to investigate the effects of foods and a dietary supplement on two migraine-related mechanisms in vitro: CGRP secretion from neuroendocrine CA77 cells, and calcium uptake by differentiated PC12 cells. Ginger and grape pomace extracts were selected for their anecdotal connections to reducing or promoting migraine. S-petasin was selected as a suspected active constituent of butterbur extract, the migraine prophylactic dietary supplement. Results showed a statistically significant decrease in stimulated CGRP secretion from CA77 cells following treatment with ginger (0.2 mg dry ginger equivalent/mL) and two doses of grape pomace (0.25 and 1.0 mg dry pomace equivalent/mL) extracts. Relative to vehicle control, CGRP secretion decreased by 22%, 43%, and 87%, respectively. S-petasin at 1.0 μM also decreased CGRP secretion by 24%. Meanwhile, S-petasin and ginger extract showed inhibition of calcium influx, whereas grape pomace had no effect on calcium. These results suggest that grape pomace and ginger extracts, and S-petasin may have anti-inflammatory propensity by preventing CGRP release in migraine, although potentially by different mechanisms, which future studies may elucidate further. PMID:27376323

  11. Calcitonin gene-related peptide protects rats from cerebral ischemia/reperfusion injury via a mechanism of action in the MAPK pathway

    PubMed Central

    YANG, SI; YUAN, YONGJIE; JIAO, SHAN; LUO, QI; YU, JINLU

    2016-01-01

    The aim of the present study was to investigate the protective function and underlying mechanism of calcitonin gene-related peptide (CGRP) on cerebral ischemia/reperfusion damage in rats. Adult male Wistar rats were selected for the establishment of an ischemia/reperfusion injury model through the application of a middle cerebral artery occlusion. Animals were randomly divided into 6 groups of 24 animals. Drugs were administered according to the design of each group; animals were administered CGRP, CGRP8–37, PD98059 and SB20358. The neurobehavioral scores of the rat cerebral ischemia model in each group were calculated. The infarction range of the rat brain tissues was observed by 2,3,5-triphenyltetrazolium chloride staining. The expression levels of three proteins, phosphorylated c-Jun N-terminal kinase (JNK)/JNK, phosphorylated extracellular signal-regulated protein kinase (ERK)/ERK and p-p38/p38, in the mitogen-activated protein kinase (MAPK) pathway in the brain tissues was detected by western blotting. The results showed that CGRP could improve the neurobehavioral function of the ischemic rats and reduce the infarction range. Western blotting results confirmed that the function of the CGRP was mediated mainly through the reduction of the JNK and p38 phosphorylation and the promotion of ERK phosphorylation. Therefore, the present study confirmed that an increase in the exogenous CRGP could effectively improve ischemia/reperfusion injury of the brain tissue. The mechanisms of action were achieved through a reduction in JNK and p38 phosphorylation and an increase in ERL phosphorylation in the MAPK pathway. These mechanisms were interdependent. PMID:27284409

  12. Expression of calcitonin gene-related peptide, adenosine A2a receptor and adenosine A1 receptor in experiment rat migraine models

    PubMed Central

    LU, WENXIAN; LI, BIN; CHEN, JINBO; SU, YIPENG; DONG, XIAOMENG; SU, XINYANG; GAO, LIXIANG

    2016-01-01

    A migraine is a disabling neurovascular disorder characterized by a unilateral throbbing headache that lasts from 4 to 72 h. The headache is often accompanied by nausea, vomiting, phonophobia and photophobia, and may be worsened by physical exercise. The trigeminovascular system (TVS) is speculated to have an important role in migraines, although the pathophysiology of this disorder remains to be elucidated. Trigeminal ganglion (TG) and spinal trigeminal nucleus caudalis (TNC) are important components of the TVS. Several clinical cases have provided evidence for the involvement of the brainstem in migraine initiation. Electrical stimulation of the trigeminal ganglion (ESTG) in rats can activate TVS during a migraine attack. Calcitonin gene-related peptide (CGRP) is an important vasoactive compound produced following TVS activation. Numerous studies have revealed that adenosine and its receptors have an important role in pain transmission and regulation process. However, only a few studies have examined whether adenosine A2a receptor (A2aR) and adenosine A1 receptor (A1R) are involved in migraine and nociceptive pathways. In the present study, CGRP, A2aR and A1R expression levels were detected in the TG and TNC of ESTG models through reverse transcription-quantitative polymerase chain reaction and western blot analysis. Tianshu capsule (TSC), a type of Chinese medicine, was also used in the ESTG rat models to examine its influence on the three proteins. Results demonstrated that CGRP, A2aR and A1R mediated pain transmission and the regulation process during migraine and the expression of the three proteins was regulated by TSC. PMID:26998280

  13. Decreased calcitonin gene-related peptide expression in the dorsal root ganglia of TNF-deficient mice in a monoiodoacetate-induced knee osteoarthritis model

    PubMed Central

    Taniguchi, Aya; Ishikawa, Tetsuhiro; Miyagi, Masayuki; Kamoda, Hiroto; Sakuma, Yoshihiro; Oikawa, Yasuhiro; Kubota, Go; Inage, Kazuhide; Sainoh, Takeshi; Nakamura, Junichi; Aoki, Yasuchika; Toyone, Tomoaki; Inoue, Gen; Suzuki, Miyako; Yamauchi, Kazuyo; Suzuki, Takane; Takahashi, Kazuhisa; Ohtori, Seiji; Orita, Sumihisa

    2015-01-01

    Background: The detailed mechanisms of knee osteoarthritis (OA) pain have not been clarified, but involvement of inflammatory cytokines such as tumor necrosis factor-alpha (TNF) has been suggested. The present study aimed to investigate the more detailed neurological involvement of TNF in joint pain using a TNF-knockout mouse OA model. Methods: The right knees of twelve-week-old C57BL/6J wild and TNF-deficient knockout (TNF-ko) mice (n=15, each group) were given a single intra-articular injection of 10 µg monoiodoacetate in 10 mL sterile saline. The left knees were only punctured as the control. Evaluations were performed immediately after the injection (baseline) and at 7, 14, and 28 days after the injection with a subsequent intra-articular injection of neurotracer into both knees. The animals were evaluated for immunofluorescence of the lumbar dorsal root ganglia (DRG) innervating the knee joints. The injected knees were observed macroscopically and mouse pain-related behaviors were scored. Results: Macroscopic observation showed similar knee OA development in both wild and TNF-ko mice. Calcitonin gene-related peptide (CGRP, a neuropeptide identified as a inflammatory pain-related biomarker) was significantly increased in DRG neurons innervating OA-induced knee joints with significantly less CGRP expression in TNF-ko animals. Pain-related behavior scoring showed a significant increase in pain in OA-induced joints, but there was no significant difference in pain observed between the wild and TNF-ko mice. Conclusions: The result of the present study indicates the possible association of TNF-alpha in OA pain but not OA development. PMID:26722492

  14. Calcitonin gene-related peptide does not mediate the abnormal vascular reactivity observed in a rat model of acute Pseudomonas pneumonia.

    PubMed

    Fox, G A; Paterson, N A; McCormack, D G

    1996-06-01

    Abnormal systemic and pulmonary vascular reactivity has been demonstrated in numerous models of sepsis and pneumonia. Furthermore, the attenuated hypoxic pulmonary pressor response observed in these animals probably is responsible for the ventilation/perfusion (V/Q) mismatching and consequent arterial hypoxemia. We hypothesized that excess release of endogenous vasodilators such as calcitonin gene-related peptide (CGRP) in pneumonia was responsible for the diminished hypoxic pressor response. Using the CGRP receptor antagonist CGRP (8-37), we examined the role of CGRP in the attenuated hypoxic pulmonary response in a rat model of acute Pseudomonas pneumonia. Sixteen Sprague-Dawley rats were instrumented for chronic hemodynamic monitoring and subsequently randomized to either Pneumonia (n = 8), induced by the instillation of 0.2 ml broth containing 2 x 10(8) colony-forming units (CFU)/ml Pseudomonas aeruginosa into the right lower lobe, or Sham (n = 8) procedure. Hemodynamic measurements and the hypoxic (FiO2 = 0.08) pulmonary pressor response were recorded at baseline, 48 h after the pneumonia or sham procedure and after the administration of 250 micrograms CGRP (8-37) (post-CGRP(8-37)). The regional distribution of pulmonary blood flow was determined by the injection of radioactive microspheres. Forty-eight hours after the instillation of Pseudomonas, Pneumonia animals had significantly increased cardiac output (CO) as compared with Sham (193 +/- 7 vs. 154 +/- 7 ml/min, p < 0.05), slightly decreased mean arterial pressure (MAP 109 +/- 4 vs. 118 +/- 3 mm Hg, p = NS), and reduced total systemic vascular resistance (TSVR 0.57 +/- 0.03 vs. 0.78 +/- 0.05 mm Hg.min.ml-1, p < 0.05). Pneumonia animals were further characterized by increased mean pulmonary artery pressure (MPAP) as compared with Sham (24 +/- 2 vs. 20 +/- 1 mm Hg, p < 0.05) animals, and an increased alveolar-arterial (A-a) oxygen gradient (31 +/- 3 vs. 20 +/- 4 mm Hg, p < 0.05). The administration of CGRP

  15. The effect of 17β-estradiol on gene expression of calcitonin gene-related peptide and some pro-inflammatory mediators in peripheral blood mononuclear cells from patients with pure menstrual migraine

    PubMed Central

    Karkhaneh, Azam; Ansari, Mohammad; Emamgholipour, Solaleh; Rafiee, Mohammad Hessam

    2015-01-01

    Objective(s): The neuropeptide calcitonin gene-related peptide (CGRP) has long been postulated to play an integral role in the pathophysiology of migraine. Earlier studies showed that CGRP can stimulate the synthesis and release of nitric oxide (NO) and cytokines from trigeminal ganglion glial cells. The purpose of this study was to determine the effect of 17β-estradiol in regulation of CGRP expression, inducible nitric oxide synthase (iNOS) activity, and NO and interleukin-1beta (IL-1β) release in cultured peripheral blood mononuclear cells (PBMCs) from patients with pure menstrual migraine and healthy individuals. Materials and Methods: This study was performed on twelve patients with pure menstrual migraine and twelve age-and sex-matched healthy individuals. PBMCs treated with 17β-estradiol for 24 hr at physiological and pharmacological doses. Gene expression was evaluated by real time-PCR. CGRP and IL-1β proteins in culture supernatant were determined by ELISA method. Activity of iNOS in PBMCs and total nitrite in the culture supernatant were measured by colorimetric assays. Results: Treatment with 17β-estradiol had a biphasic effect on expression of CGRP. We found that 17β-estradiol treatment at pharmacological dose significantly increases mRNA expression of CGRP in both groups (P<0.001), whereas at physiological dose it could significantly decrease CGRP mRNA expression (P<0.001), CGRP protein levels, IL-1β release, NO production and iNOS activity only in patient groups (P<0.05). Conclusion: Collectively, it appears that 17β-estradiol can exert protective effect on decrease of inflammation in migraine via decrease in levels of CGRP, IL-1β and iNOS activity; however, more studies are necessary in this regard. PMID:26526225

  16. Involvement of calcitonin gene-related peptide (CGRP) receptors in insulin-induced vasodilatation in mesenteric resistance blood vessels of rats.

    PubMed

    Mimaki, Y; Kawasaki, H; Okazaki, M; Nakatsuma, A; Araki, H; Gomita, Y

    1998-04-01

    1. The vascular effect of insulin in the mesenteric resistance blood vessel and the role of calcitonin generelated peptide (CGRP)-receptor in insulin-induced vascular responsiveness were investigated in rats. 2. The mesenteric vascular beds isolated from Wistar rats were perfused with Krebs solution, and perfusion pressure was measured with a pressure transducer. In preparations contracted by perfusion with Krebs solution containing methoxamine in the presence of guanethidine, the perfusion of insulin (from 0.1 to 3000 nM) caused a concentration-dependent decrease in perfusion pressure due to vasodilatation. The pD2 value and maximum relaxation (%) were 6.94+/-0.22 and 43.9+/-5.2, respectively. 3. This vasodilator response to insulin was unaffected by 100 nM propranolol (beta-adrenoceptor antagonist) plus 100 nM atropine (muscarinic cholinoceptor antagonist), 100 microM L-NG-nitroarginine (nitric oxide synthase inhibitor), 1 microM ouabain (Na+-K+ ATPase inhibitor), or 1 microM glibenclamide (ATP sensitive K+-channel inhibitor). 4. In preparations without endothelium, perfusion of insulin produced a marked vasodilatation. The pD2 value and maximum relaxation (%) were 7.62+/-0.21 and 81.0+/-4.6, respectively, significantly greater than in preparations with intact endothelium. 5. The vasodilator responses to insulin in the preparations without endothelium were significantly inhibited by CGRP[8 37], a CGRP receptor antagonist, whereas pretreatment with capsaisin, a toxin for CGRP-containing nerves, did not affect insulin-induced vasodilatation. 6. These results suggest that insulin induces non-adrenergic, non-cholinergic and endothelium-independent vasodilatation, which is partially mediated by CGRP receptors. PMID:9605576

  17. [PRRT2 gene-related paroxysmal disorders].

    PubMed

    Li, Jin; Mao, Xiao; Wang, Junling; Li, Nan; Tang, Beisha

    2014-10-01

    Proline-rich transmembrane protein 2 (PRRT2), the causative gene of paroxysmal kinesigenic dyskinesias (PKD), benign familial infantile seizures (BFIS) and infantile convulsions with paroxysmal choreoathetosis (ICCA), also causes a variety of neurological paroxysmal disorders. These diseases share the same characteristics which may be due to the same genetic defect. We therefore propose to name them as PRRT2-related paroxysmal disorders (PRPDs) in order to assist clinical diagnosis, treatment and prognosis. This paper has reviewed the clinical phenotype, common features and pathogenesis of the PRPDs. PMID:25297589

  18. Molecular functions of genes related to grain shape in rice

    PubMed Central

    Zheng, Jia; Zhang, Yadong; Wang, Cailin

    2015-01-01

    Because grain shape is an important component of rice grain yield, the discovery of genes related to rice grain shape has attracted much attention of rice breeding programs. In recent years, some of these genes have been cloned and studied. They have been found not only regulate grain shape by changing the shape of the spikelet hull, but also regulate endosperm development through control of cell division using different molecular mechanisms. In this paper, we review the recent research on genes related to rice grain shape and their possible regulatory mechanisms. PMID:26069441

  19. Review of Literature: Genes Related to Postaxial Polydactyly

    PubMed Central

    Verma, Prashant Kumar; El-Harouni, Ashraf A.

    2015-01-01

    Background: Postaxial polydactyly (PAP) is one of the commonest congenital malformations and usually is associated to several syndromes. There is no primary investigational strategy for PAP cases with single gene disorder in literature. PAP cases with single gene disorder can be classified according to common pathways and molecular basis. Molecular classification may help in diagnostic approach. Materials and Methods: All single gene disorders associated with PAP reported on PubMed and OMIM are analyzed and classified according to molecular basis. Results: Majority of genes related to cilia structure and functions are associated with PAP, so we classified them as ciliopathies and non-ciliopathies groups. Genes related to Shh–Gli3 pathway was the commonest group in non-ciliopathies. Conclusion: Genes related to cilia are most commonly related to PAP due to their indirect relationship to Shh–Gli3 signaling pathway. Initially, PAP may be the only clinical finding with ciliopathies so those cases need follow up. Proper diagnosis is helpful for management and genetic counseling. Molecular approach may help to define pleiotropy. PMID:25717468

  20. Genes related to high temperature tolerance during maize seed germination.

    PubMed

    Dutra, S M F; Von Pinho, E V R; Santos, H O; Lima, A C; Von Pinho, R G; Carvalho, M L M

    2015-01-01

    The identification of genes related to heat tolerance is fundamental for the development of high-quality seeds that are tolerant to heat stress condition. The objective of this study was to evaluate maize lineages and the gene expression involved in high temperature tolerance during germination using physiological tests, proteomics, and transcriptome analysis. Seeds from six maize lineages (30, 44, 54, 63, 64, and 91) with different levels of tolerance to high temperatures were used. Lineages 54 and 91 were observed to be more tolerant to high temperature conditions. The highest expression of α-amylase was observed in maize seeds from lineages 30 and 91 that were subjected to controlled deterioration. The highest expression of α-amylase was observed in maize seeds from lineages 30 and 91 that were subjected to controlled deterioration; with the controlled deterioration, the highest level of gene expression did not occur in the most tolerant materials; the association of lower expression of genes involved in heat-resistant protein systems was observed in seeds from lineage 44, which were more susceptible to high temperatures, and the highest gene expression of LEA D-34, ZmAN13, and AOX-1 was observed in seeds from lineage 64 when submitted to controlled deterioration. PMID:26782452

  1. Genes related to chromate resistance by Pseudomonas aeruginosa PAO1.

    PubMed

    Rivera, Sonia L; Vargas, Eréndira; Ramírez-Díaz, Martha I; Campos-García, Jesús; Cervantes, Carlos

    2008-08-01

    Chromate-hypersensitive mutants of the Pseudomonas aeruginosa PAO1 strain were isolated using transposon-insertion mutagenesis. Comparison of the nucleotide sequences of the regions interrupted in the mutants with the PAO1 genome revealed that the genes affected in three mutant strains were oprE (ORF PA0291), rmlA (ORF PA5163), and ftsK (ORF PA2615), respectively. A relationship of these genes with chromate tolerance has not been previously reported. No other phenotypic changes were observed in the oprE mutant but its resistance to chromate was not fully restored by expressing the ChrA protein, which extrudes chromate ions from the cytoplasm to the periplasmic space. These data suggest that OprE participates in the efflux of chromate from the periplasm to the outside. Increased susceptibility of the rmlA mutant to the metals cadmium and mercury and to the anion-superoxide generator paraquat suggests a protective role of LPS against chromate toxicity. A higher susceptibility of the ftsK mutant to compounds affecting DNA structure (ciprofloxacin, tellurite, mitomycin C) suggests a role of FtsK in the recombinational repair of DNA damage caused by chromate. In conclusion, the P. aeruginosa genome contains diverse genes related to its intrinsic resistance to chromate. Systems pertaining to the outer membrane (OprE), the cell wall (LPS), and the cytoplasm (FtsK) were identified in this work as involved in chromate protection mechanisms. PMID:18446454

  2. Leptin regulates gallbladder genes related to absorption and secretion.

    PubMed

    Swartz-Basile, Deborah A; Lu, Debao; Basile, David P; Graewin, Shannon J; Al-Azzawi, Hayder; Kiely, James M; Mathur, Abhishek; Yancey, Kyle; Pitt, Henry A

    2007-07-01

    Dysregulation of gallbladder ion and water absorption and/or secretion has been linked to cholesterol crystal and gallstone formation. We have recently demonstrated that obese, leptin-deficient (Lep(ob)) mice have enlarged gallbladder volumes and decreased gallbladder contractility and that leptin administration to these mice normalizes gallbladder function. However, the effect of leptin on gallbladder absorption/secretion is not known. Therefore, we sought to determine whether leptin would alter the expression of genes involved in water and ion transport across the gallbladder epithelium. Affymetrix oligonucleotide microarrays representing 39,000 transcripts were used to compare gallbladder gene-expression profiles from 12-wk-old control saline-treated Lep(ob) and from leptin-treated Lep(ob) female mice. Leptin administration to Lep(ob) mice decreased gallbladder volume, bile sodium concentration, and pH. Leptin repletion upregulated the expression of aquaporin 1 water channel by 1.3-fold and downregulated aquaporin 4 by 2.3-fold. A number of genes involved in sodium transport were also influenced by leptin replacement. Epithelial sodium channel-alpha and sodium hydrogen exchangers 1 and 3 were moderately downregulated by 2.0-, 1.6-, and 1.3-fold, respectively. Carbonic anhydrase-IV, which plays a role in the acidification of bile, was upregulated 3.7-fold. In addition, a number of inflammatory cytokines that are known to influence gallbladder epithelial cell absorption and secretion were upregulated. Thus leptin, an adipocyte-derived cytokine involved with satiety and energy balance, influences gallbladder bile volume, sodium, and pH as well as multiple inflammatory cytokine genes and genes related to water, sodium, chloride, and bicarbonate transport. PMID:17463181

  3. Amplification of a Gene Related to Mammalian mdr Genes in Drug-Resistant Plasmodium falciparum

    NASA Astrophysics Data System (ADS)

    Wilson, Craig M.; Serrano, Adelfa E.; Wasley, Annemarie; Bogenschutz, Michael P.; Shankar, Anuraj H.; Wirth, Dyann F.

    1989-06-01

    The malaria parasite Plasmodium falciparum contains at least two genes related to the mammalian multiple drug resistance genes, and at least one of the P. falciparum genes is expressed at a higher level and is present in higher copy number in a strain that is resistant to multiple drugs than in a strain that is sensitive to the drugs.

  4. Discovery of (5S,6S,9R)-5-amino-6-(2,3-difluorophenyl)-6,7,8,9-tetrahydro-5H-cyclohepta[b]pyridin-9-yl 4-(2-oxo-2,3-dihydro-1H-imidazo[4,5-b]pyridin-1-yl)piperidine-1-carboxylate (BMS-927711): an oral calcitonin gene-related peptide (CGRP) antagonist in clinical trials for treating migraine.

    PubMed

    Luo, Guanglin; Chen, Ling; Conway, Charles M; Denton, Rex; Keavy, Deborah; Signor, Laura; Kostich, Walter; Lentz, Kimberley A; Santone, Kenneth S; Schartman, Richard; Browning, Marc; Tong, Gary; Houston, John G; Dubowchik, Gene M; Macor, John E

    2012-12-13

    Calcitonin gene-related peptide (CGRP) receptor antagonists have demonstrated clinical efficacy in the treatment of acute migraine. Herein, we describe the design, synthesis, and preclinical characterization of a highly potent, oral CGRP receptor antagonist BMS-927711 (8). Compound 8 has good oral bioavailability in rat and cynomolgus monkey, attractive overall preclinical properties, and shows dose-dependent activity in a primate model of CGRP-induced facial blood flow. Compound 8 is presently in phase II clinical trials. PMID:23153230

  5. Titanium nanotubes activate genes related to bone formation in vitro

    PubMed Central

    Pozio, Alfonso; Palmieri, Annalisa; Girardi, Ambra; Cura, Francesca; Carinci, Francesco

    2012-01-01

    Background: Titanium is used worldwide to make osseointegrable devices, thanks to its favorable characteristics as mechanical proprieties and biocompatibility, demonstrated by in vivo studies with animal models and clinical trials over a forty-year period. However, the exact genetic effect of the titanium layer on cells is still not well characterized. Materials and Methods: To investigate how titanium nanotubes stimulate osteoblasts differentiation and proliferation, some osteoblast genes (SP7, RUNX2, COL3A1, COL1A1, ALPL, SPP1 and FOSL1) were analyzed by quantitative Real Time RT- PCR. Results: After 15 days, osteoblasts cultivated on titanium naotube showed the up-regulation of bone related genes SP7, ENG, FOSL1 and SPP1 and the down-regulation of RUNX2, COL3A1, COL1A1, and ALPL. After 30 days of treatment, the bone related genes SP7, ENG, FOSL1 and RUNX2 were up-regulated while COL3A1, COL1A1, ALPL and SPP1 were down-regulated. Conclusions: Our results, demonstrates that titanium nanotubes can lead to osteoblast differentiation and extracellular matrix deposition and mineralization in dental pulp stem cells by the activation of osteoblast related genes SPP1, FOSL1 and RUNX2. PMID:23814577

  6. A gene and protein expression study on four porcine genes related to intramuscular fat deposition.

    PubMed

    Zappaterra, Martina; Deserti, Marzia; Mazza, Roberta; Braglia, Silvia; Zambonelli, Paolo; Davoli, Roberta

    2016-11-01

    Intramuscular fat (IMF) content has a prominent role in meat quality, affecting sensory attributes such as flavour and texture. In the present research, we studied in samples of porcine Semimembranosus muscle four genes related to lipid metabolism and whose gene expressions have been associated to IMF deposition: FASN, SCD, LIPE and LPL. We analysed both mRNA and protein expressions in two groups of Italian Large White pigs divergent for Semimembranosus IMF deposition, with the aim of comparing the levels of four genes and enzymes between the two groups and identifying possible coexpression links. The obtained results suggest a prominent role of LIPE enzyme in IMF hydrolysis, as the samples with low IMF deposition show a significantly higher amount of this lipase. Finally, a poorly known correlation was found between LIPE and FASN enzymes only in female individuals. These results provide new information for the understanding of IMF deposition. PMID:27236338

  7. Identifying Novel Candidate Genes Related to Apoptosis from a Protein-Protein Interaction Network

    PubMed Central

    Wang, Baoman; Yuan, Fei; Kong, Xiangyin; Hu, Lan-Dian; Cai, Yu-Dong

    2015-01-01

    Apoptosis is the process of programmed cell death (PCD) that occurs in multicellular organisms. This process of normal cell death is required to maintain the balance of homeostasis. In addition, some diseases, such as obesity, cancer, and neurodegenerative diseases, can be cured through apoptosis, which produces few side effects. An effective comprehension of the mechanisms underlying apoptosis will be helpful to prevent and treat some diseases. The identification of genes related to apoptosis is essential to uncover its underlying mechanisms. In this study, a computational method was proposed to identify novel candidate genes related to apoptosis. First, protein-protein interaction information was used to construct a weighted graph. Second, a shortest path algorithm was applied to the graph to search for new candidate genes. Finally, the obtained genes were filtered by a permutation test. As a result, 26 genes were obtained, and we discuss their likelihood of being novel apoptosis-related genes by collecting evidence from published literature. PMID:26543496

  8. Large-scale cDNA transfection screening for genes related to cancer development and progression

    PubMed Central

    Wan, Dafang; Gong, Yi; Qin, Wenxin; Zhang, Pingping; Li, Jinjun; Wei, Lin; Zhou, Xiaomei; Li, Hongnian; Qiu, Xiaokun; Zhong, Fei; He, Liping; Yu, Jian; Yao, Genfu; Jiang, Huiqiu; Qian, Lianfang; Yu, Ye; Shu, Huiqun; Chen, Xianlian; Xu, Huili; Guo, Minglei; Pan, Zhimei; Chen, Yan; Ge, Chao; Yang, Shengli; Gu, Jianren

    2004-01-01

    A large-scale assay was performed by transfecting 29,910 individual cDNA clones derived from human placenta, fetus, and normal liver tissues into human hepatoma cells and 22,926 cDNA clones into mouse NIH 3T3 cells. Based on the results of colony formation in hepatoma cells and foci formation in NIH 3T3 cells, 3,806 cDNA species (8,237 clones) were found to possess the ability of either stimulating or inhibiting cell growth. Among them, 2,836 (6,958 clones) were known genes, 372 (384 clones) were previously unrecognized genes, and 598 (895 clones) were unigenes of uncharacterized structure and function. A comprehensive analysis of the genes and the potential mechanisms for their involvement in the regulation of cell growth is provided. The genes were classified into four categories: I, genes related to the basic cellular mechanism for growth and survival; II, genes related to the cellular microenvironment; III, genes related to host-cell systemic regulation; and IV, genes of miscellaneous function. The extensive growth-regulatory activity of genes with such highly diversified functions suggests that cancer may be related to multiple levels of cellular and systemic controls. The present assay provides a direct genomewide functional screening method. It offers a better understanding of the basic machinery of oncogenesis, including previously undescribed systemic regulatory mechanisms, and also provides a tool for gene discovery with potential clinical applications. PMID:15498874

  9. Negative pressure wound therapy-associated tissue trauma and pain: a controlled in vivo study comparing foam and gauze dressing removal by immunohistochemistry for substance P and calcitonin gene-related peptide in the wound edge.

    PubMed

    Malmsjö, Malin; Gustafsson, Lotta; Lindstedt, Sandra; Ingemansson, Richard

    2011-12-01

    Pain upon negative pressure wound therapy (NPWT) dressing removal has been reported and is believed to be associated with the observation that granulation tissue grows into foam. Wound tissue damage upon removal of the foam may cause the reported pain. Calcitonin gene-related peptide (CGRP) and substance P are neuropeptides that cause inflammation and signal pain and are known to be released when tissue trauma occurs. The aim of this controlled in vivo study was to compare the expression of CGRP and substance P in the wound bed in control wounds and following NPWT and foam or gauze dressing removal. Eight pigs with two wounds each were treated with open-pore structure polyurethane foam or AMD gauze and NPWT of 0 (control) or -80 mm Hg for 72 hours. Following removal of the wound filler, the expression of CGRP and substance P was measured, using arbitrary units, in sections of biopsies from the wound bed using immunofluorescence techniques. Substance P and CGRP were more abundant in the wound edge following the removal of foam than of gauze dressings and least abundant in control wounds. The immunofluorescence staining of the wound edge for CGRP was 52 ± 3 au after the removal of gauze and 97 ± 5 au after the removal of foam (P <0.001). For substance P, the staining was 55 ± 3 au after gauze removal and 95 ± 4 au after foam removal (P <0.001). CGRP and substance P staining was primarily located to nerves and leukocytes. The increase in CGRP and substance P immunofluorescence was especially prominent in the dermis but also was seen in subcutaneous and muscle tissue. Using gauze may be one way of reducing NPWT dressing change-related pain. New wound fillers designed to optimize granulation tissue formation and minimize pain issues presumably will be developed in the near future. PMID:22156176

  10. Identification of genes related to Parkinson's disease using expressed sequence tags.

    PubMed

    Kim, Jeong-Min; Lee, Kyu-Hwa; Jeon, Yeo-Jin; Oh, Jung-Hwa; Jeong, So-Young; Song, In-Sung; Kim, Jin-Man; Lee, Dong-Seok; Kim, Nam-Soon

    2006-12-31

    In a search for novel target genes related to Parkinson's disease (PD), two full-length cDNA libraries were constructed from a human normal substantia nigra (SN) and a PD patient's SN. An analysis of the gene expression profiles between them was done using the expressed sequence tags (ESTs) frequency. Data for the differently expressed genes were verified by quantitative real-time RT-PCR, immunohistochemical analysis and a cell death assay. Among the 76 genes identified with a significant difference (P > 0.9), 21 upregulated genes and 13 downregulated genes were confirmed to be differentially expressed in human PD tissues and/or in an MPTP-treated mice model by quantitative real-time RT-PCR. Among those genes, an immunohistochemical analysis using an MPTP mice model for alpha-tubulin including TUBA3 and TUBA6 showed that the protein levels are downregulated, as well as the RNA levels. In addition, MBP, PBP and GNAS were confirmed to accelerate cell death activity, whereas SPP1 and TUBA3 to retard this process. Using an analysis of ESTs frequency, it was possible to identify a large number of genes related to human PD. These new genes, MBP, PBP, GNAS, SPP1 and TUBA3 in particular, represent potential biomarkers for PD and could serve as useful targets for elucidating the molecular mechanisms associated with PD. PMID:17213182

  11. Utilization of digital differential display to identify differentially expressed genes related to rumen development.

    PubMed

    Kato, Daichi; Suzuki, Yutaka; Haga, Satoshi; So, KyoungHa; Yamauchi, Eri; Nakano, Miwa; Ishizaki, Hiroshi; Choi, Kichoon; Katoh, Kazuo; Roh, Sang-Gun

    2016-04-01

    This study aimed to identify the genes associated with the development of the rumen epithelium by screening for candidate genes by digital differential display (DDD) in silico. Using DDD in NCBI's UniGene database, expressed sequence tag (EST)-based gene expression profiles were analyzed in rumen, reticulum, omasum, abomasum and other tissues in cattle. One hundred and ten candidate genes with high expression in the rumen were derived from a library of all tissues. The expression levels of 11 genes in all candidate genes were analyzed in the rumen, reticulum, omasum and abomasum of nine Japanese Black male calves (5-week-old pre-weaning: n = 3; 15-week-old weaned calves: n = 6). Among the 11 genes, only 3-hydroxy-3-methylglutaryl-CoA synthase 2 (HMGCS2), aldo-keto reductase family 1, member C1-like (AKR1C1), and fatty acid binding protein 3 (FABP3) showed significant changes in the levels of gene expression in the rumen between the pre- and post-weaning of calves. These results indicate that DDD analysis in silico can be useful for screening candidate genes related to rumen development, and that the changes in expression levels of three genes in the rumen may have been caused by weaning, aging or both. © 2015 Japanese Society of Animal Science. PMID:26388291

  12. Identification of differentially expressed genes related to aphid resistance in cucumber (Cucumis sativus L.)

    PubMed Central

    Liang, Danna; Liu, Min; Hu, Qijing; He, Min; Qi, Xiaohua; Xu, Qiang; Zhou, Fucai; Chen, Xuehao

    2015-01-01

    Cucumber, a very important vegetable crop worldwide, is easily damaged by pests. Aphids (Aphis gossypii Glover) are among the most serious pests in cucumber production and often cause severe loss of yield and make fruit quality get worse. Identifying genes that render cucumbers resistant to aphid-induced damage and breeding aphid-resistant cucumber varieties have become the most promising control strategies. In this study, a Illumina Genome Analyzer platform was applied to monitor changes in gene expression in the whole genome of the cucumber cultivar ‘EP6392’ which is resistant to aphids. Nine DGE libraries were constructed from infected and uninfected leaves. In total, 49 differentially expressed genes related to cucumber aphid resistance were screened during the treatment period. These genes are mainly associated with signal transduction, plant-pathogen interactions, flavonoid biosynthesis, amino acid metabolism and sugar metabolism pathways. Eight of the 49 genes may be associated with aphid resistance. Finally, expression of 9 randomly selected genes was evaluated by qRT-PCR to verify the results for the tag-mapped genes. With the exception of 1-aminocyclopropane-1-carboxylate oxidase homolog 6, the expression of the chosen genes was in agreement with the results of the tag-sequencing analysis patterns. PMID:25959296

  13. Identification of Genes Related to Paulownia Witches’ Broom by AFLP and MSAP

    PubMed Central

    Cao, Xibing; Fan, Guoqiang; Deng, Minjie; Zhao, Zhenli; Dong, Yanpeng

    2014-01-01

    DNA methylation is believed to play important roles in regulating gene expression in plant growth and development. Paulownia witches’ broom (PaWB) infection has been reported to be related to gene expression changes in paulownia plantlets. To determine whether DNA methylation is associated with gene expression changes in response to phytoplasma, we investigated variations in genomic DNA sequence and methylation in PaWB plantlets treated with methyl methane sulfonate (MMS) using amplified fragment length polymorphism (AFLP) and methylation-sensitive amplification polymorphism (MSAP) techniques, respectively. The results indicated that PaWB seedings recovered a normal morphology after treatment with more than 15 mg·L−1 MMS. PaWB infection did not cause changes of the paulownia DNA sequence at the AFLP level; However, DNA methylation levels and patterns were altered. Quantitative real-time PCR (qRT-PCR) showed that three of the methylated genes were up-regulated and three were down-regulated in the MMS-treated PaWB plantlets that had regained healthy morphology. These six genes might be involved in transcriptional regulation, plant defense, signal transduction and energy. The possible roles of these genes in PaWB are discussed. The results showed that changes of DNA methylation altered gene expression levels, and that MSAP might help identify genes related to PaWB. PMID:25196603

  14. Identification of differentially expressed genes related to aphid resistance in cucumber (Cucumis sativus L.).

    PubMed

    Liang, Danna; Liu, Min; Hu, Qijing; He, Min; Qi, Xiaohua; Xu, Qiang; Zhou, Fucai; Chen, Xuehao

    2015-01-01

    Cucumber, a very important vegetable crop worldwide, is easily damaged by pests. Aphids (Aphis gossypii Glover) are among the most serious pests in cucumber production and often cause severe loss of yield and make fruit quality get worse. Identifying genes that render cucumbers resistant to aphid-induced damage and breeding aphid-resistant cucumber varieties have become the most promising control strategies. In this study, a Illumina Genome Analyzer platform was applied to monitor changes in gene expression in the whole genome of the cucumber cultivar 'EP6392' which is resistant to aphids. Nine DGE libraries were constructed from infected and uninfected leaves. In total, 49 differentially expressed genes related to cucumber aphid resistance were screened during the treatment period. These genes are mainly associated with signal transduction, plant-pathogen interactions, flavonoid biosynthesis, amino acid metabolism and sugar metabolism pathways. Eight of the 49 genes may be associated with aphid resistance. Finally, expression of 9 randomly selected genes was evaluated by qRT-PCR to verify the results for the tag-mapped genes. With the exception of 1-aminocyclopropane-1-carboxylate oxidase homolog 6, the expression of the chosen genes was in agreement with the results of the tag-sequencing analysis patterns. PMID:25959296

  15. Evolution of African swine fever virus genes related to evasion of host immune response.

    PubMed

    Frączyk, Magdalena; Woźniakowski, Grzegorz; Kowalczyk, Andrzej; Bocian, Łukasz; Kozak, Edyta; Niemczuk, Krzysztof; Pejsak, Zygmunt

    2016-09-25

    African swine fever (ASF) is a notifiable and one of the most complex and devastating infectious disease of pigs, wild boars and other representatives of Suidae family. African swine fever virus (ASFV) developed various molecular mechanisms to evade host immune response including alteration of interferon production by multigene family protein (MGF505-2R), inhibition of NF-κB and nuclear activating factor in T-cells by the A238L protein, or modulation of host defense by CD2v lectin-like protein encoded by EP402R and EP153R genes. The current situation concerning ASF in Poland seems to be stable in comparison to other eastern European countries but up-to-date in total 106 ASF cases in wild boar and 5 outbreaks in pigs were identified. The presented study aimed to reveal and summarize the genetic variability of genes related to inhibition or modulation of infected host response among 67 field ASF isolates collected from wild boar and pigs. The nucleotide sequences derived from the analysed A238L and EP153R regions showed 100% identity. However, minor but remarkable genetic diversity was found within EP402R and MGF505-2R genes suggesting slow molecular evolution of circulating ASFV isolates and the important role of this gene in modulation of interferon I production and hemadsorption phenomenon. The obtained nucleotide sequences of Polish ASFV isolates were closely related to Georgia 2007/1 and Odintsovo 02/14 isolates suggesting their common Caucasian origin. In the case of EP402R and partially in MGF505-2R gene the identified genetic variability was related to spatio-temporal occurrence of particular cases and outbreaks what may facilitate evolution tracing of ASFV isolates. This is the first report indicating identification of genetic variability within the genes related to evasion of host immune system which may be used to trace the direction of ASFV isolates molecular evolution. PMID:27599940

  16. miRTex: A Text Mining System for miRNA-Gene Relation Extraction

    PubMed Central

    Li, Gang; Ross, Karen E.; Arighi, Cecilia N.; Peng, Yifan; Wu, Cathy H.; Vijay-Shanker, K.

    2015-01-01

    MicroRNAs (miRNAs) regulate a wide range of cellular and developmental processes through gene expression suppression or mRNA degradation. Experimentally validated miRNA gene targets are often reported in the literature. In this paper, we describe miRTex, a text mining system that extracts miRNA-target relations, as well as miRNA-gene and gene-miRNA regulation relations. The system achieves good precision and recall when evaluated on a literature corpus of 150 abstracts with F-scores close to 0.90 on the three different types of relations. We conducted full-scale text mining using miRTex to process all the Medline abstracts and all the full-length articles in the PubMed Central Open Access Subset. The results for all the Medline abstracts are stored in a database for interactive query and file download via the website at http://proteininformationresource.org/mirtex. Using miRTex, we identified genes potentially regulated by miRNAs in Triple Negative Breast Cancer, as well as miRNA-gene relations that, in conjunction with kinase-substrate relations, regulate the response to abiotic stress in Arabidopsis thaliana. These two use cases demonstrate the usefulness of miRTex text mining in the analysis of miRNA-regulated biological processes. PMID:26407127

  17. Deregulation of genes related to iron and mitochondrial metabolism in refractory anemia with ring sideroblasts.

    PubMed

    del Rey, Mónica; Benito, Rocío; Fontanillo, Celia; Campos-Laborie, Francisco J; Janusz, Kamila; Velasco-Hernández, Talía; Abáigar, María; Hernández, María; Cuello, Rebeca; Borrego, Daniel; Martín-Zanca, Dionisio; De Las Rivas, Javier; Mills, Ken I; Hernández-Rivas, Jesús M

    2015-01-01

    The presence of SF3B1 gene mutations is a hallmark of refractory anemia with ring sideroblasts (RARS). However, the mechanisms responsible for iron accumulation that characterize the Myelodysplastic Syndrome with ring sideroblasts (MDS-RS) are not completely understood. In order to gain insight in the molecular basis of MDS-RS, an integrative study of the expression and mutational status of genes related to iron and mitochondrial metabolism was carried out. A total of 231 low-risk MDS patients and 81 controls were studied. Gene expression analysis revealed that iron metabolism and mitochondrial function had the highest number of genes deregulated in RARS patients compared to controls and the refractory cytopenias with unilineage dysplasia (RCUD). Thus mitochondrial transporters SLC25 (SLC25A37 and SLC25A38) and ALAD genes were over-expressed in RARS. Moreover, significant differences were observed between patients with SF3B1 mutations and patients without the mutations. The deregulation of genes involved in iron and mitochondrial metabolism provides new insights in our knowledge of MDS-RS. New variants that could be involved in the pathogenesis of these diseases have been identified. PMID:25955609

  18. Deregulation of Genes Related to Iron and Mitochondrial Metabolism in Refractory Anemia with Ring Sideroblasts

    PubMed Central

    del Rey, Mónica; Benito, Rocío; Fontanillo, Celia; Campos-Laborie, Francisco J.; Janusz, Kamila; Velasco-Hernández, Talía; Abáigar, María; Hernández, María; Cuello, Rebeca; Borrego, Daniel; Martín-Zanca, Dionisio; De Las Rivas, Javier; Mills, Ken I.; Hernández-Rivas, Jesús M.

    2015-01-01

    The presence of SF3B1 gene mutations is a hallmark of refractory anemia with ring sideroblasts (RARS). However, the mechanisms responsible for iron accumulation that characterize the Myelodysplastic Syndrome with ring sideroblasts (MDS-RS) are not completely understood. In order to gain insight in the molecular basis of MDS-RS, an integrative study of the expression and mutational status of genes related to iron and mitochondrial metabolism was carried out. A total of 231 low-risk MDS patients and 81 controls were studied. Gene expression analysis revealed that iron metabolism and mitochondrial function had the highest number of genes deregulated in RARS patients compared to controls and the refractory cytopenias with unilineage dysplasia (RCUD). Thus mitochondrial transporters SLC25 (SLC25A37 and SLC25A38) and ALAD genes were over-expressed in RARS. Moreover, significant differences were observed between patients with SF3B1 mutations and patients without the mutations. The deregulation of genes involved in iron and mitochondrial metabolism provides new insights in our knowledge of MDS-RS. New variants that could be involved in the pathogenesis of these diseases have been identified. PMID:25955609

  19. Expression of genes related to the regulation of muscle protein turnover in Angus and Nellore bulls.

    PubMed

    Busato, K C; Gomes, R A; Ladeira, M M; Duarte, M S; Freitas, N C; Rodrigues, A C; Chalfun-Junior, A; Paiva, L V; Chizzotti, M L

    2016-04-01

    We aimed to evaluate the expression of genes related to the regulation of muscle protein turnover in the longissimus dorsi (LD) muscle of Angus and Nellore bulls and to estimate the within-breed correlations of gene expression and performance traits. Thirteen genes related to the IGF-1 and myostatin pathways were studied. Thirteen animals, with an initial average BW of 381.2 ± 11.8 kg, from each breed were used in a completely randomized 2 × 2 factorial design (2 breeds and 2 feeding levels). The diet consisted of corn silage and a corn-soybean meal concentrate in a roughage-to-concentrate ratio of 30:70. Cattle were fed ad libitum (with 9 animals from each breed) or feed restricted (a 55% restriction of total DMI of ad libitum-fed animals, calculated as percentage of metabolic BW, with 4 animals of each breed). The experimental period lasted for 82 d and it was preceded by a 28-d adaptation period. The performance traits evaluated were slaughter body weight, total ADG (from d 1 to 82 of the trial), initial ADG (from d 1 to 41 of the trial), final ADG (from d 42 to 82 of the trial), total DMI (from d 1 to 82 of the trial), initial DMI (from d 1 to 41 of the trial), final DMI (from d 42 to 82 of the trial), HCW, LD weight (LDW), and rib eye area (REA). After slaughter, samples were taken from the LD muscle between the 12th and 13th ribs for gene expression analysis by quantitative reverse transcription PCR. There was no difference ( > 0.05) in the expression of any of the genes studied between ad libitum-fed Angus and ad libitum-fed Nellore, whereas feed restriction increased the expression of (; < 0.001), (; = 0.05), and (; = 0.04) and decreased the expression of ( < 0.01). The REA was negatively correlated to (; = 0.01), (; = 0.02), and ( = 0.05). The HCW was negatively correlated to ( = 0.01) and ( = 0.01) and tended to be negatively correlated to ( = 0.07), whereas the LDW tended to be negatively correlated to ( = 0.08). The genes , , and seem to be important

  20. Study of the influence of genes related to muscle oxidative processes on beef color.

    PubMed

    Falomir-Lockhart, A H; Rogberg-Muñoz, A; Papaleo-Mazzucco, J; Goszczynski, D E; Lirón, J P; Fernández, M E; Añon, M C; Melucci, L M; Giovambattista, G

    2015-10-01

    The biochemical bases of meat color are determined by the concentration and redox state of myoglobin, hemoglobin, cytochromes, and other pigments. Post-mortem depletion of cellular oxygen results in oxidative stresses that consume NADH and affects reducing activity, while enzymatic detoxification influences the cellular oxidative processes, both affecting meat color. The aim of this work was to study the influence of several genes related to cellular oxidative processes that could affect CIELAB meat color parameters. The study was performed in steers that received a grass-based diet combined with grain, hays and silages. Results suggest a possible link between colorimetric parameters (a*, b* and chroma) and SNPs in the GSTP1 gene (P<0.05). Although the influence of the enzymes, encoded by GSTP1 gene, on meat color has been proposed previously at biochemical level and protein expression level, further association studies in different populations and functional studies of proteins are needed to confirm the genetic determination of that gene on meat color. PMID:26010991

  1. Arsenic-induced alteration in the expression of genes related to type 2 diabetes mellitus

    SciTech Connect

    Diaz-Villasenor, Andrea Burns, Anna L.; Hiriart, Marcia; Cebrian, Mariano E.; Ostrosky-Wegman, Patricia

    2007-12-01

    Chronic exposure to high concentrations of arsenic in drinking water is associated with an increased risk for developing type 2 diabetes. The present revision focuses on the effect of arsenic on tissues that participate directly in glucose homeostasis, integrating the most important published information about the impairment of the expression of genes related to type 2 diabetes by arsenic as one of the possible mechanisms by which it leads to the disease. Many factors are involved in the manner in which arsenic contributes to the occurrence of diabetes. The reviewed studies suggest that arsenic might increase the risk for type 2 diabetes via multiple mechanisms, affecting a cluster of regulated events, which in conjunction trigger the disease. Arsenic affects insulin sensitivity in peripheral tissue by modifying the expression of genes involved in insulin resistance and shifting away cells from differentiation to the proliferation pathway. In the liver arsenic disturbs glucose production, whereas in pancreatic beta-cells arsenic decreases insulin synthesis and secretion and reduces the expression of antioxidant enzymes. The consequences of these changes in gene expression include the reduction of insulin secretion, induction of oxidative stress in the pancreas, alteration of gluconeogenesis, abnormal proliferation and differentiation pattern of muscle and adipocytes as well as peripheral insulin resistance.

  2. Transcriptional and functional studies of Acidithiobacillus ferrooxidans genes related to survival in the presence of copper.

    PubMed

    Navarro, Claudio A; Orellana, Luis H; Mauriaca, Cecilia; Jerez, Carlos A

    2009-10-01

    The acidophilic Acidithiobacillus ferrooxidans can resist exceptionally high copper (Cu) concentrations. This property is important for its use in biomining processes, where Cu and other metal levels range usually between 15 and 100 mM. To learn about the mechanisms that allow A. ferrooxidans cells to survive in this environment, a bioinformatic search of its genome showed the presence of at least 10 genes that are possibly related to Cu homeostasis. Among them are three genes coding for putative ATPases related to the transport of Cu (A. ferrooxidans copA1 [copA1(Af)], copA2(Af), and copB(Af)), three genes related to a system of the resistance nodulation cell division family involved in the extraction of Cu from the cell (cusA(Af), cusB(Af), and cusC(Af)), and two genes coding for periplasmic chaperones for this metal (cusF(Af) and copC(Af)). The expression of most of these open reading frames was studied by real-time reverse transcriptase PCR using A. ferrooxidans cells adapted for growth in the presence of high concentrations of Cu. The putative A. ferrooxidans Cu resistance determinants were found to be upregulated when this bacterium was exposed to Cu in the range of 5 to 25 mM. These A. ferrooxidans genes conferred to Escherichia coli a greater Cu resistance than wild-type cells, supporting their functionality. The results reported here and previously published data strongly suggest that the high resistance of the extremophilic A. ferrooxidans to Cu may be due to part or all of the following key elements: (i) a wide repertoire of Cu resistance determinants, (ii) the duplication of some of these Cu resistance determinants, (iii) the existence of novel Cu chaperones, and (iv) a polyP-based Cu resistance system. PMID:19666734

  3. De novo transcriptome sequencing and discovery of genes related to copper tolerance in Paeonia ostii.

    PubMed

    Wang, Yanjie; Dong, Chunlan; Xue, Zeyun; Jin, Qijiang; Xu, Yingchun

    2016-01-15

    Paeonia ostii, an important ornamental and medicinal plant, grows normally on copper (Cu) mines with widespread Cu contamination of soils, and it has the ability to lower Cu contents in the Cu-contaminated soils. However, very little molecular information concerned with Cu resistance of P. ostii is available. In this study, high-throughput de novo transcriptome sequencing was carried out for P. ostii with and without Cu treatment using Illumina HiSeq 2000 platform. A total of 77,704 All-unigenes were obtained with a mean length of 710 bp. Of these unigenes, 47,461 were annotated with public databases based on sequence similarities. Comparative transcript profiling allowed the discovery of 4324 differentially expressed genes (DEGs), with 2207 up-regulated and 2117 down-regulated unigenes in Cu-treated library as compared to the control counterpart. Based on these DEGs, Gene Ontology (GO) enrichment analysis indicated Cu stress-relevant terms, such as 'membrane' and 'antioxidant activity'. Meanwhile, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis uncovered some important pathways, including 'biosynthesis of secondary metabolites' and 'metabolic pathways'. In addition, expression patterns of 12 selected DEGs derived from quantitative real-time polymerase chain reaction (qRT-PCR) were consistent with their transcript abundance changes obtained by transcriptomic analyses, suggesting that all the 12 genes were authentically involved in Cu tolerance in P. ostii. This is the first report to identify genes related to Cu stress responses in P. ostii, which could offer valuable information on the molecular mechanisms of Cu resistance, and provide a basis for further genomics research on this and related ornamental species for phytoremediation. PMID:26435192

  4. Gene Ontology and KEGG Enrichment Analyses of Genes Related to Age-Related Macular Degeneration

    PubMed Central

    Zhang, Jian; Xing, ZhiHao; Ma, Mingming; Wang, Ning; Cai, Yu-Dong; Chen, Lei; Xu, Xun

    2014-01-01

    Identifying disease genes is one of the most important topics in biomedicine and may facilitate studies on the mechanisms underlying disease. Age-related macular degeneration (AMD) is a serious eye disease; it typically affects older adults and results in a loss of vision due to retina damage. In this study, we attempt to develop an effective method for distinguishing AMD-related genes. Gene ontology and KEGG enrichment analyses of known AMD-related genes were performed, and a classification system was established. In detail, each gene was encoded into a vector by extracting enrichment scores of the gene set, including it and its direct neighbors in STRING, and gene ontology terms or KEGG pathways. Then certain feature-selection methods, including minimum redundancy maximum relevance and incremental feature selection, were adopted to extract key features for the classification system. As a result, 720 GO terms and 11 KEGG pathways were deemed the most important factors for predicting AMD-related genes. PMID:25165703

  5. Identification of genes related to beak deformity of chickens using digital gene expression profiling.

    PubMed

    Bai, Hao; Zhu, Jing; Sun, Yanyan; Liu, Ranran; Liu, Nian; Li, Dongli; Wen, Jie; Chen, Jilan

    2014-01-01

    Frequencies of up to 3% of beak deformity (normally a crossed beak) occur in some indigenous chickens in China, such as and Beijing-You. Chickens with deformed beaks have reduced feed intake, growth rate, and abnormal behaviors. Beak deformity represents an economic as well as an animal welfare problem in the poultry industry. Because the genetic basis of beak deformity remains incompletely understood, the present study sought to identify important genes and metabolic pathways involved in this phenotype. Digital gene expression analysis was performed on deformed and normal beaks collected from Beijing-You chickens to detect global gene expression differences. A total of >11 million cDNA tags were sequenced, and 5,864,499 and 5,648,877 clean tags were obtained in the libraries of deformed and normal beaks, respectively. In total, 1,156 differentially expressed genes (DEG) were identified in the deformed beak with 409 being up-regulated and 747 down-regulated in the deformed beaks. qRT-PCR using eight genes was performed to verify the results of DGE profiling. Gene ontology (GO) analysis highlighted that genes of the keratin family on GGA25 were abundant among the DEGs. Pathway analysis showed that many DEGs were linked to the biosynthesis of unsaturated fatty acids and glycerolipid metabolism. Combining the analyses, 11 genes (MUC, LOC426217, BMP4, ACAA1, LPL, ALDH7A1, GLA, RETSAT, SDR16C5, WWOX, and MOGAT1) were highlighted as potential candidate genes for beak deformity in chickens. Some of these genes have been identified previously, while others have unknown function with respect to thus phenotype. To the best of our knowledge, this is the first genome-wide study to investigate the transcriptome differences in the deformed and normal beaks of chickens. The DEGs identified here are worthy of further functional characterization. PMID:25198128

  6. Genome-Wide Association Study to Identify Genes Related to Renal Mercury Concentrations in Mice

    PubMed Central

    Alkaissi, Hammoudi; Ekstrand, Jimmy; Jawad, Aksa; Nielsen, Jesper Bo; Havarinasab, Said; Soderkvist, Peter; Hultman, Per

    2016-01-01

    , Hultman P. 2016. Genome-wide association study to identify genes related to renal mercury concentrations in mice. Environ Health Perspect 124:920–926; http://dx.doi.org/10.1289/ehp.1409284 PMID:26942574

  7. Identification of Genes Related to Growth and Lipid Deposition from Transcriptome Profiles of Pig Muscle Tissue

    PubMed Central

    Chamba, Yangzom; Zhang, Bo; Shang, Peng; Zhang, Hao; Wu, Changxin

    2015-01-01

    Transcriptome profiles established using high-throughput sequencing can be effectively used for screening genome-wide differentially expressed genes (DEGs). RNA sequences (from RNA-seq) and microRNA sequences (from miRNA-seq) from the tissues of longissimus dorsi muscle of two indigenous Chinese pig breeds (Diannan Small-ear pig [DSP] and Tibetan pig [TP]) and two introduced pig breeds (Landrace [LL] and Yorkshire [YY]) were examined using HiSeq 2000 to identify and compare the differential expression of functional genes related to muscle growth and lipid deposition. We obtained 27.18 G clean data through the RNA-seq and detected that 18,208 genes were positively expressed and 14,633 of them were co-expressed in the muscle tissues of the four samples. In all, 315 DEGs were found between the Chinese pig group and the introduced pig group, 240 of which were enriched with functional annotations from the David database and significantly enriched in 27 Gene Ontology (GO) terms that were mainly associated with muscle fiber contraction, cadmium ion binding, response to organic substance and contractile fiber part. Based on functional annotation, we identified 85 DEGs related to growth traits that were mainly involved in muscle tissue development, muscle system process, regulation of cell development, and growth factor binding, and 27 DEGs related to lipid deposition that were mainly involved in lipid metabolic process and fatty acid biosynthetic process. With miRNA-seq, we obtained 23.78 M reads and 320 positively expressed miRNAs from muscle tissues, including 271 known pig miRNAs and 49 novel miRNAs. In those 271 known miRNAs, 20 were higher and 10 lower expressed in DSP-TP than in LL-YY. The target genes of the 30 miRNAs were mainly participated in MAPK, GnRH, insulin and Calcium signaling pathway and others involved cell development, growth and proliferation, etc. Combining the DEGs and the differentially expressed (DE) miRNAs, we drafted a network of 46 genes and 18

  8. Hypospadias and variants in genes related to sex hormone biosynthesis and metabolism

    PubMed Central

    Carmichael, SL; Witte, JS; Ma, C; Lammer, EJ; Shaw, GM

    2013-01-01

    We examined whether variants in genes related to sex hormone biosynthesis and metabolism were associated with hypospadias in humans. We examined 332 relatively common tagSNPs in 20 genes. Analyses included 633 cases (84 mild, 322 moderate, 212 severe, 15 undetermined severity) and 855 population-based non-malformed male controls born in California from 1990–2003. We used logistic regression models to estimate odds ratios (OR) and 95 percent confidence intervals (CI) for each SNP. Several of the 332 studied SNPs had p<0.01: one in CYP3A4, four in HSD17B3, one in HSD3B1, 2 in STARD3 10 in SRD5A2 and seven in STS. In addition, haplotype analyses gave several associations with p<0.01. For HSD17B3, 14-SNP and 5-SNP blocks had ORs of 1.5 (95% CI 1.1, 2.0, p<0.001) and 2.8 (95% CI 1.6, 4.8, p<0.001), respectively. For SRD5A2, 9-SNP, 3-SNP and 8-SNP blocks had ORs of 1.7 (95% CI 1.3, 2.2, p<0.001), 1.4 (95% CI 1.1, 1.8, p=0.008) and 1.5 (95% CI 1.2, 1.9, p=0.002), respectively. Our study indicates that several genes that contribute to sex hormone biosynthesis and metabolism are associated with hypospadias risk. PMID:24281767

  9. Tenm, a Drosophila gene related to tenascin, is a new pair-rule gene.

    PubMed Central

    Baumgartner, S; Martin, D; Hagios, C; Chiquet-Ehrismann, R

    1994-01-01

    We describe the molecular characterization of the Drosophila gene tenm, a large transcription unit spanning > 110 kb of DNA. tenm encodes a large extracellular protein of 2515 amino acids related to the extracellular matrix molecule tenascin. The Tenm protein is found in seven stripes during the blastoderm stage, and each stripe overlaps with the even-skipped stripes. tenm mutants show a phenotype resembling that of odd-paired (opa), a member of the pair-rule class of segmentation genes. Thus, Tenm is the first example of a pair-rule gene product acting from outside the cell. While the Tenm protein is under the control of fushi tarazu and even-skipped, but not of opa, at least two pair-rule genes, paired (prd) and sloppy paired (slp), and all segment-polarity genes analysed to date are under the control of tenm. Our data suggest that Tenm initiates a signal transduction cascade which acts, via or in concert with opa, on downstream targets such as prd, slp, gooseberry, engrailed and wingless, leading to an opa-like phenotype. Images PMID:8070401

  10. Comparative Transcriptome Analysis Identifies Candidate Genes Related to Skin Color Differentiation in Red Tilapia.

    PubMed

    Zhu, Wenbin; Wang, Lanmei; Dong, Zaijie; Chen, Xingting; Song, Feibiao; Liu, Nian; Yang, Hui; Fu, Jianjun

    2016-01-01

    Red tilapia is becoming more popular for aquaculture production in China in recent years. However, the pigmentation differentiation in genetic breeding is the main problem limiting its development of commercial red tilapia culture and the genetic basis of skin color variation is still unknown. In this study, we conducted Illumina sequencing of transcriptome on three color variety red tilapia. A total of 224,895,758 reads were generated, resulting in 160,762 assembled contigs that were used as reference contigs. The contigs of red tilapia transcriptome had hits in the range of 53.4% to 86.7% of the unique proteins of zebrafish, fugu, medaka, three-spined stickleback and tilapia. And 44,723 contigs containing 77,423 simple sequence repeats (SSRs) were identified, with 16,646 contigs containing more than one SSR. Three skin transcriptomes were compared pairwise and the results revealed that there were 148 common significantly differentially expressed unigenes and several key genes related to pigment synthesis, i.e. tyr, tyrp1, silv, sox10, slc24a5, cbs and slc7a11, were included. The results will facilitate understanding the molecular mechanisms of skin pigmentation differentiation in red tilapia and accelerate the molecular selection of the specific strain with consistent skin colors. PMID:27511178

  11. Comparative Transcriptome Analysis Identifies Candidate Genes Related to Skin Color Differentiation in Red Tilapia

    PubMed Central

    Zhu, Wenbin; Wang, Lanmei; Dong, Zaijie; Chen, Xingting; Song, Feibiao; Liu, Nian; Yang, Hui; Fu, Jianjun

    2016-01-01

    Red tilapia is becoming more popular for aquaculture production in China in recent years. However, the pigmentation differentiation in genetic breeding is the main problem limiting its development of commercial red tilapia culture and the genetic basis of skin color variation is still unknown. In this study, we conducted Illumina sequencing of transcriptome on three color variety red tilapia. A total of 224,895,758 reads were generated, resulting in 160,762 assembled contigs that were used as reference contigs. The contigs of red tilapia transcriptome had hits in the range of 53.4% to 86.7% of the unique proteins of zebrafish, fugu, medaka, three-spined stickleback and tilapia. And 44,723 contigs containing 77,423 simple sequence repeats (SSRs) were identified, with 16,646 contigs containing more than one SSR. Three skin transcriptomes were compared pairwise and the results revealed that there were 148 common significantly differentially expressed unigenes and several key genes related to pigment synthesis, i.e. tyr, tyrp1, silv, sox10, slc24a5, cbs and slc7a11, were included. The results will facilitate understanding the molecular mechanisms of skin pigmentation differentiation in red tilapia and accelerate the molecular selection of the specific strain with consistent skin colors. PMID:27511178

  12. DIA1R Is an X-Linked Gene Related to Deleted In Autism-1

    PubMed Central

    Aziz, Azhari; Harrop, Sean P.; Bishop, Naomi E.

    2011-01-01

    Background Autism spectrum disorders (ASDs) are frequently occurring disorders diagnosed by deficits in three core functional areas: social skills, communication, and behaviours and/or interests. Mental retardation frequently accompanies the most severe forms of ASDs, while overall ASDs are more commonly diagnosed in males. Most ASDs have a genetic origin and one gene recently implicated in the etiology of autism is the Deleted-In-Autism-1 (DIA1) gene. Methodology/Principal Findings Using a bioinformatics-based approach, we have identified a human gene closely related to DIA1, we term DIA1R (DIA1-Related). While DIA1 is autosomal (chromosome 3, position 3q24), DIA1R localizes to the X chromosome at position Xp11.3 and is known to escape X-inactivation. The gene products are of similar size, with DIA1 encoding 430, and DIA1R 433, residues. At the amino acid level, DIA1 and DIA1R are 62% similar overall (28% identical), and both encode signal peptides for targeting to the secretory pathway. Both genes are ubiquitously expressed, including in fetal and adult brain tissue. Conclusions/Significance Examination of published literature revealed point mutations in DIA1R are associated with X-linked mental retardation (XLMR) and DIA1R deletion is associated with syndromes with ASD-like traits and/or XLMR. Together, these results support a model where the DIA1 and DIA1R gene products regulate molecular traffic through the cellular secretory pathway or affect the function of secreted factors, and functional deficits cause disorders with ASD-like symptoms and/or mental retardation. PMID:21264219

  13. 6-Gingerol Protects against Nutritional Steatohepatitis by Regulating Key Genes Related to Inflammation and Lipid Metabolism

    PubMed Central

    Tzeng, Thing-Fong; Liou, Shorong-Shii; Chang, Chia Ju; Liu, I-Min

    2015-01-01

    Non-alcoholic fatty liver disease, including non-alcoholic steatohepatitis (NASH), appears to be increasingly common worldwide. The aim of the study was to investigate the effects of 6-gingerol ((S)-5-hydroxy-1-(4-hydroxy-3-methoxyphenyl)-3-decanone), a bioactive ingredient of plants belonging to the Zingiberaceae family, on experimental models of NASH. In HepG2 cells, 6-gingerol (100 μmol/L) treatment inhibited free fatty acids mixture (0.33 mmol/L palmitate and 0.66 mmol/L oleate)-induced triglyceride and inflammatory marker accumulations. Male C57BL/6 mice were fed with a methionine and choline-deficient (MCD) diet to induce steatohepatitis. After four weeks of MCD diet feeding, the mice were dosed orally with 6-gingerol (25, 50 or 100 mg/kg/day) once daily for another four weeks. 6-Gingerol (100 mg/kg/day) attenuated liver steatosis and necro-inflammation in MCD diet-fed mice. The expressions of inflammatory cytokine genes, including those for monocyte chemoattractant protein-1, tumor necrosis factor-α, and interleukin-6, and nuclear transcription factor (NF-κB), which were increased in the livers of MCD diet-fed mice, were attenuated by 6-gingerol. 6-Gingerol possesses a repressive property on hepatic steatosis, which is associated with induction of peroxisome proliferator-activated receptor α. Our study demonstrated the protective role of 6-gingerol in ameliorating nutritional steatohepatitis. The effect was mediated through regulating key genes related to lipid metabolism and inflammation. PMID:25658238

  14. [Screening genes related to 'black' character in wu-ding chicken by delta differential display].

    PubMed

    Wu, Ge-Min; Liu, Xiao-Chun; Lou, Yi-Zhou; Shi, Qi-Shun

    2005-01-01

    The 'black' character is a cardinal character of black chicken that is very important to the medical value and economical value of black chicken. The study compared gene expression between black chicken and non-black chicken by delta differential display method, and obtained twenty-nine expressed sequence tags(ESTs) by extraction, amplification, purification, northern blotting, cloning and sequencing of gel-embedded DNA. Of twenty-nine ESTs, eight ESTs have high similarity rate with known gene in chicken-beta-trepomyosin gene, cardiac myosin alkali light chain gene, insertionally activated c-Ha-ras oncogene, fra-2 oncogene, 16S rRNA gene and mitochondrion genome sequence, respectively, with similarity rate 97%,100%, 98%, 98%, 98%, 99%, 99% and 97%. Five ESTs have high similarity rate with human TTN gene, human phosphoglucomutase 5 gene, human or mouse signal recongnition particle 54 kD gene, human or mouse ribonuclease/angiogenin inhibitor gene, respectively, with similarity rate 82%, 82%, 87%, 99% and 99%. Their homologous genes in chicken are still not cloned. Thirteen ESTs are novel gene fragments. Of thirteen ESTs, nine ESTs have high similarity with ESTs in database. Four ESTs have not high similarity with any EST in database. The results of gene expression analysis show that one known gene (insertionally activated c-Ha-ras oncogene) fragment and one novel gene fragment are expressed in all non-black chicken and are not expressed in all black chicken, and that one known gene (signal recongnition particle 54 kD gene) fragment is expressed in all black chicken and is not expressed in all non-black chicken. According to these results, we conclude that the expression of insertionally activated c-Ha-ras oncogene, signal recongnition particle 54 kD gene and one novel gene are related to the 'black' character in chicken. PMID:15715435

  15. Characterization and expression analysis of dirigent family genes related to stresses in Brassica.

    PubMed

    Thamil Arasan, Senthil Kumar; Park, Jong-In; Ahmed, Nasar Uddin; Jung, Hee-Jeong; Hur, Yoonkang; Kang, Kwon-Kyoo; Lim, Yong-Pyo; Nou, Ill-Sup

    2013-06-01

    The dirigent (DIR) genes are playing a vital role in enhancing stress resistance in different crop plants. In this study, we collected 29 DIR like genes, two from a Brassica rapa cv. Osome full length cDNA library and 27 from the B. rapa database designated as B. rapa Dirigent (BrDIR) like genes. Sequence analysis and a comparison study of these genes confirmed that seven were dirigent and the remaining 22 were dirigent like genes. Expression analysis revealed an organ specific expression of these genes. BrDIR2 showed differential responses after Fusarium oxysporum f.sp. conglutinans infection in cabbage. Four Brassica oleracea dirigent like genes highly homologous to BrDIR2 also showed similar responses in cabbage plants infected with this fungus. Moreover, several BrDIR like genes showed significant responses after water, ABA and cold stress treatments in Chinese cabbage. Under water stress, most responsive genes showed the highest expression at 24 h, at which time the acid soluble lignin content of samples under the same stress condition were also highest, indicating a possible relationship between BrDIR like genes and lignin content. Taken together, our results indicate a protective role of BrDIR genes against biotic and abiotic stresses in Brassica. PMID:23562798

  16. [Estimation of the recombination fraction by the maximum likelihood method in mapping interacting genes relative to marker loci].

    PubMed

    Priiatkina, S N

    2002-05-01

    For mapping nonlinked interacting genes relative to marker loci, the recombination fractions can be calculated by using the log-likelihood functions were derived that permit estimation of recombinant fractions by solving the ML equations on the basis of F2 data at various types of interaction. In some cases, the recombinant fraction estimates are obtained in the analytical form while in others they are numerically calculated from concrete experimental data. With the same type of epistasis the log-functions were shown to differ depending on the functional role (suppression or epistasis) of the mapped gene. Methods for testing the correspondence of the model and the recombination fraction estimates to the experimental data are discussed. In ambiguous cases, analysis of the linked marker behavior makes it possible to differentiate gene interaction from distorted single-locus segregation, which at some forms of interaction imitate phenotypic ratios. PMID:12068553

  17. Integrating Genetic and Network Analysis to Characterize Genes Related to Mouse Weight

    PubMed Central

    Zhang, Bin; Wang, Susanna; Plaisier, Christopher; Castellanos, Ruth; Brozell, Alec; Schadt, Eric E; Drake, Thomas A

    2006-01-01

    Systems biology approaches that are based on the genetics of gene expression have been fruitful in identifying genetic regulatory loci related to complex traits. We use microarray and genetic marker data from an F2 mouse intercross to examine the large-scale organization of the gene co-expression network in liver, and annotate several gene modules in terms of 22 physiological traits. We identify chromosomal loci (referred to as module quantitative trait loci, mQTL) that perturb the modules and describe a novel approach that integrates network properties with genetic marker information to model gene/trait relationships. Specifically, using the mQTL and the intramodular connectivity of a body weight–related module, we describe which factors determine the relationship between gene expression profiles and weight. Our approach results in the identification of genetic targets that influence gene modules (pathways) that are related to the clinical phenotypes of interest. PMID:16934000

  18. Integrating genetic and network analysis to characterize genes related to mouse weight.

    PubMed

    Ghazalpour, Anatole; Doss, Sudheer; Zhang, Bin; Wang, Susanna; Plaisier, Christopher; Castellanos, Ruth; Brozell, Alec; Schadt, Eric E; Drake, Thomas A; Lusis, Aldons J; Horvath, Steve

    2006-08-18

    Systems biology approaches that are based on the genetics of gene expression have been fruitful in identifying genetic regulatory loci related to complex traits. We use microarray and genetic marker data from an F2 mouse intercross to examine the large-scale organization of the gene co-expression network in liver, and annotate several gene modules in terms of 22 physiological traits. We identify chromosomal loci (referred to as module quantitative trait loci, mQTL) that perturb the modules and describe a novel approach that integrates network properties with genetic marker information to model gene/trait relationships. Specifically, using the mQTL and the intramodular connectivity of a body weight-related module, we describe which factors determine the relationship between gene expression profiles and weight. Our approach results in the identification of genetic targets that influence gene modules (pathways) that are related to the clinical phenotypes of interest. PMID:16934000

  19. Transcriptome Analysis of Blunt Snout Bream (Megalobrama amblycephala) Reveals Putative Differential Expression Genes Related to Growth and Hypoxia

    PubMed Central

    Li, Fu-Gui; Chen, Jie; Jiang, Xia-Yun; Zou, Shu-Ming

    2015-01-01

    The blunt snout bream (Megalobrama amblycephala) is an important freshwater aquaculture species, but it is sensitive to hypoxia. No transcriptome data related to growth and hypoxia response are available for this species. In this study, we performed de novo transcriptome sequencing for the liver and gills of the fast-growth family and slow-growth family derived from ‘Pujiang No.1’ F10 blunt snout bream that were under hypoxic stress and normoxia, respectively. The fish were divided into the following 4 groups: fast-growth family under hypoxic stress, FH; slow-growth family under hypoxic stress, SH; fast-growth family under normoxia, FN; and slow-growth family under normoxia, SN. A total of 185 million high-quality reads were obtained from the normalized cDNA of the pooled samples, which were assembled into 465,582 contigs and 237,172 transcripts. A total of 31,338 transcripts from the same locus (unigenes) were annotated and assigned to 104 functional groups, and 23,103 unigenes were classified into seven main categories, including 45 secondary KEGG pathways. A total of 22,255 (71%) known putative unigenes were found to be shared across the genomes of five model fish species and mammals, and a substantial number (9.4%) of potentially novel genes were identified. When 6,639 unigenes were used in the analysis of differential expression (DE) genes, the number of putative DE genes related to growth pathways in FH, SH, SN and FN was 159, 118, 92 and 65 in both the liver and gills, respectively, and the number of DE genes related to hypoxic response was 57, 33, 23 and 21 in FH, FN, SH and SN, respectively. Our results suggest that growth performance of the fast-growth family should be due to complex mutual gene regulatory mechanisms of these putative DE genes between growth and hypoxia. PMID:26554582

  20. A patient with PMP22-related hereditary neuropathy and DBH-gene-related dysautonomia.

    PubMed

    Bartoletti-Stella, Anna; Chiaro, Giacomo; Calandra-Buonaura, Giovanna; Contin, Manuela; Scaglione, Cesa; Barletta, Giorgio; Cecere, Annagrazia; Garagnani, Paolo; Tieri, Paolo; Ferrarini, Alberto; Piras, Silvia; Franceschi, Claudio; Delledonne, Massimo; Cortelli, Pietro; Capellari, Sabina

    2015-10-01

    Recurrent focal neuropathy with liability to pressure palsies is a relatively frequent autosomal-dominant demyelinating neuropathy linked to peripheral myelin protein 22 (PMP22) gene deletions. The combination of PMP22 gene mutations with other genetic variants is known to cause a more severe phenotype than expected. We present the case of a patient with severe orthostatic hypotension since 12 years of age, who inherited a PMP22 gene deletion from his father. Genetic double trouble was suspected because of selective sympathetic autonomic disturbances. Through exome-sequencing analysis, we identified two novel mutations in the dopamine beta hydroxylase gene. Moreover, with interactome analysis, we excluded a further influence on the origin of the disease by variants in other genes. This case increases the number of unique patients presenting with dopamine-β-hydroxylase deficiency and of cases with genetically proven double trouble. Finding the right, complete diagnosis is crucial to obtain adequate medical care and appropriate genetic counseling. PMID:26410747

  1. Discovery of genes related to witches broom disease in Paulownia tomentosa × Paulownia fortunei by a De Novo assembled transcriptome.

    PubMed

    Liu, Rongning; Dong, Yanpeng; Fan, Guoqiang; Zhao, Zhenli; Deng, Minjie; Cao, Xibing; Niu, Suyan

    2013-01-01

    In spite of its economic importance, very little molecular genetics and genomic research has been targeted at the family Paulownia spp. The little genetic information on this plant is a big obstacle to studying the mechanisms of its ability to resist Paulownia Witches' Broom (PaWB) disease. Analysis of the Paulownia transcriptome and its expression profile data are essential to extending the genetic resources on this species, thus will greatly improves our studies on Paulownia. In the current study, we performed the de novo assembly of a transcriptome on P. tomentosa × P. fortunei using the short-read sequencing technology (Illumina). 203,664 unigenes with a mean length of 1,328 bp was obtained. Of these unigenes, 32,976 (30% of all unigenes) containing complete structures were chosen. Eukaryotic clusters of orthologous groups, gene orthology, and the Kyoto Encyclopedia of Genes and Genomes annotations were performed of these unigenes. Genes related to PaWB disease resistance were analyzed in detail. To our knowledge, this is the first study to elucidate the genetic makeup of Paulownia. This transcriptome provides a quick way to understanding Paulownia, increases the number of gene sequences available for further functional genomics studies and provides clues to the identification of potential PaWB disease resistance genes. This study has provided a comprehensive insight into gene expression profiles at different states, which facilitates the study of each gene's roles in the developmental process and in PaWB disease resistance. PMID:24278262

  2. Genetic Association Analysis of 30 Genes Related to Obesity in a European American Population

    PubMed Central

    Li, Peng; Tiwari, Hemant K.; Lin, Wan-Yu; Allison, David B.; Chung, Wendy K.; Leibel, Rudolph L.; Yi, Nengjun; Liu, Nianjun

    2013-01-01

    Objective Obesity, which is frequently associated with diabetes, hypertension, and cardiovascular diseases, is primarily the result of a net excess of caloric intake over energy expenditure. Human obesity is highly heritable, but the specific genes mediating susceptibility in non-syndromic obesity remain unclear. We tested candidate genes in pathways related to food intake and energy expenditure for association with body mass index (BMI). Methods We re-analyzed 355 common genetic variants of 30 candidate genes in 7 molecular pathways related to obesity in 1,982 unrelated European Americans from the New York Health Project. Data were analyzed by using a Bayesian hierarchical generalized linear model. The BMIs were log-transformed and then adjusted for covariates including age, age2, gender, and diabetes status. The single nucleotide polymorphisms (SNPs) were modeled as additive effects. Results With the stipulated adjustments, nine SNPs in eight genes were significantly associated with BMI: GHRL (rs35683), AGRP (rs5030980), CPE (rs1946816 and rs4481204), GLP1R (rs2268641), HTR2A (rs912127), NPY5R (Y5R1c52), SOCS3 (rs4969170), and STAT3 (rs4796793). We also found a gender-by-SNP interaction (rs1745837 in HTR2A), which indicated that variants in the gene HTR2A had a stronger association with BMI in males. In addition, NPY1R was detected as having a significant gene effect even though none of the SNPs in this gene was significant. Conclusion Variations in genes AGRP, CPE, GHRL, GLP1R, HTR2A, NPY1R, NPY5R, SOCS3, and STAT3 showed modest associations with BMI in European Americans. The pathways in which these genes participate regulate energy intake and thus these associations are mechanistically plausible in this context. PMID:23900445

  3. Transcriptome sequencing of Hydrangea macrophylla to uncover genes related to reblooming and powdery mildew resistance

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Massively parallel pyrosequencing technology has been used extensively on agronomic crop and model plants. Transcriptome sequencing is a useful first step in functional genomic studies, microarray and gene expression studies, single nucleotide polymorphism (SNP) surveys, quantitative trait loci (QT...

  4. A Drosophila muscle-specific gene related to the mouse quaking locus.

    PubMed

    Fyrberg, C; Becker, J; Barthmaier, P; Mahaffey, J; Fyrberg, E

    1997-09-15

    We have characterized a novel muscle-specific gene of Drosophila melanogaster, defined by enhancer trap strain 24B of Brand and Perrimon (1993). We show that transcripts of the gene accumulate within presumptive mesoderm and persist within developing muscles, strongly suggesting that the encoded protein is involved in muscle cell determination and differentiation. cDNA sequences reveal that the Drosophila protein is similar to quaking (64% identity over 210 amino acids), a protein essential for mouse embryogenesis, and gld-1 (53% identity over 162 amino acids) a germ-line-specific tumor suppressing protein of the nematode, Caenorhabditis elegans. We demonstrate that the Drosophila gene resides within the 93F chromosome subdivision, and describe its physical map. Finally, we have used the gene, which we have named quaking-related 93F (qkr93F), to identify a family of closely related KH domains. PMID:9332381

  5. Genes related to xylose fermentation and methods of using same for enhanced biofuel production

    DOEpatents

    Wohlbach, Dana J.; Gasch, Audrey P.

    2015-09-29

    The present invention provides isolated gene sequences involved in xylose fermentation and related recombinant yeast which are useful in methods of enhanced biofuel production, particularly ethanol production. Methods of bioengineering recombinant yeast useful for biofuel production are also provided.

  6. Genes related to xylose fermentation and methods of using same for enhanced biofuel production

    DOEpatents

    Wohlbach, Dana J.; Gasch, Audrey P.

    2014-08-05

    The present invention provides isolated gene sequences involved in xylose fermentation and related recombinant yeast which are useful in methods of enhanced biofuel production, particularly ethanol production. Methods of bioengineering recombinant yeast useful for biofuel production are also provided.

  7. Identification and expression analysis of chitinase genes related to biotic stress resistance in Brassica.

    PubMed

    Ahmed, Nasar Uddin; Park, Jong-In; Seo, Mi-Suk; Kumar, Thamilarasan Senthil; Lee, In-Ho; Park, Beom-Seok; Nou, Ill-Sup

    2012-04-01

    Brassica is a very important vegetable group because of its contribution to human nutrition and consequent economic benefits. However, biotic stress is a major concern for these crops and molecular biology techniques offer the most efficient of approaches to address this concern. Chitinase is an important biotic stress resistance-related gene. We identified three genes designated as Brassica chitinase like protein (BrCLP1), BrCLP2 and BrCLP3 from a full-length cDNA library of Brassica rapa cv. Osome. Sequence analysis of these genes confirmed that BrCLP1 was a class IV chitinase, and BrCLP2 and BrCLP3 were class VII chitinases. Also, these genes showed a high degree of homology with other biotic stress resistance-related plant chitinases. In expression analysis, organ-specific expression of all three genes was high except BrCLP1 in all the organs tested and BrCLP2 showed the highest expression compared to the other genes in flower buds. All these genes also showed expression during all developmental growth stages of Chinese cabbage. In addition, BrCLP1 was up-regulated with certain time of infection by Pectobacterium carotovorum subsp. carotovorum in Chinese cabbage plants during microarray expression analysis. On the other hand, expression of BrCLP2 and BrCLP3 were increased after 6 h post inoculation (hpi) but decreased from 12 hpi. All these data suggest that these three chitinase genes may be involved in plant resistance against biotic stresses. PMID:21720758

  8. Liver metal levels and expression of genes related to iron homeostasis in rhesus monkeys after inhalational manganese exposure

    PubMed Central

    Pettiglio, Michael A.; Herrera, Carolina; Foster, Melanie L.; Dorman, David C.; Bartnikas, Thomas B.

    2016-01-01

    Here we present data on liver metal levels and expression of genes related to iron homeostasis in rhesus monkeys after inhalational manganese exposure. Archived liver samples from rhesus monkeys exposed to 0 (n=6), 0.06 (n=6), 0.3 (n=4) and 1.5 (n=4) mg/m3 manganese inhalation for 65 days were obtained from a published study (“Tissue manganese concentrations in young male rhesus monkeys following subchronic manganese sulfate inhalation” [1]). Samples were analyzed by spectroscopy, immunoblotting and quantitative PCR to assess metal levels and gene expression. Liver manganese and iron levels were linearly correlated although only the intermediate manganese exposure level (0.3 mg Mn/m3) led to a statistically significant increase in liver iron levels. PMID:26958631

  9. Liver metal levels and expression of genes related to iron homeostasis in rhesus monkeys after inhalational manganese exposure.

    PubMed

    Pettiglio, Michael A; Herrera, Carolina; Foster, Melanie L; Dorman, David C; Bartnikas, Thomas B

    2016-03-01

    Here we present data on liver metal levels and expression of genes related to iron homeostasis in rhesus monkeys after inhalational manganese exposure. Archived liver samples from rhesus monkeys exposed to 0 (n=6), 0.06 (n=6), 0.3 (n=4) and 1.5 (n=4) mg/m(3) manganese inhalation for 65 days were obtained from a published study ("Tissue manganese concentrations in young male rhesus monkeys following subchronic manganese sulfate inhalation" [1]). Samples were analyzed by spectroscopy, immunoblotting and quantitative PCR to assess metal levels and gene expression. Liver manganese and iron levels were linearly correlated although only the intermediate manganese exposure level (0.3 mg Mn/m(3)) led to a statistically significant increase in liver iron levels. PMID:26958631

  10. Identifying genes related to choriogenesis in insect panoistic ovaries by Suppression Subtractive Hybridization

    PubMed Central

    Irles, Paula; Bellés, Xavier; Piulachs, M Dolors

    2009-01-01

    Background Insect ovarioles are classified into two categories: panoistic and meroistic, the later having apparently evolved from an ancestral panoistic type. Molecular data on oogenesis is practically restricted to meroistic ovaries. If we aim at studying the evolutionary transition from panoistic to meroistic, data on panoistic ovaries should be gathered. To this end, we planned the construction of a Suppression Subtractive Hybridization (SSH) library to identify genes involved in panoistic choriogenesis, using the cockroach Blattella germanica as model. Results We constructed a post-vitellogenic ovary library by SSH to isolate genes involved in choriogenesis in B. germanica. The tester library was prepared with an ovary pool from 6- to 7-day-old females, whereas the driver library was prepared with an ovary pool from 3- to 4-day-old females. From the SSH library, we obtained 258 high quality sequences which clustered into 34 unique sequences grouped in 19 contigs and 15 singlets. The sequences were compared against non-redundant NCBI databases using BLAST. We found that 44% of the unique sequences had homologous sequences in known genes of other organisms, whereas 56% had no significant similarity to any of the databases entries. A Gene Ontology analysis was carried out, classifying the 34 sequences into different functional categories. Seven of these gene sequences, representative of different categories and processes, were chosen to perform expression studies during the first gonadotrophic cycle by real-time PCR. Results showed that they were mainly expressed during post-vitellogenesis, which validates the SSH technique. In two of them corresponding to novel genes, we demonstrated that they are specifically expressed in the cytoplasm of follicular cells in basal oocytes at the time of choriogenesis. Conclusion The SSH approach has proven to be useful in identifying ovarian genes expressed after vitellogenesis in B. germanica. For most of the genes, functions

  11. Discovery of Genes Related to Witches Broom Disease in Paulownia tomentosa × Paulownia fortunei by a De Novo Assembled Transcriptome

    PubMed Central

    Liu, Rongning; Dong, Yanpeng; Fan, Guoqiang; Zhao, Zhenli; Deng, Minjie; Cao, Xibing; Niu, Suyan

    2013-01-01

    In spite of its economic importance, very little molecular genetics and genomic research has been targeted at the family Paulownia spp. The little genetic information on this plant is a big obstacle to studying the mechanisms of its ability to resist Paulownia Witches’ Broom (PaWB) disease. Analysis of the Paulownia transcriptome and its expression profile data are essential to extending the genetic resources on this species, thus will greatly improves our studies on Paulownia. In the current study, we performed the de novo assembly of a transcriptome on P. tomentosa × P. fortunei using the short-read sequencing technology (Illumina). 203,664 unigenes with a mean length of 1,328 bp was obtained. Of these unigenes, 32,976 (30% of all unigenes) containing complete structures were chosen. Eukaryotic clusters of orthologous groups, gene orthology, and the Kyoto Encyclopedia of Genes and Genomes annotations were performed of these unigenes. Genes related to PaWB disease resistance were analyzed in detail. To our knowledge, this is the first study to elucidate the genetic makeup of Paulownia. This transcriptome provides a quick way to understanding Paulownia, increases the number of gene sequences available for further functional genomics studies and provides clues to the identification of potential PaWB disease resistance genes. This study has provided a comprehensive insight into gene expression profiles at different states, which facilitates the study of each gene’s roles in the developmental process and in PaWB disease resistance. PMID:24278262

  12. Single nucleotide polymorphisms in candidate genes related to daughter pregnancy rate in Holstein cows

    Technology Transfer Automated Retrieval System (TEKTRAN)

    ABSTRACT: Previously, a candidate gene approach identified 40 SNPs associated with daughter pregnancy rate (DPR) in dairy bulls. We evaluated 39 of these SNPs for relationship to DPR in a separate population of Holstein cows grouped on their predicted transmitting ability for DPR: <= -1 (n=1266) a...

  13. Effect of Human Ovarian Tissue Vitrification/Warming on the Expression of Genes Related to Folliculogenesis

    PubMed Central

    Shams Mofarahe, Zahra; Ghaffari Novin, Marefat; Jafarabadi, Mina; Salehnia, Mojdeh; Noroozian, Mohsen; Ghorbanmehr, Nassim

    2015-01-01

    Background: Ovarian tissue cryopreservation is an alternative strategy to preserve the fertility of women predicted to undergo premature ovarian failure. This study was designed to evaluate the expression of folliculogenesis-related genes, including factor in the germline alpha (FIGLA), growth differentiation factor-9 (GDF-9), follicle-stimulating hormone receptor (FSHR), and KIT LIGAND after vitrification/warming of human ovarian tissue. Methods: Human ovarian tissue samples were collected from five transsexual women. In the laboratory, the ovarian medullary part was removed by a surgical blade, and the cortical tissue was cut into small pieces. Some pieces were vitrified and warmed and the others were considered as non-vitrified group (control). Follicular normality was assessed with morphological observation by a light microscope, and the expression of FIGLA, KIT LIGAND, GDF-9,, and FSHR genes was examined using real-time RT-PCR in both the vitrified and non-vitrified groups. Results: Overall, 85% of the follicles preserved their normal morphologic feature after warming. The percentage of normal follicles and the expression of FIGLA, KIT LIGAND, GDF-9, and FSHR genes were similar in both vitrified and non-vitrified groups (P > 0.05). Conclusion: Vitrification/warming of human ovarian tissue had no remarkable effect on the expression of folliculogenesis-related genes. PMID:26175108

  14. TEMPERATURE CONDITIONING ALTERS TRANSCRIPT ABUNDANCE OF GENES RELATED TO CHILLING STRESS IN GRAPEFRUIT.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Grapefruit (Citrus paradisi) are susceptible to chilling injury (CI) if held at temperatures below about 10C. Changes in transcript abundance for a number of genes have been correlated with chilling stress in citrus fruit. We tested the hypothesis that conditioning affects transcript abundance of ...

  15. Molecular cloning and characterization of beta-expansin gene related to root hair formation in barley.

    PubMed

    Kwasniewski, Miroslaw; Szarejko, Iwona

    2006-07-01

    Root hairs are specialized epidermal cells that play a role in the uptake of water and nutrients from the rhizosphere and serve as a site of interaction with soil microorganisms. The process of root hair formation is well characterized in Arabidopsis (Arabidopsis thaliana); however, there is a very little information about the genetic and molecular basis of root hair development in monocots. Here, we report on isolation and cloning of the beta-expansin (EXPB) gene HvEXPB1, tightly related to root hair initiation in barley (Hordeum vulgare). Using root transcriptome differentiation in the wild-type/root-hairless mutant system, a cDNA fragment present in roots of wild-type plants only was identified. After cloning of full-length cDNA and genomic sequences flanking the identified fragment, the subsequent bioinformatics analyses revealed homology of the protein coded by the identified gene to the EXPB family. Reverse transcription-PCR showed that expression of HvEXPB1 cosegregated with the root hair phenotype in F2 progeny of the cross between the hairless mutant rhl1.a and the wild-type Karat parent variety. Expression of the HvEXPB1 gene was root specific; it was expressed in roots of wild-type forms, but not in coleoptiles, leaves, tillers, and spikes. The identified gene was active in roots of two other analyzed root hair mutants: rhp1.a developing root hair primordia only and rhs1.a with very short root hairs. Contrary to this, a complete lack of HvEXPB1 expression was observed in roots of the spontaneous root-hairless mutant bald root barley. All these observations suggest a role of the HvEXPB1 gene in the process of root hair formation in barley. PMID:16679418

  16. Novel Genes Related to Ceftriaxone Resistance Found among Ceftriaxone-Resistant Neisseria gonorrhoeae Strains Selected In Vitro.

    PubMed

    Gong, Zijian; Lai, Wei; Liu, Min; Hua, Zhengshuang; Sun, Yayin; Xu, Qingfang; Xia, Yue; Zhao, Yue; Xie, Xiaoyuan

    2016-04-01

    The emergence of ceftriaxone-resistantNeisseria gonorrhoeaeis currently a global public health concern. However, the mechanism of ceftriaxone resistance is not yet fully understood. To investigate the potential genes related to ceftriaxone resistance inNeisseria gonorrhoeae, we subcultured six gonococcal strains with increasing concentrations of ceftriaxone and isolated the strains that became resistant. After analyzing several frequently reported genes involved in ceftriaxone resistance, we found only a single mutation inpenA(A501V). However, differential analysis of the genomes and transcriptomes between pre- and postselection strains revealed many other mutated genes as well as up- and downregulated genes. Transformation of the mutatedpenAgene into nonresistant strains increased the MIC between 2.0- and 5.3-fold, and transformation of mutatedftsXincreased the MIC between 3.3- and 13.3-fold. Genes encoding the ABC transporters FarB, Tfq, Hfq, and ExbB were overexpressed, whilepilM,pilN, andpilQwere downregulated. Furthermore, the resistant strain developed cross-resistance to penicillin and cefuroxime, had an increased biochemical metabolic rate, and presented fitness defects such as prolonged growth time and downregulated PilMNQ. In conclusion, antimicrobial pressure could result in the emergence of ceftriaxone resistance, and the evolution of resistance ofNeisseria gonorrhoeaeto ceftriaxone is a complicated process at both the pretranscriptional and posttranscriptional levels, involving several resistance mechanisms of increased efflux and decreased entry. PMID:26787702

  17. EYA4, a novel vertebrate gene related to Drosophila eyes absent.

    PubMed

    Borsani, G; DeGrandi, A; Ballabio, A; Bulfone, A; Bernard, L; Banfi, S; Gattuso, C; Mariani, M; Dixon, M; Donnai, D; Metcalfe, K; Winter, R; Robertson, M; Axton, R; Brown, A; van Heyningen, V; Hanson, I

    1999-01-01

    We have isolated a family of four vertebrate genes homologous to eyes absent (eya), a key regulator of ocular development in Drosophila. Here we present the detailed characterization of the EYA4 gene in human and mouse. EYA4 encodes a 640 amino acid protein containing a highly conserved C-terminal domain of 271 amino acids which in Drosophila eya is known to mediate developmentally important protein-protein interactions. Human EYA4 maps to 6q23 and mouse Eya4 maps to the predicted homology region near the centromere of chromosome 10. In the developing mouse embryo, Eya4 is expressed primarily in the craniofacial mesenchyme, the dermamyotome and the limb. On the basis of map position and expression pattern, EYA4 is a candidate for oculo-dento-digital (ODD) syndrome, but no EYA4 mutations were found in a panel of ODD patients. PMID:9887327

  18. Molecular cloning of genes related to aflatoxin biosynthesis by differential screening.

    PubMed Central

    Feng, G H; Chu, F S; Leonard, T J

    1992-01-01

    A differential hybridization strategy was used to clone genes associated with aflatoxin biosynthesis. A genomic library, formed between nuclear DNA and the pUC19 plasmid, was screened with three different cDNA probes by the colony hybridization procedure. Nineteen clones were selected; all were positively correlated with and presumably enriched with genes associated with aflatoxin production. Some of these clones were further characterized by using them as probes in Northern (RNA blot) hybridizations. Five clones hybridized strongly with some polyadenylated RNAs formed during the transition to or during idiophase when aflatoxin was produced. However, little or no corresponding hybridization occurred with polyadenylated RNAs formed in early and mid-log growth phase. Two of the clones were further used as probes to hybridize with polyadenylated RNAs formed under aflatoxin-permissive and nonpermissive temperatures. Hybridization occurred with RNA species formed under the permissive temperature only. Images PMID:1610169

  19. Cloning of murine G1RP, a novel gene related to Drosophila melanogaster g1.

    PubMed

    Baker, S J; Reddy, E P

    2000-05-01

    To study the nature of genes that are induced during the apoptotic death of myeloid precursor cells, we performed representational difference analysis (RDA) using 32Dcl3 myeloblastic cells that were deprived of IL-3 for 24h. We have isolated a novel cDNA (g1-related protein, G1RP) that is homologous to g1, a Drosophila melanogaster zinc-finger protein that is expressed in the mesoderm. Northern blot analysis using RNAs derived from 32Dcl3 cells that have been grown in the absence of IL-3 demonstrates that the G1RP message is upregulated in these cells following the removal of IL-3, suggesting that this gene may regulate growth factor withdrawal-induced apoptosis of myeloid precursor cells. PMID:10806348

  20. Odd-skipped related 2 regulates genes related to proliferation and development

    SciTech Connect

    Kawai, Shinji; Abiko, Yoshimitsu; Amano, Atsuo

    2010-07-23

    Cell proliferation is a biological process in which chromosomes replicate in one cell and equally divide into two daughter cells. Our previous findings suggested that Odd-skipped related 2 (Osr2) plays an important role in cellular quiescence and proliferation under epigenetic regulation. However, the mechanism used by Osr2 to establish and maintain proliferation is unknown. To examine the functional role of Osr2 in cell proliferation, we analyzed its downstream target genes using microarray analysis following adenovirus-induced overexpression of Osr2 as well as knockdown with Osr2 siRNA, which showed that Osr2 regulates a multitude of genes involved in proliferation and the cell cycle, as well as development. Additional proliferation assays also indicated that Osr2 likely functions to elicit cell proliferation. Together, these results suggest that Osr2 plays important roles in proliferation and development.

  1. Expression Profile of Genes Related to Drug Metabolism in Human Brain Tumors

    PubMed Central

    Stavrinou, Pantelis; Mavrogiorgou, Maria-Christina; Polyzoidis, Konstantinos; Kreft-Kerekes, Vincenzo; Timmer, Marco; Marselos, Marios; Pappas, Periklis

    2015-01-01

    Background Endogenous and exogenous compounds as well as carcinogens are metabolized and detoxified by phase I and II enzymes, the activity of which could be crucial to the inactivation and hence susceptibility to carcinogenic factors. The expression of these enzymes in human brain tumor tissue has not been investigated sufficiently. We studied the association between tumor pathology and the expression profile of seven phase I and II drug metabolizing genes (CYP1A1, CYP1B1, ALDH3A1, AOX1, GSTP1, GSTT1 and GSTM3) and some of their proteins. Methods Using qRT-PCR and western blotting analysis the gene and protein expression in a cohort of 77 tumors were investigated. The major tumor subtypes were meningioma, astrocytoma and brain metastases, -the later all adenocarcinomas from a lung primary. Results Meningeal tumors showed higher expression levels for AOX1, CYP1B1, GSTM3 and GSTP1. For AOX1, GSTM and GSTP1 this could be verified on a protein level as well. A negative correlation between the WHO degree of malignancy and the strength of expression was identified on both transcriptional and translational level for AOX1, GSTM3 and GSTP1, although the results could have been biased by the prevalence of meningiomas and glioblastomas in the inevitably bipolar distribution of the WHO grades. A correlation between the gene expression and the protein product was observed for AOX1, GSTP1 and GSTM3 in astrocytomas. Conclusions The various CNS tumors show different patterns of drug metabolizing gene expression. Our results suggest that the most important factor governing the expression of these enzymes is the histological subtype and to a far lesser extent the degree of malignancy itself. PMID:26580399

  2. Transcriptomic Analysis Reveals Key Genes Related to Betalain Biosynthesis in Pulp Coloration of Hylocereus polyrhizus

    PubMed Central

    Qingzhu, Hua; Chengjie, Chen; Zhe, Chen; Pengkun, Chen; Yuewen, Ma; Jingyu, Wu; Jian, Zheng; Guibing, Hu; Jietang, Zhao; Yonghua, Qin

    2016-01-01

    Betalains have high nutritional value and bioactivities. Red pulp pitaya (Hylocereus polyrhizus) is the only fruit containing abundant betalains for consumer. However, no information is available about genes involved in betalain biosynthesis in H. polyrhizus. Herein, two cDNA libraries of pitaya pulps with two different coloration stages (white and red pulp stages) of Guanhuahong (H. polyrhizus) were constructed. A total of about 12 Gb raw RNA-Seq data was generated and was de novo assembled into 122,677 transcripts with an average length of 1183 bp and an N50 value of 2008. Approximately 99.99% of all transcripts were annotated based on seven public databases. A total of 8871 transcripts were significantly regulated. Thirty-three candidate transcripts related to betalain biosynthesis were obtained from the transcriptome data. Transcripts encoding enzymes involved in betalain biosynthesis were analyzed using RT-qPCR at the whole pulp coloration stages of H. polyrhizus (7-1) and H. undatus (132-4). Nine key transcripts of betalain biosynthesis were identified. They were assigned to four kinds of genes in betalain biosynthetic pathway, including tyrosinase, 4, 5-DOPA dioxygenase extradiol, cytochrome P450 and glucosyltransferase. Ultimately, a preliminary betalain biosynthetic pathway for pitaya was proposed based on betalain analyses, gene expression profiles and published documents. PMID:26779215

  3. Low cadmium (LCD), a novel gene related to cadmium tolerance and accumulation in rice.

    PubMed

    Shimo, Hugo; Ishimaru, Yasuhiro; An, Gynheung; Yamakawa, Takashi; Nakanishi, Hiromi; Nishizawa, Naoko K

    2011-11-01

    The contamination of food crops by cadmium (Cd) is a major concern in food production because it can reduce crop yields and threaten human health. In this study, knockout rice plants (Oryza sativa) tagged with the gene trap vector pGA2707 were screened for Cd tolerance, and the tolerant line lcd was obtained. The lcd mutant showed tolerance to Cd on agar plates and in hydroponic culture during early plant development. Metal concentration measurements in hydroponically grown plants revealed significantly less Cd in the shoots of lcd plants compared with wild-type (WT) shoots. When cultured in the field in soil artificially contaminated with low levels of Cd, lcd showed no significant difference in the Cd content of its leaf blades; however, the Cd concentration in the grains was 55% lower in 2009 and 43% lower in 2010. There were no significant differences in plant dry weight or seed yield between lcd and wild-type plants. LCD, a novel gene, is not homologous to any other known gene. LCD localized to the cytoplasm and nucleus, and was expressed mainly in the vascular tissues in the roots and phloem companion cells in the leaves. These data indicate that lcd may be useful for understanding Cd transport mechanisms and is a promising candidate rice line for use in combating the threat of Cd to human health. PMID:21908474

  4. Steroids and genes related to steroid biosynthesis in the female giant freshwater prawn, Macrobrachium rosenbergii.

    PubMed

    Thongbuakaew, Tipsuda; Siangcham, Tanapan; Suwansa-ard, Saowaros; Elizur, Abigail; Cummins, Scott F; Sobhon, Prasert; Sretarugsa, Prapee

    2016-03-01

    The giant freshwater prawn, Macrobrachium rosenbergii, is important to many Asian countries due to its high economic value as an aquaculture product. With demand increasing, there is requirement for a better understanding of the biosynthetic components that regulate its growth and reproduction, including steroids, in order to help increase production. Vertebrate-type steroids and their receptors were identified in crustaceans and implicated in reproduction. In this study, we presented the sex steroids estradiol and progesterone by LC-MS/MS in female M. rosenbergii, and reveal steroidogenic-related genes by in silico analysis of de novo assembled transcriptomes. Comparative analysis with other species was performed to confirm their putative role, as well as tissue-specific and quantitative gene expression. We reveal 29 transcripts that encode for steroidogenic-related proteins, including steroidogenic enzymes, a nuclear steroid hormone receptors, and a steroidogenic factor. Moreover, we identified for the first time the presence of steroidogenic factor 1, StAR-related lipid transfer protein, estradiol receptor- and progesterone-like protein in M. rosenbergii. Those targeted for gene expression analysis (3 beta-hydroxysteroid dehydrogenase, 17 beta-hydroxysteroid dehydrogenase, estrogen sulfotransferase and progesterone receptor-like) showed widespread expression within many tissues, and at relatively high levels in the central nervous system (CNS) during ovarian maturation. In summary, we provide further evidence for the existence of steroidogenic pathways in crustaceans, which may be useful for advancing prawn aquaculture. PMID:26774430

  5. Sugarcane Giant Borer Transcriptome Analysis and Identification of Genes Related to Digestion

    PubMed Central

    de Assis Fonseca, Fernando Campos; Firmino, Alexandre Augusto Pereira; de Macedo, Leonardo Lima Pepino; Coelho, Roberta Ramos; de Sousa Júnior, José Dijair Antonino; Silva-Junior, Orzenil Bonfim; Togawa, Roberto Coiti; Pappas, Georgios Joannis; de Góis, Luiz Avelar Brandão; da Silva, Maria Cristina Mattar; Grossi-de-Sá, Maria Fátima

    2015-01-01

    Sugarcane is a widely cultivated plant that serves primarily as a source of sugar and ethanol. Its annual yield can be significantly reduced by the action of several insect pests including the sugarcane giant borer (Telchin licus licus), a lepidopteran that presents a long life cycle and which efforts to control it using pesticides have been inefficient. Although its economical relevance, only a few DNA sequences are available for this species in the GenBank. Pyrosequencing technology was used to investigate the transcriptome of several developmental stages of the insect. To maximize transcript diversity, a pool of total RNA was extracted from whole body insects and used to construct a normalized cDNA database. Sequencing produced over 650,000 reads, which were de novo assembled to generate a reference library of 23,824 contigs. After quality score and annotation, 43% of the contigs had at least one BLAST hit against the NCBI non-redundant database, and 40% showed similarities with the lepidopteran Bombyx mori. In a further analysis, we conducted a comparison with Manduca sexta midgut sequences to identify transcripts of genes involved in digestion. Of these transcripts, many presented an expansion or depletion in gene number, compared to B. mori genome. From the sugarcane giant borer (SGB) transcriptome, a number of aminopeptidase N (APN) cDNAs were characterized based on homology to those reported as Cry toxin receptors. This is the first report that provides a large-scale EST database for the species. Transcriptome analysis will certainly be useful to identify novel developmental genes, to better understand the insect’s biology and to guide the development of new strategies for insect-pest control. PMID:25706301

  6. Transcriptome Analysis of Medicinal Plant Salvia miltiorrhiza and Identification of Genes Related to Tanshinone Biosynthesis

    PubMed Central

    Yang, Lei; Ding, Guohui; Lin, Haiyan; Cheng, Haining; Kong, Yu; Wei, Yukun; Fang, Xin; Liu, Renyi; Wang, Lingiian; Chen, Xiaoya; Yang, Changqing

    2013-01-01

    Salvia miltiorrhiza Bunge, a perennial plant of Lamiaceae, accumulates abietane-type diterpenoids of tanshinones in root, which have been used as traditional Chinese medicine to treat neuroasthenic insomnia and cardiovascular diseases. However, to date the biosynthetic pathway of tanshinones is only partially elucidated and the mechanism for their root-specific accumulation remains unknown. To identify enzymes and transcriptional regulators involved in the biosynthesis of tanshinones, we conducted transcriptome profiling of S. miltiorrhiza root and leaf tissues using the 454 GS-FLX pyrosequencing platform, which generated 550,546 and 525,292 reads, respectively. RNA sequencing reads were assembled and clustered into 64,139 unigenes (29,883 isotigs and 34,256 singletons). NCBI non-redundant protein databases (NR) and Swiss-Prot database searches anchored 32,096 unigenes (50%) with functional annotations based on sequence similarities. Further assignments with Gene Ontology (GO) terms and KEGG biochemical pathways identified 168 unigenes referring to the terpenoid backbone biosynthesis (including 144 MEP and MVA pathway genes and 24 terpene synthases). Comparative analysis of the transcriptomes identified 2,863 unigenes that were highly expressed in roots, including those encoding enzymes of early steps of tanshinone biosynthetic pathway, such as copalyl diphosphate synthase (SmCPS), kaurene synthase-like (SmKSL) and CYP76AH1. Other differentially expressed unigenes predicted to be related to tanshinone biosynthesis fall into cytochrome P450 monooxygenases, dehydrogenases and reductases, as well as regulatory factors. In addition, 21 P450 genes were selectively confirmed by real-time PCR. Thus we have generated a large unigene dataset which provides a valuable resource for further investigation of the radix development and biosynthesis of tanshinones. PMID:24260395

  7. Gene-related strain variation of Staphylococcus aureus for homologous resistance response to acid stress.

    PubMed

    Lee, Soomin; Ahn, Sooyeon; Lee, Heeyoung; Kim, Won-Il; Kim, Hwang-Yong; Ryu, Jae-Gee; Kim, Se-Ri; Choi, Kyoung-Hee; Yoon, Yohan

    2014-10-01

    This study investigated the effect of adaptation of Staphylococcus aureus strains to the acidic condition of tomato in response to environmental stresses, such as heat and acid. S. aureus ATCC 13565, ATCC 14458, ATCC 23235, ATCC 27664, and NCCP10826 habituated in tomato extract at 35°C for 24 h were inoculated in tryptic soy broth. The culture suspensions were then subjected to heat challenge or acid challenge at 60°C and pH 3.0, respectively, for 60 min. In addition, transcriptional analysis using quantitative real-time PCR was performed to evaluate the expression level of acid-shock genes, such as clpB, zwf, nuoF, and gnd, from five S. aureus strains after the acid habituation of strains in tomato at 35°C for 15 min and 60 min in comparison with that of the nonhabituated strains. In comparison with the nonhabituated strains, the five tomato-habituated S. aureus strains did not show cross protection to heat, but tomato-habituated S. aureus ATCC 23235 showed acid resistance. In quantitative real-time-PCR analysis, the relative expression levels of acid-shock genes (clpB, zwf, nuoF, and gnd) were increased the most in S. aureus ATCC 23235 after 60 min of tomato habituation, but there was little difference in the expression levels among the five S. aureus strains after 15 min of tomato habituation. These results indicate that the variation of acid resistance of S. aureus is related to the expression of acid-shock genes during acid habituation. PMID:25285500

  8. Expression of genes related to tolerance to low temperature for maize seed germination.

    PubMed

    Silva-Neta, I C; Pinho, E V; Veiga, A D; Pìnho, R G; Guimarães, R M; Caixeta, F; Santos, H O; Marques, T L

    2015-01-01

    The aim of this study was to characterize maize lines tolerant to cold temperatures during the germination process. Seeds from lines with different levels of tolerance to low temperatures were used; 3 lines were classified as tolerant and 3 as susceptible to low germination temperatures. A field was set up to multiply seeds from selected lines. After the seeds were harvested and classified, we conducted physiological tests and analyzed fatty acid content of palmitic, stearic, oleic, linoleic, linolenic, and eicosenoic acids. In proteomic analysis, the expression of heat-resistant proteins, including catalase, peroxidase, esterase, superoxide dismutase, and α-amylase, were evaluated. Transcript analysis was used to measure the expression of the genes AOX1, AOX2, ZmMPK-17, and ZmAN-13. The material showing the highest susceptibility to low germination temperatures contained high saturated fatty acid content. Expression of α-amylase in seeds soaked for 72 h at a temperature of 10°C was lower than expression of α-amylase when soaked at 25°C for the same amount of time. We observed variation in the expression of heat-resistant proteins in seeds of the lines evaluated. The genes AOX and Zm-AN13 were promising for use in identifying maize materials that are tolerant to low germination temperatures. PMID:25867416

  9. The molecular signature of AML mesenchymal stromal cells reveals candidate genes related to the leukemogenic process.

    PubMed

    Binato, Renata; de Almeida Oliveira, Nathalia Correa; Du Rocher, Barbara; Abdelhay, Eliana

    2015-12-01

    Acute myeloid leukemia (AML) is a heterogeneous disease characterized by myeloid precursor proliferation in the bone marrow, apoptosis reduction and differentiation arrest. Although there are several studies in this field, events related to disease initiation and progression remain unknown. The malignant transformation of hematopoietic stem cells (HSC) is thought to generate leukemic stem cells, and this transformation could be related to changes in mesenchymal stromal cell (hMSC) signaling. Thus, the aim of this work was to analyze the gene expression profile of hMSC from AML patients (hMSC-AML) compared to healthy donors hMSCs (hMSC-HD). The results showed a common molecular signature for all hMSC-AML. Other assays were performed with a large number of patients and the results confirmed a molecular signature that is capable of distinguishing hMSC-AML from hMSC-HD. Moreover, CCL2 and BMP4 genes encode secreted proteins that could affect HSCs. To verify whether these proteins are differentially expressed in AML patients, ELISA was performed with plasma samples. CCL2 and BMP4 proteins are differentially expressed in AML patients, indicating changes in hMSC-AML signaling. Altogether, hMSCs-AML signaling alterations could be an important factor in the leukemic transformation process. PMID:26279521

  10. Screening of genes related to ovarian development in the swimming crab, Portunus trituberculatus, by suppression subtractive hybridization.

    PubMed

    Yu, Z B; Mu, C K; Song, W W; Li, R H; Chen, Y E; Wang, C L

    2015-01-01

    The swimming crab, Portunus trituberculatus, is an important marine animal and is widely cultured in China. In the present study, suppression subtractive hybridization was applied to identify the differentially expressed genes in the ovaries of mature and immature P. trituberculatus. One hundred and seventy six expressed sequence tag (ESTs) were identified, of which 100 were down-regulated, and 76 up-regulated. BLAST analysis identified 51 unigenes, of which 27 were down-regulated, and 24 up-regulated. Quantitative real-time reverse transcriptase polymerase chain reaction results indicated that the SSH technique is valuable in screening genes related to ovarian development. Genes identified in this study encoded proteins corresponding to a wide range of functions and included immune response protein, transcription initiation factor, metabolic proteins, chromosome, histone h3, ovarian development-related protein, and vitellogenin. In addition, 64 metabolic pathways were annotated in differentially expressed ESTs by using the Kyoto Encyclopedia of Genes and Genomes pathway. Four annotated pathways (oxidative phosphorylation, carbon metabolism, fatty acid degradation, and protein digestion and absorption) appeared to be involved in ovarian development. In ontology analysis, 5.83% of the cellular process genes in reverse subtraction cDNA library are involved in reproduction, and 5.88% involved in developmental process. In up-regulated genes, myosin II-expressed polehole-like protein; histone h3; ovigerous-hair stripping substance; peritrophin 48; and ovarian development-related protein appeared to be involved in ovarian development. Identification of differentially expressed genes in the mature and immature ovary of the swimming crab provides new insights for further studies on the mechanism underlying ovarian development in this species. PMID:26782517

  11. Identification of differentially expressed genes related to metabolic syndrome induced with high-fat diet in E3 rats.

    PubMed

    Lan, Xi; Li, Dongmin; Zhong, Bo; Ren, Juan; Wang, Xuan; Sun, Qingzhu; Li, Yue; Liu, Lee; Liu, Li; Lu, Shemin

    2015-02-01

    Understanding the genes differentially expressing in aberrant organs of metabolic syndrome (MetS) facilitates the uncovering of molecular mechanisms and the identification of novel therapeutic targets for the disease. This study aimed to identify differentially expressed genes related to MetS in livers of E3 rats with high-fat-diet-induced metabolic syndrome (HFD-MetS). E3 rats were fed with high-fat diet for 24 weeks to induce MetS. Then, suppression subtractive hybridization (SSH) technology was used to identify the genes differentially expressed between HFD-MetS and control E3 rat livers. Twenty positive recombinant clones were chosen randomly from forward subtractive library and sent to sequence. BLAST analysis in GenBank database was used to determine the property of each cDNA fragment. In total, 11 annotated genes, 3 ESTs, and 2 novel gene fragments were identified by SSH technology. The expression of four genes (Alb, Pip4k2a, Scd1, and Tf) known to be associated with MetS and other five genes (Eif1, Rnase4, Rps12, Rup2, and Tmsb4) unknown to be relevant to MetS was significantly up-regulated in the livers of HFD-MetS E3 rats compared with control rats using real-time quantitative PCR (RT-qPCR). By analyzing the correlations between the expression of these nine genes and serum concentrations of TG, Tch, HDL-C, and LDL-C, we found that there were significant positive correlations between TG and the expression of five genes (Alb, Eif1, Pip4k2a, Rps12, and Tmsb4x), Tch and three genes (Rnase4, Scd1, and Tmsb4x), and LDL-C and two genes (Rnase4 and Scd1), as well there were significant negative correlations between HDL-C and the expression of three genes (Rup2, Scd1, and Tf). This study provides important clues for unraveling the molecular mechanisms of MetS. PMID:25294893

  12. Transcriptome Sequencing of Chemically Induced Aquilaria sinensis to Identify Genes Related to Agarwood Formation

    PubMed Central

    Ye, Wei; Wu, Hongqing; He, Xin; Wang, Lei; Zhang, Weimin; Li, Haohua; Fan, Yunfei; Tan, Guohui; Liu, Taomei; Gao, Xiaoxia

    2016-01-01

    Background Agarwood is a traditional Chinese medicine used as a clinical sedative, carminative, and antiemetic drug. Agarwood is formed in Aquilaria sinensis when A. sinensis trees are threatened by external physical, chemical injury or endophytic fungal irritation. However, the mechanism of agarwood formation via chemical induction remains unclear. In this study, we characterized the transcriptome of different parts of a chemically induced A. sinensis trunk sample with agarwood. The Illumina sequencing platform was used to identify the genes involved in agarwood formation. Methodology/Principal Findings A five-year-old Aquilaria sinensis treated by formic acid was selected. The white wood part (B1 sample), the transition part between agarwood and white wood (W2 sample), the agarwood part (J3 sample), and the rotten wood part (F5 sample) were collected for transcriptome sequencing. Accordingly, 54,685,634 clean reads, which were assembled into 83,467 unigenes, were obtained with a Q20 value of 97.5%. A total of 50,565 unigenes were annotated using the Nr, Nt, SWISS-PROT, KEGG, COG, and GO databases. In particular, 171,331,352 unigenes were annotated by various pathways, including the sesquiterpenoid (ko00909) and plant–pathogen interaction (ko03040) pathways. These pathways were related to sesquiterpenoid biosynthesis and defensive responses to chemical stimulation. Conclusions/Significance The transcriptome data of the different parts of the chemically induced A. sinensis trunk provide a rich source of materials for discovering and identifying the genes involved in sesquiterpenoid production and in defensive responses to chemical stimulation. This study is the first to use de novo sequencing and transcriptome assembly for different parts of chemically induced A. sinensis. Results demonstrate that the sesquiterpenoid biosynthesis pathway and WRKY transcription factor play important roles in agarwood formation via chemical induction. The comparative analysis of

  13. EXPRESSION PATTERNS OF THE GLYCOGEN PHOSPHORYLASE GENE RELATED TO LARVAL DIAPAUSE IN Ostrinia furnacalis.

    PubMed

    Guo, Jianqing; Zhang, Honggang; Edwards, Martin; Wang, Zhenying; Bai, Shuxiong; He, Kanglai

    2016-04-01

    Glycogen phosphorylase (GP) acts in the first step in release of glucose from glycogen, a form of energy storage for most organisms. To investigate the characteristics and expression pattern of GP gene (Ofgp) in the Asian corn borer, Ostrinia furnacalis (Guenée), larvae, we cloned and analyzed tissue transcription of Ofgp. The results indicate that the open reading frame (ORF) is 2,526 bp, encoding 841 amino acid. The calculated three-dimensional structure shows 33 α-helices and 24 β-sheets. Ofgp transcription levels varied significantly during the second to fifth instars under long-day (28 °C, 16:8 L:D photoperiod, and 70-80% relative humidity (RH)) and short-day (24.5 °C, 11:13 L:D photoperiod, and 70-80% RH) conditions, remained low during the prediapause phase, and then increased after about 36 d under short-day photoperiod. In the larvae reared under long-day condition, hemolymph ranked the highest in the transcript level of Ofgp. The highest transcription was recorded in the fat body and was lower in the other tissues in larvae reared under short-day condition. We found that Ofgp transcription increased linearly from October 2012 to January 2013. The transcript level was negatively correlated with environmental temperature. We infer the higher Ofgp transcription may enhance the cold hardiness of the diapause larvae. PMID:26748939

  14. Cognitive development in females with PCDH19 gene-related epilepsy.

    PubMed

    Cappelletti, Simona; Specchio, Nicola; Moavero, Romina; Terracciano, Alessandra; Trivisano, Marina; Pontrelli, Giuseppe; Gentile, Simonetta; Vigevano, Federico; Cusmai, Raffaella

    2015-01-01

    Mutations in the PCDH19 gene are now recognized to cause epilepsy in females and are claiming increasing interest in the scientific world. Clinical features and seizure semiology have been described as heterogeneous. Intellectual disability might be present, ranging from mild to severe; behavioral and psychiatric problems are a common feature of the disorder, including aggressiveness, depressed mood, and psychotic traits. The purpose of our study was to describe the cognitive development in 11 girls with a de novo mutation in PCDH19 and early-onset epilepsy. Six patients had average mental development or mild intellectual disability regardless of persistence of seizures in clusters. Five patients presented moderate or severe intellectual disability and autistic features. In younger patients, we found that despite an average developmental quotient, they all presented a delay of expressive language acquisition and lower scores at follow-up testing completed at older ages, underlining that subtle dysfunctions might be present. Larger cohort and long-term follow-up might be useful in defining cognitive features and in improving the care of patients with PCDH19. PMID:25499160

  15. Use of Galerina marginata genes and proteins for peptide production

    DOEpatents

    Hallen-Adams, Heather E.; Scott-Craig, John S.; Walton, Jonathan D.; Luo, Hong

    2016-03-01

    The present invention relates to compositions and methods comprising genes and peptides associated with cyclic peptides and cyclic peptide production in mushrooms. In particular, the present invention relates to using genes and proteins from Galerina species encoding peptides specifically relating to amatoxins in addition to proteins involved with processing cyclic peptide toxins. In a preferred embodiment, the present invention also relates to methods for making small peptides and small cyclic peptides including peptides similar to amanitin. Further, the present inventions relate to providing kits for making small peptides.

  16. Aerosolized Medications for Gene and Peptide Therapy.

    PubMed

    Laube, Beth L

    2015-06-01

    Inhalation therapy has matured to include drugs that: (1) deliver nucleic acids that either lead to the restoration of a gene construct or protein coding sequence in a population of cells or suppress or disrupt production of an abnormal gene product (gene therapy); (2) deliver peptides that target lung diseases such as asthma, sarcoidosis, pulmonary hypertension, and cystic fibrosis; and (3) deliver peptides to treat diseases outside the lung whose target is the systemic circulation (systemic drug delivery). These newer applications for aerosol therapy are the focus of this paper, and I discuss the status of each and the challenges that remain to their successful development. Drugs that are highlighted include: small interfering ribonucleic acid to treat lung cancer and Mycobacterium tuberculosis; vectors carrying the normal alpha-1 antitrypsin gene to treat alpha-1 antitrypsin deficiency; vectors carrying the normal cystic fibrosis transmembrane conductance regulator gene to treat cystic fibrosis; vasoactive intestinal peptide to treat asthma, pulmonary hypertension, and sarcoidosis; glutathione to treat cystic fibrosis; granulocyte-macrophage colony-stimulating factor to treat pulmonary alveolar proteinosis; calcitonin for postmenopausal osteoporosis; and insulin to treat diabetes. The success of these new aerosol applications will depend on many factors, such as: (1) developing gene therapy formulations that are safe for acute and chronic administrations to the lung, (2) improving the delivery of the genetic material beyond the airway mucus barrier and cell membrane and transferring the material to the cell cytoplasm or the cell nucleus, (3) developing aerosol devices that efficiently deliver genetic material and peptides to their lung targets over a short period of time, (4) developing devices that increase aerosol delivery to the lungs of infants, (5) optimizing the bioavailability of systemically delivered peptides, and (6) developing peptide formulations for

  17. Identification of Aadnr1, a novel gene related to innate immunity and apoptosis in Aedes albopictus.

    PubMed

    Li, Xiaomei; Meng, Kun; Qiao, Jialu; Liu, Hao; Zhong, Chunyan; Liu, Qingzhen

    2016-08-01

    Innate immunity and apoptosis play critical roles in defending pathogens in insects. In Drosophila, Dnr1 was reported as a negative regulator of apoptosis and immune deficiency (Imd) pathway which belongs to innate immunity. Aedes albopictus is an important kind of arbovirus vector and becoming a significant threat to public health due to its rapid global expansion. Here we identified an ortholog of dnr1 from A. albopictus, named as Aadnr1. Aadnr1 encoded a putative protein containing an N-terminal FERM domain and a C-terminal RING domain. AaDnr1 shared high identity with dipteran insects Dnr1 orthologs. Phylogenetic analyses showed that the closest relative of AaDnr1 was Aedes aegypti Dnr1. Real-time PCR proved that Aadnr1 mRNA was expressed ubiquitously during developmental and adult stages. Transcriptional levels of Aadnr1 were decreased drastically in C6/36 cells underwent apoptosis induced by Actinomycin D (Act D) treatment. Partial silence of Aadnr1 enhanced Act D-induced caspase activity. When challenged by heat-inactivated E. coli, transcriptional level of Aadnr1 was also decreased dramatically in C6/36 cells. While when C6/36 cells were infected with Sindbis virus TE/GFP, transcriptional level of Aadnr1 was reduced and recovered repeatedly, with an overall decreasing trend. It was also shown in this study that similar to Drosophila Dnr1, RING domain destabilized AaDnr1 protein. Taken together, the study identified an innate immunity and apoptosis related gene Aadnr1 in A. albopictus. PMID:27045774

  18. Peptide Synthetase Gene in Trichoderma virens

    PubMed Central

    Wilhite, S. E.; Lumsden, R. D.; Straney, D. C.

    2001-01-01

    Trichoderma virens (synonym, Gliocladium virens), a deuteromycete fungus, suppresses soilborne plant diseases caused by a number of fungi and is used as a biocontrol agent. Several traits that may contribute to the antagonistic interactions of T. virens with disease-causing fungi involve the production of peptide metabolites (e.g., the antibiotic gliotoxin and siderophores used for iron acquisition). We cloned a 5,056-bp partial cDNA encoding a putative peptide synthetase (Psy1) from T. virens using conserved motifs found within the adenylate domain of peptide synthetases. Sequence similarities with conserved motifs of the adenylation domain, acyl transfer, and two condensation domains support identification of the Psy1 gene as a gene that encodes a peptide synthetase. Disruption of the native Psy1 gene through gene replacement was used to identify the function of this gene. Psy1 disruptants produced normal amounts of gliotoxin but grew poorly under low-iron conditions, suggesting that Psy1 plays a role in siderophore production. Psy1 disruptants cannot produce the major T. virens siderophore dimerum acid, a dipetide of acylated Nδ-hydroxyornithine. Biocontrol activity against damping-off diseases caused by Pythium ultimum and Rhizoctonia solani was not reduced by the Psy1 disruption, suggesting that iron competition through dimerum acid production does not contribute significantly to disease suppression activity under the conditions used. PMID:11679326

  19. The potential for prazosin and calcitonin gene-related peptide (CGRP) in causing hypoxia in tumours.

    PubMed Central

    Burney, I. A.; Maxwell, R. J.; Griffiths, J. R.; Field, S. B.

    1991-01-01

    Using 31P NMR spectroscopy, changes in tumour metabolic status were studied in a transplanted rat fibrosarcoma following the administration of vasodilators. Mean Arterial Blood Pressure (MABP) was monitored simultaneously. Two vasodilators were studied, prazosin and CGRP, which altered the NMR parameters Pi/sigma P, beta NTP,Pi, PCr/Pi and PME/Pi in a dose dependent manner. There was a good correlation between the various NMR parameters; for analysis, Pi/sigma P was used for convenience. With increasing doses of vasodilator, Pi/sigma P increased and the MABP decreased. Reduction in pHNMR showed a correlation with decreasing MABP following the administration of prazosin but not after CGRP. Both prazosin and CGRP produced changes in 31P NMR spectra consistent with a reduction in tumour blood flow. The results for prazosin and CGRP were comparable and showed a 15-20% increase in Pi/sigma P for a 20% reduction in MABP. These results were compared with those from hydralazine. With hydralazine an acceptable reduction in blood pressure (up to approximately 25%) has little effect and may even alter NMR parameters consistent with an increase in blood flow, a reduction of approximately 40% is required for a significant decrease in flow. Both prazosin and CGRP are shown to be far more effective than hydralazine in causing tumour hypoxia at a clinically acceptable reduction in blood pressure. CGRP may be the more suitable for clinical use because of its short half life, its capability to achieve controlled hypotension and the relatively few side effects associated with its use. PMID:1911217

  20. Flupirtine inhibits calcitonin-gene related peptide release from rat brainstem in vitro.

    PubMed

    Tringali, Giuseppe; Greco, Maria Cristina; Capuano, Alessandro; Guerriero, Giuseppe; Currò, Diego; Navarra, Pierluigi

    2012-01-11

    We have previously shown that the nonopioid analgesic flupirtine possesses analgesic activity in the orofacial formalin test in vivo in the rat. However, this paradigm does not allow to distinguish between central and peripheral site of action of the drug. In this study we used a recently characterized in vitro model, consisting in acute rat brainstem explants, to investigate whether flupirtine analgesia may be, at least in part, attributed to interference with neurotransmission between the first and the second order neurons of the trigeminal system, occurring within the brainstem. We used acute rat brainstem explants; CGRP released into the incubation medium was taken as a marker of CGRP release from central terminals of trigeminal ganglion afferent neurons within the brainstem. CGRP levels were measured by radioimmunoassay under basal conditions or in the presence of flupirtine, alone or with putative antagonist XE-991. We found that flupirtine inhibits in a concentration-dependent manner both basal and capsaicin-stimulated CGRP release from rat brainstem. This effect is mimicked by the flupirtine analogue retigabine, and is counteracted by the Kv7 blocker XE-991. These findings provide in vitro evidence that the analgesic activity of flupirtine may be related to interference with pain neurotransmission at the brainstem level. Pharmacological data suggests that such effect is related to opening of Kv7 channels on first-order neuronal nerve ending, and the subsequent inhibition of neurotransmitter release, since the effect is mimicked by the Kv7 opener retigabine and is counteracted by the Kv7 blocker XE-991. PMID:22155095

  1. Transcriptomic Analysis of a Tertiary Relict Plant, Extreme Xerophyte Reaumuria soongorica to Identify Genes Related to Drought Adaptation

    PubMed Central

    Zhao, Pengshan; Yin, Hengxia; Zhao, Xin; Xiao, Honglang; Li, Xinrong; Chen, Guoxiong; Ma, Xiao-Fei

    2013-01-01

    Background Reaumuria soongorica is an extreme xerophyte shrub widely distributed in the desert regions including sand dune, Gobi and marginal loess of central Asia which plays a crucial role to sustain and restore fragile desert ecosystems. However, due to the lacking of the genomic sequences, studies on R. soongorica had mainly limited in physiological responses to drought stress. Here, a deep transcriptomic sequencing of R. soongorica will facilitate molecular functional studies and pave the path to understand drought adaptation for a desert plant. Methodology/Principal Findings A total of 53,193,660 clean paired-end reads was generated from the Illumina HiSeq™ 2000 platform. By assembly with Trinity, we got 173,700 contigs and 77,647 unigenes with mean length of 677 bp and N50 of 1109 bp. Over 55% (43,054) unigenes were successfully annotated based on sequence similarity against public databases as well as Rfam and Pfam database. Local BLAST and Kyoto Encyclopedia of Genes and Genomes (KEGG) maps were used to further exhausting seek for candidate genes related to drought adaptation and a set of 123 putative candidate genes were identified. Moreover, all the C4 photosynthesis genes existed and were active in R. soongorica, which has been regarded as a typical C3 plant. Conclusion/Significance The assembled unigenes in present work provide abundant genomic information for the functional assignments in an extreme xerophyte R. soongorica, and will help us exploit the genetic basis of how desert plants adapt to drought environment in the near future. PMID:23717523

  2. Identification of Hub Genes Related to the Recovery Phase of Irradiation Injury by Microarray and Integrated Gene Network Analysis

    PubMed Central

    Zhang, Jing; Yang, Yue; Wang, Yin; Zhang, Jinyuan; Wang, Zejian; Yin, Ming; Shen, Xudong

    2011-01-01

    Background Irradiation commonly causes long-term bone marrow injury charactertized by defective HSC self-renewal and a decrease in HSC reserve. However, the effect of high-dose IR on global gene expression during bone marrow recovery remains unknown. Methodology Microarray analysis was used to identify differentially expressed genes that are likely to be critical for bone marrow recovery. Multiple bioinformatics analyses were conducted to identify key hub genes, pathways and biological processes. Principal Findings 1) We identified 1302 differentially expressed genes in murine bone marrow at 3, 7, 11 and 21 days after irradiation. Eleven of these genes are known to be HSC self-renewal associated genes, including Adipoq, Ccl3, Ccnd1, Ccnd2, Cdkn1a, Cxcl12, Junb, Pten, Tal1, Thy1 and Tnf; 2) These 1302 differentially expressed genes function in multiple biological processes of immunity, including hematopoiesis and response to stimuli, and cellular processes including cell proliferation, differentiation, adhesion and signaling; 3) Dynamic Gene Network analysis identified a subgroup of 25 core genes that participate in immune response, regulation of transcription and nucleosome assembly; 4) A comparison of our data with known irradiation-related genes extracted from literature showed 42 genes that matched the results of our microarray analysis, thus demonstrated consistency between studies; 5) Protein-protein interaction network and pathway analyses indicated several essential protein-protein interactions and signaling pathways, including focal adhesion and several immune-related signaling pathways. Conclusions Comparisons to other gene array datasets indicate that global gene expression profiles of irradiation damaged bone marrow show significant differences between injury and recovery phases. Our data suggest that immune response (including hematopoiesis) can be considered as a critical biological process in bone marrow recovery. Several critical hub genes that are

  3. Glucagon-Like Peptide-1 Gene Therapy

    PubMed Central

    Rowzee, Anne M.; Cawley, Niamh X.; Chiorini, John A.; Di Pasquale, Giovanni

    2011-01-01

    Glucagon-like peptide 1 (GLP-1) is a small peptide component of the prohormone, proglucagon, that is produced in the gut. Exendin-4, a GLP-1 receptor agonist originally isolated from the saliva of H. suspectum or Gila monster, is a peptide that shares sequence and functional homology with GLP-1. Both peptides have been demonstrated to stimulate insulin secretion, inhibit glucagon secretion, promote satiety and slow gastric emptying. As such, GLP-1 and Exendin-4 have become attractive pharmaceutical targets as an adjunctive therapy for individuals with type II diabetes mellitus, with several products currently available clinically. Herein we summarize the cell biology leading to GLP-1 production and secretion from intestinal L-cells and the endocrine functions of this peptide and Exendin-4 in humans. Additionally, gene therapeutic applications of GLP-1 and Exendin-4 are discussed with a focus on recent work using the salivary gland as a gene therapy target organ for the treatment of diabetes mellitus. PMID:21747830

  4. Mapping of DNA sex-specific markers and genes related to sex differentiation in turbot (Scophthalmus maximus).

    PubMed

    Viñas, Ana; Taboada, Xoana; Vale, Luis; Robledo, Diego; Hermida, Miguel; Vera, Manel; Martínez, Paulino

    2012-10-01

    Production of all-female populations in turbot can increase farmer's benefits since sexual dimorphism in growth in this species is among the highest within marine fish, turbot females reaching commercial size 3-6 months earlier than males. Puberty in males occurs earlier than in females, which additionally slows their growth. Thus, elucidating the mechanisms of sex determination and gonad differentiation is a relevant goal for turbot production. A ZZ/ZW sex determination mechanism has been suggested for this species, and four sex-related quantitative trait loci (QTL) were detected, the major one located in linkage group (LG) 5 and the three minor ones in LG6, LG8, and LG21. In the present work, we carried out a linkage analysis for several sex-related markers: (1) three anonymous sex-associated RAPD and (2) several candidate genes related to sex determination and gonad differentiation in other species (Sox3, Sox6, Sox8, Sox9, Sox17, Sox19, Amh, Dmrta2, Cyp19a, Cyp19b). We focused our attention on their co-localization with the major and minor sex-related QTL trying to approach to the master sex-determining gene of this species. Previously described growth-related QTL were also considered since the association observed between growth and sex determination in fish. Amh, Dmrta2, and one RAPD were located in LG5, while Sox9 and Sox17 (LG21), Cyp19b (LG6), and a second RAPD (LG8) co-mapped with suggestive sex-related QTL, thus supporting further analyses on these genes to elucidate the genetic basis of this relevant trait for turbot farming. PMID:22552957

  5. Nicotine mediates expression of genes related to antioxidant capacity and oxidative stress response in HIV-1 transgenic rat brain.

    PubMed

    Song, Guohua; Nesil, Tanseli; Cao, Junran; Yang, Zhongli; Chang, Sulie L; Li, Ming D

    2016-02-01

    Oxidative stress plays an important role in the progression of HIV-1 infection. Nicotine can either protect neurons from neurodegeneration or induce oxidative stress, depending on its dose and degree of oxidative stress impairment. However, the relationship between nicotine and oxidative stress in the HIV-1-infected individuals remains largely unknown. The purpose of this study was to determine the effect of nicotine on expression of genes related to the glutathione (GSH)-centered antioxidant system and oxidative stress in the nucleus accumbens (NAc) and ventral tegmental area (VTA) of HIV-1 transgenic (HIV-1Tg) and F344 control rats. Adult HIV-1Tg and F344 rats received nicotine (0.4 mg/kg, base, s.c.) or saline injections once per day for 27 days. At the end of treatment, various brain regions including the NAc and VTA were collected from each rat. Following total RNA extraction and complementary DNA (cDNA) synthesis of each sample, quantitative reverse transcription PCR (RT-PCR) analysis was performed for 43 oxidative-stress-related genes. Compared with F344 control rats, HIV-1Tg rats showed a significant downregulation of genes involved in ATPase and cyctochrome oxidase at the messenger RNA (mRNA) level in both regions. Further, we found a significant downregulation of Gstm5 in the NAc and upregulation of Cox1, Cox3, and Gsta6 in the VTA of HIV-1Tg rats. HIV-1Tg rats showed brain-region-specific responses to chronic nicotine treatment. This response resulted in a change in the expression of genes involved in antioxidant mechanisms including the downregulation of genes such as Atp5h, Calml1, Gpx7, Gstm5, Gsr, and Gsta6 and upregulation of Sod1 in the NAc, as well as downregulation of genes like Cox5a, Gpx4, Gpx6, Gpx7, Gstm5, and Sod1 in the VTA of HIV-1Tg rats. Together, we conclude that chronic nicotine treatment has a dual effect on the antioxidant defense system and oxidative-stress-induced apoptosis signaling in HIV-1Tg rats. These findings suggest that

  6. Molecular cloning and long terminal repeat sequences of human endogenous retrovirus genes related to types A and B retrovirus genes

    SciTech Connect

    Ono, M.

    1986-06-01

    By using a DNA fragment primarily encoding the reverse transcriptase (pol) region of the Syrian hamster intracisternal A particle (IAP; type A retrovirus) gene as a probe, human endogenous retrovirus genes, tentatively termed HERV-K genes, were cloned from a fetal human liver gene library. Typical HERV-K genes were 9.1 or 9.4 kilobases in length, having long terminal repeats (LTRs) of ca. 970 base pairs. Many structural features commonly observed on the retrovirus LTRs, such as the TATAA box, polyadenylation signal, and terminal inverted repeats, were present on each LTR, and a lysine (K) tRNA having a CUU anticodon was identified as a presumed primer tRNA. The HERV-K LTR, however, had little sequence homology to either the IAP LTR or other typical oncovirus LTRs. By filter hybridization, the number of HERV-K genes was estimated to be ca. 50 copies per haploid human genome. The cloned mouse mammary tumor virus (type B) gene was found to hybridize with both the HERV-K and IAP genes to essentially the same extent.

  7. Expression and network analysis of genes related to melanocyte development in the Silky Fowl and White Leghorn embryos.

    PubMed

    Li, Yulin; Zhu, Xuping; Yang, Liu; Li, Junying; Lian, Zhengxing; Li, Ning; Deng, Xuemei

    2011-02-01

    Silky Fowl is a natural mutant with hyperpigmentation of various internal tissues. Although the mechanism of hyperpigmentation remains unclear, recent studies have shown that the abnormal migration of melanoblast and the absence of environmental barrier molecules are responsible for the hyperpigmentation in Silky Fowl. In this study, 13 genes related to melanocyte development were selected to detect expression changes between Silky Fowl and White Leghorn [including SRY-box 10 (Sox10), paired box (Pax3), stem cell factor (Scf), v-kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog (Kit), endothelin type-B receptor (Ednrb), endothelin 3 (Edn3), microphthalmia-associated transcription factor (Mitf), tyrosinase (Tyr), tyrosinase-related protein-1 (Trp1), tyrosinase-related protein-2 (Trp2), melanocortin-1 receptor (Mc1r), Agouti-related proteins (Agrp), and Proopiomelanocortin (Pomc)]. Transcript expression was detected in 11 stages from 2.5 to 15 days of incubation. In these embryonic periods, Mitf, Kit, Scf, and Agrp expressed earlier in Silky Fowl than in White Leghorn. Sox10, Ednrb, Kit, Mc1r, and Agrp, associating with the proliferation and differentiation of melanoblast, expressed higher (P < 0.05) in Silky Fowl than White Leghorn during 5-6 days of incubation. After day 8 of incubation, Mitf, Tyr, Trp1, Trp2, and Mc1r expressed higher (P < 0.05) in Silky Fowl than White Leghorn, while Agrp expressed higher (P < 0.05) in White Leghorn than Silky Fowl. Moreover, a regulatory network for melanocyte development was constructed based on the expression data. The network predicted novel regulatory relationships and confirmed relationships that have been reported. These results provide biological insight into the molecular mechanism of hyperpigmentation in the Silky Fowl. However, further investigation is needed to confirm these regulatory relationships. PMID:20848220

  8. Functional identification of a Leishmania gene related to the peroxin 2 gene reveals common ancestry of glycosomes and peroxisomes.

    PubMed Central

    Flaspohler, J A; Rickoll, W L; Beverley, S M; Parsons, M

    1997-01-01

    Glycosomes are membrane-bounded microbody organelles that compartmentalize glycolysis as well as other important metabolic processes in trypanosomatids. The compartmentalization of these enzymatic reactions is hypothesized to play a crucial role in parasite physiology. Although the metabolic role of glycosomes differs substantially from that of the peroxisomes that are found in other eukaryotes, similarities in signals targeting proteins to these organelles suggest that glycosomes and peroxisomes may have evolved from a common ancestor. To examine this hypothesis, as well as gain insights into the function of the glycosome, we used a positive genetic selection procedure to isolate the first Leishmania mutant (gim1-1 [glycosome import] mutant) with a defect in the import of glycosomal proteins. The mutant retains glycosomes but mislocalizes a subset glycosomal proteins to the cytoplasm. Unexpectedly, the gim1-1 mutant lacks lipid bodies, suggesting a heretofore unknown role of the glycosome. We used genetic approaches to identify a gene, GIM1, that is able to restore import and lipid bodies. A nonsense mutation was found in one allele of this gene in the mutant line. The predicted Gim1 protein is related the peroxin 2 family of integral membrane proteins, which are required for peroxisome biogenesis. The similarities in sequence and function provide strong support for the common origin model of glycosomes and peroxisomes. The novel phenotype of gim1-1 and distinctive role of Leishmania glycosomes suggest that future studies of this system will provide a new perspective on microbody biogenesis and function. PMID:9032236

  9. Adaptation of Mycoplasmas to Antimicrobial Agents: Acholeplasma laidlawii Extracellular Vesicles Mediate the Export of Ciprofloxacin and a Mutant Gene Related to the Antibiotic Target

    PubMed Central

    Medvedeva, Elena S.; Baranova, Natalia B.; Mouzykantov, Alexey A.; Grigorieva, Tatiana Yu.; Davydova, Marina N.; Trushin, Maxim V.; Chernova, Olga A.; Chernov, Vladislav M.

    2014-01-01

    This study demonstrated that extracellular membrane vesicles are involved with the development of resistance to fluoroquinolones by mycoplasmas (class Mollicutes). This study assessed the differences in susceptibility to ciprofloxacin among strains of Acholeplasma laidlawii PG8. The mechanisms of mycoplasma resistance to antibiotics may be associated with a mutation in a gene related to the target of quinolones, which could modulate the vesiculation level. A. laidlawii extracellular vesicles mediated the export of the nucleotide sequences of the antibiotic target gene as well as the traffic of ciprofloxacin. These results may facilitate the development of effective approaches to control mycoplasma infections, as well as the contamination of cell cultures and vaccine preparations. PMID:24605048

  10. Mining for Candidate Genes Related to Pancreatic Cancer Using Protein-Protein Interactions and a Shortest Path Approach

    PubMed Central

    Yuan, Fei; Zhang, Yu-Hang; Wan, Sibao; Wang, ShaoPeng; Kong, Xiang-Yin

    2015-01-01

    Pancreatic cancer (PC) is a highly malignant tumor derived from pancreas tissue and is one of the leading causes of death from cancer. Its molecular mechanism has been partially revealed by validating its oncogenes and tumor suppressor genes; however, the available data remain insufficient for medical workers to design effective treatments. Large-scale identification of PC-related genes can promote studies on PC. In this study, we propose a computational method for mining new candidate PC-related genes. A large network was constructed using protein-protein interaction information, and a shortest path approach was applied to mine new candidate genes based on validated PC-related genes. In addition, a permutation test was adopted to further select key candidate genes. Finally, for all discovered candidate genes, the likelihood that the genes are novel PC-related genes is discussed based on their currently known functions. PMID:26613085

  11. Genetic Analysis of Genes Related to Tight Junction Function in the Korean Population with Non-Syndromic Hearing Loss

    PubMed Central

    Sagong, Borum; Cho, Hyun-Ju; Bae, Jae Woong; Kim, Jeongho; Lee, Jinwook; Park, Hong-Joon; Choi, Jae Young; Lee, Kyu-Yup; Kim, Un-Kyung

    2014-01-01

    Tight junctions (TJs) are essential components of eukaryotic cells, and serve as paracellular barriers and zippers between adjacent tissues. TJs are critical for normal functioning of the organ of Corti, a part of the inner ear that causes loss of sensorineural hearing when damaged. To investigate the relation between genes involved in TJ function and hereditary loss of sensorineural hearing in the Korean population, we selected the TJP2 and CLDN14 genes as candidates for gene screening of 135 Korean individuals. The TJP2 gene, mutation of which causes autosomal dominant non-syndromic hearing loss (ADNSHL), lies at the DFNA51 locus on chromosome 9. The CLDN14 gene, mutation of which causes autosomal recessive non-syndromic hearing loss (ARNSHL), lies at the DFNB29 locus on chromosome 21. In the present study, we conducted genetic analyses of the TJP2 and CLDN14 genes in 87 unrelated patients with ADNSHL and 48 unrelated patients with either ARNSHL or potentially sporadic hearing loss. We identified two pathogenic variations, c.334G>A (p.A112T) and c.3562A>G (p.T1188A), and ten single nucleotide polymorphisms (SNPs) in the TJP2 gene. We found eight non-pathogenic variations in the CLDN14 gene. These findings indicate that, whereas mutation of the TJP2 gene might cause ADNSHL, CLDN14 is not a major causative gene for ARNSHL in the Korean population studied. Our findings may improve the understanding of the genetic cause of non-syndromic hearing loss in the Korean population. PMID:24752540

  12. Retinoic Acid Induced 1, RAI1: A Dosage Sensitive Gene Related to Neurobehavioral Alterations Including Autistic Behavior

    PubMed Central

    Carmona-Mora, Paulina; Walz, Katherina

    2010-01-01

    Genomic structural changes, such as gene Copy Number Variations (CNVs) are extremely abundant in the human genome. An enormous effort is currently ongoing to recognize and catalogue human CNVs and their associations with abnormal phenotypic outcomes. Recently, several reports related neuropsychiatric diseases (i.e. autism spectrum disorders, schizophrenia, mental retardation, behavioral problems, epilepsy) with specific CNV. Moreover, for some conditions, both the deletion and duplication of the same genomic segment are related to the phenotype. Syndromes associated with CNVs (microdeletion and microduplication) have long been known to display specific neurobehavioral traits. It is important to note that not every gene is susceptible to gene dosage changes and there are only a few dosage sensitive genes. Smith-Magenis (SMS) and Potocki-Lupski (PTLS) syndromes are associated with a reciprocal microdeletion and microduplication within chromosome 17p11.2. in humans. The dosage sensitive gene responsible for most phenotypes in SMS has been identified: the Retinoic Acid Induced 1 (RAI1). Studies on mouse models and humans suggest that RAI1 is likely the dosage sensitive gene responsible for clinical features in PTLS. In addition, the human RAI1 gene has been implicated in several neurobehavioral traits as spinocerebellar ataxia (SCA2), schizophrenia and non syndromic autism. In this review we discuss the evidence of RAI1 as a dosage sensitive gene, its relationship with different neurobehavioral traits, gene structure and mutations, and what is known about its molecular and cellular function, as a first step in the elucidation of the mechanisms that relate dosage sensitive genes with abnormal neurobehavioral outcomes. PMID:21629438

  13. Pathway-Based Analysis of a Melanoma Genome-Wide Association Study: Analysis of Genes Related to Tumour-Immunosuppression

    PubMed Central

    Schoof, Nils; Iles, Mark M.; Bishop, D. Timothy; Newton-Bishop, Julia A.; Barrett, Jennifer H.; consortium, GenoMEL

    2011-01-01

    Systemic immunosuppression is a risk factor for melanoma, and sunburn-induced immunosuppression is thought to be causal. Genes in immunosuppression pathways are therefore candidate melanoma-susceptibility genes. If variants within these genes individually have a small effect on disease risk, the association may be undetected in genome-wide association (GWA) studies due to low power to reach a high significance level. Pathway-based approaches have been suggested as a method of incorporating a priori knowledge into the analysis of GWA studies. In this study, the association of 1113 single nucleotide polymorphisms (SNPs) in 43 genes (39 genomic regions) related to immunosuppression have been analysed using a gene-set approach in 1539 melanoma cases and 3917 controls from the GenoMEL consortium GWA study. The association between melanoma susceptibility and the whole set of tumour-immunosuppression genes, and also predefined functional subgroups of genes, was considered. The analysis was based on a measure formed by summing the evidence from the most significant SNP in each gene, and significance was evaluated empirically by case-control label permutation. An association was found between melanoma and the complete set of genes (pemp = 0.002), as well as the subgroups related to the generation of tolerogenic dendritic cells (pemp = 0.006) and secretion of suppressive factors (pemp = 0.0004), thus providing preliminary evidence of involvement of tumour-immunosuppression gene polymorphisms in melanoma susceptibility. The analysis was repeated on a second phase of the GenoMEL study, which showed no evidence of an association. As one of the first attempts to replicate a pathway-level association, our results suggest that low power and heterogeneity may present challenges. PMID:22216283

  14. Characterization of Antimicrobial Susceptibility and Its Association with Virulence Genes Related to Adherence, Invasion, and Cytotoxicity in Campylobacter jejuni and Campylobacter coli Isolates from Animals, Meat, and Humans.

    PubMed

    Lapierre, Lisette; Gatica, María A; Riquelme, Víctor; Vergara, Constanza; Yañez, José Manuel; San Martín, Betty; Sáenz, Leonardo; Vidal, Maricel; Martínez, María Cristina; Araya, Pamela; Flores, Roberto; Duery, Oscar; Vidal, Roberto

    2016-07-01

    The aim of this research was to statistically analyze the association between antimicrobial susceptibility/resistance to erythromycine, gentamicin, ciprofloxacin, and tetracycline and 11 virulence genes associated with adherence, invasion, and cytotoxicity in 528 isolates of Campylobacter coli and Campylobacter jejuni obtained from retail meat and fecal samples from food-producing animals and human patients. A high percentage of Campylobacter strains were resistant to antimicrobials, specifically ciprofloxacin and tetracycline. Moreover, we observed a wide distribution of virulence genes within the analyzed strains. C. jejuni strains were more susceptible to antimicrobials, and showed greater number of virulence genes than C. coli strains. Genes related to invasion capability, such as racR, ciaB, and pldA, were associated with antimicrobial-susceptible strains in both species. The genes cdtA and dnaJ, a citotoxin unit and an adherence-related gene, respectively, were associated with antimicrobial-resistant strains in both species. In conclusion, Campylobacter strains show a statistically significant association between antimicrobial susceptibility and the presence of virulence genes. PMID:26779841

  15. Deletion of the Small RNA Chaperone Protein Hfq down Regulates Genes Related to Virulence and Confers Protection against Wild-Type Brucella Challenge in Mice

    PubMed Central

    Lei, Shuangshuang; Zhong, Zhijun; Ke, Yuehua; Yang, Mingjuan; Xu, Xiaoyang; Ren, Hang; An, Chang; Yuan, Jiuyun; Yu, Jiuxuan; Xu, Jie; Qiu, Yefeng; Shi, Yanchun; Wang, Yufei; Peng, Guangneng; Chen, Zeliang

    2016-01-01

    Brucellosis is one of the most common zoonotic epidemics worldwide. Brucella, the etiological pathogen of brucellosis, has unique virulence characteristics, including the ability to survive within the host cell. Hfq is a bacterial chaperone protein that is involved in the survival of the pathogen under stress conditions. Moreover, hfq affects the expression of a large number of target genes. In the present study, we characterized the expression and regulatory patterns of the target genes of Hfq during brucellosis. The results revealed that hfq expression is highly induced in macrophages at the early infection stage and at the late stage of mouse infection. Several genes related to virulence, including omp25, omp31, vjbR, htrA, gntR, and dnaK, were found to be regulated by hfq during infection in BALB/c mice. Gene expression and cytokine secretion analysis revealed that an hfq-deletion mutant induced different cytokine profiles compared with that induced by 16M. Infection with the hfq-deletion mutant induced protective immune responses against 16M challenge. Together, these results suggest that hfq is induced during infection and its deletion results in significant attenuation which affects the host immune response caused by Brucella infection. By regulating genes related to virulence, hfq promotes the virulence of Brucella. The unique characteristics of the hfq-deletion mutant, including its decreased virulence and the ability to induce protective immune response upon infection, suggest that it represents an attractive candidate for the design of a live attenuated vaccine against Brucella. PMID:26834720

  16. Genetic polymorphisms located in genes related to immune and inflammatory processes are associated with end-stage renal disease: a preliminary study

    PubMed Central

    2012-01-01

    Background Chronic kidney disease progression has been linked to pro-inflammatory cytokines and markers of inflammation. These markers are also elevated in end-stage renal disease (ESRD), which constitutes a serious public health problem. Objective To investigate whether single nucleotide polymorphisms (SNPs) located in genes related to immune and inflammatory processes, could be associated with ESRD development. Design and methods A retrospective case-control study was carried out on 276 patients with ESRD and 288 control subjects. Forty-eight SNPs were genotyped via SNPlex platform. Logistic regression was used to assess the relationship between each sigle polymorphism and the development of ESRD. Results Four polymorphisms showed association with ESRD: rs1801275 in the interleukin 4 receptor (IL4R) gene (OR: 0.66 (95%CI = 0.46-0.95); p = 0.025; overdominant model), rs4586 in chemokine (C-C motif) ligand 2 (CCL2) gene (OR: 0.70 (95%CI = 0.54-0.90); p = 0.005; additive model), rs301640 located in an intergenic binding site for signal transducer and activator of transcription 4 (STAT4) (OR: 1.82 (95%CI = 1.17-2.83); p = 0.006; additive model) and rs7830 in the nitric oxide synthase 3 (NOS3) gene (OR: 1.31 (95%CI = 1.01-1.71); p = 0.043; additive model). After adjusting for multiple testing, results lost significance. Conclusion Our preliminary data suggest that four genetic polymorphisms located in genes related to inflammation and immune processes could help to predict the risk of developing ESRD. PMID:22817530

  17. Expressions of ECE-CYC2 clade genes relating to abortion of both dorsal and ventral stamens in Opithandra (Gesneriaceae)

    PubMed Central

    Song, Chun-Feng; Lin, Qi-Bing; Liang, Rong-Hua; Wang, Yin-Zheng

    2009-01-01

    Background ECE-CYC2 clade genes known in patterning floral dorsoventral asymmetry (zygomorphy) in Antirrhinum majus are conserved in the dorsal identity function including arresting the dorsal stamen. However, it remains uncertain whether the same mechanism underlies abortion of the ventral stamens, an important morphological trait related to evolution and diversification of zygomorphy in Lamiales sensu lato, a major clade of predominantly zygomorphically flowered angiosperms. Opithandra (Gesneriaceae) is of particular interests in addressing this question as it is in the base of Lamiales s.l., an early representative of this type zygomorphy. Results We investigated the expression patterns of four ECE-CYC2 clade genes and two putative target cyclinD3 genes in Opithandra using RNA in situ hybridization and RT-PCR. OpdCYC gene expressions were correlated with abortion of both dorsal and ventral stamens in Opithandra, strengthened by the negatively correlated expression of their putative target OpdcyclinD3 genes. The complement of OpdcyclinD3 to OpdCYC expressions further indicated that OpdCYC expressions were related to the dorsal and ventral stamen abortion through negative effects on OpdcyclinD3 genes. Conclusion These results suggest that ECE-CYC2 clade TCP genes are not only functionally conserved in the dorsal stamen repression, but also involved in arresting ventral stamens, a genetic mechanism underlying the establishment of zygomorphy with abortion of both the dorsal and ventral stamens evolved in angiosperms, especially within Lamiales s.l. PMID:19811633

  18. Transcriptome de novo assembly and analysis of differentially expressed genes related to cytoplasmic male sterility in cabbage.

    PubMed

    Wang, Shuai; Wang, Chao; Zhang, Xiao-Xuan; Chen, Xin; Liu, Jiao-Jiao; Jia, Xue-Fang; Jia, Si-Qi

    2016-08-01

    Cytoplasmic male sterility (CMS) is a maternally inherited trait producing abnormal pollen during anther development. To identify the critical genes and pathways that are involved in the sterility and to better understand the underlying mechanisms, cabbage anthers at different developmental stages were cytologically examined and the transcriptomes were analyzed in CMS line and its maintainer line using the next-generation sequencing (NGS) technology. Microscopy showed that anther development in the CMS line was abnormal in the tetrad stage and failed to produce fertile pollen. We obtained 55,663,594 and 54,801,384 raw transcriptome reads from the sterile and maintainer lines, respectively, and assembled these reads into 68,851 unigenes with an average size of 1028 bp. By using the fragments assigned per kilobase of target per million mapped reads (FPKM) method, 5592 differentially expressed genes were identified, consisting of 3403 up- and 2089 down-regulated genes. Furthermore, there were 1011 and 45 genes specifically expressed in the maintainer or sterile line, respectively. Gene Ontology (GO) functional annotation and enrichment analysis of metabolic pathways were performed to map and analyze the candidate genes that may be involved in male sterility. Expression of eighteen genes was examined using qRT-PCR and their expression patterns were found to be same as the sequencing data. A clear cytological difference exists between the sterile and maintainer lines. The differentially expressed genes are associated with carbohydrate and energy metabolisms, or encode transcription factors, heat shock proteins and other stress proteins. Identification of these candidate genes provides a comprehensive understanding of the mechanism underlying CMS in cabbage. PMID:27116370

  19. Extreme obesity is associated with variation in genes related to the circadian rhythm of food intake and hypothalamic signaling.

    PubMed

    Mariman, Edwin C M; Bouwman, Freek G; Aller, Erik E J G; van Baak, Marleen A; Wang, Ping

    2015-06-01

    The hypothalamus is important for regulation of energy intake. Mutations in genes involved in the function of the hypothalamus can lead to early-onset severe obesity. To look further into this, we have followed a strategy that allowed us to identify rare and common gene variants as candidates for the background of extreme obesity from a relatively small cohort. For that we focused on subjects with a well-selected phenotype and on a defined gene set and used a rich source of genetic data with stringent cut-off values. A list of 166 genes functionally related to the hypothalamus was generated. In those genes complete exome sequence data from 30 extreme obese subjects (60 genomes) were screened for novel rare indel, nonsense, and missense variants with a predicted negative impact on protein function. In addition, (moderately) common variants in those genes were analyzed for allelic association using the general population as reference (false discovery rate<0.05). Six novel rare deleterious missense variants were found in the genes for BAIAP3, NBEA, PRRC2A, RYR1, SIM1, and TRH, and a novel indel variant in LEPR. Common variants in the six genes for MBOAT4, NPC1, NPW, NUCB2, PER1, and PRRC2A showed significant allelic association with extreme obesity. Our findings underscore the complexity of the genetic background of extreme obesity involving rare and common variants of genes from defined metabolic and physiologic processes, in particular regulation of the circadian rhythm of food intake and hypothalamic signaling. PMID:25805767

  20. Explorative study to identify novel candidate genes related to oxaliplatin efficacy and toxicity using a DNA repair array

    PubMed Central

    Kweekel, D M; Antonini, N F; Nortier, J W R; Punt, C J A; Gelderblom, H; Guchelaar, H-J

    2009-01-01

    Purpose: To identify new polymorphisms (single nucleotide polymorphisms, SNPs) in DNA repair pathways that are associated with efficacy and toxicity in patients receiving oxaliplatin and capecitabine for advanced colorectal cancer (ACC). Methods: We studied progression-free survival (PFS) in 91 ACC patients, of whom germ-line DNA was isolated and genotyped using an Asper Biotech array. Overall survival (OS) and toxicity were studied as secondary end points. A step-wise selection of SNPs was performed, involving univariate and multivariate log-rank tests and Cox regression analysis, with age and performance status as covariates. Results: A total of 81 SNPs in 46 genes on the array were selected for further analysis, based on genotyping success rates and minor allele frequencies. After step-wise selection, we found that homozygosity for the ataxia telangiectasia mutated gene (ATM) rs1801516 or excision repair cross-complementing gene (ERCC5) rs1047768 SNPs was associated with shorter PFS; however there were no significant associations (P>0.01) with OS or toxicity. Discussion: This is the first study describing the pathway gene approach for the selection of new candidate genes involved in oxaliplatin efficacy and toxicity. The results suggest that the ATM and ERCC5 genes may be associated with oxaliplatin efficacy in ACC. PMID:19536092

  1. ‘Obesity’ is healthy for cetaceans? Evidence from pervasive positive selection in genes related to triacylglycerol metabolism

    PubMed Central

    Wang, Zhengfei; Chen, Zhuo; Xu, Shixia; Ren, Wenhua; Zhou, Kaiya; Yang, Guang

    2015-01-01

    Cetaceans are a group of secondarily adapted marine mammals with an enigmatic history of transition from terrestrial to fully aquatic habitat and subsequent adaptive radiation in waters around the world. Numerous physiological and morphological cetacean characteristics have been acquired in response to this drastic habitat transition; for example, the thickened blubber is one of the most striking changes that increases their buoyancy, supports locomotion, and provides thermal insulation. However, the genetic basis underlying the blubber thickening in cetaceans remains poorly explored. Here, 88 candidate genes associated with triacylglycerol metabolism were investigated in representative cetaceans and other mammals to test whether the thickened blubber matched adaptive evolution of triacylglycerol metabolism-related genes. Positive selection was detected in 41 of the 88 candidate genes, and functional characterization of these genes indicated that these are involved mainly in triacylglycerol synthesis and lipolysis processes. In addition, some essential regulatory genes underwent significant positive selection in cetacean-specific lineages, whereas no selection signal was detected in the counterpart terrestrial mammals. The extensive occurrence of positive selection in triacylglycerol metabolism-related genes is suggestive of their essential role in secondary adaptation to an aquatic life, and further implying that ‘obesity’ might be an indicator of good health for cetaceans. PMID:26381091

  2. Transcriptome-based discovery of pathways and genes related to resistance against Fusarium head blight in wheat landrace Wangshuibai

    PubMed Central

    2013-01-01

    Background Fusarium head blight (FHB), caused mainly by Fusarium graminearum (Fg) Schwabe (teleomorph: Gibberellazeae Schwble), brings serious damage to wheat production. Chinese wheat landrace Wangshuibai is one of the most important resistance sources in the world. The knowledge of mechanism underlying its resistance to FHB is still limited. Results To get an overview of transcriptome characteristics of Wangshuibai during infection by Fg, a high-throughput RNA sequencing based on next generation sequencing (NGS) technology (Illumina) were performed. Totally, 165,499 unigenes were generated and assigned to known protein databases including NCBI non-redundant protein database (nr) (82,721, 50.0%), Gene Ontology (GO) (38,184, 23.1%), Swiss-Prot (50,702, 30.6%), Clusters of orthologous groups (COG) (51,566, 31.2%) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) (30,657, 18.5%), as determined by Blastx search. With another NGS based platform, a digital gene expression (DGE) system, gene expression in Wangshuibai and its FHB susceptible mutant NAUH117 was profiled and compared at two infection stages by inoculation of Fg at 24 and 48 hour, with the aim of identifying genes involved in FHB resistance. Conclusion Pathogen-related proteins such as PR5, PR14 and ABC transporter and JA signaling pathway were crucial for FHB resistance, especially that mediated by Fhb1. ET pathway and ROS/NO pathway were not activated in Wangshuibai and may be not pivotal in defense to FHB. Consistent with the fact that in NAUH117 there presented a chromosome fragment deletion, which led to its increased FHB susceptibility, in Wangshuibai, twenty out of eighty-nine genes showed changed expression patterns upon the infection of Fg. The up-regulation of eight of them was confirmed by qRT-PCR, revealing they may be candidate genes for Fhb1 and need further functional analysis to confirm their roles in FHB resistance. PMID:23514540

  3. Effects of concurrent strength and endurance training on genes related to myostatin signaling pathway and muscle fiber responses.

    PubMed

    de Souza, Eduardo O; Tricoli, Valmor; Aoki, Marcelo S; Roschel, Hamilton; Brum, Patrícia C; Bacurau, Aline V N; Silva-Batista, Carla; Wilson, Jacob M; Neves, Manoel; Soares, Antonio G; Ugrinowitsch, Carlos

    2014-11-01

    Concurrent training (CT) seems to impair training-induced muscle hypertrophy. This study compared the effects of CT, strength training (ST) and interval training (IT) on the muscle fiber cross-sectional area (CSA) response, and on the expression of selected genes involved in the myostatin (MSTN) signaling mRNA levels. Thirty-seven physically active men were randomly divided into 4 groups: CT (n = 11), ST (n = 11), IT (n = 8), and control group (C) (n = 7) and underwent an 8-week training period. Vastus lateralis biopsy muscle samples were obtained at baseline and 48 hours after the last training session. Muscle fiber CSA, selected genes expression, and maximum dynamic ST (1 repetition maximum) were evaluated before and after training. Type IIa and type I muscle fiber CSA increased from pre- to posttest only in the ST group (17.08 and 17.9%, respectively). The SMAD-7 gene expression significantly increased at the posttest in the ST (53.9%) and CT groups (39.3%). The MSTN and its regulatory genes ActIIb, FLST-3, FOXO-3a, and GASP-1 mRNA levels remained unchanged across time and groups. One repetition maximum increased from pre- to posttest in both the ST and CT groups (ST = 18.5%; CT = 17.6%). Our findings are suggestive that MSTN and their regulatory genes at transcript level cannot differentiate muscle fiber CSA responses between CT and ST regimens in humans. PMID:24832980

  4. DBDiaSNP: An Open-Source Knowledgebase of Genetic Polymorphisms and Resistance Genes Related to Diarrheal Pathogens.

    PubMed

    Mehla, Kusum; Ramana, Jayashree

    2015-06-01

    Diarrhea is a highly common infection among children, responsible for significant morbidity and mortality rate worldwide. After pneumonia, diarrhea remains the second leading cause of neonatal deaths. Numerous viral, bacterial, and parasitic enteric pathogens are associated with diarrhea. With increasing antibiotic resistance among enteric pathogens, there is an urgent need for global surveillance of the mutations and resistance genes primarily responsible for resistance to antibiotic treatment. Single Nucleotide Polymorphisms are important in this regard as they have a vast potential to be utilized as molecular diagnostics for gene-disease or pharmacogenomics association studies linking genotype to phenotype. DBDiaSNP is a comprehensive repository of mutations and resistance genes among various diarrheal pathogens and hosts to advance breakthroughs that will find applications from development of sequence-based diagnostic tools to drug discovery. It contains information about 946 mutations and 326 resistance genes compiled from literature and various web resources. As of March 2015, it houses various pathogen genes and the mutations responsible for antibiotic resistance. The pathogens include, for example, DEC (Diarrheagenic E.coli), Salmonella spp., Campylobacter spp., Shigella spp., Clostridium difficile, Aeromonas spp., Helicobacter pylori, Entamoeba histolytica, Vibrio cholera, and viruses. It also includes mutations from hosts (e.g., humans, pigs, others) that render them either susceptible or resistant to a certain type of diarrhea. DBDiaSNP is therefore intended as an integrated open access database for researchers and clinicians working on diarrheal diseases. Additionally, we note that the DBDiaSNP is one of the first antibiotic resistance databases for the diarrheal pathogens covering mutations and resistance genes that have clinical relevance from a broad range of pathogens and hosts. For future translational research involving integrative biology and

  5. Integration of genome-wide association studies with biological knowledge identifies six novel genes related to kidney function

    PubMed Central

    Chasman, Daniel I.; Fuchsberger, Christian; Pattaro, Cristian; Teumer, Alexander; Böger, Carsten A.; Endlich, Karlhans; Olden, Matthias; Chen, Ming-Huei; Tin, Adrienne; Taliun, Daniel; Li, Man; Gao, Xiaoyi; Gorski, Mathias; Yang, Qiong; Hundertmark, Claudia; Foster, Meredith C.; O'Seaghdha, Conall M.; Glazer, Nicole; Isaacs, Aaron; Liu, Ching-Ti; Smith, Albert V.; O'Connell, Jeffrey R.; Struchalin, Maksim; Tanaka, Toshiko; Li, Guo; Johnson, Andrew D.; Gierman, Hinco J.; Feitosa, Mary F.; Hwang, Shih-Jen; Atkinson, Elizabeth J.; Lohman, Kurt; Cornelis, Marilyn C.; Johansson, Åsa; Tönjes, Anke; Dehghan, Abbas; Lambert, Jean-Charles; Holliday, Elizabeth G.; Sorice, Rossella; Kutalik, Zoltan; Lehtimäki, Terho; Esko, Tõnu; Deshmukh, Harshal; Ulivi, Sheila; Chu, Audrey Y.; Murgia, Federico; Trompet, Stella; Imboden, Medea; Coassin, Stefan; Pistis, Giorgio; Harris, Tamara B.; Launer, Lenore J.; Aspelund, Thor; Eiriksdottir, Gudny; Mitchell, Braxton D.; Boerwinkle, Eric; Schmidt, Helena; Cavalieri, Margherita; Rao, Madhumathi; Hu, Frank; Demirkan, Ayse; Oostra, Ben A.; de Andrade, Mariza; Turner, Stephen T.; Ding, Jingzhong; Andrews, Jeanette S.; Freedman, Barry I.; Giulianini, Franco; Koenig, Wolfgang; Illig, Thomas; Meisinger, Christa; Gieger, Christian; Zgaga, Lina; Zemunik, Tatijana; Boban, Mladen; Minelli, Cosetta; Wheeler, Heather E.; Igl, Wilmar; Zaboli, Ghazal; Wild, Sarah H.; Wright, Alan F.; Campbell, Harry; Ellinghaus, David; Nöthlings, Ute; Jacobs, Gunnar; Biffar, Reiner; Ernst, Florian; Homuth, Georg; Kroemer, Heyo K.; Nauck, Matthias; Stracke, Sylvia; Völker, Uwe; Völzke, Henry; Kovacs, Peter; Stumvoll, Michael; Mägi, Reedik; Hofman, Albert; Uitterlinden, Andre G.; Rivadeneira, Fernando; Aulchenko, Yurii S.; Polasek, Ozren; Hastie, Nick; Vitart, Veronique; Helmer, Catherine; Wang, Jie Jin; Stengel, Bénédicte; Ruggiero, Daniela; Bergmann, Sven; Kähönen, Mika; Viikari, Jorma; Nikopensius, Tiit; Province, Michael; Ketkar, Shamika; Colhoun, Helen; Doney, Alex; Robino, Antonietta; Krämer, Bernhard K.; Portas, Laura; Ford, Ian; Buckley, Brendan M.; Adam, Martin; Thun, Gian-Andri; Paulweber, Bernhard; Haun, Margot; Sala, Cinzia; Mitchell, Paul; Ciullo, Marina; Kim, Stuart K.; Vollenweider, Peter; Raitakari, Olli; Metspalu, Andres; Palmer, Colin; Gasparini, Paolo; Pirastu, Mario; Jukema, J. Wouter; Probst-Hensch, Nicole M.; Kronenberg, Florian; Toniolo, Daniela; Gudnason, Vilmundur; Shuldiner, Alan R.; Coresh, Josef; Schmidt, Reinhold; Ferrucci, Luigi; Siscovick, David S.; van Duijn, Cornelia M.; Borecki, Ingrid B.; Kardia, Sharon L.R.; Liu, Yongmei; Curhan, Gary C.; Rudan, Igor; Gyllensten, Ulf; Wilson, James F.; Franke, Andre; Pramstaller, Peter P.; Rettig, Rainer; Prokopenko, Inga; Witteman, Jacqueline; Hayward, Caroline; Ridker, Paul M; Parsa, Afshin; Bochud, Murielle; Heid, Iris M.; Kao, W.H. Linda; Fox, Caroline S.; Köttgen, Anna

    2012-01-01

    In conducting genome-wide association studies (GWAS), analytical approaches leveraging biological information may further understanding of the pathophysiology of clinical traits. To discover novel associations with estimated glomerular filtration rate (eGFR), a measure of kidney function, we developed a strategy for integrating prior biological knowledge into the existing GWAS data for eGFR from the CKDGen Consortium. Our strategy focuses on single nucleotide polymorphism (SNPs) in genes that are connected by functional evidence, determined by literature mining and gene ontology (GO) hierarchies, to genes near previously validated eGFR associations. It then requires association thresholds consistent with multiple testing, and finally evaluates novel candidates by independent replication. Among the samples of European ancestry, we identified a genome-wide significant SNP in FBXL20 (P = 5.6 × 10−9) in meta-analysis of all available data, and additional SNPs at the INHBC, LRP2, PLEKHA1, SLC3A2 and SLC7A6 genes meeting multiple-testing corrected significance for replication and overall P-values of 4.5 × 10−4–2.2 × 10−7. Neither the novel PLEKHA1 nor FBXL20 associations, both further supported by association with eGFR among African Americans and with transcript abundance, would have been implicated by eGFR candidate gene approaches. LRP2, encoding the megalin receptor, was identified through connection with the previously known eGFR gene DAB2 and extends understanding of the megalin system in kidney function. These findings highlight integration of existing genome-wide association data with independent biological knowledge to uncover novel candidate eGFR associations, including candidates lacking known connections to kidney-specific pathways. The strategy may also be applicable to other clinical phenotypes, although more testing will be needed to assess its potential for discovery in general. PMID:22962313

  6. Comparison of Muscle Transcriptome between Pigs with Divergent Meat Quality Phenotypes Identifies Genes Related to Muscle Metabolism and Structure

    PubMed Central

    Damon, Marie; Wyszynska-Koko, Joanna; Vincent, Annie; Hérault, Frédéric; Lebret, Bénédicte

    2012-01-01

    Background Meat quality depends on physiological processes taking place in muscle tissue, which could involve a large pattern of genes associated with both muscle structural and metabolic features. Understanding the biological phenomena underlying muscle phenotype at slaughter is necessary to uncover meat quality development. Therefore, a muscle transcriptome analysis was undertaken to compare gene expression profiles between two highly contrasted pig breeds, Large White (LW) and Basque (B), reared in two different housing systems themselves influencing meat quality. LW is the most predominant breed used in pig industry, which exhibits standard meat quality attributes. B is an indigenous breed with low lean meat and high fat contents, high meat quality characteristics, and is genetically distant from other European pig breeds. Methodology/Principal Findings Transcriptome analysis undertaken using a custom 15 K microarray, highlighted 1233 genes differentially expressed between breeds (multiple-test adjusted P-value<0.05), out of which 635 were highly expressed in the B and 598 highly expressed in the LW pigs. No difference in gene expression was found between housing systems. Besides, expression level of 12 differentially expressed genes quantified by real-time RT-PCR validated microarray data. Functional annotation clustering emphasized four main clusters associated to transcriptome breed differences: metabolic processes, skeletal muscle structure and organization, extracellular matrix, lysosome, and proteolysis, thereby highlighting many genes involved in muscle physiology and meat quality development. Conclusions/Significance Altogether, these results will contribute to a better understanding of muscle physiology and of the biological and molecular processes underlying meat quality. Besides, this study is a first step towards the identification of molecular markers of pork quality and the subsequent development of control tools. PMID:22470472

  7. De novo RNA sequencing and transcriptome analysis of Colletotrichum gloeosporioides ES026 reveal genes related to biosynthesis of huperzine A.

    PubMed

    Zhang, Guowei; Wang, Wenjuan; Zhang, Xiangmei; Xia, Qianqian; Zhao, Xinmei; Ahn, Youngjoon; Ahmed, Nevin; Cosoveanu, Andreea; Wang, Mo; Wang, Jialu; Shu, Shaohua

    2015-01-01

    Huperzine A is important in the treatment of Alzheimer's disease. There are major challenges for the mass production of huperzine A from plants due to the limited number of huperzine-A-producing plants, as well as the low content of huperzine A in these plants. Various endophytic fungi produce huperzine A. Colletotrichum gloeosporioides ES026 was previously isolated from a huperzine-A-producing plant Huperzia serrata, and this fungus also produces huperzine A. In this study, de novo RNA sequencing of C. gloeosporioides ES026 was carried out with an Illumina HiSeq2000. A total of 4,324,299,051 bp from 50,442,617 high-quality sequence reads of ES026 were obtained. These raw data were assembled into 24,998 unigenes, 40,536,684 residues and 19,790 genes. The majority of the unique sequences were assigned to corresponding putative functions based on BLAST searches of public databases. The molecular functions, biological processes and biochemical pathways of these unique sequences were determined using gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) assignments. A gene encoding copper amine oxidase (CAO) (unigene 9322) was annotated for the conversion of cadaverine to 5-aminopentanal in the biosynthesis of huperzine A. This gene was also detected in the root, stem and leaf of H. serrata. Furthermore, a close relationship was observed between expression of the CAO gene (unigene 9322) and quantity of crude huperzine A extracted from ES026. Therefore, CAO might be involved in the biosynthesis of huperzine A and it most likely plays a key role in regulating the content of huperzine A in ES026. PMID:25799531

  8. Shared genes related to aggression, rather than chemical communication, are associated with reproductive dominance in paper wasps (Polistes metricus)

    PubMed Central

    2014-01-01

    Background In social groups, dominant individuals may socially inhibit reproduction of subordinates using aggressive interactions or, in the case of highly eusocial insects, pheromonal communication. It has been hypothesized these two modes of reproductive inhibition utilize conserved pathways. Here, we use a comparative framework to investigate the chemical and genomic underpinnings of reproductive dominance in the primitively eusocial wasp Polistes metricus. Our goals were to first characterize transcriptomic and chemical correlates of reproductive dominance and second, to test whether dominance-associated mechanisms in paper wasps overlapped with aggression or pheromone-related gene expression patterns in other species. To explore whether conserved molecular pathways relate to dominance, we compared wasp transcriptomic data to previous studies of gene expression associated with pheromonal communication and queen-worker differences in honey bees, and aggressive behavior in bees, Drosophila, and mice. Results By examining dominant and subordinate females from queen and worker castes in early and late season colonies, we found that cuticular hydrocarbon profiles and genome-wide patterns of brain gene expression were primarily associated with season/social environment rather than dominance status. In contrast, gene expression patterns in the ovaries were associated primarily with caste and ovary activation. Comparative analyses suggest genes identified as differentially expressed in wasp brains are not related to queen pheromonal communication or caste in bees, but were significantly more likely to be associated with aggression in other insects (bees, flies), and even a mammal (mice). Conclusions This study provides the first comprehensive chemical and molecular analysis of reproductive dominance in paper wasps. We found little evidence for a chemical basis for reproductive dominance in P. metricus, and our transcriptomic analyses suggest that different pathways

  9. Chromatin structures of the rat tyrosine aminotransferase gene relate to the function of its cis-acting elements.

    PubMed Central

    Nitsch, D; Stewart, A F; Boshart, M; Mestril, R; Weih, F; Schütz, G

    1990-01-01

    The relationship between DNase I-hypersensitive sites (HSs) and transcriptional enhancers of the rat tyrosine aminotransferase (TAT) gene was examined by comparing HSs in and around the TAT gene with the activity of the corresponding DNA sequences in transient transfection assays. In this manner, we identified two HSs as liver-specific enhancers. Of three hepatoma cell lines examined, only one sustained TAT mRNA levels comparable to those of liver. In this cell line, both enhancers were strongly active, and strong hypersensitivity in chromatin over the enhancers was evident. The other two hepatoma cell lines had reduced levels of TAT mRNA and no or altered hypersensitivity over either the enhancers or the promoter. One of these lines carried a negative regulator of the TAT gene, the tissue specific extinguisher Tse-1. This cell line exhibited all HSs characteristic of the strongly active gene except at the promoter; however, one enhancer was inactive even though hypersensitive in chromatin. In a TAT-nonexpressing cell line, inactivity of both enhancers correlated with absence of the respective HSs. We conclude that although hypersensitivity in chromatin necessarily accompanies cell-type-specific enhancer activity, the occurrence of cell-type-specific HSs does not imply that the underlying sequences harbor enhancers active in transient transfection assays. Images PMID:1972541

  10. Fungal endophytes of Catharanthus roseus enhance vindoline content by modulating structural and regulatory genes related to terpenoid indole alkaloid biosynthesis

    PubMed Central

    Pandey, Shiv S.; Singh, Sucheta; Babu, C. S. Vivek; Shanker, Karuna; Srivastava, N. K.; Shukla, Ashutosh K.; Kalra, Alok

    2016-01-01

    Not much is known about the mechanism of endophyte-mediated induction of secondary metabolite production in Catharanthus roseus. In the present study two fungal endophytes, Curvularia sp. CATDLF5 and Choanephora infundibulifera CATDLF6 were isolated from the leaves of the plant that were found to enhance vindoline content by 229–403%. The isolated endophytes did not affect the primary metabolism of the plant as the maximum quantum efficiency of PSII, net CO2 assimilation, plant biomass and starch content of endophyte-inoculated plants was similar to endophyte-free control plants. Expression of terpenoid indole alkaloid (TIA) pathway genes, geraniol 10-hydroxylase (G10H), tryptophan decarboxylase (TDC), strictosidine synthase (STR), 16-hydoxytabersonine-O-methyltransferase (16OMT), desacetoxyvindoline-4-hydroxylase (D4H), deacetylvindoline-4-O-acetyltransferase (DAT) were upregulated in endophyte-inoculated plants. Endophyte inoculation upregulated the expression of the gene for transcriptional activator octadecanoid-responsive Catharanthus AP2-domain protein (ORCA3) and downregulated the expression of Cys2/His2-type zinc finger protein family transcriptional repressors (ZCTs). The gene for the vacuolar class III peroxidase (PRX1), responsible for coupling vindoline and catharanthine, was upregulated in endophyte-inoculated plants. These endophytes may enhance vindoline production by modulating the expression of key structural and regulatory genes of vindoline biosynthesis without affecting the primary metabolism of the host plant. PMID:27220774

  11. Genes Related to Long Polar Fimbriae of Pathogenic Escherichia coli Strains as Reliable Markers To Identify Virulent Isolates▿ †

    PubMed Central

    Torres, Alfredo G.; Blanco, Miguel; Valenzuela, Patricio; Slater, Terry M.; Patel, Shilpa D.; Dahbi, Ghizlane; López, Cecilia; Barriga, Ximena Fernández; Blanco, Jesús E.; Gomes, Tânia A. T.; Vidal, Roberto; Blanco, Jorge

    2009-01-01

    Lpf (stands for long polar fimbriae) is one of the few adhesive factors of enterohemorrhagic Escherichia coli O157:H7 associated with colonization of the intestine. E. coli O157:H7 strains possess two lpf loci encoding highly regulated fimbrial structures. Database analysis of the genes encoding the major fimbrial subunits demonstrated that they are present in commensal as well as pathogenic (both intestinal and extraintestinal) E. coli strains and in Salmonella strains and that the lpfA1 and lpfA2 genes are highly prevalent among LEE (locus of enterocyte effacement)-positive E. coli strains associated with severe and/or epidemic disease. Further DNA sequence analysis of the lpfA1 and lpfA2 genes from different attaching-and-effacing E. coli strains has led us to the identification of several polymorphisms and the classification of the major fimbrial subunits into distinct variants. Using collections of pathogenic E. coli isolates from Europe and Latin America, we demonstrated that the different lpfA types are associated with the presence of specific intimin (eae) adhesin variants and, most importantly, that they are found in specific E. coli pathotypes. Our results showed that the use of these fimbrial genes as markers, in combination with the different intimin types, resulted in a specific test for the identification of E. coli O157:H7, distinguishing it from other pathogenic E. coli strains. PMID:19494071

  12. Fungal endophytes of Catharanthus roseus enhance vindoline content by modulating structural and regulatory genes related to terpenoid indole alkaloid biosynthesis.

    PubMed

    Pandey, Shiv S; Singh, Sucheta; Babu, C S Vivek; Shanker, Karuna; Srivastava, N K; Shukla, Ashutosh K; Kalra, Alok

    2016-01-01

    Not much is known about the mechanism of endophyte-mediated induction of secondary metabolite production in Catharanthus roseus. In the present study two fungal endophytes, Curvularia sp. CATDLF5 and Choanephora infundibulifera CATDLF6 were isolated from the leaves of the plant that were found to enhance vindoline content by 229-403%. The isolated endophytes did not affect the primary metabolism of the plant as the maximum quantum efficiency of PSII, net CO2 assimilation, plant biomass and starch content of endophyte-inoculated plants was similar to endophyte-free control plants. Expression of terpenoid indole alkaloid (TIA) pathway genes, geraniol 10-hydroxylase (G10H), tryptophan decarboxylase (TDC), strictosidine synthase (STR), 16-hydoxytabersonine-O-methyltransferase (16OMT), desacetoxyvindoline-4-hydroxylase (D4H), deacetylvindoline-4-O-acetyltransferase (DAT) were upregulated in endophyte-inoculated plants. Endophyte inoculation upregulated the expression of the gene for transcriptional activator octadecanoid-responsive Catharanthus AP2-domain protein (ORCA3) and downregulated the expression of Cys2/His2-type zinc finger protein family transcriptional repressors (ZCTs). The gene for the vacuolar class III peroxidase (PRX1), responsible for coupling vindoline and catharanthine, was upregulated in endophyte-inoculated plants. These endophytes may enhance vindoline production by modulating the expression of key structural and regulatory genes of vindoline biosynthesis without affecting the primary metabolism of the host plant. PMID:27220774

  13. Dietary lysine affected the expression of genes related to lipid metabolism in skeletal muscle of finishing pigs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    It has been reported that some amino acids can function as signaling molecules to regulate skeletal muscle growth in mammals. This study was conducted to identify those genes that may be regulated by amino acid lysine and responsible for muscle growth and meat quality of pigs. Nine crossbred barrows...

  14. Genes related to suppression of malignant phenotype induced by Maitake D-Fraction in breast cancer cells.

    PubMed

    Alonso, Eliana Noelia; Orozco, Manuela; Eloy Nieto, Alvaro; Balogh, Gabriela Andrea

    2013-07-01

    It is already known that the Maitake (D-Fraction) mushroom is involved in stimulating the immune system and activating certain cells that attack cancer, including macrophages, T-cells, and natural killer cells. According to the U.S. National Cancer Institute, polysaccharide complexes present in Maitake mushrooms appear to have significant anticancer activity. However, the exact molecular mechanism of the Maitake antitumoral effect is still unclear. Previously, we have reported that Maitake (D-Fraction) induces apoptosis in breast cancer cells by activation of BCL2-antagonist/killer 1 (BAK1) gene expression. At the present work, we are identifying which genes are responsible for the suppression of the tumoral phenotype mechanism induced by Maitake (D-Fraction) in breast cancer cells. Human breast cancer MCF-7 cells were treated with and without increased concentrations of Maitake D-Fraction (36, 91, 183, 367 μg/mL) for 24 h. Total RNA were isolated and cDNA microarrays were hybridized containing 25,000 human genes. Employing the cDNA microarray analysis, we found that Maitake D-Fraction modified the expression of 4068 genes (2420 were upmodulated and 1648 were downmodulated) in MCF-7 breast cancer cells in a dose-dependent manner during 24 h of treatment. The present data shows that Maitake D-Fraction suppresses the breast tumoral phenotype through a putative molecular mechanism modifying the expression of certain genes (such as IGFBP-7, ITGA2, ICAM3, SOD2, CAV-1, Cul-3, NRF2, Cycline E, ST7, and SPARC) that are involved in apoptosis stimulation, inhibition of cell growth and proliferation, cell cycle arrest, blocking migration and metastasis of tumoral cells, and inducing multidrug sensitivity. Altogether, these results suggest that Maitake D-Fraction could be a potential new target for breast cancer chemoprevention and treatment. PMID:23875900

  15. Genes Related to Suppression of Malignant Phenotype Induced by Maitake D-Fraction in Breast Cancer Cells

    PubMed Central

    Alonso, Eliana Noelia; Orozco, Manuela; Nieto, Alvaro Eloy

    2013-01-01

    Abstract It is already known that the Maitake (D-Fraction) mushroom is involved in stimulating the immune system and activating certain cells that attack cancer, including macrophages, T-cells, and natural killer cells. According to the U.S. National Cancer Institute, polysaccharide complexes present in Maitake mushrooms appear to have significant anticancer activity. However, the exact molecular mechanism of the Maitake antitumoral effect is still unclear. Previously, we have reported that Maitake (D-Fraction) induces apoptosis in breast cancer cells by activation of BCL2-antagonist/killer 1 (BAK1) gene expression. At the present work, we are identifying which genes are responsible for the suppression of the tumoral phenotype mechanism induced by Maitake (D-Fraction) in breast cancer cells. Human breast cancer MCF-7 cells were treated with and without increased concentrations of Maitake D-Fraction (36, 91, 183, 367 μg/mL) for 24 h. Total RNA were isolated and cDNA microarrays were hybridized containing 25,000 human genes. Employing the cDNA microarray analysis, we found that Maitake D-Fraction modified the expression of 4068 genes (2420 were upmodulated and 1648 were downmodulated) in MCF-7 breast cancer cells in a dose-dependent manner during 24 h of treatment. The present data shows that Maitake D-Fraction suppresses the breast tumoral phenotype through a putative molecular mechanism modifying the expression of certain genes (such as IGFBP-7, ITGA2, ICAM3, SOD2, CAV-1, Cul-3, NRF2, Cycline E, ST7, and SPARC) that are involved in apoptosis stimulation, inhibition of cell growth and proliferation, cell cycle arrest, blocking migration and metastasis of tumoral cells, and inducing multidrug sensitivity. Altogether, these results suggest that Maitake D-Fraction could be a potential new target for breast cancer chemoprevention and treatment. PMID:23875900

  16. Fish oil supplementation induces expression of genes related to cell cycle, endoplasmic reticulum stress and apoptosis in peripheral blood mononuclear cells: a transcriptomic approach

    PubMed Central

    Myhrstad, M C W; Ulven, S M; Günther, C-C; Ottestad, I; Holden, M; Ryeng, E; Borge, G I; Kohler, A; Brønner, K W; Thoresen, M; Holven, K B

    2014-01-01

    Background Fish oil supplementation has been shown to alter gene expression of mononuclear cells both in vitro and in vivo. However, little is known about the total transcriptome profile in healthy subjects after intake of fish oil. We therefore investigated the gene expression profile in peripheral blood mononuclear cells (PBMCs) after intake of fish oil for 7 weeks using transcriptome analyses. Design In a 7-week, double-blinded, randomized, controlled, parallel-group study, healthy subjects received 8 g day−1 fish oil (1.6 g day−1 eicosapentaenoic acid + docosahexaenoic acid) (n = 17) or 8 g day−1 high oleic sunflower oil (n = 19). Microarray analyses of RNA isolated from PBMCs were performed at baseline and after 7 weeks of intervention. Results Cell cycle, DNA packaging and chromosome organization are biological processes found to be upregulated after intake of fish oil compared to high oleic sunflower oil using a moderated t-test. In addition, gene set enrichment analysis identified several enriched gene sets after intake of fish oil. The genes contributing to the significantly different gene sets in the subjects given fish oil compared with the control group are involved in cell cycle, endoplasmic reticulum (ER) stress and apoptosis. Gene transcripts with common motifs for 35 known transcription factors including E2F, TP53 and ATF4 were upregulated after intake of fish oil. Conclusion We have shown that intake of fish oil for 7 weeks modulates gene expression in PBMCs of healthy subjects. The increased expression of genes related to cell cycle, ER stress and apoptosis suggests that intake of fish oil may modulate basic cellular processes involved in normal cellular function. PMID:24641624

  17. Diversity in the carotenoid profiles and the expression of genes related to carotenoid accumulation among citrus genotypes.

    PubMed

    Ikoma, Yoshinori; Matsumoto, Hikaru; Kato, Masaya

    2016-01-01

    Carotenoids are not only important to the plants themselves but also are beneficial to human health. Since citrus fruit is a good source of carotenoids for the human diet, it is important to study carotenoid profiles and the accumulation mechanism in citrus fruit. Thus, in the present paper, we describe the diversity in the carotenoid profiles of fruit among citrus genotypes. In regard to carotenoids, such as β-cryptoxanthin, violaxanthin, lycopene, and β-citraurin, the relationship between the carotenoid profile and the expression of carotenoid-biosynthetic genes is discussed. Finally, recent results of quantitative trait locus (QTL) analyses of carotenoid contents and expression levels of carotenoid-biosynthetic genes in citrus fruit are shown. PMID:27069398

  18. Diversity in the carotenoid profiles and the expression of genes related to carotenoid accumulation among citrus genotypes

    PubMed Central

    Ikoma, Yoshinori; Matsumoto, Hikaru; Kato, Masaya

    2016-01-01

    Carotenoids are not only important to the plants themselves but also are beneficial to human health. Since citrus fruit is a good source of carotenoids for the human diet, it is important to study carotenoid profiles and the accumulation mechanism in citrus fruit. Thus, in the present paper, we describe the diversity in the carotenoid profiles of fruit among citrus genotypes. In regard to carotenoids, such as β-cryptoxanthin, violaxanthin, lycopene, and β-citraurin, the relationship between the carotenoid profile and the expression of carotenoid-biosynthetic genes is discussed. Finally, recent results of quantitative trait locus (QTL) analyses of carotenoid contents and expression levels of carotenoid-biosynthetic genes in citrus fruit are shown. PMID:27069398

  19. Psychological factors and DNA methylation of genes related to immune/inflammatory system markers: the VA Normative Aging Study

    PubMed Central

    Kim, Daniel; Kubzansky, Laura D; Baccarelli, Andrea; Sparrow, David; Spiro, Avron; Tarantini, Letizia; Cantone, Laura; Vokonas, Pantel; Schwartz, Joel

    2016-01-01

    Objectives Although psychological factors have been associated with chronic diseases such as coronary heart disease (CHD), the underlying pathways for these associations have yet to be elucidated. DNA methylation has been posited as a mechanism linking psychological factors to CHD risk. In a cohort of community-dwelling elderly men, we explored the associations between positive and negative psychological factors with DNA methylation in promoter regions of multiple genes involved in immune/inflammatory processes related to atherosclerosis. Design Prospective cohort study. Setting Greater Boston, Massachusetts area. Participants Samples of 538 to 669 men participating in the Normative Aging Study cohort with psychological measures and DNA methylation measures, collected on 1–4 visits between 1999 and 2006 (mean age=72.7 years at first visit). Outcome measures We examined anxiety, depression, hostility and life satisfaction as predictors of leucocyte gene-specific DNA methylation. We estimated repeated measures linear mixed models, controlling for age, smoking, education, history of heart disease, stroke or diabetes, % lymphocytes, % monocytes and plasma folate. Results Psychological distress measured by anxiety, depression and hostility was positively associated, and happiness and life satisfaction were inversely associated with average Intercellular Adhesion Molecule-1 (ICAM-1) and coagulation factor III (F3) promoter methylation levels. There was some evidence that hostility was positively associated with toll-like receptor 2 (TLR-2) promoter methylation, and that life satisfaction was inversely associated with TLR-2 and inducible nitric oxide synthase (iNOS) promoter methylation. We observed less consistent and significant associations between psychological factors and average methylation for promoters of the genes for glucocorticoid receptor (NR3C1), interferon-γ (IFN-γ) and interleukin 6 (IL-6). Conclusions These findings suggest that positive and negative

  20. Induction of pseudohyphal growth by overexpression of PHD1, a Saccharomyces cerevisiae gene related to transcriptional regulators of fungal development.

    PubMed Central

    Gimeno, C J; Fink, G R

    1994-01-01

    When starved for nitrogen, MATa/MAT alpha cells of the budding yeast Saccharomyces cerevisiae undergo a dimorphic transition to pseudohyphal growth. A visual genetic screen, called PHD (pseudohyphal determinant), for S. cerevisiae pseudohyphal growth mutants was developed. The PHD screen was used to identify seven S. cerevisiae genes that when overexpressed in MATa/MAT alpha cells growing on nitrogen starvation medium cause precocious and unusually vigorous pseudohyphal growth. PHD1, a gene whose overexpression induced invasive pseudohyphal growth on a nutritionally rich medium, was characterized. PHD1 maps to chromosome XI and is predicted to encode a 366-amino-acid protein. PHD1 has a SWI4- and MBP1-like DNA binding motif that is 73% identical over 100 amino acids to a region of Aspergillus nidulans StuA. StuA regulates two pseudohyphal growth-like cell divisions during conidiophore morphogenesis. Epitope-tagged PHD1 was localized to the nucleus by indirect immunofluorescence. These facts suggest that PHD1 may function as a transcriptional regulatory protein. Overexpression of PHD1 in wild-type haploid strains does not induce pseudohyphal growth. Interestingly, PHD1 overexpression enhances pseudohyphal growth in a haploid strain that has the diploid polar budding pattern because of a mutation in the BUD4 gene. In addition, wild-type diploid strains lacking PHD1 undergo pseudohyphal growth when starved for nitrogen. The possible functions of PHD1 in pseudohyphal growth and the uses of the PHD screen to identify morphogenetic regulatory genes from heterologous organisms are discussed. Images PMID:8114741

  1. Comparative Transcriptome Analysis of Fetal Skin Reveals Key Genes Related to Hair Follicle Morphogenesis in Cashmere Goats.

    PubMed

    Gao, Ye; Wang, Xiaolong; Yan, Hailong; Zeng, Jie; Ma, Sen; Niu, Yiyuan; Zhou, Guangxian; Jiang, Yu; Chen, Yulin

    2016-01-01

    Cashmere goat skin contains two types of hair follicles (HF): primary hair follicles (PHF) and secondary hair follicles (SHF). Although multiple genetic determinants associated with HF formation have been identified, the molecules that determine the independent morphogenesis of HF in cashmere goats remain elusive. The growth and development of SHF directly influence the quantity and quality of cashmere production. Here, we report the transcriptome profiling analysis of nine skin samples from cashmere goats using 60- and 120-day-old embryos (E60 and E120, respectively), as well as newborns (NB), through RNA-sequencing (RNA-seq). HF morphological changes indicated that PHF were initiated at E60, with maturation from E120, while differentiation of SHF was identified at E120 until formation of cashmere occurred after birth (NB). The RNA-sequencing analysis generated over 20.6 million clean reads from each mRNA library. The number of differentially expressed genes (DEGs) in E60 vs. E120, E120 vs. NB, and E60 vs. NB were 1,024, 0 and 1,801, respectively, indicating that no significant differences were found at transcriptomic levels between E120 and NB. Key genes including B4GALT4, TNC, a-integrin, and FGFR1, were up-regulated and expressed in HF initiation from E60 to E120, while regulatory genes such as GPRC5D, PAD3, HOXC13, PRR9, VSIG8, LRRC15, LHX2, MSX-2, and FOXN1 were up-regulated and expressed in HF keratinisation and hair shaft differentiation from E120 and NB to E60. Several genes belonging to the KRT and KRTAP gene families were detected throughout the three HF developmental stages. The transcriptional trajectory analyses of all DEGs indicated that immune privilege, glycosaminoglycan biosynthesis, extracellular matrix receptor interaction, and growth factor receptors all played dominant roles in the epithelial-mesenchymal interface and HF formation. We found that the Wnt, transforming growth factor-beta/bone morphogenetic protein, and Notch family members

  2. Comparative Transcriptome Analysis of Fetal Skin Reveals Key Genes Related to Hair Follicle Morphogenesis in Cashmere Goats

    PubMed Central

    Yan, Hailong; Zeng, Jie; Ma, Sen; Niu, Yiyuan; Zhou, Guangxian; Jiang, Yu; Chen, Yulin

    2016-01-01

    Cashmere goat skin contains two types of hair follicles (HF): primary hair follicles (PHF) and secondary hair follicles (SHF). Although multiple genetic determinants associated with HF formation have been identified, the molecules that determine the independent morphogenesis of HF in cashmere goats remain elusive. The growth and development of SHF directly influence the quantity and quality of cashmere production. Here, we report the transcriptome profiling analysis of nine skin samples from cashmere goats using 60- and 120-day-old embryos (E60 and E120, respectively), as well as newborns (NB), through RNA-sequencing (RNA-seq). HF morphological changes indicated that PHF were initiated at E60, with maturation from E120, while differentiation of SHF was identified at E120 until formation of cashmere occurred after birth (NB). The RNA-sequencing analysis generated over 20.6 million clean reads from each mRNA library. The number of differentially expressed genes (DEGs) in E60 vs. E120, E120 vs. NB, and E60 vs. NB were 1,024, 0 and 1,801, respectively, indicating that no significant differences were found at transcriptomic levels between E120 and NB. Key genes including B4GALT4, TNC, a-integrin, and FGFR1, were up-regulated and expressed in HF initiation from E60 to E120, while regulatory genes such as GPRC5D, PAD3, HOXC13, PRR9, VSIG8, LRRC15, LHX2, MSX-2, and FOXN1 were up-regulated and expressed in HF keratinisation and hair shaft differentiation from E120 and NB to E60. Several genes belonging to the KRT and KRTAP gene families were detected throughout the three HF developmental stages. The transcriptional trajectory analyses of all DEGs indicated that immune privilege, glycosaminoglycan biosynthesis, extracellular matrix receptor interaction, and growth factor receptors all played dominant roles in the epithelial-mesenchymal interface and HF formation. We found that the Wnt, transforming growth factor-beta/bone morphogenetic protein, and Notch family members

  3. Transcriptome Analysis of the Oil-Rich Tea Plant, Camellia oleifera, Reveals Candidate Genes Related to Lipid Metabolism

    PubMed Central

    Huang, Hui; Zhang, Li-Ping; Zhang, Hai-Bin; Gao, Li-Zhi

    2014-01-01

    Background Rapidly driven by the need for developing sustainable sources of nutritionally important fatty acids and the rising concerns about environmental impacts after using fossil oil, oil-plants have received increasing awareness nowadays. As an important oil-rich plant in China, Camellia oleifera has played a vital role in providing nutritional applications, biofuel productions and chemical feedstocks. However, the lack of C. oleifera genome sequences and little genetic information have largely hampered the urgent needs for efficient utilization of the abundant germplasms towards modern breeding efforts of this woody oil-plant. Results Here, using the 454 GS-FLX sequencing platform, we generated approximately 600,000 RNA-Seq reads from four tissues of C. oleifera. These reads were trimmed and assembled into 104,842 non-redundant putative transcripts with a total length of ∼38.9 Mb, representing more than 218-fold of all the C. oleifera sequences currently deposited in the GenBank (as of March 2014). Based on the BLAST similarity searches, nearly 42.6% transcripts could be annotated with known genes, conserved domains, or Gene Ontology (GO) terms. Comparisons with the cultivated tea tree, C. sinensis, identified 3,022 pairs of orthologs, of which 211 exhibited the evidence under positive selection. Pathway analysis detected the majority of genes potentially related to lipid metabolism. Evolutionary analysis of omega-6 fatty acid desaturase (FAD2) genes among 20 oil-plants unexpectedly suggests that a parallel evolution may occur between C. oleifera and Olea oleifera. Additionally, more than 2,300 simple sequence repeats (SSRs) and 20,200 single-nucleotide polymorphisms (SNPs) were detected in the C. oleifera transcriptome. Conclusions The generated transcriptome represents a considerable increase in the number of sequences deposited in the public databases, providing an unprecedented opportunity to discover all related-genes associated with lipid metabolic

  4. PI3K-Akt-mTOR signal inhibition affects expression of genes related to endoplasmic reticulum stress.

    PubMed

    Song, Q; Han, C C; Xiong, X P; He, F; Gan, W; Wei, S H; Liu, H H; Li, L; Xu, H Y

    2016-01-01

    PI3K-Akt-mTOR signaling pathway is associated with endoplasmic reticulum (ER) stress. However, it is not clear how this signaling pathway affects the ER stress. The present study aimed to determine whether the PI3K-Akt-mTOR signaling pathway regulates tunicamycin (TM)-induced increases in mRNA levels of genes involved in the ER stress, to help elucidate the mechanism by which this pathway affects the ER stress in primary goose hepatocytes. Primary hepatocytes were isolated from geese and cultured in vitro. After 12 h in a serum-free medium, the hepatocytes were incubated for 24 h in a medium with either no addition (control) or with supplementation of TM or TM together with PI3K-Akt-mTOR signaling pathway inhibitors (LY294002, rapamycin, NVP-BEZ235). Thereafter, the expression levels of genes involved in the ER stress (BIP, EIF2a, ATF6, and XBP1) were assessed. The results indicated that the mRNA level of BIP was up-regulated in 0.2, 2, and 20 μM TM treatment group (P < 0.05), whereas the mRNA levels of EIF2a, ATF6, and XBP1 were up-regulated in the 2 μM TM treatment group (P < 0.05). However, the TM mediated induction of mRNA levels of genes involved in the ER stress (BIP, EIF2a, ATF6, and XBP1) was down-regulated after the treatment with PI3K-Akt-mTOR pathway inhibitors (LY294002, NVP-BEZ235, and rapamycin). Therefore, our results strongly suggest that the PI3K-Akt-mTOR signaling pathway might be involved in the down-regulation of the TM-induced ER stress in primary goose hepatocytes. PMID:27525855

  5. Yeast Vps55p, a functional homolog of human obesity receptor gene-related protein, is involved in late endosome to vacuole trafficking.

    PubMed

    Belgareh-Touzé, Naïma; Avaro, Sandrine; Rouillé, Yves; Hoflack, Bernard; Haguenauer-Tsapis, Rosine

    2002-05-01

    The Saccharomyces cerevisiae VPS55 (YJR044c) gene encodes a small protein of 140 amino acids with four potential transmembrane domains. VPS55 belongs to a family of genes of unknown function, including the human gene encoding the obesity receptor gene-related protein (OB-RGRP). Yeast cells with a disrupted VPS55 present normal vacuolar morphology, but exhibit an abnormal secretion of the Golgi form of the soluble vacuolar carboxypeptidase Y. However, trafficking of the membrane-bound vacuolar alkaline phosphatase remains normal. The endocytosis of uracil permease, used as an endocytic marker, is normal in vps55Delta cells, but its degradation is delayed and this marker transiently accumulates in late endosomal compartments. We also found that Vps55p is mainly localized in the late endosomes. Collectively, these results indicate that Vps55p is involved in late endosome to vacuole trafficking. Finally, we show that human OB-RGRP displays the same distribution as Vps55p and corrects the phenotypic defects of the vps55Delta strain. Therefore, the function of Vps55p has been conserved throughout evolution. This study highlights the importance of the multispanning Vps55p and OB-RGRP in membrane trafficking to the vacuole/lysosome of eukaryotic cells. PMID:12006663

  6. Yeast Vps55p, a Functional Homolog of Human Obesity Receptor Gene-related Protein, Is Involved in Late Endosome to Vacuole Trafficking

    PubMed Central

    Belgareh-Touzé, Naïma; Avaro, Sandrine; Rouillé, Yves; Hoflack, Bernard; Haguenauer-Tsapis, Rosine

    2002-01-01

    The Saccharomyces cerevisiae VPS55 (YJR044c) gene encodes a small protein of 140 amino acids with four potential transmembrane domains. VPS55 belongs to a family of genes of unknown function, including the human gene encoding the obesity receptor gene-related protein (OB-RGRP). Yeast cells with a disrupted VPS55 present normal vacuolar morphology, but exhibit an abnormal secretion of the Golgi form of the soluble vacuolar carboxypeptidase Y. However, trafficking of the membrane-bound vacuolar alkaline phosphatase remains normal. The endocytosis of uracil permease, used as an endocytic marker, is normal in vps55Δ cells, but its degradation is delayed and this marker transiently accumulates in late endosomal compartments. We also found that Vps55p is mainly localized in the late endosomes. Collectively, these results indicate that Vps55p is involved in late endosome to vacuole trafficking. Finally, we show that human OB-RGRP displays the same distribution as Vps55p and corrects the phenotypic defects of the vps55Δ strain. Therefore, the function of Vps55p has been conserved throughout evolution. This study highlights the importance of the multispanning Vps55p and OB-RGRP in membrane trafficking to the vacuole/lysosome of eukaryotic cells. PMID:12006663

  7. Differential Effect of Lactobacillus johnsonii BFE 6128 on Expression of Genes Related to TLR Pathways and Innate Immunity in Intestinal Epithelial Cells.

    PubMed

    Seifert, Stephanie; Rodriguez Gómez, Manuel; Watzl, Bernhard; Holzapfel, Wilhelm H; Franz, Charles M A P; Vizoso Pinto, María G

    2010-12-01

    Probiotics have been shown to enhance immune defenses, but their mechanisms of action are only partially understood. We investigated the modulation of signal pathways involved in innate immunity in enterocytes by Lactobacillus johnsonii BFE 6128 isolated from 'Kule naoto', a Maasai traditional fermented milk product. This lactobacillus sensitized HT29 intestinal epithelial cells toward recognition of Salmonella enterica serovar Typhimurium by increasing the IL-8 levels released after challenge with this pathogen and by differentially modulating genes related to toll-like receptor (TLR) pathways and innate immunity. Thus, the modulation of pro-inflammatory mediators and TLR-pathway-related molecules may be an important mechanism contributing to the potential stimulation of innate immunity by lactobacilli at the intestinal epithelial level. PMID:26781315

  8. Expression of genes related to apoptosis, cell cycle and signaling pathways are independent of TP53 status in urinary bladder cancer cells.

    PubMed

    da Silva, Glenda N; Evangelista, Adriane F; Magalhães, Danielle A; Macedo, Cláudia; Búfalo, Michelle C; Sakamoto-Hojo, Elza T; Passos, Geraldo A S; Salvadori, Daisy M F

    2011-08-01

    Urinary bladder cancer is the fourth most common malignancy in the Western world. Transitional cell carcinoma (TCC) is the most common subtype, accounting for about 90% of all bladder cancers. The TP53 gene plays an essential role in the regulation of the cell cycle and apoptosis and therefore contributes to cellular transformation and malignancy; however, little is known about the differential gene expression patterns in human tumors that present with the wild-type or mutated TP53 gene. Therefore, because gene profiling can provide new insights into the molecular biology of bladder cancer, the present study aimed to compare the molecular profiles of bladder cancer cell lines with different TP53 alleles, including the wild type (RT4) and two mutants (5637, with mutations in codons 280 and 72; and T24, a TP53 allele encoding an in-frame deletion of tyrosine 126). Unsupervised hierarchical clustering and gene networks were constructed based on data generated by cDNA microarrays using mRNA from the three cell lines. Differentially expressed genes related to the cell cycle, cell division, cell death, and cell proliferation were observed in the three cell lines. However, the cDNA microarray data did not cluster cell lines based on their TP53 allele. The gene profiles of the RT4 cells were more similar to those of T24 than to those of the 5637 cells. While the deregulation of both the cell cycle and the apoptotic pathways was particularly related to TCC, these alterations were not associated with the TP53 status. PMID:21116856

  9. The impact of mitochondrial DNA and nuclear genes related to mitochondrial functioning on the risk of Parkinson's disease.

    PubMed

    Gaweda-Walerych, Katarzyna; Zekanowski, Cezary

    2013-12-01

    Mitochondrial dysfunction and oxidative stress are the major factors implicated in Parkinson's disease (PD) pathogenesis. The maintenance of healthy mitochondria is a very complex process coordinated bi-genomically. Here, we review association studies on mitochondrial haplogroups and subhaplogroups, discussing the underlying molecular mechanisms. We also focus on variation in the nuclear genes (NDUFV2, PGC-1alpha, HSPA9, LRPPRC, MTIF3, POLG1, and TFAM encoding NADH dehydrogenase (ubiquinone) flavoprotein 2, peroxisome proliferator-activated receptor gamma coactivator 1-alpha, mortalin, leucine-rich pentatricopeptide repeat containing protein, translation initiation factor 3, mitochondrial DNA polymerase gamma, and mitochondrial transcription factor A, respectively) primarily linked to regulation of mitochondrial functioning that recently have been associated with PD risk. Possible interactions between mitochondrial and nuclear genetic variants and related proteins are discussed. PMID:24532986

  10. The Impact of Mitochondrial DNA and Nuclear Genes Related to Mitochondrial Functioning on the Risk of Parkinson’s Disease

    PubMed Central

    Gaweda-Walerych, Katarzyna; Zekanowski, Cezary

    2013-01-01

    Mitochondrial dysfunction and oxidative stress are the major factors implicated in Parkinson’s disease (PD) pathogenesis. The maintenance of healthy mitochondria is a very complex process coordinated bi-genomically. Here, we review association studies on mitochondrial haplogroups and subhaplogroups, discussing the underlying molecular mechanisms. We also focus on variation in the nuclear genes (NDUFV2, PGC-1alpha, HSPA9, LRPPRC, MTIF3, POLG1, and TFAM encoding NADH dehydrogenase (ubiquinone) flavoprotein 2, peroxisome proliferator-activated receptor gamma coactivator 1-alpha, mortalin, leucine-rich pentatricopeptide repeat containing protein, translation initiation factor 3, mitochondrial DNA polymerase gamma, and mitochondrial transcription factor A, respectively) primarily linked to regulation of mitochondrial functioning that recently have been associated with PD risk. Possible interactions between mitochondrial and nuclear genetic variants and related proteins are discussed. PMID:24532986

  11. Association study of genes related to bone formation and resorption and the extent of radiographic change in ankylosing spondylitis

    PubMed Central

    Cortes, A; Maksymowych, W P; Wordsworth, B P; Inman, R D; Danoy, P; Rahman, P; Stone, M A; Corr, M; Gensler, Lianne S; Gladman, D; Morgan, A; Marzo-Ortega, H; Ward, M M; Learch, T J; Reveille, J D; Brown, M A; Weisman, M H

    2014-01-01

    Objective To identify genetic associations with severity of radiographic damage in ankylosing spondylitis (AS). Method We studied 1537 AS cases of European descent; all fulfilled the modified New York Criteria. Radiographic severity was assessed from digitised lateral radiographs of the cervical and lumbar spine using the modified Stoke Ankylosing Spondylitis Spinal Score (mSASSS). A two-phase genotyping design was used. In phase 1, 498 single nucleotide polymorphisms (SNPs) were genotyped in 688 cases; these were selected to capture >90% of the common haplotypic variation in the exons, exon–intron boundaries, and 5 kb flanking DNA in the 5′ and 3′ UTR of 74 genes involved in anabolic or catabolic bone pathways. In phase 2, 15 SNPs exhibiting p<0.05 were genotyped in a further cohort of 830 AS cases; results were analysed both separately and in combination with the discovery phase data. Association was tested by contingency tables after separating the samples into ‘mild’ and ‘severe’ groups, defined as the bottom and top 40% by mSASSS, adjusted for gender and disease duration. Results Experiment-wise association was observed with the SNP rs8092336 (combined OR 0.32, p=1.2×10−5), which lies within RANK (receptor activator of NFκB), a gene involved in osteoclastogenesis, and in the interaction between T cells and dendritic cells. Association was also found with the SNP rs1236913 in PTGS1 (prostaglandin-endoperoxide synthase 1, cyclooxygenase 1), giving an OR of 0.53 (p=2.6×10−3). There was no observed association between radiographic severity and HLA-B*27. Conclusions These findings support roles for bone resorption and prostaglandins pathways in the osteoproliferative changes in AS. PMID:24651623

  12. Practicality of Intermittent Fasting in Humans and its Effect on Oxidative Stress and Genes Related to Aging and Metabolism

    PubMed Central

    Wegman, Martin P.; Guo, Michael H.; Bennion, Douglas M.; Shankar, Meena N.; Chrzanowski, Stephen M.; Goldberg, Leslie A.; Xu, Jinze; Williams, Tiffany A.; Lu, Xiaomin; Hsu, Stephen I.; Anton, Stephen D.; Leeuwenburgh, Christiaan

    2015-01-01

    Abstract Caloric restriction has consistently been shown to extend life span and ameliorate aging-related diseases. These effects may be due to diet-induced reactive oxygen species acting to up-regulate sirtuins and related protective pathways, which research suggests may be partially inhibited by dietary anti-oxidant supplementation. Because caloric restriction is not sustainable long term for most humans, we investigated an alternative dietary approach, intermittent fasting (IF), which is proposed to act on similar biological pathways. We hypothesized that a modified IF diet, where participants maintain overall energy balance by alternating between days of fasting (25% of normal caloric intake) and feasting (175% of normal), would increase expression of genes associated with aging and reduce oxidative stress and that these effects would be suppressed by anti-oxidant supplementation. To assess the tolerability of the diet and to explore effects on biological mechanisms related to aging and metabolism, we recruited a cohort of 24 healthy individuals in a double-crossover, double-blinded, randomized clinical trial. Study participants underwent two 3-week treatment periods—IF and IF with anti-oxidant (vitamins C and E) supplementation. We found strict adherence to study-provided diets and that participants found the diet tolerable, with no adverse clinical findings or weight change. We detected a marginal increase (2.7%) in SIRT3 expression due to the IF diet, but no change in expression of other genes or oxidative stress markers analyzed. We also found that IF decreased plasma insulin levels (1.01 μU/mL). Although our study suggests that the IF dieting paradigm is acceptable in healthy individuals, additional research is needed to further assess the potential benefits and risks. PMID:25546413

  13. Genome-Wide Analysis of MicroRNAs and Their Target Genes Related to Leaf Senescence of Rice

    PubMed Central

    Liu, Chaoping; Chen, Eryong; Chen, Qifeng; Zhuang, Jieyun; Shen, Bo

    2014-01-01

    Grain production of rice (Oryza sativa L.) is a top priority in ensuring food security for human beings. One of the approaches to increase yield is to delay leaf senescence and to extend the available time for photosynthesis. MicroRNAs (miRNAs) are key regulators of aging and cellular senescence in eukaryotes. Here, to help understand their biological role in rice leaf senescence, we report identification of miRNAs and their putative target genes by deep sequencing of six small RNA libraries, six RNA-seq libraries and two degradome libraries from the leaves of two super hybrid rice, Nei-2-You 6 (N2Y6, age-resistant rice) and Liang-You-Pei 9 (LYP9, age-sensitive rice). In total 372 known miRNAs, 162 miRNA candidates and 1145 targets were identified. Compared with the expression of miRNAs in the leaves of LYP9, the numbers of miRNAs up-regulated and down-regulated in the leaves of N2Y6 were 47 and 30 at early stage of grain-filling, 21 and 17 at the middle stage, and 11 and 37 at the late stage, respectively. Six miRNA families, osa-miR159, osa-miR160 osa-miR164, osa-miR167, osa-miR172 and osa-miR1848, targeting the genes encoding APETALA2 (AP2), zinc finger proteins, salicylic acid-induced protein 19 (SIP19), auxin response factors (ARF) and NAC transcription factors, respectively, were found to be involved in leaf senescence through phytohormone signaling pathways. These results provided valuable information for understanding the miRNA-mediated leaf senescence of rice, and offered an important foundation for rice breeding. PMID:25479006

  14. Practicality of intermittent fasting in humans and its effect on oxidative stress and genes related to aging and metabolism.

    PubMed

    Wegman, Martin P; Guo, Michael H; Bennion, Douglas M; Shankar, Meena N; Chrzanowski, Stephen M; Goldberg, Leslie A; Xu, Jinze; Williams, Tiffany A; Lu, Xiaomin; Hsu, Stephen I; Anton, Stephen D; Leeuwenburgh, Christiaan; Brantly, Mark L

    2015-04-01

    Caloric restriction has consistently been shown to extend life span and ameliorate aging-related diseases. These effects may be due to diet-induced reactive oxygen species acting to up-regulate sirtuins and related protective pathways, which research suggests may be partially inhibited by dietary anti-oxidant supplementation. Because caloric restriction is not sustainable long term for most humans, we investigated an alternative dietary approach, intermittent fasting (IF), which is proposed to act on similar biological pathways. We hypothesized that a modified IF diet, where participants maintain overall energy balance by alternating between days of fasting (25% of normal caloric intake) and feasting (175% of normal), would increase expression of genes associated with aging and reduce oxidative stress and that these effects would be suppressed by anti-oxidant supplementation. To assess the tolerability of the diet and to explore effects on biological mechanisms related to aging and metabolism, we recruited a cohort of 24 healthy individuals in a double-crossover, double-blinded, randomized clinical trial. Study participants underwent two 3-week treatment periods-IF and IF with anti-oxidant (vitamins C and E) supplementation. We found strict adherence to study-provided diets and that participants found the diet tolerable, with no adverse clinical findings or weight change. We detected a marginal increase (2.7%) in SIRT3 expression due to the IF diet, but no change in expression of other genes or oxidative stress markers analyzed. We also found that IF decreased plasma insulin levels (1.01 μU/mL). Although our study suggests that the IF dieting paradigm is acceptable in healthy individuals, additional research is needed to further assess the potential benefits and risks. PMID:25546413

  15. Embryonic exposure to carbendazim induces the transcription of genes related to apoptosis, immunotoxicity and endocrine disruption in zebrafish (Danio rerio).

    PubMed

    Jiang, Jinhua; Wu, Shenggan; Wu, Changxing; An, Xuehua; Cai, Leiming; Zhao, Xueping

    2014-12-01

    Carbendazim is one of the most widespread environmental contaminant that can cause major concern to human and animal reproductive system. To date, very few studies have been conducted on the toxic effect of carbendazim in the non-target organism zebrafish (Danio rerio). The study presented here aimed to assess how carbendazim triggers apoptosis, immunotoxicity and endocrine disruption pathways in zebrafish during its embryo development. Our results demonstrated that the expression patterns of many key genes involved in cell apoptosis pathway (e.g. P53, Mdm2, Bbc3 and Cas8) were significantly up-regulated upon the exposure to carbendazim at the concentration of 500 μg/L, while the Bcl2 and Cas3 were down-regulated at the same concentration, interestingly, the expression level of Ogg1 decreased at all the exposure concentrations. It was also observed that the mRNA levels of CXCL-C1C, CCL1, IL-1b and TNFα which were closely related to the innate immune system, were affected in newly hatched zebrafish after exposed to different concentrations of carbendazim. Moreover, the expression of genes that are involved in the hypothalamic-pituitary-gonadal/thyroid (HPG/HPT) axis including VTG, ERα, ERβ2, Dio1, Dio2, Thraa and Thrb were all down-regulated significantly after the exposure to carbendazim. The expression levels of two cytochrome P450 aromatases CYP19a and CYP19b were increased significantly after 20 and 100 μg/L carbendazim exposure, respectively. Taken together, our results indicated that carbendazim had the potential to induce cell apoptosis and cause immune toxicity as well as endocrine disruption in zebrafish during the embryo developmental stage. The information presented here also help to elucidate the environmental risks caused by the carbendazim-induced toxicity in aquatic organisms. PMID:25304545

  16. Lipoprotein Lipase, Tissue Expression and Effects on Genes Related to Fatty Acid Synthesis in Goat Mammary Epithelial Cells

    PubMed Central

    Zhao, Wang-Sheng; Hu, Shi-Liang; Yu, Kang; Wang, Hui; Wang, Wei; Loor, Juan; Luo, Jun

    2014-01-01

    Lipoprotein lipase (LPL) serves as a central factor in hydrolysis of triacylglycerol and uptake of free fatty acids from the plasma. However, there are limited data concerning the action of LPL on the regulation of milk fat synthesis in goat mammary gland. In this investigation, we describe the cloning and sequencing of the LPL gene from Xinong Saanen dairy goat mammary gland, along with a study of its phylogenetic relationships. Sequence analysis showed that goat LPL shares similarities with other species including sheep, bovine, human and mouse. LPL mRNA expression in various tissues determined by RT-qPCR revealed the highest expression in white adipose tissue, with lower expression in heart, lung, spleen, rumen, small intestine, mammary gland, and kidney. Expression was almost undetectable in liver and muscle. The expression profiles of LPL gene in mammary gland at early, peak, mid, late lactation, and the dry period were also measured. Compared with the dry period, LPL mRNA expression was markedly greater at early lactation. However, compared with early lactation, the expression was lower at peak lactation and mid lactation. Despite those differences, LPL mRNA expression was still greater at peak, mid, and late lactation compared with the dry period. Using goat mammary epithelial cells (GMEC), the in vitro knockdown of LPL via shRNA or with Orlistat resulted in a similar degree of down-regulation of LPL (respectively). Furthermore, knockdown of LPL was associated with reduced mRNA expression of SREBF1, FASN, LIPE and PPARG but greater expression of FFAR3. There was no effect on ACACA expression. Orlistat decreased expression of LIPE, FASN, ACACA, and PPARG, and increased FFAR3 and SREBF1 expression. The pattern of LPL expression was similar to the changes in milk fat percentage in lactating goats. Taken together, results suggest that LPL may play a crucial role in fatty acid synthesis. PMID:25501331

  17. Significance of Polymorphisms and Expression of Enzyme-Encoding Genes Related to Glutathione in Hematopoietic Cancers and Solid Tumors

    PubMed Central

    Zmorzyński, Szymon; Świderska-Kołacz, Grażyna; Koczkodaj, Dorota; Filip, Agata Anna

    2015-01-01

    Antioxidant compounds such as glutathione and its enzymes have become the focus of attention of medical sciences. Glutathione, a specific tripeptide, is involved in many intercellular processes. The glutathione concentration is determined by the number of GAG repeats in gamma-glutamylcysteine synthetase. GAG polymorphisms are associated with an increased risk of schizophrenia, berylliosis, diabetes, lung cancer, and nasopharyngeal tumors. Cancer cells with high glutathione concentration are resistant to chemotherapy treatment. The oxidized form of glutathione is formed by glutathione peroxidases (GPXs). The changes in activity of GPX1, GPX2, and GPX3 isoforms may be associated with the development of cancers, for example, prostate cancer or even colon cancer. Detoxification of glutathione conjugates is possible due to activity of glutathione S-transferases (GSTs). Polymorphisms in GSTM1, GSTP1, and GSTO1 enzymes increase the risk of developing breast cancer and hepatocellular carcinoma. Gamma-glutamyl transpeptidases (GGTs) are responsible for glutathione degradation. Increased activity of GGT correlates with adverse prognosis in patients with breast cancer. Studies on genes encoding glutathione enzymes are continued in order to determine the correlation between DNA polymorphisms in cancer patients. PMID:26682223

  18. Identification of genes related to agarwood formation: transcriptome analysis of healthy and wounded tissues of Aquilaria sinensis

    PubMed Central

    2013-01-01

    Background Agarwood is an expensive resinous heartwood derived from Aquilaria plants that is widely used in traditional medicines, incense and perfume. Only wounded trees can produce agarwood, and the huge demand for the agarwood products has led all Aquilaria spp. being endangered and listed in the Appendix II of the CITES (http://www.cites.org). The major components of agarwood are sesquiterpenes and phenylethyl chromones. Owing to a lack of genomic information, the molecular basis of wound-induced sesquiterpenes biosynthesis and agarwood formation remains unknown. Results To identify the primary genes that maybe related to agarwood formation, we sequenced 2 cDNA libraries generated from healthy and wounded A. sinensis (Lour.) Gilg. A total of 89,137 unigenes with an average length of 678.65 bp were obtained, and they were annotated in detail at bioinformatics levels. Of those associated with agarwood formation, 30 putatively encoded enzymes in the sesquiterpene biosynthesis pathway, and a handful of transcription factors and protein kinases were related to wound signal transduction. Three full-length cDNAs of sesquiterpene synthases (ASS1-3) were cloned and expressed in Escherichia coli, and enzyme assays revealed that they are active enzymes, with the major products being δ-guaiene. A methyl jasmonate (MJ) induction experiment revealed that the expression of ASS was significantly induced by MJ, and the production of sesquiterpenes was elevated accordingly. The expression of some transcription factors and protein kinases, especially MYB4, WRKY4, MPKK2 and MAPK2, was also induced by MJ and coordinated with ASS expression, suggesting they maybe positive regulators of ASS. Conclusions This study provides extensive transcriptome information for Aquilaria spp. and valuable clues for elucidating the mechanism of wound-induced agarwood sesquiterpenes biosynthesis and their regulation. PMID:23565705

  19. Effect of the ratios of unsaturated fatty acids on the expressions of genes related to fat and protein in the bovine mammary epithelial cells.

    PubMed

    Sheng, R; Yan, S M; Qi, L Z; Zhao, Y L

    2015-04-01

    The objective of this study was to evaluate the effects of the different ratios of unsaturated fatty acids (UFAs) (oleic acid, linoleic acid, and linolenic acid) on the cell viability and triacylglycerol (TAG) content, as well as the mRNA expression of the genes related to lipid and protein synthesis in bovine mammary epithelial cells (BMECs). Primary cells were isolated from the mammary glands of Holstein dairy cows and were passaged twice. Afterward, the cells were randomly allocated to six treatments, five UFA-treated groups, and one control group. For all of the treatments, the the fetal bovine serum in the culture solution was replaced with fatty acid-free BSA (1 g/L), and the cells were treated with different ratios of oleic, linoleic, and linolenic acids (0.75:4:1, 1.5:10:1, 2:13.3:1, 3:20:1, and 4:26.7:1) for 48 h, which were group 1 to group 5. The control culture solution contained only fatty acid-free BSA without UFAs (0 μM). The results indicated that the cell viability was not affected by adding different ratios of UFAs, but the accumulation of TAG was significantly influenced by supplementing with different ratios of UFAs. Adding different ratios of UFAs suppressed the expression of ACACA and FASN but had the opposite effect on the abundances of FABP3 and CD36 mRNA. The expression levels of PPARG, SPEBF1, CSN1S1, and CSN3 mRNA in the BMECs were affected significantly after adding different ratios of UFAs. Our results suggested that groups 1, 2, and 3 (0.75:4:1, 1.5:10:1, and 2:13.3:1) had stronger auxo-action on fat synthesis in the BMECs, where group 3 (2:13.3:1) was the best, followed by group 4 (3:20:1). However, group 5 (4:26.7:1) was the worst. Genes related to protein synthesis in the BMECs were better promoted in groups 2 and 3, and group 3 had the strongest auxo-action, whereas the present study only partly examined the regulation of protein synthesis at the transcriptional level; more studies on translation level are needed in the future

  20. Effects of different dwarfing interstocks on key enzyme activities and the expression of genes related to malic acid metabolism in Red Fuji apples.

    PubMed

    Shi, J; Li, F F; Ma, H; Li, Z Y; Xu, J Z

    2015-01-01

    In this experiment, the test materials were 'Red Fuji' apple trees grafted onto three interstocks (No. 53, No. 111, and No. 236), which were chosen from SH40 seeding interstocks. The content of malic acid, the enzyme activities, and the expression of genes related to malic acid metabolism were determined during fruit development.The results showed that malic acid content in the ripe fruit on interstock No. 53 was higher than that in the interstock No. 111 fruit. The malate dehydrogenase (NAD-MDH) activity in apples on interstock No. 53 was highest on Day 30, Day 100, and Day 160 after bloom, and the malic enzyme (NADP-ME) activity in apples on interstock No. 111 was higher than in the interstock No. 53 fruit from Day 70 to Day 100 after bloom. The relative expression of NAD-MDH genes in interstock No. 53 fruit was higher than in No. 236 fruit on Day 100 after bloom, but the relative expression of NADP-ME in No. 236 interstock fruit was lower than in No. 53 fruit. The relative expression of NAD-MDH genes in No. 53 interstock fruit was highest on Day 160 after bloom. This might have been the main reason for the difference in the accumulation of malic acid in the ripe apples.There was a positive correlation between the relative expression of phosphoenolpyruvate carboxylase (PEPC) and the malic acid content of the fruit, and the content of malic acid in the apples was affected by the PEPC activity during the early developmental stage. PMID:26782412

  1. Increasing levels of dietary crystalline methionine affect plasma methionine profiles, ammonia excretion, and the expression of genes related to the hepatic intermediary metabolism in rainbow trout (Oncorhynchus mykiss).

    PubMed

    Rolland, Marine; Skov, Peter V; Larsen, Bodil K; Holm, Jørgen; Gómez-Requeni, Pedro; Dalsgaard, Johanne

    2016-08-01

    Strictly carnivorous fish with high requirements for dietary protein, such as rainbow trout (Oncorhynchus mykiss) are interesting models for studying the role of amino acids as key regulators of intermediary metabolism. Methionine is an essential amino acid for rainbow trout, and works as a signalling factor in different metabolic pathways. The study investigated the effect of increasing dietary methionine intake on the intermediary metabolism in the liver of juvenile rainbow trout. For this purpose, five diets were formulated with increasing methionine levels from 0.60 to 1.29% dry matter. The diets were fed in excess for six weeks before three sampling campaigns carried out successively to elucidate (i) the hepatic expression of selected genes involved in lipid, glucose and amino acid metabolism; (ii) the postprandial ammonia excretion; and (iii) the postprandial plasma methionine concentrations. The transcript levels of enzymes involved in lipid metabolism (fatty acid synthase, glucose 6 phosphate dehydrogenase and carnitine palmitoyl transferase 1 a), gluconeogenesis (fructose-1,6-biphosphatase) and amino acid catabolism (alanine amino transferase and glutamate dehydrogenase) were significantly affected by the increase in dietary methionine. Changes in gene expression reflected to some extent the decrease in ammonia excretion (P=0.022) and in the hepatosomatic index (HSI; P<0.001) when dietary methionine increased. Postprandial plasma methionine concentrations correlated positively with the dietary level (P<0.001) at the different sampling points. The study shows that the expression of several genes related to the hepatic intermediary metabolism in rainbow trout responded in a dose-dependent manner to increasing levels of dietary methionine. PMID:27105833

  2. B.E.A.R. GeneInfo: A tool for identifying gene-related biomedical publications through user modifiable queries

    PubMed Central

    Zhou, Guohui; Wen, Xinyu; Liu, Hang; Schlicht, Michael J; Hessner, Martin J; Tonellato, Peter J; Datta, Milton W

    2004-01-01

    Background Once specific genes are identified through high throughput genomics technologies there is a need to sort the final gene list to a manageable size for validation studies. The triaging and sorting of genes often relies on the use of supplemental information related to gene structure, metabolic pathways, and chromosomal location. Yet in disease states where the genes may not have identifiable structural elements, poorly defined metabolic pathways, or limited chromosomal data, flexible systems for obtaining additional data are necessary. In these situations having a tool for searching the biomedical literature using the list of identified genes while simultaneously defining additional search terms would be useful. Results We have built a tool, BEAR GeneInfo, that allows flexible searches based on the investigators knowledge of the biological process, thus allowing for data mining that is specific to the scientist's strengths and interests. This tool allows a user to upload a series of GenBank accession numbers, Unigene Ids, Locuslink Ids, or gene names. BEAR GeneInfo takes these IDs and identifies the associated gene names, and uses the lists of gene names to query PubMed. The investigator can add additional modifying search terms to the query. The subsequent output provides a list of publications, along with the associated reference hyperlinks, for reviewing the identified articles for relevance and interest. An example of the use of this tool in the study of human prostate cancer cells treated with Selenium is presented. Conclusions This tool can be used to further define a list of genes that have been identified through genomic or genetic studies. Through the use of targeted searches with additional search terms the investigator can limit the list to genes that match their specific research interests or needs. The tool is freely available on the web at [1], and the authors will provide scripts and database components if requested mdatta@mcw.edu PMID

  3. A low protein diet during pregnancy provokes a lasting shift of hepatic expression of genes related to cell cycle throughout ontogenesis in a porcine model

    PubMed Central

    2012-01-01

    Background In rodent models and in humans the impact of gestational diets on the offspring's phenotype was shown experimentally and epidemiologically. Adverse environmental conditions during fetal development provoke an intrauterine adaptive response termed 'fetal programming', which may lead to both persistently biased responsiveness to extrinsic factors and permanent consequences for the organismal phenotype. This leads to the hypothesis that the offspring's transcriptome exhibits short-term and long-term changes, depending on the maternal diet. In order to contribute to a comprehensive inventory of genes and functional networks that are targets of nutritional programming initiated during fetal life, we applied whole-genome microarrays for expression profiling in a longitudinal experimental design covering prenatal, perinatal, juvenile, and adult ontogenetic stages in a porcine model. Pregnant sows were fed either a gestational low protein diet (LP, 6% CP) or an adequate protein diet (AP, 12% CP). All offspring was nursed by foster sows receiving standard diets. After weaning, all offspring was fed standard diets ad libitum. Results Analyses of the hepatic gene expression of the offspring at prenatal (94 dies post conceptionem, dpc) and postnatal stages (1, 28, 188 dies post natum, dpn) included comparisons between dietary groups within stages as well as comparisons between ontogenetic stages within diets to separate diet-specific transcriptional changes and maturation processes. We observed differential expression of genes related to lipid metabolism (e.g. Fatty acid metabolism, Biosynthesis of steroids, Synthesis and degradation of ketone bodies, FA elongation in mitochondria, Bile acid synthesis) and cell cycle regulation (e.g. Mitotic roles of PLK, G1/S checkpoint regulation, G2/M DNA damage checkpoint regulation). Notably, at stage 1 dpn no regulation of a distinct pathway was found in LP offspring. Conclusions The transcriptomic modulations point to

  4. Isolation and characterization of a Saccharomyces cerevisiae peptide transport gene.

    PubMed Central

    Perry, J R; Basrai, M A; Steiner, H Y; Naider, F; Becker, J M

    1994-01-01

    We have cloned and characterized a Saccharomyces cerevisiae peptide transport gene (PTR2) isolated from a genomic DNA library by directly selecting for functional complementation of a peptide transport-deficient mutant. Deletion and frameshift mutageneses were used to localize the complementing activity to a 3.1-kbp region on the transforming plasmid. DNA sequencing of the complementing region identified an open reading frame spanning 1,803 bp. The deduced amino acid sequence predicts a hydrophobic peptide consisting of 601 amino acids, having a molecular mass of 68.1 kDa, composed in part of 12 hydrophobic segments, and sharing significant similarities with a nitrate transport protein encoded by the CHL1 gene of Arabidopsis thaliana. Northern (RNA) hybridization experiments demonstrated a single transcript that was 1.8 kb in length and that was transiently induced by the addition of L-leucine to the growth medium. The PTR2 gene was localized to the right arm of chromosome XI by contour-clamped homogeneous electric field gel chromosome blotting and by hybridization to known chromosome XI lambda phage clones of S. cerevisiae DNA. PTR2 was tightly linked to the UBI2 gene, with the coding sequences being separated by a 466-bp region and oriented so that the genes were transcribed convergently. A chromosomal disruption of the PTR2 gene in a haploid strain was not lethal under standard growth conditions. The cloning of PTR2 represents the first example of the molecular genetic characterization of a eucaryotic peptide transport gene. Images PMID:8264579

  5. Rat Hepatocytes Weighted Gene Co-Expression Network Analysis Identifies Specific Modules and Hub Genes Related to Liver Regeneration after Partial Hepatectomy

    PubMed Central

    Zhou, Yun; Xu, Jiucheng; Liu, Yunqing; Li, Juntao; Chang, Cuifang; Xu, Cunshuan

    2014-01-01

    The recovery of liver mass is mainly mediated by proliferation of hepatocytes after 2/3 partial hepatectomy (PH) in rats. Studying the gene expression profiles of hepatocytes after 2/3 PH will be helpful to investigate the molecular mechanisms of liver regeneration (LR). We report here the first application of weighted gene co-expression network analysis (WGCNA) to analyze the biological implications of gene expression changes associated with LR. WGCNA identifies 12 specific gene modules and some hub genes from hepatocytes genome-scale microarray data in rat LR. The results suggest that upregulated MCM5 may promote hepatocytes proliferation during LR; BCL3 may play an important role by activating or inhibiting NF-kB pathway; MAPK9 may play a permissible role in DNA replication by p38 MAPK inactivation in hepatocytes proliferation stage. Thus, WGCNA can provide novel insight into understanding the molecular mechanisms of LR. PMID:24743545

  6. Characterization and developmental expression of xSim, a Xenopus bHLH/PAS gene related to the Drosophila neurogenic master gene single-minded.

    PubMed

    Coumailleau, P; Penrad-Mobayed, M; Lecomte, C; Bollérot, K; Simon, F; Poellinger, L; Angelier, N

    2000-12-01

    We have isolated a novel gene from Xenopus, denominated xSim, which encodes a protein of 760 amino acids containing a basic helix-loop-helix (bHLH) motif contiguous to a PAS domain characteristic of an emerging family of transcriptional regulators so called bHLH/PAS. xSim shares a strong amino acid sequence identity with the Drosophila Single-minded (dSim) and with the murine Sim1 and Sim2 proteins. Phylogenetic analysis reveals that xSim gene is an ortholog gene of the mSim2 gene. Spatio-temporal analysis shows a maternal and a zygotic expression of xSim throughout early Xenopus development. In situ hybridization assays reveal that the transcripts are enriched in the animal hemisphere until blastula stage and extend to the marginal zone at early gastrula stage. As development proceeds, xSim is mainly restricted to the central nervous system. PMID:11091086

  7. Chokeberry attenuates the expression of genes related to de novo lipogenesis in the hepatocytes of mice with nonalcoholic fatty liver disease.

    PubMed

    Park, Hyunjin; Liu, Yanan; Kim, Hyun-Sook; Shin, Jung-Hee

    2016-01-01

    Nonalcoholic fatty liver disease (NAFLD), which is characterized by steatosis, is a major public health concern. Previous studies have shown that chokeberry has anti-inflammatory, antimutagenic, hepatoprotective, cardioprotective, and antidiabetic effects. In this study, we hypothesized that chokeberry powder can attenuate the expression of genes related to de novo lipogenesis and the triglyceride levels in the hepatocytes of mice with high-fat diet-induced NAFLD. After coadministering chokeberry powder for 8weeks (0.5% and 1% powder) with a high-fat diet, mice that consumed chokeberry powder diets, regardless of the dose, had significantly lower liver triglyceride levels than control mice that were fed a high-fat diet (P=.0145 and P<.0012, respectively). Compared with mice that were fed a high-fat diet, mice that were given 1% chokeberry powder exhibited significantly decreased mRNA expression of sterol regulatory element-binding protein (P=.009) and acetyl-CoA carboxylase (P=.0032) in the liver. Compared with mice in the control group, fatty acid synthase (FAS) expression significantly increased in the mice that were fed a high-fat diet, but both chokeberry powder-treated groups had significantly decreased FAS expression (P=.0157 and P<.0001, respectively). The size of the fat droplets was decreased in the livers of the chokeberry-supplemented groups. In summary, the administration of chokeberry powder may help attenuate high-fat diet-induced NAFLD by regulating the expression levels of sterol regulatory element-binding protein, acetyl-CoA carboxylase, and FAS and by decreasing the size of the fat droplets in the liver. PMID:26773781

  8. Expression of essential genes for biosynthesis of antimicrobial peptides of Bacillus is modulated by inactivated cells of target microorganisms.

    PubMed

    Leães, Fernanda Leal; Velho, Renata Voltolini; Caldas, Danielle Gregório Gomes; Ritter, Ana Carolina; Tsai, Siu Mui; Brandelli, Adriano

    2016-01-01

    Certain Bacillus strains are important producers of antimicrobial peptides with great potential for biological control. Antimicrobial peptide production by Bacillus amyloliquefaciens P11 was investigated in the presence of heat-inactivated cells of bacteria and fungi. B. amyloliquefaciens P11 exhibited higher antimicrobial activity in the presence of inactivated cells of Staphylococcus aureus and Aspergillus parasiticus compared to other conditions tested. Expression of essential genes related to biosynthesis of the antimicrobial peptides surfactin (sfp), iturin A (lpa-14 and ituD), subtilosin A (sboA) and fengycin (fenA) was investigated by quantitative real-time PCR (qRT-PCR). The genes lpa-14 and ituD were highly expressed in the presence of S. aureus (inactivated cells), indicating induction of iturin A production by B. amyloliquefaciens P11. The other inducing condition (inactivated cells of A. parasiticus) suppressed expression of lpa-14, but increased expression of ituD. A twofold increase in fenA expression was observed for both conditions, while strong suppression of sboA expression was observed in the presence of inactivated cells of S. aureus. An increase in antimicrobial activity was observed, indicating that synthesis of antimicrobial peptides may be induced by target microorganisms. PMID:26577655

  9. Gene expression for peptides in neurons of the petrosal and nodose ganglia in rat.

    PubMed

    Czyzyk-Krzeska, M F; Bayliss, D A; Seroogy, K B; Millhorn, D E

    1991-01-01

    In situ hybridization was used to determine whether genes for neuropeptides [substance P/neurokinin A (SP/NKA), calcitonin gene-related peptide (CGRP), somatostatin (SOM), neuropeptide tyrosine (NPY) and cholecystokinin (CCK)] are expressed in inferior ganglia of the vagus (nodose) and glossopharyngeal (petrosal) nerves. Synthetic oligodeoxyribonucleotides, complementary to the cognate, mRNAs were labeled with [32P] or [35S], and hybridized to 10 microns thick sections of unperfused tissue which were then processed for film and emulsion autoradiography. We found numerous, clustered neuronal perikarya throughout the nodose and petrosal ganglia that expressed preprotachykinin A (SP/NKA) and CGRP mRNAs to varying degrees. Neurons expressing preproSOM mRNA were less abundant and more scattered throughout both ganglia. Notably, we found mRNA for NPY in cells (usually 5-10 per section) in both ganglia. To our knowledge, this is first evidence for NPY in these sensory ganglia. In contrast to previous immunohistochemical findings, we found no evidence for expression of preproCCK in either the nodose or petrosal ganglia. The present findings demonstrate that cells of the nodose and petrosal ganglia express the genes for a number of neuropeptides that are presumably involved with transmission of visceral sensory afferent information to higher order neurons of the central nervous system. PMID:1708726

  10. Genome-wide analysis of CrRLK1L gene family in Gossypium and identification of candidate CrRLK1L genes related to fiber development.

    PubMed

    Niu, Erli; Cai, Caiping; Zheng, Yongjie; Shang, Xiaoguang; Fang, Lei; Guo, Wangzhen

    2016-06-01

    Members of the CrRLK1L family, a subgroup of the receptor-like kinase (RLK) gene family, are thought to act as sensors for the integrity of the cell wall and regulators of polar elongation. To better understand the various functions in fiber development, we conducted genome-wide identification and characterization analyses of CrRLK1L family in cotton. Here 44, 40, and 79 CrRLK1L genes were identified from three cotton species: diploid G. raimondii (D5), diploid G. arboreum (A2), and tetraploid G. hirsutum TM-1 (AD1), respectively. The 44 CrRLK1Ls in G. raimondii were anchored to the 12 chromosomes unevenly and were classified into six groups (I-VI), with group II and group IV being further divided into two subgroups (groups IIa and IIb, and IVa and IVb, respectively). These CrRLK1Ls displayed a highly regular pattern of developmental and spatial regulation in cotton. Using the transcriptome data of five chromosomal segment introgression lines (CSILs) and the physical integration of CrRLK1Ls with the quantitative trait loci (QTLs) related to fiber quality traits, we revealed that six CrRLK1L genes were highly associated with fiber development. This study brings new insights into the integrated genome-wide identification of CrRLK1Ls in cotton and provides references for the genetic improvement of cotton fiber. PMID:26833484

  11. Cloning of genes related to exo-beta-glucanase production in Saccharomyces cerevisiae: characterization of an exo-beta-glucanase structural gene.

    PubMed

    Nebreda, A R; Villa, T G; Villanueva, J R; del Rey, F

    1986-01-01

    The EXG1 gene of Saccharomyces cerevisiae was cloned and identified by complementation of a mutant strain (exg1-2) with highly reduced extracellular exo-beta-1,3-glucanase (EXG) activity. Two recombinant plasmids containing an overlapping region of 5.2 kb were isolated from a genomic DNA library and characterized by restriction mapping. The coding region was located by subcloning the original DNA inserts in a 2.7-kb HindIII-XhoI fragment. Exg+ strains and Exg- mutants transformed with yeast multicopy plasmids containing this DNA fragment showed an EXG activity 5- to 20-fold higher than for the untransformed Exg+ wild-type (wt) strains. The overproduced EXG had the same enzymic activity on different substrates, and showed the same electrophoretic behaviour on polyacrylamide gels and identical properties upon filtration through Sephacryl S-200 as those of the main EXG from Exg+ wt strains. The EXG1 gene transformed Schizosaccharomyces pombe, yielding extracellular EXG activity which showed cross-reactivity with anti-S. cervisiae EXG antibodies. A fragment including only a part of the EXG1 region was subcloned into the integrating vector YIp5, and the resulting plasmid was used to transform an Exg+ strain. Genetic and Southern analysis of several stable Exg- transformants showed that the fragment integrated by homology with the EXG1 locus. The chromosomal DNA fragment into which the plasmid integrated has a restriction pattern identical to that of the fragment on which we had previously identified the putative EXG1 gene. Only one copy of the EXG1 gene per genome was found in several strains tested by Southern analysis. Furthermore, two additional recombinant plasmids sharing a yeast DNA fragment of about 4.1 kb, which partially complements the exg1-2 mutation but which shows no homology with the 2.7-kb fragment containing the EXG1 gene, were also identified in this study. This 4.1-kb DNA fragment does not appear to contain an extragenic suppressor and could be related

  12. Novel endosomolytic peptides for enhancing gene delivery in nanoparticles.

    PubMed

    Ahmad, Aqeel; Ranjan, Sanjeev; Zhang, Weikai; Zou, Jing; Pyykkö, Ilmari; Kinnunen, Paavo K J

    2015-02-01

    Trapping in the endosomes is currently believed to represent the main barrier for transfection. Peptides, which allow endosomal escape have been demonstrated to overcome this barrier, similarly to the entry of viruses. However, the design principles of such endosomolytic peptides remain unclear. We characterized three analogs derived from membrane disrupting antimicrobial peptides (AMP), viz. LL-37, melittin, and bombolitin V, with glutamic acid substituting for all basic residues. These analogs are pH-sensitive and cause negligible membrane permeabilization and insignificant cytotoxicity at pH7.4. However, at pH5.0, prevailing in endosomes, membrane binding and hemolysis of human erythrocytes become evident. We first condensed the emerald green fluorescent protein (emGFP) containing plasmid by protamine, yielding 115 nm diameter soluble nanoplexes. For coating of the nanoplex surface with a lipid bilayer we introduced a hydrophobic tether, stearyl-octa-arginine (SR8). The indicated peptides were dissolved in methanol and combined with lipid mixtures in chloroform, followed by drying at RT under a nitrogen flow. The dry residues were hydrated with nanoplexes in Hepes, pH7.4 yielding after a 30 min incubation at RT,rather monodisperse nanoparticles having an average diameter of 150-300 nm, measured by DLS and cryo-TEM. Studies with cell cultures showed the above peptides to yield expression levels comparable to those obtained using Lipofectamine 2000. However, unlike the polydisperse aggregates formed upon mixing Lipofectamine 2000 and plasmid, the procedure described yields soluble, and reasonably monodisperse nanoparticles, which can be expected to be suitable for gene delivery in vivo, using intravenous injection. PMID:25445677

  13. Antimicrobial peptide genes in Bacillus strains from plant environments.

    PubMed

    Mora, Isabel; Cabrefiga, Jordi; Montesinos, Emilio

    2011-12-01

    The presence of the antimicrobial peptide (AMP) biosynthetic genes srfAA (surfactin), bacA (bacylisin), fenD (fengycin), bmyB (bacyllomicin), spaS (subtilin), and ituC (iturin) was examined in 184 isolates of Bacillus spp. obtained from plant environments (aerial, rhizosphere, soil) in the Mediterranean land area of Spain. Most strains had between two and four AMP genes whereas strains with five genes were seldom detected and none of the strains had six genes. The most frequent AMP gene markers were srfAA, bacA, bmyB, and fenD, and the most frequent genotypes srfAA-bacA-bmyB and srfAAbacA- bmyB-fenD. The dominance of these particular genes in Bacillus strains associated with plants reinforces the competitive role of surfactin, bacyllomicin, fengycin, and bacilysin in the fitness of strains in natural environments. The use of these AMP gene markers may assist in the selection of putative biological control agents of plant pathogens. PMID:22569759

  14. Identification of peptide-specific TCR genes by in vitro peptide stimulation and CDR3 length polymorphism analysis.

    PubMed

    Shao, Hongwei; Lin, Yanmei; Wang, Teng; Ou, Yusheng; Shen, Han; Tao, Changli; Wu, Fenglin; Zhang, Wenfeng; Bo, Huaben; Wang, Hui; Huang, Shulin

    2015-07-10

    Identification of TCR genes specific for tumor-associated antigens (TAAs) is necessary for TCR gene modification of T cells, which is applied in anti-tumor adoptive T cell therapy (ACT). The usual identification methods are based on isolating single peptide-responding T cells and cloning the TCR gene by in vitro expansion or by single-cell RT-PCR. However, the long and exacting in vitro culture period and demanding operational requirements restrict the application of these methods. Immunoscope is an effective tool that profiles a repertoire of TCRs and identifies significantly expanded clones through CDR3 length analysis. In this study, a survivin-derived mutant peptide optimized for HLA-A2 binding was selected to load DCs and activate T cells. The monoclonal expansion of TCRA and TCRB genes was separately identified by Immunoscope analysis and following sequence identification, the properly paired TCR genes were transferred into T cells. Peptide recognition and cytotoxicity assays indicated that TCR-modified PBMCs could respond to both the mutant and wild type peptides and lyse target cells. These results show that combining Immunoscope with in vitro peptide stimulation provides an alternative and superior method for identifying specific TCR genes, which represents a significant advance for the application of TCR gene-modified T cells. PMID:25890221

  15. A Cell-Penetrating Peptide with a Guanidinylethyl Amine Structure Directed to Gene Delivery

    PubMed Central

    Oba, Makoto; Kato, Takuma; Furukawa, Kaori; Tanaka, Masakazu

    2016-01-01

    A peptide composed of lysine with a guanidinylethyl (GEt) amine structure in the side chain [Lys(GEt)] was developed as a cell-penetrating peptide directed to plasmid DNA (pDNA) delivery. The GEt amine adopted a diprotonated form at neutral pH, which may have led to the more efficient cellular uptake of a Lys(GEt)-peptide than an arginine-peptide at a low concentration. Lys(GEt)-peptide/pDNA complexes showed the highest transfection efficiency due to efficient endosomal escape without any cytotoxicity. Lys(GEt)-peptide may be a promising candidate as a gene delivery carrier. PMID:26814673

  16. A Cell-Penetrating Peptide with a Guanidinylethyl Amine Structure Directed to Gene Delivery

    NASA Astrophysics Data System (ADS)

    Oba, Makoto; Kato, Takuma; Furukawa, Kaori; Tanaka, Masakazu

    2016-01-01

    A peptide composed of lysine with a guanidinylethyl (GEt) amine structure in the side chain [Lys(GEt)] was developed as a cell-penetrating peptide directed to plasmid DNA (pDNA) delivery. The GEt amine adopted a diprotonated form at neutral pH, which may have led to the more efficient cellular uptake of a Lys(GEt)-peptide than an arginine-peptide at a low concentration. Lys(GEt)-peptide/pDNA complexes showed the highest transfection efficiency due to efficient endosomal escape without any cytotoxicity. Lys(GEt)-peptide may be a promising candidate as a gene delivery carrier.

  17. Epigenetic regulation of the formyl peptide receptor 2 gene.

    PubMed

    Simiele, Felice; Recchiuti, Antonio; Patruno, Sara; Plebani, Roberto; Pierdomenico, Anna Maria; Codagnone, Marilina; Romano, Mario

    2016-10-01

    Lipoxin (LX) A4, a main stop signal of inflammation, exerts potent bioactions by activating a specific G protein-coupled receptor, termed formyl peptide receptor 2 and recently renamed ALX/FPR2. Knowledge of the regulatory mechanisms that drive ALX/FPR2 gene expression is key for the development of innovative anti-inflammatory pharmacology. Here, we examined chromatin patterns of the ALX/FPR2 gene. We report that in MDA-MB231 breast cancer cells, the ALX/FPR2 gene undergoes epigenetic silencing characterized by low acetylation at lysine 27 and trimethylation at lysine 4, associated with high methylation at lysine 27 of histone 3. This pattern, which is consistent with transcriptionally inaccessible chromatin leading to low ALX/FPR2 mRNA and protein expression, is reversed in polymorphonuclear leukocytes that express high ALX/FPR2 levels. Activation of p300 histone acetyltransferase and inhibition of DNA methyltransferase restored chromatin accessibility and significantly increased ALX/FPR2 mRNA transcription and protein levels in MDA-MB231 cells, as well as in pulmonary artery endothelial cells. In both cells types, changes in the histone acetylation/methylation status enhanced ALX/FPR2 signaling in response to LXA4. Collectively, these results uncover unappreciated epigenetic regulation of ALX/FPR2 expression that can be exploited for innovative approaches to inflammatory disorders. PMID:27424221

  18. Genome-wide identification of cassava R2R3 MYB family genes related to abscission zone separation after environmental-stress-induced abscission

    PubMed Central

    Liao, Wenbin; Yang, Yiling; Li, Yayun; Wang, Gan; Peng, Ming

    2016-01-01

    Cassava plants (Manihot esculenta Crantz) resist environmental stresses by shedding leaves in leaf pulvinus abscission zones (AZs), thus leading to adaptation to new environmental conditions. Little is known about the roles of cassava R2R3 MYB factors in regulating AZ separation. Herein, 166 cassava R2R3 MYB genes were identified. Evolutionary analysis indicated that the 166 R2R3 MYB genes could be divided into 11 subfamilies. Transcriptome analysis indicated that 26 R2R3 MYB genes were expressed in AZs across six time points during both ethylene- and water-deficit stress-induced leaf abscission. Comparative expression profile analysis of similar SOTA (Self Organizing Tree Algorithm) clusters demonstrated that 10 R2R3 MYB genes had similar expression patterns at six time points in response to both treatments. GO (Gene Ontology) annotation confirmed that all 10 R2R3 MYB genes participated in the responses to stress and ethylene and auxin stimuli. Analysis of the putative 10 R2R3 MYB promoter regions showed that those genes primarily contained ethylene- and stress-related cis-elements. The expression profiles of the genes acting downstream of the selected MYBs were confirmed to be involved in cassava abscission zone separation. All these results indicated that R2R3 MYB plays an important regulatory role in AZ separation. PMID:27573926

  19. Genome-wide identification of cassava R2R3 MYB family genes related to abscission zone separation after environmental-stress-induced abscission.

    PubMed

    Liao, Wenbin; Yang, Yiling; Li, Yayun; Wang, Gan; Peng, Ming

    2016-01-01

    Cassava plants (Manihot esculenta Crantz) resist environmental stresses by shedding leaves in leaf pulvinus abscission zones (AZs), thus leading to adaptation to new environmental conditions. Little is known about the roles of cassava R2R3 MYB factors in regulating AZ separation. Herein, 166 cassava R2R3 MYB genes were identified. Evolutionary analysis indicated that the 166 R2R3 MYB genes could be divided into 11 subfamilies. Transcriptome analysis indicated that 26 R2R3 MYB genes were expressed in AZs across six time points during both ethylene- and water-deficit stress-induced leaf abscission. Comparative expression profile analysis of similar SOTA (Self Organizing Tree Algorithm) clusters demonstrated that 10 R2R3 MYB genes had similar expression patterns at six time points in response to both treatments. GO (Gene Ontology) annotation confirmed that all 10 R2R3 MYB genes participated in the responses to stress and ethylene and auxin stimuli. Analysis of the putative 10 R2R3 MYB promoter regions showed that those genes primarily contained ethylene- and stress-related cis-elements. The expression profiles of the genes acting downstream of the selected MYBs were confirmed to be involved in cassava abscission zone separation. All these results indicated that R2R3 MYB plays an important regulatory role in AZ separation. PMID:27573926

  20. Polymorphisms of genes related to the hypothalamic-pituitary-adrenal axis influence the cortisol awakening response as well as self-perceived stress.

    PubMed

    Li-Tempel, Ting; Larra, Mauro F; Winnikes, Ulrike; Tempel, Tobias; DeRijk, Roel H; Schulz, André; Schächinger, Hartmut; Meyer, Jobst; Schote, Andrea B

    2016-09-01

    The hypothalamus-pituitary-adrenal (HPA) axis is a crucial endocrine system for coping with stress. A reliable and stable marker for the basal state of that system is the cortisol awakening response (CAR). We examined the influence of variants of four relevant candidate genes; the mineralocorticoid receptor gene (MR), the glucocorticoid receptor gene (GR), the serotonin transporter gene (5-HTT) and the gene encoding the brain-derived neurotrophic factor (BDNF) on CAR and self-perceived stress in 217 healthy subjects. We found that polymorphisms of GR influenced both, the basal state of the HPA axis as well as self-perceived stress. MR only associated with self-perceived stress and 5-HTT only with CAR. BDNF did not affected any of the investigated indices. In summary, we suggest that GR variants together with the CAR and supplemented with self reports on perceived stress might be useful indicators for the basal HPA axis activity. PMID:27427534

  1. Transcription factors and anthocyanin genes related to low-temperature tolerance in rd29A:RdreB1BI transgenic strawberry.

    PubMed

    Gu, Xianbin; Chen, Yahua; Gao, Zhihong; Qiao, Yushan; Wang, Xiuyun

    2015-04-01

    Dehydration-responsive element-binding (DREB) transcription factors play critical roles in plant stress responses and signal transduction. To further understand how DREB regulates genes expression to promote cold-hardiness, Illumina/Solexa sequencing technology was used to compare the transcriptomes of non-transgenic and rd29A:RdreB1BI transgenic strawberry plants exposed to low temperatures. Approximately 3.5 million sequence tags were obtained from non-transgenic (NT) and transgenic (T) strawberry untreated (C) or low-temperature treated (LT) leaf samples. Over 1000 genes were differentially expressed between the NT-C and T-C plants, and also the NT-C and NT-LT, as well as the T-C and T-LT plants. Analysis of the genes up-regulated following low-temperature treatment revealed that the majority are linked to metabolism, biosynthesis, transcription and signal transduction. Uniquely up-regulated transcription factors as well as anthocyanin biosynthetic pathway genes are discussed. Accumulation of anthocyanin in the stolon and the base of the petiole differed between non-treated NT and T plants, and this correlated with gene expression patterns. The differentially expressed genes that encode transcription factors and anthocyanin enzymes may contribute to the cold hardiness of RdreB1BI transgenic strawberry. The transcriptome data provide a valuable resource for further studies of strawberry growth and development and DREB-mediated gene regulation under low-temperature stress. PMID:25686702

  2. Cloning and characterization of the CarbcL gene related to chlorophyll in pepper (Capsicum annuum L.) under fruit shade stress.

    PubMed

    Wang, Shu-Bin; Tian, Shi-Lin; Shah, Syed N M; Pan, Bao-Gui; Diao, Wei-Ping; Gong, Zhen-Hui

    2015-01-01

    Light is an important environmental factor for fruit development and ripening in pepper plant. Fruit bagging is a significant agrotechnology practiced for the illumination regulation of fruits; some previous researches have shown that fruit bagging could improve the appearance and external quality of fruits and cause them to mature early. However, it would decrease the intrinsic qualities of fruits; especially, fruit bagging could decrease the content of capsanthin in peppers. On the basis of these details, fruit bagging was used as the method of fruit shade stress in this study to explore the characteristics and molecular mechanisms of pepper fruit's color change under shade stress. By using cDNA-AFLP under fruit shading, a fragment related to fruit color was obtained. Next, the full-length coding sequence of the gene was cloned from the pepper fruits. Homologous gene alignment confirmed that the gene has high homology with the rbcL gene, named CarbcL. The function of the CarbcL gene was identified through virus-induced gene silencing (VIGS); it was found that the fruit color changed completely from green to red except for some residue of green fleck when CarbcL gene was silenced, and the green color of fruits had not fully faded in the control group and the empty vector group. The combine determination of chlorophyll content showed that CarbcL was involved in the metabolic control of chlorophyll in pepper fruits; subsequently, HPLC was used to determine the content of capsanthin in pepper fruit which the CarbcL gene was silencing, and it was also found that the content of capsanthin decreased appreciably. These results further confirmed that CarbcL gene was involved in the adjustment of chlorophyll and capsanthin. PMID:26528313

  3. Cloning and characterization of the CarbcL gene related to chlorophyll in pepper (Capsicum annuum L.) under fruit shade stress

    PubMed Central

    Wang, Shu-Bin; Tian, Shi-Lin; Shah, Syed N. M.; Pan, Bao-Gui; Diao, Wei-Ping; Gong, Zhen-Hui

    2015-01-01

    Light is an important environmental factor for fruit development and ripening in pepper plant. Fruit bagging is a significant agrotechnology practiced for the illumination regulation of fruits; some previous researches have shown that fruit bagging could improve the appearance and external quality of fruits and cause them to mature early. However, it would decrease the intrinsic qualities of fruits; especially, fruit bagging could decrease the content of capsanthin in peppers. On the basis of these details, fruit bagging was used as the method of fruit shade stress in this study to explore the characteristics and molecular mechanisms of pepper fruit's color change under shade stress. By using cDNA-AFLP under fruit shading, a fragment related to fruit color was obtained. Next, the full-length coding sequence of the gene was cloned from the pepper fruits. Homologous gene alignment confirmed that the gene has high homology with the rbcL gene, named CarbcL. The function of the CarbcL gene was identified through virus-induced gene silencing (VIGS); it was found that the fruit color changed completely from green to red except for some residue of green fleck when CarbcL gene was silenced, and the green color of fruits had not fully faded in the control group and the empty vector group. The combine determination of chlorophyll content showed that CarbcL was involved in the metabolic control of chlorophyll in pepper fruits; subsequently, HPLC was used to determine the content of capsanthin in pepper fruit which the CarbcL gene was silencing, and it was also found that the content of capsanthin decreased appreciably. These results further confirmed that CarbcL gene was involved in the adjustment of chlorophyll and capsanthin. PMID:26528313

  4. Determination of the key innate genes related to individual variation in carbon tetrachloride-induced hepatotoxicity using a pre-biopsy procedure

    SciTech Connect

    Yun, Jun-Won; Kim, Chae-Wook; Bae, Il-Hong; Park, Young-Ho; Chung, Jin-Ho; Lim, Kyung-Min Kang, Kyung-Sun

    2009-08-15

    High inter-individual variation in chemical-induced liver injury is a frequent observation with many hepatotoxic chemicals, yet the mechanism underlying it remains poorly understood. Even with carbon tetrachloride (CCl{sub 4}), a well-known model hepatotoxicant, substantial individual variations are observed in the severity of liver injury. Using microarray, many attempts have been made to identify the key genes in CCl{sub 4}-induced liver injury but mostly, they examined the gene expression of liver after CCl{sub 4} exposure, unable to dissect out the complicating factors from pathological changes secondary to liver injury. To more accurately identify the genes for the individual variation in CCl{sub 4}-induced hepatotoxicity, we compared the innate gene expression of the individual liver samples pre-biopsied prior to CCl{sub 4}-treatment with the severity of liver injury after CCl{sub 4}-treatment. Effect of biopsy procedure and 3 week recovery period on liver function and gene expression were confirmed to be insignificant. Using this design, we found that the expression of genes associated with immunity and defense, lipid metabolism, transport and complement-mediated immunity, which are previously known to be suppressed by CCl{sub 4}-treatment, were innately lower in the susceptible animals than resistant animals. Moreover, we demonstrated that the genes such as Gsta2, Sult2a1, Fgl1 and C6 were newly found to be innately lower in the susceptible animals to CCl{sub 4}-hepatotoxicity. These naturally lower gene expression patterns were further confirmed by RT-PCR. We believe that this pre-biopsy design may provide a useful tool for understanding the cause of variability of hepatotoxicity and for the prediction and pre-screening of the susceptible individual to drug-induced hepatotoxicity.

  5. Cancer-associated fibroblasts up-regulate CCL2, CCL26, IL6 and LOXL2 genes related to promotion of cancer progression in hepatocellular carcinoma cells.

    PubMed

    Lin, Zu-Yau; Chuang, Yen-Hwang; Chuang, Wan-Long

    2012-10-01

    Impact of different cancer-associated fibroblast (CAF) cell lines on proliferation, migration, invasion and differential expressions of genes in different hepatocellular carcinoma (HCC) cell lines was investigated. Two human CAF cell lines (F26/KMUH, F28/KMUH) and two human HCC cell lines (HCC24/KMUH, HCC38/KMUH) were studied. Influence of F28/KMUH cells on expressions of genes in HCC38/KMUH cells was detected by microarray to select genes for further analysis. Both CAF cell lines promoted proliferation (all P<0.05), migration (all P<0.05) and Matrigel invasion (all P<0.0001) of both HCC cell lines. F26/KMUH cells showed stronger promoted effects on, firstly, proliferation of HCC24/KMUH cells (P=0.0064) and, secondly, migration of both HCC cell lines than F28/KMUH cells did (all P<0.002). Ten up-regulated genes (APLN, CCL2, CCL26, CXCR4, IL6, MUC1, LOXL2, PDGFA, PGK1, VEGFA) related to proliferation, migration, invasion and angiogenesis of HCC detected by microarray were selected for quantitative reverse transcriptase-polymerase chain reaction analysis. Both CAF cell lines had same tendency of effects on differential expressions of genes in same HCC cell line, but expressions of genes between different HCC cell lines were not consistent. Only CCL2, CCL26, IL6 and LOXL2 genes were consistently up-regulated in both HCC cell lines. In conclusion, the effects of CAFs to promote proliferation, migration and invasion of HCC cells are influenced by the characteristics of both CAFs and HCC cells. Up-regulations of CCL2, CCL26, IL6 and LOXL2 genes in cancer cells are part of the common effects of CAFs on HCC cells. PMID:22739041

  6. Effects of Sodium Butyrate Treatment on Histone Modifications and the Expression of Genes Related to Epigenetic Regulatory Mechanisms and Immune Response in European Sea Bass (Dicentrarchus Labrax) Fed a Plant-Based Diet

    PubMed Central

    Díaz, Noelia; Rimoldi, Simona; Ceccotti, Chiara; Gliozheni, Emi; Piferrer, Francesc

    2016-01-01

    Bacteria that inhabit the epithelium of the animals’ digestive tract provide the essential biochemical pathways for fermenting otherwise indigestible dietary fibers, leading to the production of short-chain fatty acids (SCFAs). Of the major SCFAs, butyrate has received particular attention due to its numerous positive effects on the health of the intestinal tract and peripheral tissues. The mechanisms of action of this four-carbon chain organic acid are different; many of these are related to its potent regulatory effect on gene expression since butyrate is a histone deacetylase inhibitor that play a predominant role in the epigenetic regulation of gene expression and cell function. In the present work, we investigated in the European sea bass (Dicentrarchus labrax) the effects of butyrate used as a feed additive on fish epigenetics as well as its regulatory role in mucosal protection and immune homeostasis through impact on gene expression. Seven target genes related to inflammatory response and reinforcement of the epithelial defense barrier [tnfα (tumor necrosis factor alpha) il1β, (interleukin 1beta), il-6, il-8, il-10, and muc2 (mucin 2)] and five target genes related to epigenetic modifications [dicer1(double-stranded RNA-specific endoribonuclease), ehmt2 (euchromatic histone-lysine-N-methyltransferase 2), pcgf2 (polycomb group ring finger 2), hdac11 (histone deacetylase-11), and jarid2a (jumonji)] were analyzed in fish intestine and liver. We also investigated the effect of dietary butyrate supplementation on histone acetylation, by performing an immunoblotting analysis on liver core histone extracts. Results of the eight-week-long feeding trial showed no significant differences in weight gain or SGR (specific growth rate) of sea bass that received 0.2% sodium butyrate supplementation in the diet in comparison to control fish that received a diet without Na-butyrate. Dietary butyrate led to a twofold increase in the acetylation level of histone H4 at

  7. Effects of Sodium Butyrate Treatment on Histone Modifications and the Expression of Genes Related to Epigenetic Regulatory Mechanisms and Immune Response in European Sea Bass (Dicentrarchus Labrax) Fed a Plant-Based Diet.

    PubMed

    Terova, Genciana; Díaz, Noelia; Rimoldi, Simona; Ceccotti, Chiara; Gliozheni, Emi; Piferrer, Francesc

    2016-01-01

    Bacteria that inhabit the epithelium of the animals' digestive tract provide the essential biochemical pathways for fermenting otherwise indigestible dietary fibers, leading to the production of short-chain fatty acids (SCFAs). Of the major SCFAs, butyrate has received particular attention due to its numerous positive effects on the health of the intestinal tract and peripheral tissues. The mechanisms of action of this four-carbon chain organic acid are different; many of these are related to its potent regulatory effect on gene expression since butyrate is a histone deacetylase inhibitor that play a predominant role in the epigenetic regulation of gene expression and cell function. In the present work, we investigated in the European sea bass (Dicentrarchus labrax) the effects of butyrate used as a feed additive on fish epigenetics as well as its regulatory role in mucosal protection and immune homeostasis through impact on gene expression. Seven target genes related to inflammatory response and reinforcement of the epithelial defense barrier [tnfα (tumor necrosis factor alpha) il1β, (interleukin 1beta), il-6, il-8, il-10, and muc2 (mucin 2)] and five target genes related to epigenetic modifications [dicer1(double-stranded RNA-specific endoribonuclease), ehmt2 (euchromatic histone-lysine-N-methyltransferase 2), pcgf2 (polycomb group ring finger 2), hdac11 (histone deacetylase-11), and jarid2a (jumonji)] were analyzed in fish intestine and liver. We also investigated the effect of dietary butyrate supplementation on histone acetylation, by performing an immunoblotting analysis on liver core histone extracts. Results of the eight-week-long feeding trial showed no significant differences in weight gain or SGR (specific growth rate) of sea bass that received 0.2% sodium butyrate supplementation in the diet in comparison to control fish that received a diet without Na-butyrate. Dietary butyrate led to a twofold increase in the acetylation level of histone H4 at

  8. Evolution of paired domains: Isolation and sequencing of jellyfish and hydra Pax genes related to Pax-5 and Pax-6

    PubMed Central

    Sun, Hongmin; Rodin, Andrei; Zhou, Yihong; Dickinson, Douglas P.; Harper, Donald E.; Hewett-Emmett, David; Li, Wen-Hsiung

    1997-01-01

    Pax proteins are a family of transcription factors with a highly conserved paired domain; many members also contain a paired-type homeodomain and/or an octapeptide. Nine mammalian Pax genes are known and classified into four subgroups: Pax-1/9, Pax-2/5/8, Pax-3/7, and Pax-4/6. Most of these genes are involved in nervous system development. In particular, Pax-6 is a key regulator that controls eye development in vertebrates and Drosophila. Although the Pax-4/6 subgroup seems to be more closely related to Pax-2/5/8 than to Pax-3/7 or Pax-1/9, its evolutionary origin is unknown. We therefore searched for a Pax-6 homolog and related genes in Cnidaria, which is the lowest phylum of animals that possess a nervous system and eyes. A sea nettle (a jellyfish) genomic library was constructed and two pax genes (Pax-A and -B) were isolated and partially sequenced. Surprisingly, unlike most known Pax genes, the paired box in these two genes contains no intron. In addition, the complete cDNA sequences of hydra Pax-A and -B were obtained. Hydra Pax-B contains both the homeodomain and the octapeptide, whereas hydra Pax-A contains neither. DNA binding assays showed that sea nettle Pax-A and -B and hydra Pax-A paired domains bound to a Pax-5/6 site and a Pax-5 site, although hydra Pax-B paired domain bound neither. An alignment of all available paired domain sequences revealed two highly conserved regions, which cover the DNA binding contact positions. Phylogenetic analysis showed that Pax-A and especially Pax-B were more closely related to Pax-2/5/8 and Pax-4/6 than to Pax-1/9 or Pax-3/7 and that the Pax genes can be classified into two supergroups: Pax-A/Pax-B/Pax-2/5/8/4/6 and Pax-1/9/3/7. From this analysis and the gene structure, we propose that modern Pax-4/6 and Pax-2/5/8 genes evolved from an ancestral gene similar to cnidarian Pax-B, having both the homeodomain and the octapeptide. PMID:9144207

  9. De Novo Characterization of Fall Dormant and Nondormant Alfalfa (Medicago sativa L.) Leaf Transcriptome and Identification of Candidate Genes Related to Fall Dormancy

    PubMed Central

    Cheng, Ningning; Du, Hongqi; Fan, Wenna; Wang, Chengzhang

    2015-01-01

    Alfalfa (Medicago sativa L.) is one of the most widely cultivated perennial forage legumes worldwide. Fall dormancy is an adaptive character related to the biomass production and winter survival in alfalfa. The physiological, biochemical and molecular mechanisms causing fall dormancy and the related genes have not been well studied. In this study, we sequenced two standard varieties of alfalfa (dormant and non-dormant) at two time points and generated approximately 160 million high quality paired-end sequence reads using sequencing by synthesis (SBS) technology. The de novo transcriptome assembly generated a set of 192,875 transcripts with an average length of 856 bp representing about 165.1 Mb of the alfalfa leaf transcriptome. After assembly, 111,062 (57.6%) transcripts were annotated against the NCBI non-redundant database. A total of 30,165 (15.6%) transcripts were mapped to 323 Kyoto Encyclopedia of Genes and Genomes pathways. We also identified 41,973 simple sequence repeats, which can be used to generate markers for alfalfa, and 1,541 transcription factors were identified across 1,350 transcripts. Gene expression between dormant and non-dormant alfalfa at different time points were performed, and we identified several differentially expressed genes potentially related to fall dormancy. The Gene Ontology and pathways information were also identified. We sequenced and assembled the leaf transcriptome of alfalfa related to fall dormancy, and also identified some genes of interest involved in the fall dormancy mechanism. Thus, our research focused on studying fall dormancy in alfalfa through transcriptome sequencing. The sequencing and gene expression data generated in this study may be used further to elucidate the complete mechanisms governing fall dormancy in alfalfa. PMID:25799491

  10. Transcriptomic Analysis Identifies Candidate Genes Related to Intramuscular Fat Deposition and Fatty Acid Composition in the Breast Muscle of Squabs (Columba).

    PubMed

    Ye, Manhong; Zhou, Bin; Wei, Shanshan; Ding, MengMeng; Lu, Xinghui; Shi, Xuehao; Ding, Jiatong; Yang, Shengmei; Wei, Wanhong

    2016-01-01

    Despite the fact that squab is consumed throughout the world because of its high nutritional value and appreciated sensory attributes, aspects related to its characterization, and in particular genetic issues, have rarely been studied. In this study, meat traits in terms of pH, water-holding capacity, intramuscular fat content, and fatty acid profile of the breast muscle of squabs from two meat pigeon breeds were determined. Breed-specific differences were detected in fat-related traits of intramuscular fat content and fatty acid composition. RNA-Sequencing was applied to compare the transcriptomes of muscle and liver tissues between squabs of two breeds to identify candidate genes associated with the differences in the capacity of fat deposition. A total of 27 differentially expressed genes assigned to pathways of lipid metabolism were identified, of which, six genes belonged to the peroxisome proliferator-activated receptor signaling pathway along with four other genes. Our results confirmed in part previous reports in livestock and provided also a number of genes which had not been related to fat deposition so far. These genes can serve as a basis for further investigations to screen markers closely associated with intramuscular fat content and fatty acid composition in squabs. The data from this study were deposited in the National Center for Biotechnology Information (NCBI)'s Sequence Read Archive under the accession numbers SRX1680021 and SRX1680022. This is the first transcriptome analysis of the muscle and liver tissue in Columba using next generation sequencing technology. Data provided here are of potential value to dissect functional genes influencing fat deposition in squabs. PMID:27175015

  11. Identification of Differentially Expressed Genes Related to Dehydration Resistance in a Highly Drought-Tolerant Pear, Pyrus betulaefolia, as through RNA-Seq.

    PubMed

    Li, Kong-Qing; Xu, Xiao-Yong; Huang, Xiao-San

    2016-01-01

    Drought is a major abiotic stress that affects plant growth, development and productivity. Pear is one of the most important deciduous fruit trees in the world, but the mechanisms of drought tolerance in this plant are still unclear. To better understand the molecular basis regarding drought stress response, RNA-seq was performed on samples collected before and after dehydration in Pyrus betulaefolia. In total, 19,532 differentially expressed genes (DEGs) were identified. These genes were annotated into 144 Gene Ontology (GO) terms and 18 clusters of orthologous groups (COG) involved in 129 Kyoto Encyclopedia of Genes and Genomes (KEGG) defined pathways. These DEGs comprised 49 (26 up-regulated, 23 down-regulated), 248 (166 up-regulated, 82 down-regulated), 3483 (1295 up-regulated, 2188 down-regulated), 1455 (1065 up-regulated, 390 down-regulated) genes from the 1 h, 3 h and 6 h dehydration-treated samples and a 24 h recovery samples, respectively. RNA-seq was validated by analyzing the expresson patterns of randomly selected 16 DEGs by quantitative real-time PCR. Photosynthesis, signal transduction, innate immune response, protein phosphorylation, response to water, response to biotic stimulus, and plant hormone signal transduction were the most significantly enriched GO categories amongst the DEGs. A total of 637 transcription factors were shown to be dehydration responsive. In addition, a number of genes involved in the metabolism and signaling of hormones were significantly affected by the dehydration stress. This dataset provides valuable information regarding the Pyrus betulaefolia transcriptome changes in response to dehydration and may promote identification and functional analysis of potential genes that could be used for improving drought tolerance via genetic engineering of non-model, but economically-important, perennial species. PMID:26900681

  12. Transcriptomic Analysis Identifies Candidate Genes Related to Intramuscular Fat Deposition and Fatty Acid Composition in the Breast Muscle of Squabs (Columba)

    PubMed Central

    Ye, Manhong; Zhou, Bin; Wei, Shanshan; Ding, MengMeng; Lu, Xinghui; Shi, Xuehao; Ding, Jiatong; Yang, Shengmei; Wei, Wanhong

    2016-01-01

    Despite the fact that squab is consumed throughout the world because of its high nutritional value and appreciated sensory attributes, aspects related to its characterization, and in particular genetic issues, have rarely been studied. In this study, meat traits in terms of pH, water-holding capacity, intramuscular fat content, and fatty acid profile of the breast muscle of squabs from two meat pigeon breeds were determined. Breed-specific differences were detected in fat-related traits of intramuscular fat content and fatty acid composition. RNA-Sequencing was applied to compare the transcriptomes of muscle and liver tissues between squabs of two breeds to identify candidate genes associated with the differences in the capacity of fat deposition. A total of 27 differentially expressed genes assigned to pathways of lipid metabolism were identified, of which, six genes belonged to the peroxisome proliferator-activated receptor signaling pathway along with four other genes. Our results confirmed in part previous reports in livestock and provided also a number of genes which had not been related to fat deposition so far. These genes can serve as a basis for further investigations to screen markers closely associated with intramuscular fat content and fatty acid composition in squabs. The data from this study were deposited in the National Center for Biotechnology Information (NCBI)’s Sequence Read Archive under the accession numbers SRX1680021 and SRX1680022. This is the first transcriptome analysis of the muscle and liver tissue in Columba using next generation sequencing technology. Data provided here are of potential value to dissect functional genes influencing fat deposition in squabs. PMID:27175015

  13. Identification of Differentially Expressed Genes Related to Dehydration Resistance in a Highly Drought-Tolerant Pear, Pyrus betulaefolia, as through RNA-Seq

    PubMed Central

    Li, Kong-Qing; Xu, Xiao-Yong; Huang, Xiao-San

    2016-01-01

    Drought is a major abiotic stress that affects plant growth, development and productivity. Pear is one of the most important deciduous fruit trees in the world, but the mechanisms of drought tolerance in this plant are still unclear. To better understand the molecular basis regarding drought stress response, RNA-seq was performed on samples collected before and after dehydration in Pyrus betulaefolia. In total, 19,532 differentially expressed genes (DEGs) were identified. These genes were annotated into 144 Gene Ontology (GO) terms and 18 clusters of orthologous groups (COG) involved in 129 Kyoto Encyclopedia of Genes and Genomes (KEGG) defined pathways. These DEGs comprised 49 (26 up-regulated, 23 down-regulated), 248 (166 up-regulated, 82 down-regulated), 3483 (1295 up-regulated, 2188 down-regulated), 1455 (1065 up-regulated, 390 down-regulated) genes from the 1 h, 3 h and 6 h dehydration-treated samples and a 24 h recovery samples, respectively. RNA-seq was validated by analyzing the expresson patterns of randomly selected 16 DEGs by quantitative real-time PCR. Photosynthesis, signal transduction, innate immune response, protein phosphorylation, response to water, response to biotic stimulus, and plant hormone signal transduction were the most significantly enriched GO categories amongst the DEGs. A total of 637 transcription factors were shown to be dehydration responsive. In addition, a number of genes involved in the metabolism and signaling of hormones were significantly affected by the dehydration stress. This dataset provides valuable information regarding the Pyrus betulaefolia transcriptome changes in response to dehydration and may promote identification and functional analysis of potential genes that could be used for improving drought tolerance via genetic engineering of non-model, but economically-important, perennial species. PMID:26900681

  14. Expression profiling of sodium butyrate (NaB)-treated cells: identification of regulation of genes related to cytokine signaling and cancer metastasis by NaB.

    PubMed

    Joseph, Jeena; Mudduluru, Giridhar; Antony, Sini; Vashistha, Surabhi; Ajitkumar, Parthasarathi; Somasundaram, Kumaravel

    2004-08-19

    Histone deacetylase (HDAC) inhibitors induce growth arrest and apoptosis in a variety of human cancer cells. Sodium butyrate (NaB), a short chain fatty acid, is a HDAC inhibitor and is produced in the colonic lumen as a consequence of microbial degradation of dietary fibers. In order to dissect out the mechanism of NaB-induced growth inhibition of cancer cells, we carried out expression profiling of a human lung carcinoma cell line (H460) treated with NaB using a cDNA microarray. Of the total 1728 genes analysed, there were 32 genes with a mean expression value of 2.0-fold and higher and 66 genes with a mean expression value 3.0-fold and lower in NaB-treated cells. For a few selected genes, we demonstrate that their expression pattern by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis is matching with the results obtained by microarray analysis. Closer view at the expression profile of NaB-treated cells revealed the downregulation of a total of 16 genes associated with cytokine signaling, in particular, interferon gamma (IFNgamma) pathway. In good correlation, NaB-pretreated cells failed to induce interferon regulatory factor 1, an INFgamma target gene, efficiently upon IFNgamma addition. These results suggest that NaB inhibits proinflammatory cytokine signaling pathway, thus providing proof of mechanism for its anti-inflammatory activity. We also found that NaB induced three genes, which are known metastatic suppressors, and downregulated 11 genes, which have been shown to promote metastasis. Upregulation of metastatic suppressor Kangai 1 (KAI1) by NaB in a time-dependent manner was confirmed by RT-PCR analysis. The differential regulation of metastasis-associated genes by NaB provides explanation for the anti-invasive properties of NaB. Therefore, our study presents new evidence for pathways regulated by NaB, thus providing evidence for the mechanism behind anti-inflammatory and antimetastatic activities of NaB. PMID:15318170

  15. Screening of genes related to sulfide metabolism in Urechis unicinctus (Echiura, Urechidae) using suppression subtractive hybridization and cDNA microarray analysis.

    PubMed

    Shi, Xiaoli; Shao, Mingyu; Zhang, Litao; Ma, Yubin; Zhang, Zhifeng

    2012-09-01

    Exogenous sulfide can generally induce metabolic injuries in most organisms and even cause death. However, organisms inhabiting intertidal zones, hydrothermal vents, and cold seeps, can tolerate, metabolize, and utilize sulfide. In this study, both suppression subtractive hybridization and cDNA microarray analysis were employed to screen sulfide metabolism-related genes from the body wall in echiuran worm Urechis unicinctus, a marine sediment species. A total of 3456 monoclones were isolated and 82 were identified as differentially expressed genes in worms exposed to 50 μM sulfide for 24 h, compared to controls. The identified genes encoded proteins with multiple processes, including metabolism, cellular process, biological regulation, response to stimulus, multicellular organismal process, localization, development, and cellular component organization. Eight genes, serase, vacuolar protein, src tyrosine kinase, sulfide oxidase-like oxidoreductase, aprataxin, SN-RNP, aminopeptidase, and predicted protein, were selected to verify expression in the worm using qRT-PCR. The agreement of gene expression evaluation was 62.5% between the results of microarray analysis and qRT-PCR. These new data will provide clues for further probing of the molecular mechanism of sulfide metabolism. PMID:22591583

  16. Knockout of the adp gene related with colonization in Bacillus nematocida B16 using customized transcription activator-like effectors nucleases.

    PubMed

    Niu, Qiuhong; Zheng, Haoying; Zhang, Lin; Qin, Fujun; Facemire, Loryn; Zhang, Guo; Cao, Feng; Zhang, Ke-Qin; Huang, Xiaowei; Yang, Jianwei; He, Lei; Liu, Chanjuan

    2015-07-01

    Bacillus nematocida B16 is able to dominate in the intestines of the worm Caenorhabditis elegans in 'Trojan horse' pathogenic mechanism. The adp is one candidate gene which potentially play a vital role in the colonization from our previous random mutagenesis screening results. To analyse the functional role of this gene, we constructed the adp knockout mutant through customized transcription activator-like effectors nucleases (TALEN), which has been successfully used in yeasts, nematodes, zebrafish and human pluripotent cells. Here, we first time report this knockout method in bacteria on this paper. Bioassay experiments demonstrated that the adp knockout mutant of B16 showed considerably lower colonization activity, reduced numbers of intestines and less than 80% nematocidal activity compared with the wild-type strain when infected for 48 h. However, no obvious change on proteolytic activity was observed in the mutant. Conversely, the complementation of adp gene restored most of the above deficient phenotypes. These results indicated that the adp gene was involved in surface adhesion and played a comparatively important role in colonizing host nematodes. Moreover, TALENs successfully disrupt target genes in bacteria. PMID:25912819

  17. Knockout of the adp gene related with colonization in Bacillus nematocida B16 using customized transcription activator-like effectors nucleases

    PubMed Central

    Niu, Qiuhong; Zheng, Haoying; Zhang, Lin; Qin, Fujun; Facemire, Loryn; Zhang, Guo; Cao, Feng; Zhang, Ke-qin; Huang, Xiaowei; Yang, Jianwei; He, Lei; Liu, Chanjuan

    2015-01-01

    Bacillus nematocida B16 is able to dominate in the intestines of the worm Caenorhabditis elegans in ‘Trojan horse’ pathogenic mechanism. The adp is one candidate gene which potentially play a vital role in the colonization from our previous random mutagenesis screening results. To analyse the functional role of this gene, we constructed the adp knockout mutant through customized transcription activator-like effectors nucleases (TALEN), which has been successfully used in yeasts, nematodes, zebrafish and human pluripotent cells. Here, we first time report this knockout method in bacteria on this paper. Bioassay experiments demonstrated that the adp knockout mutant of B16 showed considerably lower colonization activity, reduced numbers of intestines and less than 80% nematocidal activity compared with the wild-type strain when infected for 48 h. However, no obvious change on proteolytic activity was observed in the mutant. Conversely, the complementation of adp gene restored most of the above deficient phenotypes. These results indicated that the adp gene was involved in surface adhesion and played a comparatively important role in colonizing host nematodes. Moreover, TALENs successfully disrupt target genes in bacteria. PMID:25912819

  18. A genome-wide linkage and association study of musical aptitude identifies loci containing genes related to inner ear development and neurocognitive functions.

    PubMed

    Oikkonen, J; Huang, Y; Onkamo, P; Ukkola-Vuoti, L; Raijas, P; Karma, K; Vieland, V J; Järvelä, I

    2015-02-01

    Humans have developed the perception, production and processing of sounds into the art of music. A genetic contribution to these skills of musical aptitude has long been suggested. We performed a genome-wide scan in 76 pedigrees (767 individuals) characterized for the ability to discriminate pitch (SP), duration (ST) and sound patterns (KMT), which are primary capacities for music perception. Using the Bayesian linkage and association approach implemented in program package KELVIN, especially designed for complex pedigrees, several single nucleotide polymorphisms (SNPs) near genes affecting the functions of the auditory pathway and neurocognitive processes were identified. The strongest association was found at 3q21.3 (rs9854612) with combined SP, ST and KMT test scores (COMB). This region is located a few dozen kilobases upstream of the GATA binding protein 2 (GATA2) gene. GATA2 regulates the development of cochlear hair cells and the inferior colliculus (IC), which are important in tonotopic mapping. The highest probability of linkage was obtained for phenotype SP at 4p14, located next to the region harboring the protocadherin 7 gene, PCDH7. Two SNPs rs13146789 and rs13109270 of PCDH7 showed strong association. PCDH7 has been suggested to play a role in cochlear and amygdaloid complexes. Functional class analysis showed that inner ear and schizophrenia-related genes were enriched inside the linked regions. This study is the first to show the importance of auditory pathway genes in musical aptitude. PMID:24614497

  19. Transcriptome Analysis Identifies Candidate Genes Related to Triacylglycerol and Pigment Biosynthesis and Photoperiodic Flowering in the Ornamental and Oil-Producing Plant, Camellia reticulata (Theaceae)

    PubMed Central

    Yao, Qiu-Yang; Huang, Hui; Tong, Yan; Xia, En-Hua; Gao, Li-Zhi

    2016-01-01

    Camellia reticulata, which is native to Southwest China, is famous for its ornamental flowers and high-quality seed oil. However, the lack of genomic information for this species has largely hampered our understanding of its key pathways related to oil production, photoperiodic flowering process and pigment biosynthesis. Here, we first sequenced and characterized the transcriptome of a diploid C. reticulata in an attempt to identify genes potentially involved in triacylglycerol biosynthesis (TAGBS), photoperiodic flowering, flavonoid biosynthesis (FlaBS), carotenoid biosynthesis (CrtBS) pathways. De novo assembly of the transcriptome provided a catalog of 141,460 unigenes with a total length of ~96.1 million nucleotides (Mnt) and an N50 of 1080 nt. Of them, 22,229 unigenes were defined as differentially expressed genes (DEGs) across five sequenced tissues. A large number of annotated genes in C. reticulata were found to have been duplicated, and differential expression patterns of these duplicated genes were commonly observed across tissues, such as the differential expression of SOC1_a, SOC1_b, and SOC1_c in the photoperiodic flowering pathway. Up-regulation of SAD_a and FATA genes and down-regulation of FAD2_a gene in the TAGBS pathway in seeds may be relevant to the ratio of monounsaturated fatty acid (MUFAs) to polyunsaturated fatty acid (PUFAs) in seed oil. MYBF1, a transcription regulator gene of the FlaBS pathway, was found with great sequence variation and alteration of expression patterns, probably resulting in functionally evolutionary differentiation in C. reticulata. MYBA1_a and some anthocyanin-specific biosynthetic genes in the FlaBS pathway were highly expressed in both flower buds and flowers, suggesting important roles of anthocyanin biosynthesis in flower development. Besides, a total of 40,823 expressed sequence tag simple sequence repeats (EST-SSRs) were identified in the C. reticulata transcriptome, providing valuable marker resources for

  20. Transcriptome Analysis Identifies Candidate Genes Related to Triacylglycerol and Pigment Biosynthesis and Photoperiodic Flowering in the Ornamental and Oil-Producing Plant, Camellia reticulata (Theaceae).

    PubMed

    Yao, Qiu-Yang; Huang, Hui; Tong, Yan; Xia, En-Hua; Gao, Li-Zhi

    2016-01-01

    Camellia reticulata, which is native to Southwest China, is famous for its ornamental flowers and high-quality seed oil. However, the lack of genomic information for this species has largely hampered our understanding of its key pathways related to oil production, photoperiodic flowering process and pigment biosynthesis. Here, we first sequenced and characterized the transcriptome of a diploid C. reticulata in an attempt to identify genes potentially involved in triacylglycerol biosynthesis (TAGBS), photoperiodic flowering, flavonoid biosynthesis (FlaBS), carotenoid biosynthesis (CrtBS) pathways. De novo assembly of the transcriptome provided a catalog of 141,460 unigenes with a total length of ~96.1 million nucleotides (Mnt) and an N50 of 1080 nt. Of them, 22,229 unigenes were defined as differentially expressed genes (DEGs) across five sequenced tissues. A large number of annotated genes in C. reticulata were found to have been duplicated, and differential expression patterns of these duplicated genes were commonly observed across tissues, such as the differential expression of SOC1_a, SOC1_b, and SOC1_c in the photoperiodic flowering pathway. Up-regulation of SAD_a and FATA genes and down-regulation of FAD2_a gene in the TAGBS pathway in seeds may be relevant to the ratio of monounsaturated fatty acid (MUFAs) to polyunsaturated fatty acid (PUFAs) in seed oil. MYBF1, a transcription regulator gene of the FlaBS pathway, was found with great sequence variation and alteration of expression patterns, probably resulting in functionally evolutionary differentiation in C. reticulata. MYBA1_a and some anthocyanin-specific biosynthetic genes in the FlaBS pathway were highly expressed in both flower buds and flowers, suggesting important roles of anthocyanin biosynthesis in flower development. Besides, a total of 40,823 expressed sequence tag simple sequence repeats (EST-SSRs) were identified in the C. reticulata transcriptome, providing valuable marker resources for

  1. Multiple Genes Related to Muscle Identified through a Joint Analysis of a Two-stage Genome-wide Association Study for Racing Performance of 1,156 Thoroughbreds

    PubMed Central

    Shin, Dong-Hyun; Lee, Jin Woo; Park, Jong-Eun; Choi, Ik-Young; Oh, Hee-Seok; Kim, Hyeon Jeong; Kim, Heebal

    2015-01-01

    Thoroughbred, a relatively recent horse breed, is best known for its use in horse racing. Although myostatin (MSTN) variants have been reported to be highly associated with horse racing performance, the trait is more likely to be polygenic in nature. The purpose of this study was to identify genetic variants strongly associated with racing performance by using estimated breeding value (EBV) for race time as a phenotype. We conducted a two-stage genome-wide association study to search for genetic variants associated with the EBV. In the first stage of genome-wide association study, a relatively large number of markers (~54,000 single-nucleotide polymorphisms, SNPs) were evaluated in a small number of samples (240 horses). In the second stage, a relatively small number of markers identified to have large effects (170 SNPs) were evaluated in a much larger number of samples (1,156 horses). We also validated the SNPs related to MSTN known to have large effects on racing performance and found significant associations in the stage two analysis, but not in stage one. We identified 28 significant SNPs related to 17 genes. Among these, six genes have a function related to myogenesis and five genes are involved in muscle maintenance. To our knowledge, these genes are newly reported for the genetic association with racing performance of Thoroughbreds. It complements a recent horse genome-wide association studies of racing performance that identified other SNPs and genes as the most significant variants. These results will help to expand our knowledge of the polygenic nature of racing performance in Thoroughbreds. PMID:25925054

  2. Deer Antler Extract Improves Fatigue Effect through Altering the Expression of Genes Related to Muscle Strength in Skeletal Muscle of Mice

    PubMed Central

    Hsiang, Chien-Yun; Lin, Yung-Chang; Ho, Tin-Yun

    2014-01-01

    Deer antler is a well-known traditional Chinese medicine used in Asian countries for the tonic and the improvement of aging symptoms. The present study was designed to investigate the antifatigue effect and mechanism of Formosan sambar deer tip antler extract (FSDTAE). The swimming times to exhaustion of mice administered FSDTAE (8.2 mg/day) for 28 days were apparently longer than those of the vehicle-treated mice in forced swim test. However, the indicators of fatigue, such as the reduction in glucose level and the increases in blood urea nitrogen and lactic acid levels, were not significantly inhibited by FSDTAE. Therefore, microarray analysis was further used to examine the anti-fatigue mechanism of FSDTAE. We selected genes with fold changes >2 or <−2 in skeletal muscle for pathway analysis. FSDTAE-affected genes were involved in 9 different signaling pathways, such as GnRH signaling pathway and insulin signaling pathway. All of the significantly expressed genes were classified into 8 different categories by their functions. The most enriched category was muscular system, and 6 upregulated genes, such as troponin I, troponin T1, cysteine and glycine-rich protein 2, myosin heavy polypeptide 7, tropomyosin 2, and myomesin family member 3, were responsible for the development and contraction of muscle. Real-time PCR analysis indicated that FSDTAE increased troponins mRNA expression in skeletal muscle. In conclusion, our findings suggested that FSDTAE might increase the muscle strength through the upregulation of genes responsible for muscle contraction and consequently exhibited the anti-fatigue effect in mice. PMID:24701242

  3. Multiple Genes Related to Muscle Identified through a Joint Analysis of a Two-stage Genome-wide Association Study for Racing Performance of 1,156 Thoroughbreds.

    PubMed

    Shin, Dong-Hyun; Lee, Jin Woo; Park, Jong-Eun; Choi, Ik-Young; Oh, Hee-Seok; Kim, Hyeon Jeong; Kim, Heebal

    2015-06-01

    Thoroughbred, a relatively recent horse breed, is best known for its use in horse racing. Although myostatin (MSTN) variants have been reported to be highly associated with horse racing performance, the trait is more likely to be polygenic in nature. The purpose of this study was to identify genetic variants strongly associated with racing performance by using estimated breeding value (EBV) for race time as a phenotype. We conducted a two-stage genome-wide association study to search for genetic variants associated with the EBV. In the first stage of genome-wide association study, a relatively large number of markers (~54,000 single-nucleotide polymorphisms, SNPs) were evaluated in a small number of samples (240 horses). In the second stage, a relatively small number of markers identified to have large effects (170 SNPs) were evaluated in a much larger number of samples (1,156 horses). We also validated the SNPs related to MSTN known to have large effects on racing performance and found significant associations in the stage two analysis, but not in stage one. We identified 28 significant SNPs related to 17 genes. Among these, six genes have a function related to myogenesis and five genes are involved in muscle maintenance. To our knowledge, these genes are newly reported for the genetic association with racing performance of Thoroughbreds. It complements a recent horse genome-wide association studies of racing performance that identified other SNPs and genes as the most significant variants. These results will help to expand our knowledge of the polygenic nature of racing performance in Thoroughbreds. PMID:25925054

  4. Isolation and characterization of the human parathyroid hormone-like peptide gene

    SciTech Connect

    Mangin, M.; Ikeda, K.; Dreyer, B.E.; Broadus, A.E. )

    1989-04-01

    A parathyroid hormone-like peptide (PTH-LP) has recently been identified in human tumors associated with the syndrome of humoral hypercalcemia of malignancy. The peptide appears to be encoded by a single-copy gene that gives rise to multiple mRNAs that are heterogeneous at both their 5{prime} and their 3{prime} ends. Alternative RNA splicing is responsible for the 3{prime} heterogeneity and results in mRNAs encoding three different peptides, each with a unique C terminus. The authors have isolated and characterized the human PTHLP gene. The gene is a complex transcriptional unit spanning more than 12 kilobases of DNA and containing six exons. Two 5{prime} exons encode distinct 5{prime} untranslated regions and are separated by a putative promoter element, indicating that the gene either has two promoters or is alternatively spliced from a single promoter upstream of the first exon. The middle portion of the PTHLP gene, comprising exons 2-4, has an organizational pattern of introns and exons identical to that of the parathyroid hormone gene, consistent with a common ancestral origin of these two genes. Exon 4 of the PTHLP gene encodes the region common to all three peptides and the C terminus of the shortest peptide, and exons 5 and 6 encode the unique C termini of the other two peptides. Northern analysis of mRNAs from four human tumors of different histological types reveals the preferential use of 3{prime} splicing patterns of individual tumors.

  5. Dtrk, a Drosophila gene related to the trk family of neurotrophin receptors, encodes a novel class of neural cell adhesion molecule.

    PubMed Central

    Pulido, D; Campuzano, S; Koda, T; Modolell, J; Barbacid, M

    1992-01-01

    We report the identification and molecular characterization of Dtrk, a Drosophila gene encoding a receptor tyrosine kinase highly related to the trk family of mammalian neurotrophin receptors. The product of the Dtrk gene, gp160Dtrk, is dynamically expressed during Drosophila embryogenesis in several areas of the developing nervous system, including neurons and fasciculating axons. gp160Dtrk has structural homology with neural cell adhesion molecules of the immunoglobulin superfamily and promotes cell adhesion in a homophilic, Ca2+ independent manner. More importantly, this adhesion process specifically activates its tyrosine protein kinase activity. These findings suggest that gp160Dtrk represents a new class of neural cell adhesion molecules that may regulate neuronal recognition and axonal guidance during the development of the Drosophila nervous system. Images PMID:1371458

  6. Identification and functional characterization of cDNAs coding for hydroxybenzoate/hydroxycinnamate glucosyltransferases co-expressed with genes related to proanthocyanidin biosynthesis

    PubMed Central

    Khater, F.; Fournand, D.; Vialet, S.; Meudec, E.; Cheynier, V.; Terrier, N.

    2012-01-01

    Grape proanthocyanidins (PAs) play a major role in the organoleptic properties of wine. They are accumulated mainly in grape skin and seeds during the early stages of berry development. Despite the recent progress in the identification of genes involved in PA biosynthesis, the mechanisms involved in subunit condensation, galloylation, or fine regulation of the spatio-temporal composition of grape berries in PAs are still not elucidated. Two Myb transcription factors, VvMybPA1 and VvMybPA2, controlling the PA pathway have recently been identified and ectopically over-expressed in an homologous system. In addition to already known PA genes, three genes coding for glucosyltransferases were significantly differentially expressed between hairy roots over-expressing VvMybPA1 or VvMybPA2 and control lines. The involvement of these genes in PA biosynthesis metabolism is unclear. The three glucosyltransferases display high sequence similarities with other plant glucosyltransferases able to catalyse the formation of glucose esters, which are important intermediate actors for the synthesis of different phenolic compounds. Studies of the in vitro properties of these three enzymes (Km, Vmax, substrate specificity, pH sensitivity) were performed through production of recombinant proteins in E. coli and demonstrated that they are able to catalyse the formation of 1-O-acyl-Glc esters of phenolic acids but are not active on flavonoids and stilbenes. The transcripts are expressed in the early stages of grape berry development, mainly in the berry skins and seeds. The results presented here suggest that these enzymes could be involved in vivo in PA galloylation or in the synthesis of hydroxycinnamic esters. PMID:22090445

  7. Comparison of orthologous cytochrome P450 genes relative expression patterns in the bark beetles Dendroctonus rhizophagus and Dendroctonus valens (Curculionidae: Scolytinae) during host colonization.

    PubMed

    Obregón-Molina, G; Cesar-Ayala, A K; López, M F; Cano-Ramírez, C; Zúñiga, G

    2015-12-01

    Bark beetles of the genus Dendroctonus are important components of coniferous forests. During host colonization, they must overcome the chemical defences of their host trees, which are metabolized by cytochrome P450 (CYP or P450) enzymes to compounds that are readily excreted. In this study, we report the relative expression (quantitative real-time PCR) of four orthologous cytochrome P450 genes (CYP6BW5, CYP6DG1, CYP6DJ2 and CYP9Z20) in Dendroctonus rhizophagus and Dendroctonus valens forced to attack host trees at 8 and 24 h following forced attack and in four stages during natural colonization [solitary females boring the bark (T1); both male and female members of couples before oviposition (T2); both male and female members of couples during oviposition (T3), and solitary females inside the gallery containing eggs (T4)]. For both species gene expression was different compared with that observed in insects exposed to single monoterpenes in the laboratory, and the expression patterns were significantly different amongst species, sex, gut region and exposure time or natural colonization stage. The induction of genes (CYP6BW5v1, CYP6DJ2v1 and CYP9Z20v1 from D. rhizophagus, as well as CYP6DG1v3 from D. valens) correlated with colonization stage as well as with the increase in oxygenated monoterpenes in the gut of both species throughout the colonization of the host. Our results point to different functions of these orthologous genes in both species. PMID:26537737

  8. Analysis of Sensitive CO2 Pathways and Genes Related to Carbon Uptake and Accumulation in Chlamydomonas reinhardtii through Genomic Scale Modeling and Experimental Validation

    PubMed Central

    Winck, Flavia V.; Melo, David O. Páez; Riaño-Pachón, Diego M.; Martins, Marina C. M.; Caldana, Camila; Barrios, Andrés F. González

    2016-01-01

    The development of microalgae sustainable applications needs better understanding of microalgae biology. Moreover, how cells coordinate their metabolism toward biomass accumulation is not fully understood. In this present study, flux balance analysis (FBA) was performed to identify sensitive metabolic pathways of Chlamydomonas reinhardtii under varied CO2 inputs. The metabolic network model of Chlamydomonas was updated based on the genome annotation data and sensitivity analysis revealed CO2 sensitive reactions. Biological experiments were performed with cells cultivated at 0.04% (air), 2.5, 5, 8, and 10% CO2 concentration under controlled conditions and cell growth profiles and biomass content were measured. Pigments, lipids, proteins, and starch were further quantified for the reference low (0.04%) and high (10%) CO2 conditions. The expression level of candidate genes of sensitive reactions was measured and validated by quantitative real time PCR. The sensitive analysis revealed mitochondrial compartment as the major affected by changes on the CO2 concentrations and glycolysis/gluconeogenesis, glyoxylate, and dicarboxylate metabolism among the affected metabolic pathways. Genes coding for glycerate kinase (GLYK), glycine cleavage system, H-protein (GCSH), NAD-dependent malate dehydrogenase (MDH3), low-CO2 inducible protein A (LCIA), carbonic anhydrase 5 (CAH5), E1 component, alpha subunit (PDC3), dual function alcohol dehydrogenase/acetaldehyde dehydrogenase (ADH1), and phosphoglucomutase (GPM2), were defined, among other genes, as sensitive nodes in the metabolic network simulations. These genes were experimentally responsive to the changes in the carbon fluxes in the system. We performed metabolomics analysis using mass spectrometry validating the modulation of carbon dioxide responsive pathways and metabolites. The changes on CO2 levels mostly affected the metabolism of amino acids found in the photorespiration pathway. Our updated metabolic network was

  9. Analysis of Sensitive CO2 Pathways and Genes Related to Carbon Uptake and Accumulation in Chlamydomonas reinhardtii through Genomic Scale Modeling and Experimental Validation.

    PubMed

    Winck, Flavia V; Melo, David O Páez; Riaño-Pachón, Diego M; Martins, Marina C M; Caldana, Camila; Barrios, Andrés F González

    2016-01-01

    The development of microalgae sustainable applications needs better understanding of microalgae biology. Moreover, how cells coordinate their metabolism toward biomass accumulation is not fully understood. In this present study, flux balance analysis (FBA) was performed to identify sensitive metabolic pathways of Chlamydomonas reinhardtii under varied CO2 inputs. The metabolic network model of Chlamydomonas was updated based on the genome annotation data and sensitivity analysis revealed CO2 sensitive reactions. Biological experiments were performed with cells cultivated at 0.04% (air), 2.5, 5, 8, and 10% CO2 concentration under controlled conditions and cell growth profiles and biomass content were measured. Pigments, lipids, proteins, and starch were further quantified for the reference low (0.04%) and high (10%) CO2 conditions. The expression level of candidate genes of sensitive reactions was measured and validated by quantitative real time PCR. The sensitive analysis revealed mitochondrial compartment as the major affected by changes on the CO2 concentrations and glycolysis/gluconeogenesis, glyoxylate, and dicarboxylate metabolism among the affected metabolic pathways. Genes coding for glycerate kinase (GLYK), glycine cleavage system, H-protein (GCSH), NAD-dependent malate dehydrogenase (MDH3), low-CO2 inducible protein A (LCIA), carbonic anhydrase 5 (CAH5), E1 component, alpha subunit (PDC3), dual function alcohol dehydrogenase/acetaldehyde dehydrogenase (ADH1), and phosphoglucomutase (GPM2), were defined, among other genes, as sensitive nodes in the metabolic network simulations. These genes were experimentally responsive to the changes in the carbon fluxes in the system. We performed metabolomics analysis using mass spectrometry validating the modulation of carbon dioxide responsive pathways and metabolites. The changes on CO2 levels mostly affected the metabolism of amino acids found in the photorespiration pathway. Our updated metabolic network was

  10. Effects of BPA and E2 on expression profiles of genes related to hypothalamic-pituitary-gonadal axis of half-smooth tongue sole Cynoglossus semilaevis

    NASA Astrophysics Data System (ADS)

    Li, Fengling; Li, Zhaoxin; Wang, Qingyin; Zhai, Yuxiu

    2013-05-01

    Endocrine disrupting chemicals (EDCs) are increasingly viewed as persistent pollutants, similar to natural hormones in function. This paper describes the expression profiles of 7 genes ( DMRT, VTG GnRHR FSHR CYP17A CYP19A, and CYP19B) involved in sex steroid synthesis and action as well as sexual development in adult male and female Cynoglossus semilaevis, after exposure to different concentrations of Bisphenol A (BPA) and 17β-estradiol (E2). Both BPA (1, 10, 50, 125, and 250 mg/kg) and E2 (0.5, 5, and 10 mg/kg) induced changes in target gene expression, although the estrogenic effects of E2 as a model estrogen were stronger. Among the 7 genes, VTG CYP17A and CYP19 responded strongly to BPA or E2 exposure and can thus serve as reference biomarkers for estrogenic EDCs exposure in marine teleosts. These data will provide a window to establish a hypothalamic-pituitary-gonadal model in C. semilaevis to better understand the effect pathways of EDCs.

  11. Expression of Genes Related to Germ Cell Lineage and Pluripotency in Single Cells and Colonies of Human Adult Germ Stem Cells.

    PubMed

    Conrad, Sabine; Azizi, Hossein; Hatami, Maryam; Kubista, Mikael; Bonin, Michael; Hennenlotter, Jörg; Sievert, Karl-Dietrich; Skutella, Thomas

    2016-01-01

    The aim of this study was to elucidate the molecular status of single human adult germ stem cells (haGSCs) and haGSC colonies, which spontaneously developed from the CD49f MACS and matrix- (collagen-/laminin+ binding-) selected fraction of enriched spermatogonia. Single-cell transcriptional profiling by Fluidigm BioMark system of a long-term cultured haGSCs cluster in comparison to human embryonic stem cells (hESCs) and human fibroblasts (hFibs) revealed that haGSCs showed a characteristic germ- and pluripotency-associated gene expression profile with some similarities to hESCs and with a significant distinction from somatic hFibs. Genome-wide comparisons with microarray analysis confirmed that different haGSC colonies exhibited gene expression heterogeneity with more or less pluripotency. The results of this study confirm that haGSCs are adult stem cells with a specific molecular gene expression profile in vitro, related but not identical to true pluripotent stem cells. Under ES-cell conditions haGSC colonies could be selected and maintained in a partial pluripotent state at the molecular level, which may be related to their cell plasticity and potential to differentiate into cells of all germ layers. PMID:26649052

  12. Transcriptome-Wide Mapping of Pea Seed Ageing Reveals a Pivotal Role for Genes Related to Oxidative Stress and Programmed Cell Death

    PubMed Central

    Colville, Louise; Lorenzo, Oscar; Graeber, Kai; Küster, Helge; Leubner-Metzger, Gerhard; Kranner, Ilse

    2013-01-01

    Understanding of seed ageing, which leads to viability loss during storage, is vital for ex situ plant conservation and agriculture alike. Yet the potential for regulation at the transcriptional level has not been fully investigated. Here, we studied the relationship between seed viability, gene expression and glutathione redox status during artificial ageing of pea (Pisum sativum) seeds. Transcriptome-wide analysis using microarrays was complemented with qRT-PCR analysis of selected genes and a multilevel analysis of the antioxidant glutathione. Partial degradation of DNA and RNA occurred from the onset of artificial ageing at 60% RH and 50°C, and transcriptome profiling showed that the expression of genes associated with programmed cell death, oxidative stress and protein ubiquitination were altered prior to any sign of viability loss. After 25 days of ageing viability started to decline in conjunction with progressively oxidising cellular conditions, as indicated by a shift of the glutathione redox state towards more positive values (>−190 mV). The unravelling of the molecular basis of seed ageing revealed that transcriptome reprogramming is a key component of the ageing process, which influences the progression of programmed cell death and decline in antioxidant capacity that ultimately lead to seed viability loss. PMID:24205239

  13. Genes Related to Fatty Acid β-Oxidation Play a Role in the Functional Decline of the Drosophila Brain with Age

    PubMed Central

    Laranjeira, António; Schulz, Joachim; Dotti, Carlos G.

    2016-01-01

    In living organisms, ageing is widely considered to be the result of a multifaceted process consisting of the progressive accumulation of damage over time, having implications both in terms of function and survival. The study of ageing presents several challenges, from the different mechanisms implicated to the great diversity of systems affected over time. In the current study, we set out to identify genes involved in the functional decline of the brain with age and study its relevance in a tissue dependent manner using Drosophila melanogaster as a model system. Here we report the age-dependent upregulation of genes involved in the metabolic process of fatty acid β-oxidation in the nervous tissue of female wild-type flies. Downregulation of CG10814, dHNF4 and lipid mobilizing genes bmm and dAkh rescues the functional decline of the brain with age, both at the cellular and behaviour level, while over-expression worsens performance. Our data proposes the occurrence of a metabolic alteration in the fly brain with age, whereby the process of β-oxidation of fatty acids experiences a genetic gain-of-function. This event proved to be one of the main causes contributing to the functional decline of the brain with age. PMID:27518101

  14. Expression of Genes Related to Germ Cell Lineage and Pluripotency in Single Cells and Colonies of Human Adult Germ Stem Cells

    PubMed Central

    Conrad, Sabine; Azizi, Hossein; Hatami, Maryam; Kubista, Mikael; Bonin, Michael; Hennenlotter, Jörg; Sievert, Karl-Dietrich; Skutella, Thomas

    2016-01-01

    The aim of this study was to elucidate the molecular status of single human adult germ stem cells (haGSCs) and haGSC colonies, which spontaneously developed from the CD49f MACS and matrix- (collagen−/laminin+ binding-) selected fraction of enriched spermatogonia. Single-cell transcriptional profiling by Fluidigm BioMark system of a long-term cultured haGSCs cluster in comparison to human embryonic stem cells (hESCs) and human fibroblasts (hFibs) revealed that haGSCs showed a characteristic germ- and pluripotency-associated gene expression profile with some similarities to hESCs and with a significant distinction from somatic hFibs. Genome-wide comparisons with microarray analysis confirmed that different haGSC colonies exhibited gene expression heterogeneity with more or less pluripotency. The results of this study confirm that haGSCs are adult stem cells with a specific molecular gene expression profile in vitro, related but not identical to true pluripotent stem cells. Under ES-cell conditions haGSC colonies could be selected and maintained in a partial pluripotent state at the molecular level, which may be related to their cell plasticity and potential to differentiate into cells of all germ layers. PMID:26649052

  15. Characterization of an essential Saccharomyces cerevisiae gene related to RNA processing: cloning of RNA1 and generation of a new allele with a novel phenotype.

    PubMed Central

    Atkinson, N S; Dunst, R W; Hopper, A K

    1985-01-01

    The RNA1 gene product is believed to be involved in RNA metabolism due to the phenotype of a single conditionally lethal, temperature-sensitive allele, rna1-1. We cloned the RNA1 gene and determined that it produces a 1,400-nucleotide polyadenylated transcript. On a multicopy plasmid, the mutant rna1-1 allele partially complements the rna1-1 temperature-sensitive growth defect. This suggests that the temperature-sensitive nature of the rna1-1 allele results from the synthesis of a product with lowered activity or stability at elevated temperatures or from a decrease in synthesis of the rna1-1 product at the restrictive temperature. A chromosomal disruption of RNA1 behaves as a recessive lethal mutation. Haploids bearing the disruption were isolated by sporulating a diploid heterozygous for the disrupted allele and the rna1-1 allele and possessing an episomal copy of the RNA1 gene. Analysis of the rescued haploids bearing the chromosomal disruption indicated that the recessive lethal phenotype of the RNA1 disruption is not merely due to a block in spore germination. Unexpectedly, diploids heterozygous for the disruption and the rna1-1 alleles become aneuploid for chromosome XIII at a frequency of 2 to 5%. It appears that the disrupted RNA1 allele on a multicopy plasmid also promotes aneuploidy for chromosome XIII. Promotion of aneuploidy seems to be a phenotype of this particular allele of RNA1. Images PMID:3889591

  16. A Homeobox Gene Related to Drosophila Distal-Less Promotes Ovarian Tumorigenicity by Inducing Expression of Vascular Endothelial Growth Factor and Fibroblast Growth Factor-2

    PubMed Central

    Hara, Fumikata; Samuel, Shaija; Liu, Jinsong; Rosen, Daniel; Langley, Robert R.; Naora, Honami

    2007-01-01

    Homeobox genes control developmental patterning and are increasingly being found to be deregulated in tumors. The DLX4 homeobox gene maps to the 17q21.3-q22 region that is amplified in some epithelial ovarian cancers. Because amplification of this region correlates with poor prognosis, we investigated whether DLX4 overexpression contributes to aggressive behavior of this disease. DLX4 was not detected in normal ovary and cystadenomas, whereas its expression in ovarian carcinomas was strongly associated with high tumor grade and advanced disease stage. Overexpression of DLX4 in ovarian cancer cells promoted growth in low serum and colony formation. Imaging of mice bearing intraperitoneal tumors revealed that DLX4 overexpression substantially increased tumor burden. Tumors that overexpressed DLX4 were more vascularized than vector-control tumors. Conditioned medium of DLX4-overexpressing tumor cells was more effective than medium conditioned by vector-control cells in stimulating endothelial cell growth. These observations were associated with the ability of DLX4 to induce expression of vascular endothelial growth factor as well as intracellular and secreted isoforms of fibroblast growth factor-2. Moreover, increased levels of these fibroblast growth factor-2 isoforms induced vascular endothelial growth factor expression in tumor cells. This study reveals a novel role for a homeobox gene in ovarian tumorigenicity by its induction of a proangiogenic, growth-stimulatory molecular program. PMID:17456765

  17. A homeobox gene related to Drosophila distal-less promotes ovarian tumorigenicity by inducing expression of vascular endothelial growth factor and fibroblast growth factor-2.

    PubMed

    Hara, Fumikata; Samuel, Shaija; Liu, Jinsong; Rosen, Daniel; Langley, Robert R; Naora, Honami

    2007-05-01

    Homeobox genes control developmental patterning and are increasingly being found to be deregulated in tumors. The DLX4 homeobox gene maps to the 17q21.3-q22 region that is amplified in some epithelial ovarian cancers. Because amplification of this region correlates with poor prognosis, we investigated whether DLX4 overexpression contributes to aggressive behavior of this disease. DLX4 was not detected in normal ovary and cystadenomas, whereas its expression in ovarian carcinomas was strongly associated with high tumor grade and advanced disease stage. Overexpression of DLX4 in ovarian cancer cells promoted growth in low serum and colony formation. Imaging of mice bearing intraperitoneal tumors revealed that DLX4 overexpression substantially increased tumor burden. Tumors that overexpressed DLX4 were more vascularized than vector-control tumors. Conditioned medium of DLX4-overexpressing tumor cells was more effective than medium conditioned by vector-control cells in stimulating endothelial cell growth. These observations were associated with the ability of DLX4 to induce expression of vascular endothelial growth factor as well as intracellular and secreted isoforms of fibroblast growth factor-2. Moreover, increased levels of these fibroblast growth factor-2 isoforms induced vascular endothelial growth factor expression in tumor cells. This study reveals a novel role for a homeobox gene in ovarian tumorigenicity by its induction of a proangiogenic, growth-stimulatory molecular program. PMID:17456765

  18. Effects of steroid treatment on growth, nutrient partitioning, and expression of genes related to growth and nutrient metabolism in adult triploid rainbow trout (Oncorhynchus mykiss).

    PubMed

    Cleveland, B M; Weber, G M

    2016-07-01

    The contribution of sex steroids to nutrient partitioning and energy balance during gonad development was studied in rainbow trout. Specifically, 19-mo old triploid (3N) female rainbow trout were fed treatment diets supplemented with estradiol-17β (E2), testosterone (T), or dihydrotestosterone at 30-mg steroid/kg diet for a 1-mo period. Growth performance, nutrient partitioning, and expression of genes central to growth and nutrient metabolism were compared with 3N and age-matched diploid (2N) female fish consuming a control diet not supplemented with steroids. Only 2 N fish exhibited active gonad development, with gonad weights increasing from 3.7% to 5.5% of body weight throughout the study, whereas gonad weights in 3N fish remained at 0.03%. Triploid fish consuming dihydrotestosterone exhibited faster specific growth rates than 3N-controls (P < 0.05). Consumption of E2 in 3N fish reduced fillet growth and caused lower fillet yield compared with all other treatment groups (P < 0.05). In contrast, viscera fat gain was not affected by steroid consumption (P > 0.05). Gene transcripts associated with physiological pathways were identified in maturing 2N and E2-treated 3N fish that differed in abundance from 3N-control fish (P < 0.05). In liver these mechanisms included the growth hormone/insulin-like growth factor (IGF) axis (igf1, igf2), IGF binding proteins (igfbp1b1, igfbp2b1, igfbp5b1, igfbp6b1), and genes associated with lipid binding and transport (fabp3, fabp4, lpl, cd36), fatty acid oxidation (cpt1a), and the pparg transcription factor. In muscle, these mechanisms included reductions in myogenic gene expression (fst, myog) and the proteolysis-related gene, cathepsin-L, suggesting an E2-induced reduction in the capacity for muscle growth. These findings suggest that increased E2 signaling in the sexually maturing female rainbow trout alters physiological pathways in liver, particularly those related to IGF signaling and lipid metabolism, to partition

  19. Retinoid receptor-specific agonists regulate bovine in vitro early embryonic development, differentiation and expression of genes related to cell cycle arrest and apoptosis.

    PubMed

    Rodríguez, A; Díez, C; Caamaño, J N; de Frutos, C; Royo, L J; Muñoz, M; Ikeda, S; Facal, N; Alvarez-Viejo, M; Gómez, E

    2007-11-01

    A major goal in reproductive biotechnology is the identification of pathways that regulate early embryonic development and the allocation of cells to the inner cell mass (ICM) and trophectoderm (TE). Retinoids regulate the development and differentiation of the bovine blastocyst in vitro, although the involvement of the retinoid X receptors (RXRs) remains to be clarified. This paper compares the effect of a synthetic RXR agonist (LG100268; LG) with that of the retinoic acid receptor (RAR) agonist all-trans retinoic acid (ATRA) on blastulation. In vitro-produced morulae were treated for 48 h with LG (0.1 microM, 1 microM and 10 microM), ATRA 0.7 microM, or no additives. Treatment with ATRA did not increase the rate of development; however, the LG 0.1 microM treatment increased both the blastocyst development and hatching rate. Cell numbers increased in the ICM with LG 10 microM, while a dose-dependent reduction was observed in the TE in the presence of LG. Gene expression levels of p53 and p66 did not vary with LG but increased with ATRA. Both LG and ATRA activated bax, a pro-apoptotic gene and H2A.Z, a cell cycle-related gene. The above effects suggest the existence of active p53-dependent and -independent apoptotic pathways for ATRA and LG, respectively, in the bovine embryo. The expression of p53 and H2A.Z showed a strong, positive correlation (r=0.93; p<0.0001) in all experimental groups; both proteins are linked through the cell cycle. Agonists of RXR could be used to control blastocyst development and differentiation. PMID:17869331

  20. Transcriptome analysis of genes related to resistance against powdery mildew in wheat-Thinopyrum alien addition disomic line germplasm SN6306.

    PubMed

    Li, Quanquan; Niu, Zubiao; Bao, Yinguang; Tian, Qiuju; Wang, Honggang; Kong, Lingrang; Feng, Deshun

    2016-09-15

    Wheat powdery mildew, which is mainly caused by Blumeria graminis f. sp. tritici (Bgt), seriously damages wheat production. The wheat-Thinopyrum intermedium alien addition disomic line germplasm SN6306, being one of the important sources of genes for wheat resistance, is highly resistant to Bgt E09 and to many other powdery mildew physiological races. However, knowledge on the resistance mechanism of SN6306 remains limited. Our study employed high-throughput RNA sequencing based on next-generation sequencing technology (Illumina) to obtain an overview of the transcriptome characteristics of SN6306 and its parent wheat Yannong 15 (YN15) during Bgt infection. The sequencing generated 104,773 unigenes, 9909 of which showed varied expression levels. Among the 9909 unigenes, 1678 unigenes showed 0 reads in YN15. The expression levels in Bgt-inoculated SN6306 and YN15 of exactly 39 unigenes that showed 0 or considerably low reads in YN15 were validated to identify the genes involved in Bgt resistance. Among the 39 unigenes, 12 unigenes were upregulated in SN6306 by 3-45 times. These unigenes mainly encoded kinase, synthase, proteases, and signal transduction proteins, which may play an important role in the resistance against Bgt. To confirm whether the unigenes that showed 0 reads in YN15 are really unique to SN6306, 8 unigenes were cloned and sequenced. Results showed that the selected unigenes are more similar to SN6306 and Th. intermedium than to the wheat cultivar YN15. The sequencing results further confirmed that the unigenes showing 0 reads in YN15 are unique to SN6306 and are most likely derived from Th. intermedium (Host) Nevski. Thus, the genes from Th. intermedium most probably conferred the resistance of SN6306 to Bgt. PMID:27265028

  1. Expression of genes related to the human erbB, erbA, pdgf and pdgf-r in tumors of different etiology in Xiphophorus.

    PubMed

    Zechel, C; Peters, H; Schleenbecker, U; Anders, F

    1992-08-19

    The melanoma determining Tu locus of the teleost Xiphophorus contains an accessory gene, x-erbB*a, which is closely related to the EGF receptor gene family, and is probably oncogenic. x-erbB*a exists in allelic forms that are specific for distinct Tu-loci, and shows high homology to a non-allelic non-oncogenic counterpart x-erbB*i which is transcribed into mRNA of 4.6 kb in non-tumorous and tumorous tissues of fish harboring and lacking Tu. Expression of a 4.0-kb mRNA in tumors (melanoma and fibrosarcoma) of different etiology is strictly correlated with the inheritance of X. maculatus x-erbB*a alleles; transcripts of 8.0 kb were detected in melanoma and carcinoma of fish harboring a certain x-erbB*a of X. variatus. The expression of the putative x-erbB*a transcripts parallels the stage of malignancy of the tumor. The expression of the xiphophorine EGF receptor gene (x-erbB) was detected in almost all tumors, is strongly enhanced in carcinoma, and is positively correlated with the degree of malignancy of melanoma and fibrosarcoma. Some tumors show expression of erbA-related genes. The PDGF receptor mRNA is expressed in all tumors analyzed and shows enhanced expression in malignant tumors of neurogenic, epithelial and mesenchymal origin. Expression of x-pdgf was observed in several cases of melanoma, but more frequently in carcinoma and fibrosarcoma. We conclude that x-erbB*a might be involved in initiation of tumors of different cellular origin and etiology in fish harboring Tu, as well as in the determination of the malignancy of the tumor. Furthermore, we assume that x-erbB*i, x-erbB, x-pdgf and x-pdgf-r play a role in secondary events in tumorigenesis by, e.g., conferring a selective growth advantage to the tumor cells. PMID:1323541

  2. Matrix-based three-dimensional culture of buffalo mammary epithelial cells showed higher induction of genes related to milk protein and fatty acid metabolism.

    PubMed

    Shandilya, Umesh K; Sharma, Ankita; Sodhi, Monika; Kapila, Neha; Kishore, Amit; Mohanty, Ashok; Kataria, Ranjit; Malakar, Dhruva; Mukesh, Manishi

    2016-02-01

    Demanding transcriptomic studies in livestock animal species could be replaced by good in vitro models mimicking the function of mammary gland. Mammary epithelial cells (MEC) are the functional unit of the mammary gland. Extracellular matrix is known to be a key factor providing normal homeostasis in three-dimensional (3D) environment as important signals are lost when cells are cultured in two-dimensional (2D) environment. The aims of this study were to establish a buffalo mammary epithelial cells (BMECs) in 3D culture using extracellular matrix and to determine whether such a 3D culture model has different expression pattern than 2D counterpart. The purified MEC generated after several passages were used to establish 3D culture using Geltrex matrix. The expression of milk casein genes viz., alpha S1-casein (CSN1S1), alpha S2-casein (CSN1S2), beta-casein (CSN2), kappa-casein (CSN3); and fatty acid metabolism genes viz., butyrophilin (BTN1A1), glycerol-3-phosphate acyltransferase (GPAM), fatty acid-binding protein 3 (FABP3), and stearoyl-CoA desaturase (SCD) was assessed in 3D culture in comparison to traditional monolayer culture using qRT-PCR. Notable morphological differences were observed for BMECs grown in 3D culture in comparison to 2D culture. Morphologically, epithelial structures grown in Geltrex matrix (3D) environment showed enhanced functional differentiation in comparison to 2D culture. In 3D culture, lumen and dome-like structures were formed by day 5, whereas polarized acinus-like structure were formed within 15 days of culturing. The expression data showed higher mRNA induction of milk casein and fatty acid metabolism genes in 10-day-old 3D BMECs culture in comparison to 2D monolayer culture. The result suggests that 3D organization of epithelial cells has favorable effect on induction of milk and fatty acid metabolism-related genes. Therefore, matrix-based 3D culture of MEC that recapitulate the structural and functional context of normal tissues

  3. L206W mutation of the cystic fibrosis gene, relatively frequent in French Canadians, is associated with atypical presentations of cystic fibrosis

    SciTech Connect

    Rozen, R.; Ferreira-Rajabi, L.; Robb, L.

    1995-07-03

    Cystic fibrosis is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Over 400 mutations have been reported at this locus. Although severe forms of cystic fibrosis are usually associated with pancreatic insufficiency, pulmonary dysfunction, and elevated sweat chloride, there is a wide range of phenotypes, including congenital absence of the vas deferens, observed with some of the milder mutations. The L206W mutation, which was first identified in patients from South France, is relatively frequent in French Canadians from Quebec. In this report, we document the atypical form of cystic fibrosis associated with this mutation in a cohort of 7 French Canadian probands. 20 refs.

  4. Effects of nutritional level of concentrate-based diets on meat quality and expression levels of genes related to meat quality in Hainan black goats.

    PubMed

    Wang, Dingfa; Zhou, Luli; Zhou, Hanlin; Hou, Guanyu; Shi, Liguang; Li, Mao; Huang, Xianzhou; Guan, Song

    2015-02-01

    The present study investigated the effects of the nutritional levels of diets on meat quality and related gene expression in Hainan black goat. Twenty-four goats were divided into six dietary treatments and were fed a concentrate-based diet with two levels of crude protein (CP) (15% or 17%) and three levels of digestive energy (DE) (11.72, 12.55 or 13.39 MJ/kg DM) for 90 days. Goats fed the concentrate-based diet with 17% CP had significantly (P < 0.05) higher average daily gains (ADG) and better feed conversion rates (FCR). The pH 24h value tended to decrease (P < 0.05) with increasing DE levels. The tenderness of Longissimus dorsi muscle (LD) and Semimembranosus muscle (SM) reduced with increasing CP levels (P < 0.05). With increasing DE levels, tenderness was increased (P < 0.05). The heart fatty acid-binding protein (H-FABP) mRNA expression levels in LD and SM increased with increasing DE levels (P < 0.05), but decreased with increasing CP levels (P < 0.05). The calpastatin (CAST) and μ-calpain mRNA expressions levels in LD and SM were affected significantly (P < 0.05) by CP and DE levels in the diet. Therefore, the nutritional levels of diets affect meat quality and expression levels of genes associated with meat quality in Hainan black goats. PMID:25039653

  5. Mutation in TOR1AIP1 encoding LAP1B in a form of muscular dystrophy: a novel gene related to nuclear envelopathies.

    PubMed

    Kayman-Kurekci, Gulsum; Talim, Beril; Korkusuz, Petek; Sayar, Nilufer; Sarioglu, Turkan; Oncel, Ibrahim; Sharafi, Parisa; Gundesli, Hulya; Balci-Hayta, Burcu; Purali, Nuhan; Serdaroglu-Oflazer, Piraye; Topaloglu, Haluk; Dincer, Pervin

    2014-07-01

    We performed genome-wide homozygosity mapping and mapped a novel myopathic phenotype to chromosomal region 1q25 in a consanguineous family with three affected individuals manifesting proximal and distal weakness and atrophy, rigid spine and contractures of the proximal and distal interphalangeal hand joints. Additionally, cardiomyopathy and respiratory involvement were noted. DNA sequencing of torsinA-interacting protein 1 (TOR1AIP1) gene encoding lamina-associated polypeptide 1B (LAP1B), showed a homozygous c.186delG mutation that causes a frameshift resulting in a premature stop codon (p.E62fsTer25). We observed that expression of LAP1B was absent in the patient skeletal muscle fibres. Ultrastructural examination showed intact sarcomeric organization but alterations of the nuclear envelope including nuclear fragmentation, chromatin bleb formation and naked chromatin. LAP1B is a type-2 integral membrane protein localized in the inner nuclear membrane that binds to both A- and B-type lamins, and is involved in the regulation of torsinA ATPase. Interestingly, luminal domain-like LAP1 (LULL1)-an endoplasmic reticulum-localized partner of torsinA-was overexpressed in the patient's muscle in the absence of LAP1B. Therefore, the findings suggest that LAP1 and LULL1 might have a compensatory effect on each other. This study expands the spectrum of genes associated with nuclear envelopathies and highlights the critical function for LAP1B in striated muscle. PMID:24856141

  6. Mouse cortical collecting duct cells show nonselective cation channel activity and express a gene related to the cGMP-gated rod photoreceptor channel.

    PubMed Central

    Ahmad, I; Korbmacher, C; Segal, A S; Cheung, P; Boulpaep, E L; Barnstable, C J

    1992-01-01

    Apical nonselective cation channels with an average single-channel conductance of 34 +/- 2.3 pS were found in M-1 mouse cortical collecting duct cells. Channel activity is increased by depolarization and abolished by cytoplasmic calcium removal. Cytoplasmic application of 0.1 mM cGMP decreases channel open probability by 27%. cDNAs corresponding to approximately 40% of the coding region of the photoreceptor channel were isolated by the polymerase chain reaction from M-1 cells and a rat kidney cDNA library. The rat kidney-derived sequence differs by a single base, and the M-1-cell-derived sequence differs by only two bases, from the photoreceptor sequence. A second clone from M-1 cells differs by 20 out of 426 bases from the photoreceptor sequence. In all three clones, the deduced amino acid sequence is identical to that of the rat photoreceptor channel. Northern blot analysis of poly(A)+ RNA from M-1 cells reveals the presence of a 3.2-kilobase band hybridizing with a retinal cGMP-gated cation channel probe. The results suggest the expression in M-1 cells of more than one gene coding for nonselective cation channels or channel subunits, one of which is identical to the cGMP-gated cation channel gene of rod photoreceptors. Images PMID:1279673

  7. De Novo Transcriptome Sequencing of the Orange-Fleshed Sweet Potato and Analysis of Differentially Expressed Genes Related to Carotenoid Biosynthesis

    PubMed Central

    Li, Ruijie; Zhai, Hong; Kang, Chen; Liu, Degao; He, Shaozhen; Liu, Qingchang

    2015-01-01

    Sweet potato, Ipomoea batatas (L.) Lam., is an important food crop worldwide. The orange-fleshed sweet potato is considered to be an important source of beta-carotene. In this study, the transcriptome profiles of an orange-fleshed sweet potato cultivar “Weiduoli” and its mutant “HVB-3” with high carotenoid content were determined by using the high-throughput sequencing technology. A total of 13,767,387 and 9,837,090 high-quality reads were produced from Weiduoli and HVB-3, respectively. These reads were de novo assembled into 58,277 transcripts and 35,909 unigenes with an average length of 596 bp and 533 bp, respectively. In all, 874 differentially expressed genes (DEGs) were obtained between Weiduoli and HVB-3, 401 of which were upregulated and 473 were downregulated in HVB-3 compared to Weiduoli. Of the 697 DEGs annotated, 316 DEGs had GO terms and 62 DEGs were mapped onto 50 pathways. The 22 DEGs and 31 transcription factors involved in carotenoid biosynthesis were identified between Weiduoli and HVB-3. In addition, 1,725 SSR markers were detected. This study provides the genomic resources for discovering the genes involved in carotenoid biosynthesis of sweet potato and other plants. PMID:26649293

  8. Transcriptome Analysis for Identification of Genes Related to Gonad Differentiation, Growth, Immune Response and Marker Discovery in The Turbot (Scophthalmus maximus)

    PubMed Central

    Ma, Deyou; Ma, Aijun; Huang, Zhihui; Wang, Guangning; Wang, Ting; Xia, Dandan; Ma, Benhe

    2016-01-01

    Background Turbot Scophthalmus maximus is an economically important species extensively aquacultured in China. The genetic selection program is necessary and urgent for the sustainable development of this industry, requiring more and more genome background knowledge. Transcriptome sequencing is an excellent alternative way to identify transcripts involved in specific biological processes and exploit a considerable quantity of molecular makers when no genome sequences are available. In this study, a comprehensive transcript dataset for major tissues of S. maximus was produced on basis of an Illumina platform. Results Total RNA was isolated from liver, spleen, kidney, cerebrum, gonad (testis and ovary) and muscle. Equal quantities of RNA from each type of tissues were pooled to construct two cDNA libraries (male and female). Using the Illumina paired-end sequencing technology, nearly 44.22 million clean reads in length of 100 bp were generated and then assembled into 106,643 contigs, of which 71,107 were named unigenes with an average length of 892 bp after the elimination of redundancies. Of these, 24,052 unigenes (33.83% of the total) were successfully annotated. GO, KEGG pathway mapping and COG analysis were performed to predict potential genes and their functions. Based on our sequence analysis and published documents, many candidate genes with fundamental roles in sex determination and gonad differentiation (dmrt1), growth (ghrh, myf5, prl/prlr) and immune response (TLR1/TLR21/TLR22, IL-15/IL-34), were identified for the first time in this species. In addition, a large number of credible genetic markers, including 21,192 SSRs and 8,642 SNPs, were identified in the present dataset. Conclusion This informative transcriptome provides valuable new data to increase genomic resources of Scophthalmus maximus. The future studies of corresponding gene functions will be very useful for the management of reproduction, growth and disease control in turbot aquaculture

  9. Longitudinal Profiling of the Tissue-Specific Expression of Genes Related with Insulin Sensitivity in Dairy Cows during Lactation Focusing on Different Fat Depots

    PubMed Central

    Saremi, Behnam; Winand, Sarah; Friedrichs, Paula; Kinoshita, Asako; Rehage, Jürgen; Dänicke, Sven; Häussler, Susanne; Breves, Gerhard; Mielenz, Manfred; Sauerwein, Helga

    2014-01-01

    In dairy cows the milk associated energy output in early lactation exceeds the input via voluntary feed intake. To spare glucose for mammary lactose synthesis, peripheral insulin sensitivity (IS) is reduced and fat mobilization is stimulated. For these processes a link between IS and the endocrine functions of adipose tissue (AT) is likely; we thus aimed to characterise the mRNA expression from bovine AT derived proteins and receptors that are related to IS according to the literature in metabolically active tissues plus systemic IS throughout lactation. Conjugated linoleic acids (CLA) reduce milk fat thus decreasing the milk drain of energy and potentially dampening lipolysis, but may also affect IS. Subcutaneous (s.c.) AT and liver from pluriparous cows receiving either control fat or CLA supplement (100 g/day from 1 to 182 days in milk each) were biopsied covering week −3 to 36 relative to parturition. In an additional trial with primiparous cows treated analogously and slaughtered on days in milk 1, 42 or 105, samples from liver, udder, skeletal muscle and 3 visceral and 3 s.c. AT were obtained and assayed for mRNA abundance of adiponectin, its receptors, leptin, leptin receptor, PPARγ, PPARγ2, IL-6, and TNF-α. In pluriparous animals, the mRNA abundance of most of the target genes decreased after parturition in s.c. AT but increased in liver. In primiparous cows, AT depot specific differences were mostly related to retroperitoneal AT; adiponectin receptor 1 and TNF-α were affected predominantly. CLA effects in primiparous cows were largely limited to decreased PPARγ2 mRNA abundance in udder tissue. In pluriparous cows, insulin secretion was increased by CLA resulting in decreased systemic IS but without consistent changes in tissue target mRNA abundance. The temporal gene expression profiles from the adipokines and related receptors support their coactive function in adapting to the needs of lactation. PMID:24465964

  10. Global Proteome Response to Deletion of Genes Related to Mercury Methylation and Dissimilatory Metal Reduction Reveals Changes in Respiratory Metabolism in Geobacter sulfurreducens PCA

    DOE PAGESBeta

    Qian, Chen; Johs, Alexander; Chen, Hongmei; Mann, Benjamin F.; Lu, Xia; Abraham, Paul E.; Hettich, Robert L.; Gu, Baohua

    2016-07-27

    Geobacter sulfurreducens PCA can reduce, sorb, and methylate mercury (Hg); however, the underlying biochemical mechanisms of these processes and interdependent metabolic pathways remain unknown. In this study, shotgun proteomics was used to compare global proteome profiles between wild-type G. sulfurreducens PCA and two mutant strains: a ΔhgcAB mutant, which is deficient in two genes known to be essential for Hg methylation and a ΔomcBESTZ mutant, which is deficient in five outer membrane c-type cytochromes and thus impaired in its ability for dissimilatory metal ion reduction. We were able to delineate the global response of G. sulfurreducens PCA in both mutantsmore » and identify cellular networks and metabolic pathways that were affected by the loss of these genes. Deletion of hgcAB increased the relative abundances of proteins implicated in extracellular electron transfer, including most of the c-type cytochromes, PilA-C, and OmpB, and is consistent with a previously observed increase in Hg reduction in the hgcAB mutant. Deletion of omcBESTZ was found to significantly increase relative abundances of various methyltransferases, suggesting that a loss of dissimilatory reduction capacity results in elevated activity among one-carbon metabolic pathways and thus increased methylation. We show that G. sulfurreducens PCA encodes only the folate branch of the Wood Ljungdahl pathway, and proteins associated with the folate branch were found at lower abundance in the ΔhgcAB mutant strain than the wild type. In conclusion, this observation supports the hypothesis that the function of HgcA and HgcB may be linked to one carbon metabolism through the folate branch of the Wood-Ljungdahl pathway by providing methyl groups required for Hg methylation.« less

  11. Identification and Comparative Study of Chemosensory Genes Related to Host Selection by Legs Transcriptome Analysis in the Tea Geometrid Ectropis obliqua

    PubMed Central

    Bian, Lei; Cai, Xiao-Ming; Luo, Zong-Xiu; Zhang, Yong-Jun; Chen, Zong-Mao

    2016-01-01

    Host selection by female moths is fundamental to the survival of their larvae. Detecting and perceiving the non-volatile chemicals of the plant surface involved in gustatory detection determine the host preference. In many lepidopteran species, tarsal chemosensilla are sensitive to non-volatile chemicals and responsible for taste detection. The tea geometrid Ectropis obliqua is one devastating chewing pest selectively feeding on limited plants, requiring the specialized sensors to forage certain host for oviposition. In present study, we revealed the distribution of chemosensilla in the ventral side of female fifth tarsomere in E. obliqua. To investigate its molecular mechanism of gustatory perception, we performed HiSeq 2500 sequencing of the male- and female- legs transcriptome and identified 24 candidate odorant binding proteins (OBPs), 21 chemosensory proteins (CSPs), 2 sensory neuron membrane proteins (SNMPs), 3 gustatory receptors (GRs) and 4 odorant receptors (ORs). Several leg-specific or enriched chemosensory genes were screened by tissue expression analysis, and clustered with functionally validated genes from other moths, suggesting the potential involvement in taste sensation or other physiological processes. The RPKM value analysis revealed that 9 EoblOBPs showed sex discrepancy in the leg expression, 8 being up-regulated in female and only 1 being over expressed in male. These female-biased EoblOBPs indicated an ecological adaption related with host-seeking and oviposition behaviors. Our work will provide basic knowledge for further studies on the molecular mechanism of gustatory perception, and enlighten a host-selection-based control strategy of insect pests. PMID:26930056

  12. Identification and Comparative Study of Chemosensory Genes Related to Host Selection by Legs Transcriptome Analysis in the Tea Geometrid Ectropis obliqua.

    PubMed

    Ma, Long; Li, Zhao-Qun; Bian, Lei; Cai, Xiao-Ming; Luo, Zong-Xiu; Zhang, Yong-Jun; Chen, Zong-Mao

    2016-01-01

    Host selection by female moths is fundamental to the survival of their larvae. Detecting and perceiving the non-volatile chemicals of the plant surface involved in gustatory detection determine the host preference. In many lepidopteran species, tarsal chemosensilla are sensitive to non-volatile chemicals and responsible for taste detection. The tea geometrid Ectropis obliqua is one devastating chewing pest selectively feeding on limited plants, requiring the specialized sensors to forage certain host for oviposition. In present study, we revealed the distribution of chemosensilla in the ventral side of female fifth tarsomere in E. obliqua. To investigate its molecular mechanism of gustatory perception, we performed HiSeq 2500 sequencing of the male- and female- legs transcriptome and identified 24 candidate odorant binding proteins (OBPs), 21 chemosensory proteins (CSPs), 2 sensory neuron membrane proteins (SNMPs), 3 gustatory receptors (GRs) and 4 odorant receptors (ORs). Several leg-specific or enriched chemosensory genes were screened by tissue expression analysis, and clustered with functionally validated genes from other moths, suggesting the potential involvement in taste sensation or other physiological processes. The RPKM value analysis revealed that 9 EoblOBPs showed sex discrepancy in the leg expression, 8 being up-regulated in female and only 1 being over expressed in male. These female-biased EoblOBPs indicated an ecological adaption related with host-seeking and oviposition behaviors. Our work will provide basic knowledge for further studies on the molecular mechanism of gustatory perception, and enlighten a host-selection-based control strategy of insect pests. PMID:26930056

  13. Effects of low-level laser therapy on the expression of osteogenic genes related in the initial stages of bone defects in rats

    NASA Astrophysics Data System (ADS)

    Fernandes, Kelly Rossetti; Ribeiro, Daniel Araki; Rodrigues, Natália Camargo; Tim, Carla; Santos, Anderson Amaro; Parizotto, Nivaldo Antônio; de Araujo, Heloisa Selistre; Driusso, Patrícia; Rennó, Ana Claudia Muniz

    2013-03-01

    We evaluate the effects of low-level laser therapy (LLLT) on the histological modifications and temporal osteogenic genes expression during the initial phase of bone healing in a model of bone defect in rats. Sixty-four Wistar rats were divided into control and treated groups. Noncritical size bone defects were surgically created at the upper third of the tibia. Laser irradiation (Ga-Al-As laser 830 nm, 30 mW, 0.028 cm2, 1.071 W/cm2, 1 min and 34 s, 2.8 Joules, 100 J/cm2) was performed for 1, 2, 3, and 5 sessions. Histopathology revealed that treated animals presented higher inflammatory cells recruitment, especially 12 and 36 h postsurgery. Also, a better tissue organization at the site of the injury, with the presence of granulation tissue and new bone formation was observed on days three and five postsurgery in the treated animals. The quantitative real time polymerase chain reaction showed that LLLT produced a significantly increase in mRNA expression of Runx-2, 12 h and three days post-surgery, a significant upregulation of alkaline phosphatase mRNA expression after 36 h and three days post-surgery and a significant increase of osteocalcin mRNA expression after three and five days. We concluded that LLLT modulated the inflammatory process and accelerated bone repair, and this advanced repair pattern in the laser-treated groups may be related to the higher mRNA expression of genes presented by these animals.

  14. Early post mortem expression of genes related to tenderization in two Italian Simmental young bulls' skeletal muscles differing in contractile type.

    PubMed

    Saccà, Elena; Corazzin, Mirco; Pizzutti, Nicoletta; Lippe, Giovanna; Piasentier, Edi

    2015-12-01

    The early post mortem expression of eight genes potentially involved in meat ageing process and the tenderness of two Italian Simmental young bulls' (Bos taurus) skeletal muscles differing in their contractile type were evaluated. Samples of Longissimus lumborum (LL) and Infraspinatus (IS) muscles were collected from 17 bulls. The messenger RNA (mRNA) abundances of calpain-1, calpain-2, calpastatin, caspase 3, caspase 9, heat shock protein 27 (Hsp27), Hsp40 and Hsp70 were detected by quantitative PCR. The myosin heavy chain-slow and -fast isoform content, the pH48h and the lipid content of the muscles were in line with the contractile and metabolic type. In comparison with the fast LL, the slow IS showed a lower calpain-1/calpastatin mRNA content ratio after slaughtering and a higher Warner-Bratzler Initial Yield value after 7 days of ageing. Hsp27 and Hsp70 mRNA abundances were significantly lower in LL than IS, highlighting their potential role in the ageing process of bovine muscles. PMID:26152790

  15. Effect of A One-Week Balanced Diet on Expression of Genes Related to Zinc Metabolism and Inflammation in Type 2 Diabetic Patients.

    PubMed

    Lais, Lucia Leite; de Lima Vale, Sancha Helena; Xavier, Camila Alves; de Araujo Silva, Alfredo; Aydemir, Tolunay Beker; Cousins, Robert J

    2016-01-01

    To evaluate the effect of diet on metabolic control and zinc metabolism in patients with type 2 diabetes mellitus (T2DM). One-week balanced diet was provided to 10 Brazilians patients with T2DM. Nutritional assessment, laboratorial parameters and expression of zinc transporter and inflammatory genes in peripheral blood mononuclear cells (PBMC) were performed. Healthy non-diabetic subjects of the same demographic were recruited to provide baseline data. Diabetic patients had higher body mass index and greater fasting plasma glucose, plasma tumor necrosis factor α (TNFα) and plasma interleukin 6 (IL6) levels compared with healthy subjects. In addition, the expression of transporters 4 (ZnT4) mRNA was lower and IL6 mRNA was higher in PBMC of these diabetic patients than in healthy subject. One week after a balanced diet was provided, fasting plasma glucose decreased significantly as did TNFα, IL6 and Metallothionein 1 (MT1) mRNAs. No change was observed in zinc transporter expression in PBMC after the dietary intervention. A healthy eating pattern maintained for one week was able to improve metabolic control of diabetic patients by lowering fasting plasma glucose. This metabolic control may be related to down-regulation of zinc-related transcripts from PBMCs, as TNFα, IL6 and MT1 mRNA. PMID:26839874

  16. Iron Dextran Increases Hepatic Oxidative Stress and Alters Expression of Genes Related to Lipid Metabolism Contributing to Hyperlipidaemia in Murine Model

    PubMed Central

    Silva, Maísa; Guerra, Joyce Ferreira da Costa; Sampaio, Ana Flávia Santos; de Lima, Wanderson Geraldo; Silva, Marcelo Eustáquio

    2015-01-01

    The objective of this study was to investigate the effects of iron dextran on lipid metabolism and to determine the involvement of oxidative stress. Fischer rats were divided into two groups: the standard group (S), which was fed the AIN-93M diet, and the standard plus iron group (SI), which was fed the same diet but also received iron dextran injections. Serum cholesterol and triacylglycerol levels were higher in the SI group than in the S group. Iron dextran was associated with decreased mRNA levels of pparα, and its downstream gene cpt1a, which is involved in lipid oxidation. Iron dextran also increased mRNA levels of apoB-100, MTP, and L-FABP indicating alterations in lipid secretion. Carbonyl protein and TBARS were consistently higher in the liver of the iron-treated rats. Moreover, a significant positive correlation was found between oxidative stress products, lfabp expression, and iron stores. In addition, a negative correlation was found between pparα expression, TBARS, carbonyl protein, and iron stores. In conclusion, our results suggest that the increase observed in the transport of lipids in the bloodstream and the decreased fatty acid oxidation in rats, which was promoted by iron dextran, might be attributed to increased oxidative stress. PMID:25685776

  17. Iron dextran increases hepatic oxidative stress and alters expression of genes related to lipid metabolism contributing to hyperlipidaemia in murine model.

    PubMed

    Silva, Maísa; da Costa Guerra, Joyce Ferreira; Sampaio, Ana Flávia Santos; de Lima, Wanderson Geraldo; Silva, Marcelo Eustáquio; Pedrosa, Maria Lucia

    2015-01-01

    The objective of this study was to investigate the effects of iron dextran on lipid metabolism and to determine the involvement of oxidative stress. Fischer rats were divided into two groups: the standard group (S), which was fed the AIN-93M diet, and the standard plus iron group (SI), which was fed the same diet but also received iron dextran injections. Serum cholesterol and triacylglycerol levels were higher in the SI group than in the S group. Iron dextran was associated with decreased mRNA levels of pparα, and its downstream gene cpt1a, which is involved in lipid oxidation. Iron dextran also increased mRNA levels of apoB-100, MTP, and L-FABP indicating alterations in lipid secretion. Carbonyl protein and TBARS were consistently higher in the liver of the iron-treated rats. Moreover, a significant positive correlation was found between oxidative stress products, lfabp expression, and iron stores. In addition, a negative correlation was found between pparα expression, TBARS, carbonyl protein, and iron stores. In conclusion, our results suggest that the increase observed in the transport of lipids in the bloodstream and the decreased fatty acid oxidation in rats, which was promoted by iron dextran, might be attributed to increased oxidative stress. PMID:25685776

  18. Cloning, expression, and purification of a new antimicrobial peptide gene from Musca domestica larva.

    PubMed

    Pei, Zhihua; Sun, Xiaoning; Tang, Yan; Wang, Kai; Gao, Yunhang; Ma, Hongxia

    2014-10-01

    Musca domestica (Diptera: Muscidae), the housefly, exhibits unique immune defences and can produce antimicrobial peptides upon stimulation with bacteria. Based on the cDNA library constructed using the suppression subtractive hybridization (SSH) method, a 198-bp antimicrobial peptide gene, which we named MDAP-2, was amplified by rapid amplification of cDNA ends (RACE) from M. domestica larvae stimulated with Salmonella pullorum (Enterobacteriaceae: Salmonella). In the present study, the full-length MDAP-2 gene was cloned and inserted into a His-tagged Escherichia coli prokaryotic expression system to enable production of the recombinant peptide. The recombinant MDAP-2 peptide was purified using Ni-NTA HisTrap FF crude column chromatography. The bacteriostatic activity of the recombinant purified MDAP-2 protein was assessed. The results indicated that MDAP-2 had in vitro antibacterial activity against all of the tested Gram- bacteria from clinical isolates, including E. coli (Enterobacteriaceae: Escherichia), one strain of S. pullorum (Enterobacteriaceae: Salmonella), and one strain of Pasteurella multocida. DNA sequencing and BLAST analysis showed that the MDAP-2 antimicrobial peptide gene was not homologous to any other antimicrobial peptide genes in GenBank. The antibacterial mechanisms of the newly discovered MDAP-2 peptide warrant further study. PMID:25020259

  19. Effects of supplementation with branched-chain amino acids to low-protein diets on expression of genes related to lipid metabolism in skeletal muscle of growing pigs.

    PubMed

    Duan, Yehui; Duan, Yangmiao; Li, Fengna; Li, Yinghui; Guo, Qiuping; Ji, Yujiao; Tan, Bie; Li, Tiejun; Yin, Yulong

    2016-09-01

    Branched-chain amino acids (BCAA), including leucine (Leu), isoleucine (Ile), and valine (Val), play critical roles in energy homeostasis and lipid metabolism in addition to their other functions, such as in protein metabolism. This study investigated the effects of different dietary BCAA ratios on the intramuscular fat (IMF) content and fatty acid composition in different location of skeletal muscles, including the longissimus dorsi (LD), biceps femoris (BF), and psoas major (PM) muscles of growing pigs, and also examined the mRNA expression levels of genes involved in lipid metabolism in these muscle tissues. The experiment was performed on 40 growing pigs (Large White × Landrace) with a similar initial weight (9.85 ± 0.35 kg). The pigs were randomly assigned to one of five diets: diet A was a positive control and contained 20 % crude protein (CP) with a Leu:Ile:Val ratio of 1:0.51:0.63 according to the recommendation of the National Research Council (NRC); for diets B to E, the CP level was reduced to 17 %, and the Leu:Ile:Val ratios were 1:1:1, 1:0.75:0.75, 1:0.51:0.63, and 1:0.25:0.25, respectively. No significant difference was observed in the average feed intake and feed efficiency of the pigs fed the low protein diet (17 % CP) with BCAA treatments relative to the positive control. However, there was a tendency for increased feed efficiency of the 1:0.75:0.75 group compared with the 1:1:1 group (P = 0.09). The BCAA ratio of 1:0.75:0.75 (17 % CP) increased the IMF content of BF muscle (P < 0.01). Moreover, varied dietary BCAA supplementation with a reduced protein level had different effects on the fatty acid composition of the LD, BF, and PM muscles. The BCAA ratio of 1:0.51:0.63-1:0.75:0.75 (17 % CP) significantly lowered the ratio of n-6 to n-3 polyunsaturated fatty acid in these muscles compared with the positive control group (20 % CP). This effect was associated with an increase in mRNA expression levels of acetyl-CoA carboxylase

  20. Genes related to growth regulation, DNA repair and apoptosis in an oestrogen receptor-negative (MDA-231) versus an oestrogen receptor-positive (MCF-7) breast tumour cell line.

    PubMed

    Skog, Sven; He, Qimin; Khoshnoud, Reza; Fornander, Tommy; Rutqvist, Lars-Erik

    2004-01-01

    The molecular mechanism(s) behind the development of endocrine resistance in breast cancer remains controversial. Here, we compare the capability of oestrogen receptor (ER)-negative cells (MDA-231) versus ER-positive tamoxifen-sensitive cells (MCF-7) to handle DNA repair, transmit signals from damaged DNA, initiate cell death via apoptosis, and then to control transmitted signals from the cell cycle and to synthesize growth factors and receptors. Genes related to these events were studied by cDNA micro-array. Normal human breast cells (H2F) and human lymphoblastoid tumour cells (CEM) were used as controls. Of the 18 genes investigated, 10 genes showed differences in their expression between the cell types. The ER-negative cells showed higher expressions of BRCA1, BRCA2, cdc2, cyclin B1, cyclin D1, cyclin E, IGFBP-3, TGF-alpha, TGF beta 2 and a lower expression of TGF beta R1. No differences in the expressions of bax, bcl-2, p53, p21 and GADD45 were found between the two cell lines. We found that the ER-negative cells were characterized by: (1) a stimulated expression of growth factors and cell cycle regulation compounds, (2) improved DNA repair capacity, but (3) no change in DNA damage signals and apoptotic pathways. Improved DNA repair capacity of ER-negative cells would have a growth advantage over ER-positive tumours when receiving antitumour therapy. PMID:15192311

  1. Engineering biodegradable and multifunctional peptide-based polymers for gene delivery

    PubMed Central

    2013-01-01

    The complex nature of in vivo gene transfer establishes the need for multifunctional delivery vectors capable of meeting these challenges. An additional consideration for clinical translation of synthetic delivery formulations is reproducibility and scale-up of materials. In this review, we summarize our work over the last five years in developing a modular approach for synthesizing peptide-based polymers. In these materials, bioactive peptides that address various barriers to gene delivery are copolymerized with a hydrophilic backbone of N-(2-hydroxypropyl)methacrylamide (HPMA) using reversible-addition fragmentation chain-transfer (RAFT) polymerization. We demonstrate that this synthetic approach results in well-defined, narrowly-disperse polymers with controllable composition and molecular weight. To date, we have investigated the effectiveness of various bioactive peptides for DNA condensation, endosomal escape, cell targeting, and degradability on gene transfer, as well as the impact of multivalency and polymer architecture on peptide bioactivity. PMID:24156736

  2. Correlation between algogenic effects of calcitonin-gene-related peptide (CGRP) and activation of trigeminal vascular system, in an in vivo experimental model of nitroglycerin-induced sensitization.

    PubMed

    Capuano, Alessandro; Greco, Maria Cristina; Navarra, Pierluigi; Tringali, Giuseppe

    2014-10-01

    The neural mechanism(s) underlying migraine remain poorly defined at present; preclinical and clinical studies show an involvement of CGRP in this disorder. However current evidence pointed out that CGRP does not exert an algogenic action per se, but it is able to mediate migraine pain only if the trigeminal-vascular system is sensitized. The present study was addressed to investigate CGRP-evoked behavior in nitric oxide (NO) sensitized rats, using an experimental model of nitroglycerin induced sensitization of trigeminal system, looking at neuropeptide release from different cerebral areas after the intra-peritoneal (i.p.) administration of NO-donors. CGRP injected into the rat whisker pad did not induce significant changes in face rubbing behavior compared to controls. On the contrary, CGRP injected in animals pre-treated with 10mg/kg nitroglycerin significantly increased the time spent in face rubbing. Nitroglycerin pre-treated animals did not show any rubbing behavior after locally injected saline. Furthermore, the i.p. treatment with nitroglycerin produced an increase of CGRP levels in brainstem and trigeminal ganglia, but not in the hypothalamus and hippocampus. The absolute amounts of CGRP produced in the brainstem were lower compared to those in the trigeminal ganglion; however, after nitroglycerin stimulation the percentage increase was higher in the brainstem. In conclusion, findings presented in this study suggest that CGRP induces a painful behavior in rats only after sensitization of trigeminal system; thus supporting the concept that a genetic as well as acquired predisposition to trigemino- vascular activation represents the neurobiological basis of CGRP nociceptive effects in migraineurs. PMID:24998872

  3. Indole-3-carbinol protects against cisplatin-induced acute nephrotoxicity: role of calcitonin gene-related peptide and insulin-like growth factor-1.

    PubMed

    El-Naga, Reem N; Mahran, Yasmen F

    2016-01-01

    Nephrotoxicity associated with the clinical use of the anticancer drug cisplatin is a limiting problem. Thus, searching for new protective measures is required. Indole-3-carbinol is a powerful anti-oxidant, anti-inflammatory and anti-tumor agent. The present study aimed to investigate the potential protective effect of indole-3-carbinol against cisplatin-induced acute nephrotoxicity in rats. Rats were pre-treated with 20 mg/kg indole-3-carbinol orally before giving cisplatin (7 mg/kg). Cisplatin-induced acute nephrotoxicity was demonstrated where relative kidney weight, BUN and serum creatinine were significantly increased. Increased oxidative stress was evident in cisplatin group where GSH and SOD tissue levels were significantly depleted. Also, lipid peroxidation and NOX-1 were increased as compared to the control. Additionally, renal expression of pro-inflammatory mediators was induced by cisplatin. Cisplatin-induced cell death was shown by increased caspase-3 and decreased expression of EGF, IGF-1 and IGF-1 receptor. Nephrotoxicity, oxidative stress, inflammation and apoptotic effects induced by cisplatin were significantly ameliorated by indole-3-carbinol pre-treatment. Besides, the role of CGRP in cisplatin-induced nephrotoxicity was explored. Furthermore, cisplatin cytotoxic activity was significantly enhanced by indole-3-carbinol pre-treatment in vitro. In conclusion, indole-3-carbinol provides protection against cisplatin-induced nephrotoxicity. Also, reduced expression of CGRP may play a role in the pathogenesis of cisplatin-induced renal injury. PMID:27417335

  4. Comparison of the effects of salmon calcitonin (sCT) and calcitonin gene-related peptide (CGRP) in a number of in vivo and in vitro tests

    SciTech Connect

    Welch, S.P.; Brase, D.; Cooper, C.; Dewey, W.L.

    1986-03-05

    sCT and CGRP have been shown previously to have multiple activities in the central nervous system (CNS). Recent work has shown that CGRP (15 ..mu..g) intraventricularly (IVT) produces a naloxone reversible 37% inhibition in the p-phenylquinone test (PPQ) accompanied by severe diarrhea. The ED50 of sCT in the PPQ test is 362 ng and this effect is not reversed totally by naloxone. The onset of CGRP is more rapid than that of sCT. sCT and CGRP (10/sup -6/M) both produce naloxone reversible inhibition of the electrically stimulated guinea pig ileum (GPI) (25% and 50% respectively). Both sCT and CGRP (10/sup -6/ M) produce contracture (15% and 40% respectively) of the non-stimulated GPI that is not blocked by atropine. Both sCT and CGRP block the naloxone-induced contracture of the morphine (MS04) dependent ilea (29% and 68% respectively). Both sCT and CGRP produce biphasic shifts in the MS04 acetylcholine dose-effect curves in the stimulated and nonstimulated GPI, respectively. Neither sCT nor CGRP (10/sup -9/ to 10/sup -4/ M) displaces /sup 3/H-naloxone binding to mouse brain membranes. Both sCT and CGRP may produce their effects by modulation of CA/sup +2/ fluxes in the CNS and GPI.

  5. Protective role of ellagitannins from Eucalyptus citriodora against ethanol-induced gastric ulcer in rats: impact on oxidative stress, inflammation and calcitonin-gene related peptide.

    PubMed

    Al-Sayed, Eman; El-Naga, Reem N

    2015-01-15

    The gastroprotective activity of an ellagitannin-rich fraction obtained from Eucalyptus citriodora (ECF) was investigated against ethanol-induced gastric ulceration in rats. The rats were pretreated with ECF (25, 50 and 100mg/kg) 1h before the administration of absolute ethanol to induce acute gastric ulceration. The gastric lesions were significantly reduced by all doses of ECF. Notably, pre-treatment with ECF (100mg/kg) conferred 99.6% gastroprotection, which is significantly higher than that produced by omeprazole. Moreover, ECF administration markedly increased the mucin content in a dose-dependent manner. The potent gastroprotective effect of ECF could be partly mediated by attenuating ethanol-induced oxidative stress. ECF-pre-treatment markedly increased the depleted GSH and SOD levels in a dose-dependent manner. Moreover, ECF significantly decreased the elevated MDA tissue levels induced by ethanol administration. The results demonstrated that ECF administration exerted a powerful anti-inflammatory activity as evidenced by the reduction in the pro-inflammatory markers; IL-1β, TNF-α, 5-LO and COX-2. Additionally, the caspase-3 tissue levels were significantly reduced in the groups pre-treated with ECF. These results suggest that ECF could exert a beneficial gastroprotective effect through their antioxidant, anti-inflammatory and anti-apoptotic properties. Furthermore, ECF pre-treatment significantly attenuated the ethanol-induced decrease in CGRP expression, which has a protective role against gastric ulceration. Histopathological examination revealed intact mucosal layer, absence of hemorrhage and necrosis in groups treated with ECF. Ellagitannins were identified as the major active constituents responsible for the marked antioxidant and gastroprotective properties of ECF. The HPLC-PDA-ESI/MS/MS technique was employed to identify the ellagitannins of E. citriodora. PMID:25636864

  6. Indole-3-carbinol protects against cisplatin-induced acute nephrotoxicity: role of calcitonin gene-related peptide and insulin-like growth factor-1

    PubMed Central

    El-Naga, Reem N.; Mahran, Yasmen F.

    2016-01-01

    Nephrotoxicity associated with the clinical use of the anticancer drug cisplatin is a limiting problem. Thus, searching for new protective measures is required. Indole-3-carbinol is a powerful anti-oxidant, anti-inflammatory and anti-tumor agent. The present study aimed to investigate the potential protective effect of indole-3-carbinol against cisplatin-induced acute nephrotoxicity in rats. Rats were pre-treated with 20 mg/kg indole-3-carbinol orally before giving cisplatin (7 mg/kg). Cisplatin-induced acute nephrotoxicity was demonstrated where relative kidney weight, BUN and serum creatinine were significantly increased. Increased oxidative stress was evident in cisplatin group where GSH and SOD tissue levels were significantly depleted. Also, lipid peroxidation and NOX-1 were increased as compared to the control. Additionally, renal expression of pro-inflammatory mediators was induced by cisplatin. Cisplatin-induced cell death was shown by increased caspase-3 and decreased expression of EGF, IGF-1 and IGF-1 receptor. Nephrotoxicity, oxidative stress, inflammation and apoptotic effects induced by cisplatin were significantly ameliorated by indole-3-carbinol pre-treatment. Besides, the role of CGRP in cisplatin-induced nephrotoxicity was explored. Furthermore, cisplatin cytotoxic activity was significantly enhanced by indole-3-carbinol pre-treatment in vitro. In conclusion, indole-3-carbinol provides protection against cisplatin-induced nephrotoxicity. Also, reduced expression of CGRP may play a role in the pathogenesis of cisplatin-induced renal injury. PMID:27417335

  7. In Vivo Effect of Arsenic Trioxide on Keap1-p62-Nrf2 Signaling Pathway in Mouse Liver: Expression of Antioxidant Responsive Element-Driven Genes Related to Glutathione Metabolism

    PubMed Central

    Srivastava, Ritu; Sengupta, Archya; Mukherjee, Sandip; Chatterjee, Sarmishtha; Sudarshan, Muthammal; Chakraborty, Anindita; Bhattacharya, Shelley; Chattopadhyay, Ansuman

    2013-01-01

    Arsenic is a Group I human carcinogen, and chronic arsenic exposure through drinking water is a major threat to human population. Liver is one of the major organs for the detoxification of arsenic. The present study was carried out in mice in vivo after arsenic treatment through drinking water at different doses and time of exposure. Arsenic toxicity is found to be mediated by reactive oxygen species. Nuclear factor (erythroid-2 related) factor 2 (Nrf2)/Keap1 (Kelch-like ECH-associated protein 1)/ARE (antioxidant response element)—driven target gene system protects cells against oxidative stress and maintains cellular oxidative homeostasis. Our result showed 0.4 ppm, 2 ppm, and 4 ppm arsenic trioxide treatment through drinking water for 30 days and 90 days induced damages in the liver of Swiss albino mice as evidenced by histopathology, disturbances in liver function, induction of heat shock protein 70, modulation of trace elements, alteration in reduced glutathione level, glutathione-s-transferase and catalase activity, malondialdehyde production, and induction of apoptosis. Cellular Nrf2 protein level and mRNA level increased in all treatment groups. Keap1 protein as well as mRNA level decreased concomitantly in arsenic treated mice. Our study clearly indicates the important role of Nrf2 in activating ARE driven genes related to GSH metabolic pathway and also the adaptive response mechanisms in arsenic induced hepatotoxicity. PMID:27335833

  8. Vcsa1 gene peptides for the treatment of inflammatory and allergic reactions.

    PubMed

    Morris, Katherine; Kuo, Byron; Wilkinson, Mark D; Davison, Joseph S; Befus, A Dean; Mathison, Ronald D

    2007-06-01

    The recently emerged Vcsa1 gene is one member of the variable coding sequence (VCS) multigene family of Rattus norvegicus. This gene encodes the precursor prohormone SMR1 (submandibular rat-1), which on enzymatic processing gives rise to several 5 to 11 amino acid peptides that modulate a variety of physiological functions. The analgesic pentapeptide sialorphin and anti-inflammatory heptapeptide submandibular gland peptide-T (TDIFEGG) are the most intensively studied. Although the Vcsa1 gene and its protein product are unique to rats, TDIFEGG or a derivative acts on all species examined to date, including human cells, in functions related to allergic reactions and inflammation. In this review, the patent and academic literature on SMR1 and its natural peptides and their derivatives are reviewed for consideration of biological targets and relevance to the development of novel therapeutic agents. The VCS gene family is discussed and we speculate on possible human homologs of these potent anti-inflammatory rat-derived peptides. The biologically active peptide products of SMR1 are considered and the mechanism of action and structure-activity relationships of the anti-inflammatory submandibular gland peptide-T and its derivatives are discussed. PMID:19075974

  9. Isolation, characterization, and expression analyses of plant elicitor peptides (pep) genes in maize

    Technology Transfer Automated Retrieval System (TEKTRAN)

    PROPEP1, PROPEP 2, and PROPEP3 genes appear to have roles in a feedback loop that amplifies defense signaling pathways initiated by pathogens. We present evidence to support the role of peptides derived from PROPEP genes as endogenous elicitors that are generated in response to pathogens. The preval...

  10. Expression of proglucagon and proglucagon-derived peptide hormone receptor genes in the chicken

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To better understand how the proglucagon system functions in birds, we utilized a molecular cloning strategy to sequence and characterize the chicken proglucagon gene that encodes glucagon, glucagon-like peptide(GLP)-1 and GLP-2. This gene has seven exons and six introns with evidence for an additi...

  11. Toxic peptides and genes encoding toxin gamma of the Brazilian scorpions Tityus bahiensis and Tityus stigmurus.

    PubMed

    Becerril, B; Corona, M; Coronas, F I; Zamudio, F; Calderon-Aranda, E S; Fletcher, P L; Martin, B M; Possani, L D

    1996-02-01

    Seven toxic peptides from the venom of Tityus bahiensis and Tityus stigmurus was isolated and sequenced, five of them to completion. The most abundant peptide from each of these two species of scorpion was 95% identical with that of toxin gamma from the venom of Tityus serrulatus. They were consequently named gamma-b and gamma-st respectively. The genes encoding these new gamma-like peptides were cloned and sequenced by utilizing oligonucleotides synthesized according to known cDNA sequences of toxin gamma, and amplified by PCR on templates of DNA purified from both T. bahiensis and T. stigmurus. They contain an intron of approx. 470 bp. Possible mechanisms of processing and expressing these peptides are discussed, in view of the fact that glycine is the first residue of the N-terminal sequence of T. stigmurus, whereas lysine is the residue at position 1 of toxin gamma from T. serrulatus and T. bahiensis. In addition, chemical characterization of the less abundant toxic peptides showed the presence of at least four distinct families of peptides in all three species of the genus Tityus studied. There is a large degree of similarity among peptides from different venoms of the same family. By using specific horse and rabbit antisera, the venoms of T. bahiensis, T. serrulatus and T. stigmurus were compared. They showed an extended degree of cross-reactivity. Thus these three species of scorpion have similar toxic components, the genes of which are similarly organized, processed and expressed. PMID:8611151

  12. Predicting functional regulatory SNPs in the human antimicrobial peptide genes DEFB1 and CAMP in tuberculosis and HIV/AIDS.

    PubMed

    Flores Saiffe Farías, Adolfo; Jaime Herrera López, Enrique; Moreno Vázquez, Cristopher Jorge; Li, Wentian; Prado Montes de Oca, Ernesto

    2015-12-01

    Single nucleotide polymorphisms (SNPs) in transcription factor binding sites (TFBSs) within gene promoter region or enhancers can modify the transcription rate of genes related to complex diseases. These SNPs can be called regulatory SNPs (rSNPs). Data compiled from recent projects, such as the 1000 Genomes Project and ENCODE, has revealed essential information used to perform in silico prediction of the molecular and biological repercussions of SNPs within TFBS. However, most of these studies are very limited, as they only analyze SNPs in coding regions or when applied to promoters, and do not integrate essential biological data like TFBSs, expression profiles, pathway analysis, homotypic redundancy (number of TFBSs for the same TF in a region), chromatin accessibility and others, which could lead to a more accurate prediction. Our aim was to integrate different data in a biologically coherent method to analyze the proximal promoter regions of two antimicrobial peptide genes, DEFB1 and CAMP, that are associated with tuberculosis (TB) and HIV/AIDS. We predicted SNPs within the promoter regions that are more likely to interact with transcription factors (TFs). We also assessed the impact of homotypic redundancy using a novel approach called the homotypic redundancy weight factor (HWF). Our results identified 10 SNPs, which putatively modify the binding affinity of 24 TFs previously identified as related to TB and HIV/AIDS expression profiles (e.g. KLF5, CEBPA and NFKB1 for TB; FOXP2, BRCA1, CEBPB, CREB1, EBF1 and ZNF354C for HIV/AIDS; and RUNX2, HIF1A, JUN/AP-1, NR4A2, EGR1 for both diseases). Validating with the OregAnno database and cell-specific functional/non functional SNPs from additional 13 genes, our algorithm performed 53% sensitivity and 84.6% specificity to detect functional rSNPs using the DNAseI-HUP database. We are proposing our algorithm as a novel in silico method to detect true functional rSNPs in antimicrobial peptide genes. With further

  13. Cationic polymethacrylates with covalently linked membrane destabilizing peptides as gene delivery vectors.

    PubMed

    Funhoff, Arjen M; van Nostrum, Cornelus F; Lok, Martin C; Kruijtzer, John A W; Crommelin, Daan J A; Hennink, Wim E

    2005-01-01

    A membrane-disrupting peptide derived from the influenza virus was covalently linked to different polymethacrylates (pDMAEMA, pDAMA and the degradable pHPMA-DMAE, monomers depicted in Fig. 1) using N-succinimidyl 3-(2-pyridyldithio)propionate (SPDP) as coupling agent to increase the transfection efficiency of polyplexes based on these polymers. It was shown by circular dichroism (CD) measurements that the polymer-conjugated peptide was, as the free peptide, able to undergo a conformational change of a random coil to an alpha helix upon lowering the pH to 5.0. This indicates that the property of the peptide to destabilize the endosomal membrane was preserved after its conjugation to the cationic polymers. In line herewith, a liposome leakage assay revealed that the polymer-bound peptide has comparable activity as the free peptide. The DNA condensing properties of the synthesized polymer-peptide conjugates were studied with dynamic light scattering and zeta-potential measurements, and it was shown that small (100 to 250 nm), positively charged (+15 to +20 mV) particles were formed. In vitro transfection and toxicity was tested in COS-7 cells, and these experiments showed that the polyplexes with grafted peptide had a substantially higher transfection activity than the control polyplexes, while the toxicity remained unchanged. Cellular uptake of the polyplexes was visualized with confocal laser scanning microscopy, and no differences in cellular uptake could be determined between the peptide containing systems and the control formulation. This shows that the increased transfection activity is indeed due to a better endosomal escape of the peptide grafted polyplexes. This study demonstrates that it is possible to covalently conjugate an endosome disruptive peptide to cationic gene delivery polymers with preservation of its membrane destabilization activity, making these conjugates suitable for in vivo DNA delivery. PMID:15588908

  14. The nonribosomal peptide and polyketide synthetic gene clusters in two strains of entomopathogenic fungi in Cordyceps.

    PubMed

    Wang, Wen-Jing; Vogel, Heiko; Yao, Yi-Jian; Ping, Liyan

    2012-11-01

    Species of Cordyceps Fr. are entomopathogenic fungi that parasitize the larvae or pupae of lepidopteran insects. The secondary metabolites, nonribosomal peptides and polyketides are well-known mediators of pathogenesis. The biosynthetic gene clusters of these compounds in two fungal strains (1630 and DSM 1153) formerly known as Cordyceps militaris were screened using polymerase chain reaction with degenerate primers. Two nonribosomal peptide synthetase genes, one polyketide synthetase gene and one hybrid gene cluster were identified, and certain characteristics of the structures of their potential products were predicted. All four genes were actively expressed under laboratory conditions but at markedly different levels. The gene clusters from the two fungal strains were structurally and functionally unrelated, suggesting different evolutionary origins and physiological functions. Phylogenetic and biochemical analyses confirmed that the two fungal strains are not conspecific as currently assigned. PMID:22889355

  15. BAGEL3: automated identification of genes encoding bacteriocins and (non-)bactericidal posttranslationally modified peptides

    PubMed Central

    van Heel, Auke J.; de Jong, Anne; Montalbán-López, Manuel; Kok, Jan; Kuipers, Oscar P.

    2013-01-01

    Identifying genes encoding bacteriocins and ribosomally synthesized and posttranslationally modified peptides (RiPPs) can be a challenging task. Especially those peptides that do not have strong homology to previously identified peptides can easily be overlooked. Extensive use of BAGEL2 and user feedback has led us to develop BAGEL3. BAGEL3 features genome mining of prokaryotes, which is largely independent of open reading frame (ORF) predictions and has been extended to cover more (novel) classes of posttranslationally modified peptides. BAGEL3 uses an identification approach that combines direct mining for the gene and indirect mining via context genes. Especially for heavily modified peptides like lanthipeptides, sactipeptides, glycocins and others, this genetic context harbors valuable information that is used for mining purposes. The bacteriocin and context protein databases have been updated and it is now easy for users to submit novel bacteriocins or RiPPs. The output has been simplified to allow user-friendly analysis of the results, in particular for large (meta-genomic) datasets. The genetic context of identified candidate genes is fully annotated. As input, BAGEL3 uses FASTA DNA sequences or folders containing multiple FASTA formatted files. BAGEL3 is freely accessible at http://bagel.molgenrug.nl. PMID:23677608

  16. Upregulation of genes related to bone formation by γ-amino butyric acid and γ-oryzanol in germinated brown rice is via the activation of GABAB-receptors and reduction of serum IL-6 in rats

    PubMed Central

    Muhammad, Sani Ismaila; Maznah, Ismail; Mahmud, Rozi; Zuki, Abu Bakar Zakaria; Imam, Mustapha Umar

    2013-01-01

    Background Osteoporosis and other bone degenerative diseases are among the most challenging non-communicable diseases to treat. Previous works relate bone loss due to osteoporosis with oxidative stress generated by free radicals and inflammatory cytokines. Alternative therapy to hormone replacement has been an area of interest to researchers for almost three decades due to hormone therapy-associated side effects. Methods In this study, we investigated the effects of gamma-amino butyric acid (GABA), gamma-oryzanol (ORZ), acylated steryl glucosides (ASG), and phenolic extracts from germinated brown rice (GBR) on the expression of genes related to bone metabolism, such as bone morphogenic protein-2 (BMP-2), secreted protein acidic and rich in cysteine (SPARC), runt-related transcription factor 2 (RUNX-2), osteoblast-specific transcription factor osterix (Osx), periostin, osteoblast specific factor (Postn), collagen 1&2 (Col1&2), calcitonin receptor gene (CGRP); body weight measurement and also serum interleukin-6 (IL-6) and osteocalcin, in serum and bone. Rats were treated with GBR, ORZ, GABA, and ASG at (100 and 200 mg/kg); estrogen (0.2 mg/kg), or remifemin (10 and 20 mg/kg), compared to ovariectomized non-treated group as well as non-ovariectomized non-treated (sham) group. Enzyme-linked immunosorbent assay was used to measure the IL-6 and osteocalcin levels at week 2, 4, and 8, while the gene expression in the bone tissue was determined using the Genetic Analysis System (Beckman Coulter Inc., Brea, CA, USA). Results The results indicate that groups treated with GABA (100 and 200 mg/kg) showed significant upregulation of SPARC, calcitonin receptor, and BMP-2 genes (P < 0.05), while the ORZ-treated group (100 and 200 mg/kg) revealed significant (P < 0.05) upregulation of Osx, Postn, RUNX-2, and Col1&2. Similarly, IL-6 concentration decreased, while osteocalcin levels increased significantly (P < 0.05) in the treated groups as compared to ovariectomized non

  17. Gene duplication of the human peptide YY gene (PYY) generated the pancreatic polypeptide gene (PPY) on chromosome 17q21.1

    SciTech Connect

    Hort, Y.; Shine, J.; Herzog, H.

    1995-03-01

    Neuropeptide Y (NPY), peptide YY (PYY), and pancreatic polypeptide (PP) are structurally related but functionally diverse peptides, encoded by separate genes and expressed in different tissues. Although the human NPY gene has been mapped to chromosome 7, the authors demonstrate here that the genes for human PYY and PP (PPY) are localized only 10 kb apart from each another on chromosome 17q21.1. The high degree of homology between the members of this gene family, both in primary sequence and exon/intron structure, suggests that the NYP and the PYY genes arose from an initial gene duplication event, with a subsequent tandem duplication of the PYY gene being responsible for the creation of the PPY gene. A second weaker hybridization signal also found on chromosome 17q11 and results obtained by Southern blot analysis suggest that the entire PYY-PPY region has undergone a further duplication event. 27 refs., 5 figs.

  18. Peptide nanofibrils boost retroviral gene transfer and provide a rapid means for concentrating viruses

    NASA Astrophysics Data System (ADS)

    Yolamanova, Maral; Meier, Christoph; Shaytan, Alexey K.; Vas, Virag; Bertoncini, Carlos W.; Arnold, Franziska; Zirafi, Onofrio; Usmani, Shariq M.; Müller, Janis A.; Sauter, Daniel; Goffinet, Christine; Palesch, David; Walther, Paul; Roan, Nadia R.; Geiger, Hartmut; Lunov, Oleg; Simmet, Thomas; Bohne, Jens; Schrezenmeier, Hubert; Schwarz, Klaus; Ständker, Ludger; Forssmann, Wolf-Georg; Salvatella, Xavier; Khalatur, Pavel G.; Khokhlov, Alexei R.; Knowles, Tuomas P. J.; Weil, Tanja; Kirchhoff, Frank; Münch, Jan

    2013-02-01

    Inefficient gene transfer and low virion concentrations are common limitations of retroviral transduction. We and others have previously shown that peptides derived from human semen form amyloid fibrils that boost retroviral gene delivery by promoting virion attachment to the target cells. However, application of these natural fibril-forming peptides is limited by moderate efficiencies, the high costs of peptide synthesis, and variability in fibril size and formation kinetics. Here, we report the development of nanofibrils that self-assemble in aqueous solution from a 12-residue peptide, termed enhancing factor C (EF-C). These artificial nanofibrils enhance retroviral gene transfer substantially more efficiently than semen-derived fibrils or other transduction enhancers. Moreover, EF-C nanofibrils allow the concentration of retroviral vectors by conventional low-speed centrifugation, and are safe and effective, as assessed in an ex vivo gene transfer study. Our results show that EF-C fibrils comprise a highly versatile, convenient and broadly applicable nanomaterial that holds the potential to significantly facilitate retroviral gene transfer in basic research and clinical applications.

  19. Gene introduction into the mitochondria of Arabidopsis thaliana via peptide-based carriers

    NASA Astrophysics Data System (ADS)

    Chuah, Jo-Ann; Yoshizumi, Takeshi; Kodama, Yutaka; Numata, Keiji

    2015-01-01

    Available methods in plant genetic transformation are nuclear and plastid transformations because similar procedures have not yet been established for the mitochondria. The double membrane and small size of the organelle, in addition to its large population in cells, are major obstacles in mitochondrial transfection. Here we report the intracellular delivery of exogenous DNA localized to the mitochondria of Arabidopsis thaliana using a combination of mitochondria-targeting peptide and cell-penetrating peptide. Low concentrations of peptides were sufficient to deliver DNA into the mitochondria and expression of imported DNA reached detectable levels within a short incubation period (12 h). We found that electrostatic interaction with the cell membrane is not a critical factor for complex internalization, instead, improved intracellular penetration of mitochondria-targeted complexes significantly enhanced gene transfer efficiency. Our results delineate a simple and effective peptide-based method, as a starting point for the development of more sophisticated plant mitochondrial transfection strategies.

  20. Gene introduction into the mitochondria of Arabidopsis thaliana via peptide-based carriers

    PubMed Central

    Chuah, Jo-Ann; Yoshizumi, Takeshi; Kodama, Yutaka; Numata, Keiji

    2015-01-01

    Available methods in plant genetic transformation are nuclear and plastid transformations because similar procedures have not yet been established for the mitochondria. The double membrane and small size of the organelle, in addition to its large population in cells, are major obstacles in mitochondrial transfection. Here we report the intracellular delivery of exogenous DNA localized to the mitochondria of Arabidopsis thaliana using a combination of mitochondria-targeting peptide and cell-penetrating peptide. Low concentrations of peptides were sufficient to deliver DNA into the mitochondria and expression of imported DNA reached detectable levels within a short incubation period (12 h). We found that electrostatic interaction with the cell membrane is not a critical factor for complex internalization, instead, improved intracellular penetration of mitochondria-targeted complexes significantly enhanced gene transfer efficiency. Our results delineate a simple and effective peptide-based method, as a starting point for the development of more sophisticated plant mitochondrial transfection strategies. PMID:25583214

  1. Secreted antiviral entry inhibitory (SAVE) peptides for gene therapy of HIV infection.

    PubMed

    Egerer, Lisa; Volk, Andreas; Kahle, Joerg; Kimpel, Janine; Brauer, Frances; Hermann, Felix G; von Laer, Dorothee

    2011-07-01

    Gene therapeutic strategies for human immunodeficiency virus type 1 (HIV-1) infection could potentially overcome the limitations of standard antiretroviral drug therapy (ART). However, in none of the clinical gene therapy trials published to date, therapeutic levels of genetic protection have been achieved in the target cell population for HIV-1. To improve systemic antiviral efficacy, C peptides, which are efficient inhibitors of HIV-1 entry, were engineered for high-level secretion by genetically modified cells. The size restrictions for efficient peptide export through the secretory pathway were overcome by expressing the C peptides as concatemers, which were processed into monomers by furin protease cleavage. These secreted antiviral entry inhibitory (SAVE) peptides mediated a substantial protective bystander effect on neighboring nonmodified cells, thus suppressing virus replication even if only a small fraction of cells was genetically modified. Accordingly, these SAVE peptides may provide a strong benefit to AIDS patients in future, and, if applied by direct in vivo gene delivery, could present an effective alternative to antiretroviral drug regimen. PMID:21364540

  2. Efficient Gene Delivery of Primary Human Cells Using Peptide Linked Polyethylenimine Polymer Hybrid

    PubMed Central

    Dey, Devaveena; Inayathullah, Mohammed; Lee, Andrew S; Limiuex, Melbes; Zhang, Xuexiang; Wu, Yi; Nag, Divya; De Almeida, Patricia Eliza; Han, Leng; Rajadas, Jayakumar; Wu, Joseph C

    2011-01-01

    Polyethylenimine (PEI) based polymers are efficient agents for cell transfection. However, their use has been hampered due to high cell death associated with transfection thereby resulting in low efficiency of gene delivery within the cells. To circumvent the problem of cellular toxicity, metal binding peptides were linked to PEI. Eight peptide-PEI derivatives were synthesized to improve cell survival and transfection efficiency. TAT linked PEI was used as a control polymer. Peptides linked with PEI amines formed nano gels as shown by electron microscopy and atomic force microscopic measurements. Polymers were characterized by spectroscopic methods and their ability to form complexes with plasmids was tested using electrophoretic studies. These modifications improved polymer biocompatibility as well as cell survival markedly when compared to PEI alone. A subset of the modified peptide-polymers also showed significantly higher transfection efficiency in primary human cells with respect to the widely used transfection agent, lipofectamine. Study of the underlying mechanism of the observed phenomena revealed lower levels of ‘reactive oxygen species’ (ROS) in presence of the peptide-polymers when compared to PEI alone. This was further corroborated with global gene expression analysis which showed upregulation of multiple genes and pathways involved in regulating intracellular oxidative stress. PMID:21477858

  3. Structure of the gene for porcine peptide antibiotic PR-39, a cathelin gene family member: comparative mapping of the locus for the human peptide antibiotic FALL-39.

    PubMed Central

    Gudmundsson, G H; Magnusson, K P; Chowdhary, B P; Johansson, M; Andersson, L; Boman, H G

    1995-01-01

    PR-39 is a porcine 39-aa peptide antibiotic composed of 49% proline and 24% arginine, with an activity against Gram-negative bacteria comparable to that of tetracycline. In Escherichia coli, it inhibits DNA and protein synthesis. PR-39 was originally isolated from pig small intestine, but subsequent cDNA cloning showed that the gene is expressed in the bone marrow. The open reading frame of the clone showed that PR-39 is made as 173-aa precursor whose proregion belongs to the cathelin family. The PR39 gene, which is rather compact and spans only 1784 bp has now been sequenced. The coding information is split into four exons. The first exon contains the signal sequence of 29 residues and the first 37 residues of the cathelin propart. Exons 2 and 3 contain only cathelin information, while exon 4 codes for the four C-terminal cathelin residues and the mature PR-39 peptide extended by three residues. The sequenced upstream region (1183 bp) contains four potential recognition sites for NF-IL6 and three for APRF, transcription factors known to regulate genes for both cytokines and acute phase response factors. Genomic hybridizations revealed a fairly high level of restriction fragment length polymorphism and indicated that there are at least two copies of the PR39 gene in the pig genome. PR39 was mapped to pig chromosome 13 by linkage and in situ hybridization mapping. The gene for the human peptide antibiotic FALL-39 (also a member of the cathelin family) was mapped to human chromosome 3, which is homologous to pig chromosome 13. Images Fig. 2 Fig. 3 PMID:7624374

  4. Steroid induction of a peptide hormone gene leads to orchestration of a defined behavioral sequence.

    PubMed

    Zitnan, D; Ross, L S; Zitnanova, I; Hermesman, J L; Gill, S S; Adams, M E

    1999-07-01

    At the end of each molt, insects shed the old cuticle by performing preecdysis and ecdysis behaviors. Regulation of these centrally patterned movements involves peptide signaling between endocrine Inka cells and the CNS. In Inka cells, we have identified the cDNA and gene encoding preecdysis-triggering hormone (PETH) and ecdysis-triggering hormone (ETH), which activate these behaviors. Prior to behavioral onset, rising ecdysteroid levels induce expression of the ecdysone receptor (EcR) and ETH gene in Inka cells and evoke CNS sensitivity to PETH and ETH. Subsequent ecdysteroid decline is required for peptide release, which initiates three motor patterns in specific order: PETH triggers preecdysis I, while ETH activates preecdysis II and ecdysis. The Inka cell provides a model for linking steroid regulation of peptide hormone expression and release with activation of a defined behavioral sequence. PMID:10433264

  5. Airway Epithelial Cells are the Site of Expression of a Mammalian Antimicrobial Peptide Gene

    NASA Astrophysics Data System (ADS)

    Diamond, Gill; Jones, Douglas E.; Bevins, Charles L.

    1993-05-01

    We previously reported the isolation and characterization of a broad-spectrum antimicrobial peptide from the bovine tracheal mucosa, which we called tracheal antimicrobial peptide (TAP). We now show the TAP gene is expressed throughout the adult conducting airway, from nasal to bronchiolar tissue, but not in tissues other than airway mucosa, as determined by Northern blot analysis. In situ hybridization of airway sections localizes TAP mRNA to columnar cells of the pseudostratified epithelium. We report the structural organization of the TAP gene and show that TAP is a member of a large family of related sequences with high nucleotide identity in the 5'exon. The data support the hypothesis that antimicrobial peptides contribute to host defense of the respiratory tract.

  6. Inhibition of Growth and Gene Expression by PNA-peptide Conjugates in Streptococcus pyogenes

    PubMed Central

    Patenge, Nadja; Pappesch, Roberto; Krawack, Franziska; Walda, Claudia; Mraheil, Mobarak Abu; Jacob, Anette; Hain, Torsten; Kreikemeyer, Bernd

    2013-01-01

    While Streptococcus pyogenes is consistently susceptible toward penicillin, therapeutic failure of penicillin treatment has been reported repeatedly and a considerable number of patients exhibit allergic reactions to this substance. At the same time, streptococcal resistance to alternative antibiotics, e.g., macrolides, has increased. Taken together, these facts demand the development of novel therapeutic strategies. In this study, S. pyogenes growth was inhibited by application of peptide-conjugated antisense-peptide nucleic acids (PNAs) specific for the essential gyrase A gene (gyrA). Thereby, HIV-1 Tat peptide-coupled PNAs were more efficient inhibitors of streptococcal growth as compared with (KFF)3K-coupled PNAs. Peptide-anti-gyrA PNAs decreased the abundance of gyrA transcripts in S. pyogenes. Growth inhibition by antisense interference was enhanced by combination of peptide-coupled PNAs with protein-level inhibitors. Antimicrobial synergy could be detected with levofloxacin and novobiocin, targeting the gyrase enzyme, and with spectinomycin, impeding ribosomal function. The prospective application of carrier peptide-coupled antisense PNAs in S. pyogenes covers the use as an antimicrobial agent and the employment as a knock-down strategy for the investigation of virulence factor function. PMID:24193033

  7. Efficient Gene Knockdown in Mouse Oocytes through Peptide Nanoparticle-Mediated SiRNA Transfection.

    PubMed

    Jin, Zhen; Li, Ruichao; Zhou, Chunxiang; Shi, Liya; Zhang, Xiaolan; Yang, Zhixia; Zhang, Dong

    2016-01-01

    The use of mouse oocytes as a model for studying female meiosis is very important in reproductive medicine. Gene knockdown by specific small interfering RNA (siRNA) is usually the first step in the study of the function of a target gene in mouse oocytes during in vitro maturation. Traditionally, the only way to introduce siRNA into mouse oocytes is through microinjection, which is certainly less efficient and strenuous than siRNA transfection in somatic cells. Recently, in research using somatic cells, peptide nanoparticle-mediated siRNA transfection has been gaining popularity over liposome nanoparticle-mediated methods because of its high efficiency, low toxicity, good stability, and strong serum compatibility. However, no researchers have yet tried transfecting siRNA into mouse oocytes because of the existence of the protective zona pellucida surrounding the oocyte membrane (vitelline membrane). We therefore tested whether peptide nanoparticles can introduce siRNA into mouse oocytes. In the present study, we showed for the first time that our optimized program can efficiently knock down a target gene with high specificity. Furthermore, we achieved the expected meiotic phenotypes after we knocked down a test unknown target gene TRIM75. We propose that peptide nanoparticles may be superior for preliminary functional studies of unknown genes in mouse oocytes. PMID:26974323

  8. Efficient Gene Knockdown in Mouse Oocytes through Peptide Nanoparticle-Mediated SiRNA Transfection

    PubMed Central

    Jin, Zhen; Li, Ruichao; Zhou, Chunxiang; Shi, Liya; Zhang, Xiaolan; Yang, Zhixia; Zhang, Dong

    2016-01-01

    The use of mouse oocytes as a model for studying female meiosis is very important in reproductive medicine. Gene knockdown by specific small interfering RNA (siRNA) is usually the first step in the study of the function of a target gene in mouse oocytes during in vitro maturation. Traditionally, the only way to introduce siRNA into mouse oocytes is through microinjection, which is certainly less efficient and strenuous than siRNA transfection in somatic cells. Recently, in research using somatic cells, peptide nanoparticle-mediated siRNA transfection has been gaining popularity over liposome nanoparticle-mediated methods because of its high efficiency, low toxicity, good stability, and strong serum compatibility. However, no researchers have yet tried transfecting siRNA into mouse oocytes because of the existence of the protective zona pellucida surrounding the oocyte membrane (vitelline membrane). We therefore tested whether peptide nanoparticles can introduce siRNA into mouse oocytes. In the present study, we showed for the first time that our optimized program can efficiently knock down a target gene with high specificity. Furthermore, we achieved the expected meiotic phenotypes after we knocked down a test unknown target gene TRIM75. We propose that peptide nanoparticles may be superior for preliminary functional studies of unknown genes in mouse oocytes. PMID:26974323

  9. Silk-Based Gene Carriers with Cell Membrane-Destabilizing Peptides

    PubMed Central

    Numata, Keiji; Kaplan, David L

    2010-01-01

    Complexes of recombinant silk-polylysine molecules with ppTG1 peptide, a lysine-rich cell membrane-destabilizing peptide to bind plasmid DNA (pDNA), are designed as less-cytotoxic and highly efficient gene carriers. The peptide destabilizes the cell membrane and promotes gene transfer. Our particular interest is in how ppTG1 enhances transfection efficiency of the silk-based delivery system into human cells. Genetically engineered silk proteins containing polylysine and the monomeric and dimeric ppTG1 sequences are synthesized in Escherichia coli, followed by transfection experiments. The pDNA complexes of Silk-polylysine-ppTG1 dimer recombinant proteins prepared at an N/P 2 (the ratio of number of amines/ phosphates from pDNA) shows the highest transfection efficiency into human embryonic kidney (HEK) cells, the level of which is comparable to the transfection reagent Lipofectamine 2000. The assemblies show a globular morphology with an average hydrodynamic diameter of 99 nm and almost no beta-sheet structure. Additionally, the silk-based pDNA complexes demonstrate excellent DNase resistance as well as efficient release of the pDNA by enzymes that degrade silk proteins. Also, comparison with beta-sheet induced silk-based pDNA complexes indicates that the beta-sheet structure content of the silk sequence of the pDNA complexes controls the enzymatic degradation rate of the complexes, and hence can regulate the release profile of genes from the complexes. The bioengineered silk-based gene delivery vehicles containing cell membrane-destabilizing peptides are therefore concluded to have potential for a less-toxic and controlled-release gene delivery system. PMID:20942485

  10. Engineering a Recombinant Baculovirus with a Peptide Hormone Gene and its Effect on the Corn Earworm, Helicoverpa zea

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The helicokinins are peptides identified from Helicoverpa zea that when injected into the larvae were found to cause excessive diuresis and loss of feeding activity. Of the three peptides, helicokinin II (HezK-II) was found to be most potent. A synthetic gene encoding HezK-II was constructed based o...

  11. Rapid evolution of antimicrobial peptide genes in an insect host-social parasite system.

    PubMed

    Erler, Silvio; Lhomme, Patrick; Rasmont, Pierre; Lattorff, H Michael G

    2014-04-01

    Selection, as a major driver for evolution in host-parasite interactions, may act on two levels; the virulence of the pathogen, and the hosts' defence system. Effectors of the host defence system might evolve faster than other genes e.g. those involved in adaptation to changes in life history or environmental fluctuations. Host-parasite interactions at the level of hosts and their specific social parasites, present a special setting for evolutionarily driven selection, as both share the same environmental conditions and pathogen pressures. Here, we study the evolution of antimicrobial peptide (AMP) genes, in six host bumblebee and their socially parasitic cuckoo bumblebee species. The selected AMP genes evolved much faster than non-immune genes, but only defensin-1 showed significant differences between host and social parasite. Nucleotide diversity and codon-by-codon analyses confirmed that purifying selection is the main selective force acting on bumblebee defence genes. PMID:24530902

  12. Transcriptional regulation of neuropeptide and peptide hormone expression by the Drosophila dimmed and cryptocephal genes.

    PubMed

    Gauthier, Sebastien A; Hewes, Randall S

    2006-05-01

    The regulation of neuropeptide and peptide hormone gene expression is essential for the development and function of neuroendocrine cells in integrated physiological networks. In insects, a decline in circulating ecdysteroids triggers the activation of a neuroendocrine system to stimulate ecdysis, the behaviors used to shed the old cuticle at the culmination of each molt. Here we show that two evolutionarily conserved transcription factor genes, the basic helix-loop-helix (bHLH) gene dimmed (dimm) and the basic-leucine zipper (bZIP) gene cryptocephal (crc), control expression of diverse neuropeptides and peptide hormones in Drosophila. Central nervous system expression of three neuropeptide genes, Dromyosuppressin, FMRFamide-related and Leucokinin, is activated by dimm. Expression of Ecdysis triggering hormone (ETH) in the endocrine Inka cells requires crc; homozygous crc mutant larvae display markedly reduced ETH levels and corresponding defects in ecdysis. crc activates ETH expression though a 382 bp enhancer, which completely recapitulates the ETH expression pattern. The enhancer contains two evolutionarily conserved regions, and both are imperfect matches to recognition elements for activating transcription factor-4 (ATF-4), the vertebrate ortholog of the CRC protein and an important intermediate in cellular responses to endoplasmic reticulum stress. These regions also contain a putative ecdysteroid response element and a predicted binding site for the products of the E74 ecdysone response gene. These results suggest that convergence between ATF-related signaling and an important intracellular steroid response pathway may contribute to the neuroendocrine regulation of insect molting. PMID:16651547

  13. Targeted gene delivery to human airway epithelial cells with synthetic vectors incorporating novel targeting peptides selected by phage display.

    PubMed

    Writer, Michele J; Marshall, Barry; Pilkington-Miksa, Michael A; Barker, Susie E; Jacobsen, Marianne; Kritz, Angelika; Bell, Paul C; Lester, Douglas H; Tabor, Alethea B; Hailes, Helen C; Klein, Nigel; Hart, Stephen L

    2004-05-01

    Human airway epithelial cell targeting peptides were identified by biopanning on 1HAEo-cells, a well characterised epithelial cell line. Bound phage were recovered after three rounds of binding, high stringency washing and elution, leading to the production of an enriched phage peptide population. DNA sequencing of 56 clones revealed 14 unique sequences. Subsequent binding analysis revealed that 13 of these peptides bound 1HAEo-cells with high affinity. Three peptides, SERSMNF, YGLPHKF and PSGAARA were represented at high frequency. Three clearly defined families of peptide were identified on the basis of sequence motifs including (R/K)SM, L(P/Q)HK and PSG(A/T)ARA. Two peptides, LPHKSMP and LQHKSMP contained two motifs. Further detailed sequence analysis by comparison of peptide sequences with the SWISSPROT protein database revealed that some of the peptides closely resembled the cell binding proteins of viral and bacterial pathogens including Herpes Simplex Virus, rotavirus, Mycoplasma pneumoniae and rhinovirus, the latter two being respiratory pathogens, as well as peptide YGLPHKF having similarity to a protein of unknown function from the respiratory pathogen Legionella pneumophila. Peptides were incorporated into gene delivery formulations with the cationic lipid Lipofectin and plasmid DNA and shown to confer a high degree of transfection efficiency and specificity in 1HAEo-cells. Improved transfection efficiency and specificity was also observed in human endothelial cells, fibroblasts and keratinocytes. Therefore, on the basis of clone frequency after biopanning, cell binding affinity, peptide sequence conservation and pathogenic similarity, we have identified 3 novel peptide families and 5 specific peptides that have the potential for gene transfer to respiratory epithelium in vivo as well as providing useful in vitro transfection reagents for primary human cell types of scientific and commercial interest. PMID:15506167

  14. Gene coevolution and regulation lock cyclic plant defence peptides to their targets.

    PubMed

    Gilding, Edward K; Jackson, Mark A; Poth, Aaron G; Henriques, Sónia Troeira; Prentis, Peter J; Mahatmanto, Tunjung; Craik, David J

    2016-04-01

    Plants have evolved many strategies to protect themselves from attack, including peptide toxins that are ribosomally synthesized and thus adaptable directly by genetic polymorphisms. Certain toxins in Clitoria ternatea (butterfly pea) are cyclic cystine-knot peptides of c. 30 residues, called cyclotides, which have co-opted the plant's albumin-1 gene family for their production. How butterfly pea albumin-1 genes were commandeered and how these cyclotides are utilized in defence remain unclear. The role of cyclotides in host plant ecology and biotechnological applications requires exploration. We characterized the sequence diversity and expression dynamics of precursor and processing proteins implicated in butterfly pea cyclotide biosynthesis by expression profiling through RNA-sequencing (RNA-seq). Peptide-enriched extracts from various organs were tested for activity against insect-like membranes and the model nematode Caenorhabditis elegans. We found that the evolution and deployment of cyclotides involved their diversification to exhibit different chemical properties and expression between organs facing different defensive challenges. Cyclotide-enriched fractions from soil-contacting organs were effective at killing nematodes, whereas similar enriched fractions from aerial organs contained cyclotides that exhibited stronger interactions with insect-like membrane lipids. Cyclotides are employed as versatile and combinatorial mediators of defence in C. ternatea and have specialized to affect different classes of attacking organisms. PMID:26668107

  15. abf-1 and abf-2, ASABF-type antimicrobial peptide genes in Caenorhabditis elegans.

    PubMed Central

    Kato, Yusuke; Aizawa, Tomoyasu; Hoshino, Hirokazu; Kawano, Keiichi; Nitta, Katsutoshi; Zhang, Hong

    2002-01-01

    Two genes encoding the ASABF (Ascaris suum antibacterial factor)-type antimicrobial peptide, abf-1 and abf-2, were identified in Caenorhabditis elegans. Recombinant ABF-2 exhibited potent microbicidal activity against Gram-positive and Gram-negative bacteria, and yeasts. The tissue-specific distribution estimated by immunofluorescence staining and transgenic analysis of a gfp fusion gene (where GFP corresponds to green fluorescent protein) suggested that ABF-2 contributes to surface defence in the pharynx. abf-1 contains a single intron at a conserved position, suggesting that asabf and abf originated from a common ancestor. Both transcripts for abf-1 and abf-2 were detected as two distinct forms, i.e. spliced leader (SL)1-trans-spliced with a long 5'-untranslated region (UTR) and SL-less with a short 5'-UTR. A polycistronic precursor RNA encoding ABF-1 and ABF-2 was detected, suggesting that these genes form an operon. An 'opportunistic operon' model for regulation of abf genes, including the generation of short SL-less transcripts, is proposed. In conclusion, C. elegans should have an immune defence system due to the antimicrobial peptides. C. elegans can be a novel model for innate immunity. Furthermore, the combination of biochemical identification in Ascaris suum and homologue hunting in C. elegans should be a powerful method of finding rapidly evolved proteins, such as some immune-related molecules in C. elegans. PMID:11772394

  16. Reiterated Targeting Peptides on the Nanoparticle Surface Significantly Promote Targeted Vascular Endothelial Growth Factor Gene Delivery to Stem Cells.

    PubMed

    Wang, Dong-Dong; Yang, Mingying; Zhu, Ye; Mao, Chuanbin

    2015-12-14

    Nonviral gene delivery vectors hold great promise for gene therapy due to the safety concerns with viral vectors. However, the application of nonviral vectors is hindered by their low transfection efficiency. Herein, in order to tackle this challenge, we developed a nonviral vector integrating lipids, sleeping beauty transposon system and 8-mer stem cell targeting peptides for safe and efficient gene delivery to hard-to-transfect mesenchymal stem cells (MSCs). The 8-mer MSC-targeting peptides, when synthetically reiterated in three folds and chemically presented on the surface, significantly promoted the resultant lipid-based nanoparticles (LBNs) to deliver VEGF gene into MSCs with a high transfection efficiency (∼52%) and long-lasting gene expression (for longer than 170 h) when compared to nonreiterated peptides. However, the reiterated stem cell targeting peptides do not enable the highly efficient gene transfer to other control cells. This work suggests that the surface presentation of the reiterated stem cell-targeting peptides on the nonviral vectors is a promising method for improving the efficiency of cell-specific nonviral gene transfection in stem cells. PMID:26588028

  17. Molecular Characterization of Antimicrobial Peptide Genes of the Carpenter Ant Camponotus floridanus

    PubMed Central

    Ratzka, Carolin; Förster, Frank; Liang, Chunguang; Kupper, Maria; Dandekar, Thomas; Feldhaar, Heike; Gross, Roy

    2012-01-01

    The production of antimicrobial peptides (AMPs) is a major defense mechanism against pathogen infestation and of particular importance for insects relying exclusively on an innate immune system. Here, we report on the characterization of three AMPs from the carpenter ant Camponotus floridanus. Due to sequence similarities and amino acid composition these peptides can be classified into the cysteine-rich (e.g. defensin) and glycine-rich (e.g. hymenoptaecin) AMP groups, respectively. The gene and cDNA sequences of these AMPs were established and their expression was shown to be induced by microbial challenge. We characterized two different defensin genes. The defensin-2 gene has a single intron, whereas the defensin-1 gene has two introns. The deduced amino acid sequence of the C. floridanus defensins is very similar to other known ant defensins with the exception of a short C-terminal extension of defensin-1. The hymenoptaecin gene has a single intron and a very peculiar domain structure. The corresponding precursor protein consists of a signal- and a pro-sequence followed by a hymenoptaecin-like domain and six directly repeated hymenoptaecin domains. Each of the hymenoptaecin domains is flanked by an EAEP-spacer sequence and a RR-site known to be a proteolytic processing site. Thus, proteolytic processing of the multipeptide precursor may generate several mature AMPs leading to an amplification of the immune response. Bioinformatical analyses revealed the presence of hymenoptaecin genes with similar multipeptide precursor structure in genomes of other ant species suggesting an evolutionary conserved important role of this gene in ant immunity. PMID:22912782

  18. Strategic expression of ion transport peptide gene products in central and peripheral neurons of insects.

    PubMed

    Dai, Li; Zitnan, Dusan; Adams, Michael E

    2007-01-10

    Structurally related ion transport peptides (ITP) and crustacean hyperglycemic hormones (CHH) are increasingly implicated in diverse metabolic and developmental functions in arthropods. We identified a conserved ITP gene encoding two peptides by alternative splicing in Manduca sexta, Bombyx mori, and Aedes aegypti: A C-terminally amidated ITP and a C-terminally unblocked ITP-like peptide (ITPL), which share common N-terminal sequences but have divergent C-termini. In the moth M. sexta, these peptides are expressed in two, regionally distinct neuronal populations in the central and peripheral nervous systems (CNS, PNS). MasITP expression is confined to the brain in five pairs of lateral neurosecretory cells (type Ia(2)) projecting ipsilateral axons into the retrocerebral complex and three to five pairs of adjacent small neurons that arborize extensively within the brain. Expression of MasITPL is comparatively weak in the brain but strong in the ventral ganglia and the PNS, where MasITP is absent. MasITPL occurs in bilaterally paired neurons of all thoracic and abdominal ganglia. In the PNS, MasITPL is coexpressed with crustacean cardioactive peptide in type II link nerve neurons (L1) of abdominal segments 2-7, which project axons into neurohemal transverse nerves. During metamorphosis, additional expression of MasITPL is observed in two pairs of small lateral neurons in the brain and one pair of ventromedial neurons in each of AG2-6. A similar pattern of differential ITP and ITPL expression was observed in the CNS and PNS of B. mori and Schistocerca americana. These distinctive cellular expression patterns suggest that ITP and ITPL have evolved specialized physiological functions in arthropods. PMID:17111378

  19. Alzheimer amyloid peptide aβ42 regulates gene expression of transcription and growth factors.

    PubMed

    Barucker, Christian; Sommer, Anette; Beckmann, Georg; Eravci, Murat; Harmeier, Anja; Schipke, Carola G; Brockschnieder, Damian; Dyrks, Thomas; Althoff, Veit; Fraser, Paul E; Hazrati, Lili-Naz; George-Hyslop, Peter St; Breitner, John C S; Peters, Oliver; Multhaup, Gerhard

    2015-01-01

    The pathogenesis of Alzheimer's disease (AD) is characterized by the aggregation of amyloid-β (Aβ) peptides leading to deposition of senile plaques and a progressive decline of cognitive functions, which currently remains the main criterion for its diagnosis. Robust biomarkers for AD do not yet exist, although changes in the cerebrospinal fluid levels of tau and Aβ represent promising candidates in addition to brain imaging and genetic risk profiling. Although concentrations of soluble Aβ42 correlate with symptoms of AD, less is known about the biological activities of Aβ peptides which are generated from the amyloid-β protein precursor. An unbiased DNA microarray study showed that Aβ42, at sub-lethal concentrations, specifically increases expression of several genes in neuroblastoma cells, notably the insulin-like growth factor binding proteins 3 and 5 (IGFBP3/5), the transcription regulator inhibitor of DNA binding, and the transcription factor Lim only domain protein 4. Using qRT-PCR, we confirmed that mRNA levels of the identified candidate genes were exclusively increased by the potentially neurotoxic Aβ42 wild-type peptide, as both the less toxic Aβ40 and a non-toxic substitution peptide Aβ42 G33A did not affect mRNA levels. In vivo immunohistochemistry revealed a corresponding increase in both hippocampal and cortical IGFBP5 expression in an AD mouse model. Proteomic analyses of human AD cerebrospinal fluid displayed increased in vivo concentrations of IGFBPs. IGFBPs and transcription factors, as identified here, are modulated by soluble Aβ42 and may represent useful early biomarkers. PMID:25318543

  20. Characterization of Treefoil Peptide Genes in Iron-Ion or X-Irradiated Human Cells

    NASA Technical Reports Server (NTRS)

    Balcer-Kubiczek, E. K.; Harrison, G. H.; Xu, J. F.; Zhou, X. F.

    1999-01-01

    The gastrointestinal (GI) tract is especially sensitive to ionizing radiation, probably because of its high rate of cell turn over. Most of the data in the literature concerns the histological/anatomical description of damage rather than functional studies. In fact, previous reports in humans have shown that, at doses of 2 Gy or more, functional abnormalities appear indicating that in radiation sensitive tissues the effects of radiation are not limited to cell death. GI functions are controlled in particular by GI peptides. One hypothesis is that ionizing radiation may modulate the synthesis and release of these peptides and consequently may contribute largely to abnormalities in GI function. However, no previous studies have been concerned with GI-specific gene expression in irradiated GI tissues. The family of human trefoil peptides comprises three members thus far, all of which are expressed in specific regions of the GI tract. In addition, two trefoil peptides, pS2 (TFFI) and HITF (TFF2) are expressed in breast tissue. Their exact function in GI and breast tissues is unclear but mucosal integrity, repair, mucin secretion and responsiveness to hormones have been shown. We recently isolated and characterized pS2 as a novel p53- and estrogen receptor-independent gene whose MRNA expression in several cells lines was found to be delayed 4 to 7 days after irradiation with X-rays, fission neutrons or 1 GeV/n Fe-ions. The aim of the present study was to determine whether pS2 and HITF have a similar induction kinetics in irradiated gastric and breast cell lines, and whether they have the phorbol ester (TPA) responsive element (TRE).

  1. Diversity of Nonribosomal Peptide Synthetase Genes in the Microbial Metagenomes of Marine Sponges

    PubMed Central

    Pimentel-Elardo, Sheila Marie; Grozdanov, Lubomir; Proksch, Sebastian; Hentschel, Ute

    2012-01-01

    Genomic mining revealed one major nonribosomal peptide synthetase (NRPS) phylogenetic cluster in 12 marine sponge species, one ascidian, an actinobacterial isolate and seawater. Phylogenetic analysis predicts its taxonomic affiliation to the actinomycetes and hydroxy-phenyl-glycine as a likely substrate. Additionally, a phylogenetically distinct NRPS gene cluster was discovered in the microbial metagenome of the sponge Aplysina aerophoba, which shows highest similarities to NRPS genes that were previously assigned, by ways of single cell genomics, to a Chloroflexi sponge symbiont. Genomic mining studies such as the one presented here for NRPS genes, contribute to on-going efforts to characterize the genomic potential of sponge-associated microbiota for secondary metabolite biosynthesis. PMID:22822366

  2. Gene expression of atrial natriuretic peptide in rat papillary muscle. Rapid induction by mechanical loading.

    PubMed

    Jarygin, C; Hänze, J; Lang, R E

    1994-06-13

    The effect of mechanical stretch on protein synthesis and the expression of the gene for atrial natriuretic peptide (ANP) was examined in electrically paced, isolated papillary muscles from rat heart. Incorporation of [3H]phenylalanine into protein increased only in stretched but not in unloaded muscles. Five hours of stretching increased ANP mRNA levels more than threefold as compared to freshly excised papillary muscles. A drastic fall in ANP mRNA levels was observed in unloaded muscles over this time. These data indicate that papillary muscles similar to other ventricular tissue are capable of activating ANP gene expression in response to increased load. The effect occurs in vitro and does not depend on circulating or nervous factors. The unexpected rapid induction of ANP gene expression in such a particular structure of the heart raises the possibility of local actions of ventricular ANP. PMID:8013631

  3. Diversity of nonribosomal peptide synthetase genes in the microbial metagenomes of marine sponges.

    PubMed

    Pimentel-Elardo, Sheila Marie; Grozdanov, Lubomir; Proksch, Sebastian; Hentschel, Ute

    2012-06-01

    Genomic mining revealed one major nonribosomal peptide synthetase (NRPS) phylogenetic cluster in 12 marine sponge species, one ascidian, an actinobacterial isolate and seawater. Phylogenetic analysis predicts its taxonomic affiliation to the actinomycetes and hydroxy-phenyl-glycine as a likely substrate. Additionally, a phylogenetically distinct NRPS gene cluster was discovered in the microbial metagenome of the sponge Aplysina aerophoba, which shows highest similarities to NRPS genes that were previously assigned, by ways of single cell genomics, to a Chloroflexi sponge symbiont. Genomic mining studies such as the one presented here for NRPS genes, contribute to on-going efforts to characterize the genomic potential of sponge-associated microbiota for secondary metabolite biosynthesis. PMID:22822366

  4. Verification of elicitor efficacy of lipopolysaccharides and peptidoglycans on antibacterial peptide gene expression in Bombyx mori.

    PubMed

    Ha Lee, Joon; Hee Lee, In; Noda, Hiroaki; Mita, Kazuei; Taniai, Kiyoko

    2007-12-01

    We verified the efficacy of lipopolysaccharide (LPS) in activating the cecropin B gene (CecB) in an immune-competent Bombyx mori cell line. Strong activation of CecB by the LPSs from Escherichia coli, Pseudomonas aeruginosa, and Salmonella minnesota were completely eliminated after digestion of the LPSs with muramidase. The results clearly indicate that a polymer form of PGN in the LPSs elicited CecB. An oligonucleotide microarray screen revealed that none of the 16,000 genes on the array were activated by LPS in the cells. In contrast, E. coli PGN strongly elicited five antibacterial peptide genes and numerous other genes, and PGN from Micrococcus luteus activated only several genes. Semi-quantitative RT-PCR revealed that all antibacterial genes activated by both PGNs, but the extents were 10-100 times higher with E. coli PGN. Similarly, higher elicitor activity of E. coli than M. luteus was indicated using peptidoglycan recognition protein gene, which is involved in pro-phenol oxidase cascade. PMID:17967352

  5. Three gonadotropin-releasing hormone genes in one organism suggest novel roles for an ancient peptide.

    PubMed Central

    White, S A; Kasten, T L; Bond, C T; Adelman, J P; Fernald, R D

    1995-01-01

    Gonadotropin-releasing hormone (GnRH) is known and named for its essential role in vertebrate reproduction. Release of this decapeptide from neurons in the hypothalamus controls pituitary gonadotropin levels which, in turn, regulate gonadal state. The importance of GnRH is underscored by its widespread expression and conservation across vertebrate taxa: five amino acids are invariant in all nine known forms, whereas two others show only conservative changes. In most eutherian mammals, only one form, expressed in the hypothalamus, is thought to exist, although in a recent report, antibody staining in developing primates suggests an additional form. In contrast, multiple GnRH forms and expression loci have been reported in many non-mammalian vertebrates. However, evidence based on immunological discrimination does not always agree with analysis of gene expression, since GnRH forms encoded by different genes may not be reliably distinguished by antibodies. Here we report the expression of three distinct GnRH genes in a teleost fish brain, including the sequence encoding a novel GnRH preprohormone. Using in situ hybridization, we show that this form is found only in neurons that project to the pituitary and exhibit changes in soma size depending on social and reproductive state. The other two GnRH genes are expressed in other, distinct cell populations. All three genes share the motif of encoding a polypeptide consisting of GnRH and a GnRH-associated peptide. Whereas the GnRH moiety is highly conserved, the GnRH-associated peptides are not, reflecting differential selective pressure on different parts of the gene. GnRH forms expressed in nonhypothalamic regions may serve to coordinate reproductive activities of the animal. Images Fig. 3 PMID:7667296

  6. Lipid peptide nanocomplexes for gene delivery and magnetic resonance imaging in the brain.

    PubMed

    Writer, Michele J; Kyrtatos, Panagiotis G; Bienemann, Alison S; Pugh, John A; Lowe, Andrew S; Villegas-Llerena, Claudio; Kenny, Gavin D; White, Edward A; Gill, Steven S; McLeod, Cameron W; Lythgoe, Mark F; Hart, Stephen L

    2012-09-10

    Gadolinium-labelled nanocomplexes offer prospects for the development of real-time, non-invasive imaging strategies to visualise the location of gene delivery by MRI. In this study, targeted nanoparticle formulations were prepared comprising a cationic liposome (L) containing a Gd-chelated lipid at 10, 15 and 20% by weight of total lipid, a receptor-targeted, DNA-binding peptide (P) and plasmid DNA (D), which electrostatically self-assembled into LPD nanocomplexes. The LPD formulation containing the liposome with 15% Gd-chelated lipid displayed optimal peptide-targeted, transfection efficiency. MRI conspicuity peaked at 4h after incubation of the nanocomplexes with cells, suggesting enhancement by cellular uptake and trafficking. This was supported by time course confocal microscopy analysis of transfections with fluorescently-labelled LPD nanocomplexes. Gd-LPD nanocomplexes delivered to rat brains by convection-enhanced delivery were visible by MRI at 6 h, 24 h and 48 h after administration. Histological brain sections analysed by laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) confirmed that the MRI signal was associated with the distribution of Gd(3+) moieties and differentiated MRI signals due to haemorrhage. The transfected brain cells near the injection site appeared to be mostly microglial. This study shows the potential of Gd-LPD nanocomplexes for simultaneous delivery of contrast agents and genes for real-time monitoring of gene therapy in the brain. PMID:22800579

  7. Lipid peptide nanocomplexes for gene delivery and magnetic resonance imaging in the brain

    PubMed Central

    Writer, Michele J.; Kyrtatos, Panagiotis G.; Bienemann, Alison S.; Pugh, John A.; Lowe, Andrew S.; Villegas-Llerena, Claudio; Kenny, Gavin D.; White, Edward A.; Gill, Steven S.; McLeod, Cameron W.; Lythgoe, Mark F.; Hart, Stephen L.

    2012-01-01

    Gadolinium-labelled nanocomplexes offer prospects for the development of real-time, non-invasive imaging strategies to visualise the location of gene delivery by MRI. In this study, targeted nanoparticle formulations were prepared comprising a cationic liposome (L) containing a Gd-chelated lipid at 10, 15 and 20% by weight of total lipid, a receptor-targeted, DNA-binding peptide (P) and plasmid DNA (D), which electrostatically self-assembled into LPD nanocomplexes. The LPD formulation containing the liposome with 15% Gd-chelated lipid displayed optimal peptide-targeted, transfection efficiency. MRI conspicuity peaked at 4 h after incubation of the nanocomplexes with cells, suggesting enhancement by cellular uptake and trafficking. This was supported by time course confocal microscopy analysis of transfections with fluorescently-labelled LPD nanocomplexes. Gd-LPD nanocomplexes delivered to rat brains by convection-enhanced delivery were visible by MRI at 6 h, 24 h and 48 h after administration. Histological brain sections analysed by laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) confirmed that the MRI signal was associated with the distribution of Gd3 + moieties and differentiated MRI signals due to haemorrhage. The transfected brain cells near the injection site appeared to be mostly microglial. This study shows the potential of Gd-LPD nanocomplexes for simultaneous delivery of contrast agents and genes for real-time monitoring of gene therapy in the brain. PMID:22800579

  8. Virulent Shigella flexneri subverts the host innate immune response through manipulation of antimicrobial peptide gene expression

    PubMed Central

    Sperandio, Brice; Regnault, Béatrice; Guo, Jianhua; Zhang, Zhi; Stanley, Samuel L.; Sansonetti, Philippe J.; Pédron, Thierry

    2008-01-01

    Antimicrobial factors are efficient defense components of the innate immunity, playing a crucial role in the intestinal homeostasis and protection against pathogens. In this study, we report that upon infection of polarized human intestinal cells in vitro, virulent Shigella flexneri suppress transcription of several genes encoding antimicrobial cationic peptides, particularly the human β-defensin hBD-3, which we show to be especially active against S. flexneri. This is an example of targeted survival strategy. We also identify the MxiE bacterial regulator, which controls a regulon encompassing a set of virulence plasmid-encoded effectors injected into host cells and regulating innate signaling, as being responsible for this dedicated regulatory process. In vivo, in a model of human intestinal xenotransplant, we confirm at the transcriptional and translational level, the presence of a dedicated MxiE-dependent system allowing S. flexneri to suppress expression of antimicrobial cationic peptides and promoting its deeper progression toward intestinal crypts. We demonstrate that this system is also able to down-regulate additional innate immunity genes, such as the chemokine CCL20 gene, leading to compromised recruitment of dendritic cells to the lamina propria of infected tissues. Thus, S. flexneri has developed a dedicated strategy to weaken the innate immunity to manage its survival and colonization ability in the intestine. PMID:18426984

  9. A Peptide-based Vector for Efficient Gene Transfer In Vitro and In Vivo

    PubMed Central

    Lehto, Taavi; Simonson, Oscar E; Mäger, Imre; Ezzat, Kariem; Sork, Helena; Copolovici, Dana-Maria; Viola, Joana R; Zaghloul, Eman M; Lundin, Per; Moreno, Pedro MD; Mäe, Maarja; Oskolkov, Nikita; Suhorutšenko, Julia; Smith, CI Edvard; Andaloussi, Samir EL

    2011-01-01

    Finding suitable nonviral delivery vehicles for nucleic acid–based therapeutics is a landmark goal in gene therapy. Cell-penetrating peptides (CPPs) are one class of delivery vectors that has been exploited for this purpose. However, since CPPs use endocytosis to enter cells, a large fraction of peptides remain trapped in endosomes. We have previously reported that stearylation of amphipathic CPPs, such as transportan 10 (TP10), dramatically increases transfection of oligonucleotides in vitro partially by promoting endosomal escape. Therefore, we aimed to evaluate whether stearyl-TP10 could be used for the delivery of plasmids as well. Our results demonstrate that stearyl-TP10 forms stable nanoparticles with plasmids that efficiently enter different cell-types in a ubiquitous manner, including primary cells, resulting in significantly higher gene expression levels than when using stearyl-Arg9 or unmodified CPPs. In fact, the transfection efficacy of stearyl-TP10 almost reached the levels of Lipofectamine 2000 (LF2000), however, without any of the observed lipofection-associated toxicities. Most importantly, stearyl-TP10/plasmid nanoparticles are nonimmunogenic, mediate efficient gene delivery in vivo, when administrated intramuscularly (i.m.) or intradermally (i.d.) without any associated toxicity in mice. PMID:21343913

  10. Detection of genes encoding antimicrobial peptides in Mexican strains of Trichoplusia ni (Hubner) exposed to Bacillus thuringiensis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The systemic immune response of Trichoplusia ni after Bacillus thuringiensis (Bt) exposure was evaluated by comparing the expression of genes encoding antimicrobial peptides (AMP) in Bt-susceptible and -resistant T. ni strains that were either exposed or not to XenTari® (Bt-XT). AMP genes were dete...

  11. Targeted gene correction using psoralen, chlorambucil and camptothecin conjugates of triplex forming peptide nucleic acid (PNA)

    PubMed Central

    Birkedal, Henrik

    2011-01-01

    Gene correction activation effects of a small series of triplex forming peptide nucleic acid (PNA) covalently conjugated to the DNA interacting ligands psoralen, chlorambucil and camptothecin targeted proximal to a stop codon mutation in an EGFP reporter gene were studied. A 15-mer homopyrimidine PNA conjugated to the topoisomerase I inhibitor camptothecin was found to increase the frequency of repair domain mediated gene correctional events of the EGFP reporter in an in vitro HeLa cell nuclear extract assay, whereas PNA psoralen or chlorambucil conjugates both of which form covalent and also interstrand crosslinked adducts with dsDNA dramatically decreased the frequency of targeted repair/correction. The PNA conjugates were also studied in mammalian cell lines upon transfection of PNA bound EGFP reporter vector and scoring repair of the EGFP gene by FACS analysis of functional EGFP expression. Consistent with the extract experiments, treatment with adduct forming PNA conjugates (psoralen and chlorambucil) resulted in a decrease in background correction frequencies in transiently transfected cells, whereas unmodified PNA or the PNA-camptothecin conjugate had little or no effect. These results suggest that simple triplex forming PNAs have little effect on proximal gene correctional events whereas PNA conjugates capable of forming DNA adducts and interstrand crosslinks are strong inhibitors. Most interestingly the PNA conjugated to the topoisomerase inhibitor, camptothecin enhanced repair in nuclear extract. Thus the effects and use of camptothecin conjugates in gene targeted repair merit further studies. PMID:21686249

  12. Novel gelatin siloxane nanoparticles decorated by Tat peptide as vectors for gene therapy

    NASA Astrophysics Data System (ADS)

    Wang, Zu-yong; Zhao, Yang; Ren, Lei; Jin, Li-hua; Sun, Li-ping; Yin, Pei; Zhang, Ya-fei; Zhang, Qi-Qing

    2008-11-01

    In principle, the technique of gene delivery involves taking complete or parts of genes that can code specific messages and delivering them to selected cells in the body. Such a transfer of plasmid DNA into mammalian cells has posed major challenges for gene therapy. A series of gelatin-siloxane nanoparticles (GS NPs) with controlled size and surface charge were synthesized through a two-step sol-gel process. In order to increase the efficiency of cellular uptake, HIV-derived Tat peptide was further grafted to GS NPs. In vitro co-location and endocytosis inhibition experiments suggested that the as-synthesized TG NPs may enter HeLa cells via a combined pathway of lipid-raft- and receptor-dependent endocytosis, and only cause little cell damage. Moreover, this study shows the encapsulation of a plasmid DNA in TG NPs to be obtained as a non-viral gene vector. This kind of encapsulation provides complete protection to the plasmid DNA from the external DNase and serum environment, and generates the hope that the resulting formulation can be developed into a potential vector for effective gene delivery. In order to check this potential, the reporter gene pSVβ-gal was encapsulated, and in vitro transfection efficiency of this system was found to be nearly 130% compared to the commercially available transfection reagent Lipofectamine™.

  13. Peptide portions may hold key to amplifying bone against porosis

    SciTech Connect

    Cotton, P.

    1990-02-02

    Pieces of peptides that are encoded in the calcitonin gene may explain enigmas in treatment of bone disease. Amplification of bone formation by two peptides with similar amino acid sequences was reported at the Third International Conference on the Fundamentals of Bone Growth at the University of California, Los Angeles (UCLA), schools of medicine and dentistry. One treatment enigma is that calcitonin regulates normal bone resorption but does not work as well when administered for the treatment of osteoporosis. While hormone therapy does work, it has wide-ranging effects like the potential for an increased risk of breast cancer. Bone-growth promotion by a better-known peptide, calcitonin gene-related peptide (CGRP), was described. The CGRP is usually processed in the nervous system and has a wide range of activity.

  14. Discovery of a Linear Peptide for Improving Tumor Targeting of Gene Products and Treatment of Distal Tumors by IL-12 Gene Therapy

    PubMed Central

    Cutrera, Jeffry; Dibra, Denada; Xia, Xueqing; Hasan, Azeem; Reed, Scott; Li, Shulin

    2011-01-01

    Like many effective therapeutics, interleukin-12 (IL-12) therapy often causes side effects. Tumor targeted delivery may improve the efficacy and decrease the toxicity of systemic IL-12 treatments. In this study, a novel targeting approach was investigated. A secreted alkaline phosphatase (SEAP) reporter gene-based screening process was used to identify a mini-peptide which can be produced in vivo to target gene products to tumors. The coding region for the best peptide was inserted into an IL-12 gene to determine the antitumor efficacy. Affinity chromatography, mass spectrometry analysis, and binding studies were used to identify a receptor for this peptide. We discovered that the linear peptide VNTANST increased the tumor accumulation of the reporter gene products in five independent tumor models including one human xenogeneic model. The product from VNTANST-IL-12 fusion gene therapy increased accumulation of IL-12 in the tumor environment, and in three tumor models, VNTANST-IL-12 gene therapy inhibited distal tumor growth. In a spontaneous lung metastasis model, inhibition of metastatic tumor growth was improved compared to wild-type IL-12 gene therapy, and in a squamous cell carcinoma model, toxic liver lesions were reduced. The receptor for VNTANST was identified as vimentin. These results show the promise of using VNTANST to improve IL-12 treatments. PMID:21386825

  15. Stearylated antimicrobial peptide [D]-K6L9 with cell penetrating property for efficient gene transfer.

    PubMed

    Zhang, Wei; Song, Jingjing; Liang, Ranran; Zheng, Xin; Chen, Jianbo; Li, Guolin; Zhang, Bangzhi; Wang, Kairong; Yan, Xiang; Wang, Rui

    2013-08-01

    Stearyl-cell penetrating peptides (CPPs) have been proved to be efficient nonviral gene vectors. Due to the similarities between antimicrobial peptides and CPPs, we constructed a novel type of gene vectors by introducing stearyl moiety to the N-terminus of antimicrobial peptide [D]-K6L9. In this study, stearyl-[D]-K6L9 delivered plasmids into cells by clathrin- and caveolin-mediated endocytosis. Gratifyingly, stearyl-[D]-K6L9 exhibited high transfection efficiency and almost reached the level of Lipofectamine 2000. Taken together, the combination of the stearyl moiety with [D]-K6L9 provides a novel framework for the development of excellent nonviral gene vectors. PMID:23727033

  16. Multi-species sequence comparison reveals conservation of ghrelin gene-derived splice variants encoding a truncated ghrelin peptide.

    PubMed

    Seim, Inge; Jeffery, Penny L; Thomas, Patrick B; Walpole, Carina M; Maugham, Michelle; Fung, Jenny N T; Yap, Pei-Yi; O'Keeffe, Angela J; Lai, John; Whiteside, Eliza J; Herington, Adrian C; Chopin, Lisa K

    2016-06-01

    The peptide hormone ghrelin is a potent orexigen produced predominantly in the stomach. It has a number of other biological actions, including roles in appetite stimulation, energy balance, the stimulation of growth hormone release and the regulation of cell proliferation. Recently, several ghrelin gene splice variants have been described. Here, we attempted to identify conserved alternative splicing of the ghrelin gene by cross-species sequence comparisons. We identified a novel human exon 2-deleted variant and provide preliminary evidence that this splice variant and in1-ghrelin encode a C-terminally truncated form of the ghrelin peptide, termed minighrelin. These variants are expressed in humans and mice, demonstrating conservation of alternative splicing spanning 90 million years. Minighrelin appears to have similar actions to full-length ghrelin, as treatment with exogenous minighrelin peptide stimulates appetite and feeding in mice. Forced expression of the exon 2-deleted preproghrelin variant mirrors the effect of the canonical preproghrelin, stimulating cell proliferation and migration in the PC3 prostate cancer cell line. This is the first study to characterise an exon 2-deleted preproghrelin variant and to demonstrate sequence conservation of ghrelin gene-derived splice variants that encode a truncated ghrelin peptide. This adds further impetus for studies into the alternative splicing of the ghrelin gene and the function of novel ghrelin peptides in vertebrates. PMID:26792793

  17. Genetic Analysis of the Atrial Natriuretic Peptide Gene Polymorphisms among Essential Hypertensive Patients in Malaysia

    PubMed Central

    Ghodsian, Nooshin; Ismail, Patimah; Ahmadloo, Salma; Eskandarian, Narges; Etemad, Ali

    2016-01-01

    Background. Atrial natriuretic peptide (ANP) considerably influences blood pressure regulation through water and sodium homoeostasis. Several of the studies have utilized anonymous genetic polymorphic markers and made inconsequent claims about the ANP relevant disorders. Thus, we screened Insertion/Deletion (ID) and G191A polymorphisms of ANP to discover sequence variations with potential functional significance and to specify the linkage disequilibrium pattern between polymorphisms. The relationships of detected polymorphisms with EH with or without Type 2 Diabetes Mellitus (T2DM) status were tested subsequently. Method. ANP gene polymorphisms (I/D and A191G) were specified utilizing mutagenically separated Polymerase Chain Reaction (PCR) in 320 subjects including 163 EH case subjects and 157 controls. Result. This case-control study discovered a significant association between I/D polymorphisms of ANP gene in EH patient without T2DM. However, the study determined no association between G191A polymorphisms of ANP in EH with or without T2DM. In addition, sociodemographic factors in the case and healthy subjects exhibited strong differences (P < 0.05). Conclusion. As a risk factor, ANP gene polymorphisms may affect hypertension. Despite the small sample size in this study, it is the first research assessing the ANP gene polymorphisms in both EH and T2DM patients among Malaysian population. PMID:27413750

  18. Enhancing gene delivery of adeno-associated viruses by cell-permeable peptides

    PubMed Central

    Liu, Yarong; Kim, Young Joo; Ji, Man; Fang, Jinxu; Siriwon, Natnaree; Zhang, Li I; Wang, Pin

    2014-01-01

    Adeno-associated virus type 2 (AAV2) is considered a promising gene delivery vector and has been extensively applied in several disease models; however, inefficient transduction in various cells and tissues has limited its widespread application in many areas of gene therapy. In this study, we have developed a general, but efficient, strategy to enhance viral transduction, both in vitro and in vivo, by incubating viral particles with cell-permeable peptides (CPPs). We show that CPPs increase internalization of viral particles into cells by facilitating both energy-independent and energy-dependent endocytosis. Moreover, CPPs can significantly enhance the endosomal escape process of viral particles, thus enhancing viral transduction to those cells that have exhibited very low permissiveness to AAV2 infection as a result of impaired intracellular viral processing. We also demonstrated that this approach could be applicable to other AAV serotypes. Thus, the membrane-penetrating ability of CPPs enables us to generate an efficient method for enhanced gene delivery of AAV vectors, potentially facilitating its applicability to human gene therapy. PMID:26015948

  19. Role of a molecular variant of rat atrial natriuretic Peptide gene in vascular remodeling.

    PubMed

    De Paolis, Paola; Nobili, Valerio; Lombardi, Alessia; Tarasi, David; Barbato, Daniela; Marchitti, Simona; Ganten, Ursula; Brunetti, Ercole; Volpe, Massimo; Rubattu, Speranza

    2007-01-01

    Previous studies in a hypertensive animal model of stroke and in humans showed that mutations of the atrial natriuretic peptide (ANP) gene are associated with increased risk of stroke. To elucidate the vascular disease mechanisms that result from structural modifications of the ANP gene, we investigated a coding mutation of the ANP gene in stroke-prone spontaneously hypertensive rats (SHRsp). This mutation leads to a Gly/Ser transposition in the prosegment of ANP. We found that presence of this mutation is associated with increased immunostaining of ANP in the wall of SHRsp cerebral vessels. The mutation causes a major inhibitory effect on endothelial cell proliferation, as assessed by thymidine incorporation, and on angiogenesis, as determined by an endothelial cell tube formation assay, in human umbilical vein endothelial cells (HUVEC) exposed to ANP/SHRsp. These in vitro findings show that the SHRsp-derived form of ANP has an inhibitory effect on vascular remodeling and they provide further support for a role of the ANP gene in the pathogenesis of cerebrovascular disease in the animal model. PMID:17522368

  20. Diurnal gene expression of lipolytic natriuretic peptide receptors in white adipose tissue

    PubMed Central

    Smith, Julie; Fahrenkrug, Jan; Jørgensen, Henrik L; Christoffersen, Christina; Goetze, Jens P

    2015-01-01

    Disruption of the circadian rhythm can lead to obesity and cardiovascular disease. In white adipose tissue, activation of the natriuretic peptide receptors (NPRs) stimulates lipolysis. We have previously shown that natriuretic peptides are expressed in a circadian manner in the heart, but the temporal expression profile of their cognate receptors has not been examined in white adipose tissue. We therefore collected peri-renal white adipose tissue and serum from WT mice. Tissue mRNA contents of NPRs – NPR-A and NPR-C, the clock genes Per1 and Bmal1, and transcripts involved in lipid metabolism were quantified at 4-h intervals: in the diurnal study, mice were exposed to a period of 12 h light followed by 12 h darkness (n=52). In the circadian study, mice were kept in darkness for 24 h (n=47). Concomitant serum concentrations of free fatty acids, glycerol, triglycerides (TGs), and insulin were measured. Per1 and Bmal1 mRNA contents showed reciprocal circadian profiles (P<0.0001). NPR-A mRNA contents followed a temporal pattern (P=0.01), peaking in the dark (active) period. In contrast, NPR-C mRNA was expressed in an antiphase manner with nadir in the active period (P=0.007). TG concentrations in serum peaked in the active dark period (P=0.003). In conclusion, NPR-A and NPR-C gene expression is associated with the expression of clock genes in white adipose tissue. The reciprocal expression may thus contribute to regulate lipolysis and energy homeostasis in a diurnal manner. PMID:26286623

  1. Synthesis of Electroneutralized Amphiphilic Copolymers with Peptide Dendrons for Intramuscular Gene Delivery.

    PubMed

    Pu, Linyu; Wang, Jiali; Li, Na; Chai, Qiuxia; Irache, Juan M; Wang, Gang; Tang, James Zhenggui; Gu, Zhongwei

    2016-06-01

    Intramuscular gene delivery materials are of great importance in plasmid-based gene therapy system, but there is limited information so far on how to design and synthesize them. A previous study showed that the peptide dendron-based triblock copolymer with its components arranged in a reversed biomembrane architecture could significantly increase intramuscular gene delivery and expression. Herein, we wonder whether copolymers with biomembrane-mimicking arrangement may have similar function on intramuscular gene delivery. Meanwhile, it is of great significance to uncover the influence of electric charge and molecular structure on the function of the copolymers. To address the issues, amphiphilic triblock copolymers arranged in hydrophilic-hydrophobic-hydrophilic structure were constructed despite the paradoxical characteristics and difficulties in synthesizing such hydrophilic but electroneutral molecules. The as-prepared two copolymers, dendronG2(l-lysine-OH)-poly propylene glycol2k(PPG2k)-dendronG2(l-lysine-OH) (rL2PL2) and dendronG3(l-lysine-OH)-PPG2k-dendronG3(l-lysine-OH) (rL3PL3), were in similar structure but had different hydrophilic components and surface charges, thus leading to different capabilities in gene delivery and expression in skeletal muscle. rL2PL2 was more efficient than Pluronic L64 and rL3PL3 when mediating luciferase, β-galactosidase, and fluorescent protein expressions. Furthermore, rL2PL2-mediated growth-hormone-releasing hormone expression could significantly induce mouse body weight increase in the first 21 days after injection. In addition, both rL2PL2 and rL3PL3 showed good in vivo biosafety in local and systemic administration. Altogether, rL2PL2-mediated gene expression in skeletal muscle exhibited applicable potential for gene therapy. The study revealed that the molecular structure and electric charge were critical factors governing the function of the copolymers for intramuscular gene delivery. It can be concluded that, combined

  2. Perinatal α-linolenic acid availability alters the expression of genes related to memory and to epigenetic machinery, and the Mecp2 DNA methylation in the whole brain of mouse offspring.

    PubMed

    He, Fuli; Lupu, Daniel S; Niculescu, Mihai D

    2014-08-01

    Many animal and human studies indicated that dietary ω-3 fatty acids could have beneficial roles on brain development, memory, and learning. However, the exact mechanisms involved are far from being clearly understood, especially for α-linolenic acid (ALA), which is the precursor for the ω-3 elongation and desaturation pathways. This study investigated the alterations induced by different intakes of flaxseed oil (containing 50% ALA), during gestation and lactation, upon the expression of genes involved in neurogenesis, memory-related molecular processes, and DNA methylation, in the brains of mouse offspring at the end of lactation (postnatal day 19, P19). In addition, DNA methylation status for the same genes was investigated. Maternal flaxseed oil supplementation during lactation increased the expression of Mecp2, Ppp1cc, and Reelin, while decreasing the expression of Ppp1cb and Dnmt3a. Dnmt1 expression was decreased by postnatal flaxseed oil supplementation but this effect was offset by ALA deficiency during gestation. Mecp2 DNA methylation was decreased by maternal ALA deficiency during gestation, with a more robust effect in the lactation-deficient group. In addition, linear regression analysis revealed positive correlations between Mecp2, Reelin, and Ppp1cc, between Gadd45b, Bdnf, and Creb1, and between Egr1 and Dnmt1, respectively. However, there were no correlations, in any gene, between DNA methylation and gene expression. In summary, the interplay between ALA availability during gestation and lactation differentially altered the expression of genes involved in neurogenesis and memory, in the whole brain of the offspring at the end of lactation. The Mecp2 epigenetic status was correlated with ALA availability during gestation. However, the epigenetic status of the genes investigated was not associated with transcript levels, suggesting that either the regulation of these genes is not necessarily under epigenetic control, or that the whole brain model is

  3. DNA methylation level of cyp19a1a and Foxl2 gene related to their expression patterns and reproduction traits during ovary development stages of Japanese flounder (Paralichthys olivaceus).

    PubMed

    Si, Yufeng; Ding, Yuxia; He, Feng; Wen, Haishen; Li, Jifang; Zhao, Junli; Huang, Zhengju

    2016-01-10

    Foxl2 and cyp19a1a genes are crucial for the ovarian development, and Foxl2 could play a direct regulatory role on cyp19a1a transcription. In this study, we aimed to study DNA methylation status and mRNA expression patterns of Foxl2 and cyp19a1a genes during ovarian development of female Japanese flounder. The relative expression level of cyp19a1a and Foxl2 gene during the gonadal development stages was measured by quantitative PCR. Moreover, DNA methylation status in the promoter and coding regions of the two genes was detected by bisulfite sequencing. The estradiol-17β (E2) was measured by radioimmunoassay. The results showed low expression levels of cyp19a1a and Foxl2 genes in stages II and V, while the highest expression levels were detected in stage IV. The variation trend of the methylation level of all CpG sites in promoter and exon 1 of cyp19a1a gene and three CpG rich regions in coding region of Foxl2 gene was negatively associated with their expression levels during the ovarian development. In addition, two CpG sites in promoter and seven CpG sites in exon 1 of cyp19a1a were on the putative transcription factors binding sequence. Further studies showed that the forkhead domain, which is important for Foxl2 binding to cyp19a1a was located in the F1 and F2 region. These results provide a powerful theoretical basis for the regulatory mechanism on Foxl2 regulating cyp19a1a and promoting gonadal differentiation towards the female pathway, and further reveal that Foxl2 and cyp19a1a play a vital role in the female Japanese flounder gonad development. PMID:26343797

  4. Perinatal α-linolenic acid availability alters the expression of genes related to memory and to epigenetic machinery, and the Mecp2 DNA methylation in the whole brain of mouse offspring

    PubMed Central

    He, Fuli; Lupu, Daniel S.; Niculescu, Mihai D.

    2015-01-01

    Many animal and human studies indicated that dietary ω-3 fatty acids could have beneficial roles on brain development, memory, and learning. However, the exact mechanisms involved are far from being clearly understood, especially for α-linolenic acid (ALA), which is the precursor for the ω-3 elongation and desaturation pathways. This study investigated the alterations induced by different intakes of flaxseed oil (containing 50% ALA), during gestation and lactation, upon the expression of genes involved in neurogenesis, memory-related molecular processes, and DNA methylation, in the brains of mouse offspring at the end of lactation (postnatal day 19, P19). In addition, DNA methylation status for the same genes was investigated. Maternal flaxseed oil supplementation during lactation increased the expression of Mecp2, Ppp1cc, and Reelin, while decreasing the expression of Ppp1cb and Dnmt3a. Dnmt1 expression was decreased by postnatal flaxseed oil supplementation but this effect was offset by ALA deficiency during gestation. Mecp2 DNA methylation was decreased by maternal ALA deficiency during gestation, with a more robust effect in the lactation-deficient group. In addition, linear regression analysis revealed positive correlations between Mecp2, Reelin, and Ppp1cc, between Gadd45b, Bdnf, and Creb1, and between Egr1 and Dnmt1, respectively. However, there were no correlations, in any gene, between DNA methylation and gene expression. In summary, the interplay between ALA availability during gestation and lactation differentially altered the expression of genes involved in neurogenesis and memory, in the whole brain of the offspring at the end of lactation. The Mecp2 epigenetic status was correlated with ALA availability during gestation. However, the epigenetic status of the genes investigated was not associated with transcript levels, suggesting that either the regulation of these genes is not necessarily under epigenetic control, or that the whole brain model is

  5. Rapid expansion of the protein disulfide isomerase gene family facilitates the folding of venom peptides.

    PubMed

    Safavi-Hemami, Helena; Li, Qing; Jackson, Ronneshia L; Song, Albert S; Boomsma, Wouter; Bandyopadhyay, Pradip K; Gruber, Christian W; Purcell, Anthony W; Yandell, Mark; Olivera, Baldomero M; Ellgaard, Lars

    2016-03-22

    Formation of correct disulfide bonds in the endoplasmic reticulum is a crucial step for folding proteins destined for secretion. Protein disulfide isomerases (PDIs) play a central role in this process. We report a previously unidentified, hypervariable family of PDIs that represents the most diverse gene family of oxidoreductases described in a single genus to date. These enzymes are highly expressed specifically in the venom glands of predatory cone snails, animals that synthesize a remarkably diverse set of cysteine-rich peptide toxins (conotoxins). Enzymes in this PDI family, termed conotoxin-specific PDIs, significantly and differentially accelerate the kinetics of disulfide-bond formation of several conotoxins. Our results are consistent with a unique biological scenario associated with protein folding: The diversification of a family of foldases can be correlated with the rapid evolution of an unprecedented diversity of disulfide-rich structural domains expressed by venomous marine snails in the superfamily Conoidea. PMID:26957604

  6. Bioreducible cross-linked polymers based on G1 peptide dendrimer as potential gene delivery vectors.

    PubMed

    Li, Chun-Yan; Wang, Hai-Jiao; Cao, Jing-Ming; Zhang, Ji; Yu, Xiao-Qi

    2014-11-24

    A series of cationic polymers based on low generation (G1) peptide dendrimer were synthesized with disulfide-containing linkages. The DNA binding abilities of the target polymers were studied by gel electrophoresis and fluorescence quenching assay. The bioreducible property of the disulfide-containing polymers P2 and P3 was also investigated in the presence of dithiothreitol (DTT). Results from dynamic light scattering (DLS) and transmission electron microscopy (TEM) assays reveal that these materials may condense DNA into nanoparticles with proper sizes and zeta-potentials. In vitro cell experiments show that compared to branched 25 KDa PEI, P2 and P3 may exhibit much higher gene transfection efficiency and lower cytotoxicity in both HEK293 and U-2OS cells. Additionally, polymer prepared from Michael addition gives better gene transfection ability, while polymer prepared from ring-opening reaction has better serum tolerance. Results indicate that these polymers might be promising non-viral gene vectors for their easy preparation, very low cytotoxicity, and good transfection efficiency. PMID:25282264

  7. Pear transformed with a lytic peptide gene for disease control affects nontarget organism, pear Psylla (Homoptera: Psyllidae)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Pear plants were transformed with D5C1a construct containing a gene that produces lytic peptide which is excreted into the intercellular spaces of pear leaves to impart resistance to fireblight, Erwinia amylovora (Burrill). The biology and behavior of pear psylla (Cacopsylla pyricola Foerster) was ...

  8. Monocyte-mediated regulation of genes by the amyloid and prion peptides in SH-SY5Y neuroblastoma cells.

    PubMed

    Morte, Beatriz; Martínez, Tamara; Zambrano, Alberto; Pascual, Angel

    2011-05-01

    Alzheimer's disease as well as prion-related encephalopathies are neurodegenerative disorders of the central nervous system, which cause mental deterioration and progressive dementia. Both pathologies appear to be primarily associated with the pathological accumulation and deposit of β-amyloid or prion peptides in the brain, and it has been even suggested that neurotoxicity induced by these peptides would be associated to essentially similar pathogenic mechanisms, in particular to those that follow the activation of microglial cells. To probe whether the neurotoxic effects induced by the β-amyloid and prion peptides are actually mediated by similar glial-associated mechanisms, we have examined the differential expression of genes in SH-SY5Y neuroblastoma cells incubated with conditioned media from β-amyloid or prion-stimulated THP-1 monocytic cells. According to microarray analysis, not many coincidences are observed and only four genes (Hint3, Psph, Daam1 and c-Jun) appear to be commonly upregulated by both peptides. Furthermore, c-Jun appears to be involved in the cell death mediated by both peptides. PMID:21303680

  9. Agrobacterium-Mediated Disruption of a Nonribosomal Peptide Synthetase Gene in the Invertebrate Pathogen Metarhizium anisopliae Reveals a Peptide Spore Factor▿ †

    PubMed Central

    Moon, Yong-Sun; Donzelli, Bruno G. G.; Krasnoff, Stuart B.; McLane, Heather; Griggs, Mike H.; Cooke, Peter; Vandenberg, John D.; Gibson, Donna M.; Churchill, Alice C. L.

    2008-01-01

    Numerous secondary metabolites have been isolated from the insect pathogenic fungus Metarhizium anisopliae, but the roles of these compounds as virulence factors in disease development are poorly understood. We targeted for disruption by Agrobacterium tumefaciens-mediated transformation a putative nonribosomal peptide synthetase (NPS) gene, MaNPS1. Four of six gene disruption mutants identified were examined further. Chemical analyses showed the presence of serinocyclins, cyclic heptapeptides, in the extracts of conidia of control strains, whereas the compounds were undetectable in ΔManps1 mutants treated identically or in other developmental stages, suggesting that MaNPS1 encodes a serinocyclin synthetase. Production of the cyclic depsipeptide destruxins, M. anisopliae metabolites also predicted to be synthesized by an NPS, was similar in ΔManps1 mutant and control strains, indicating that MaNPS1 does not contribute to destruxin biosynthesis. Surprisingly, a MaNPS1 fragment detected DNA polymorphisms that correlated with relative destruxin levels produced in vitro, and MaNPS1 was expressed concurrently with in vitro destruxin production. ΔManps1 mutants exhibited in vitro development and responses to external stresses comparable to control strains. No detectable differences in pathogenicity of the ΔManps1 mutants were observed in bioassays against beet armyworm and Colorado potato beetle in comparison to control strains. This is the first report of targeted disruption of a secondary metabolite gene in M. anisopliae, which revealed a novel cyclic peptide spore factor. PMID:18502925

  10. The effect of peptidoglycan enriched diets on antimicrobial peptide gene expression in rainbow trout (Oncorhynchus mykiss).

    PubMed

    Casadei, Elisa; Bird, Steve; Vecino, Jose L González; Wadsworth, Simon; Secombes, Christopher J

    2013-02-01

    The aim of this study was to investigate the effect of feeding rainbow trout (Oncorhynchus mykiss) peptidoglycan (PG) enriched diets on antimicrobial peptide (AMP) gene expression. Fish were divided into 5 groups and fed diets containing 0, 5, 10, 50 and 100 mg PG/Kg, and sampled 1, 7 and 14 days later. The expression of eight AMP genes (four defensins, two cathelicidins and two liver expressed AMPs) was determined in skin, gill, gut and liver, tissues important for first lines of defence or production of acute phase proteins. Up-regulation of many AMPs was found after feeding the PG enriched diets, with sequential expression seen over the time course studied, where defensins were typically expressed early and cathelicidins and LEAPs later on. A number of clear differences in AMP responsiveness between the tissues examined were also apparent. Of the four PG concentrations used, 5 mg PG/Kg did not always elicit AMP gene induction or to the same degree as seen with the other diets. The three higher dose groups generally showed similar trends although differences in fold change were more pronounced in the 50 and 100 mg PG/Kg groups. Curiously several AMPs were down-regulated after 14 days of feeding in gills, gut and liver. Nevertheless, overall the PG enriched diets had a positive effect on AMP expression. Further investigations now need to be undertaken to confirm whether this higher AMP gene expression correlates with protection against common bacterial diseases and if PG enriched diets have value as a means to temporarily boost the piscine immune system. PMID:23220715

  11. Nonribosomal peptide synthase gene clusters for lipopeptide biosynthesis in Bacillus subtilis 916 and their phenotypic functions.

    PubMed

    Luo, Chuping; Liu, Xuehui; Zhou, Huafei; Wang, Xiaoyu; Chen, Zhiyi

    2015-01-01

    Bacillus cyclic lipopeptides (LPs) have been well studied for their phytopathogen-antagonistic activities. Recently, research has shown that these LPs also contribute to the phenotypic features of Bacillus strains, such as hemolytic activity, swarming motility, biofilm formation, and colony morphology. Bacillus subtilis 916 not only coproduces the three families of well-known LPs, i.e., surfactins, bacillomycin Ls (iturin family), and fengycins, but also produces a new family of LP called locillomycins. The genome of B. subtilis 916 contains four nonribosomal peptide synthase (NRPS) gene clusters, srf, bmy, fen, and loc, which are responsible for the biosynthesis of surfactins, bacillomycin Ls, fengycins, and locillomycins, respectively. By studying B. subtilis 916 mutants lacking production of one, two, or three LPs, we attempted to unveil the connections between LPs and phenotypic features. We demonstrated that bacillomycin Ls and fengycins contribute mainly to antifungal activity. Although surfactins have weak antifungal activity in vitro, the strain mutated in srfAA had significantly decreased antifungal activity. This may be due to the impaired productions of fengycins and bacillomycin Ls. We also found that the disruption of any LP gene cluster other than fen resulted in a change in colony morphology. While surfactins and bacillomycin Ls play very important roles in hemolytic activity, swarming motility, and biofilm formation, the fengycins and locillomycins had little influence on these phenotypic features. In conclusion, B. subtilis 916 coproduces four families of LPs which contribute to the phenotypic features of B. subtilis 916 in an intricate way. PMID:25362061

  12. Nonribosomal Peptide Synthase Gene Clusters for Lipopeptide Biosynthesis in Bacillus subtilis 916 and Their Phenotypic Functions

    PubMed Central

    Liu, Xuehui; Zhou, Huafei; Wang, Xiaoyu

    2014-01-01

    Bacillus cyclic lipopeptides (LPs) have been well studied for their phytopathogen-antagonistic activities. Recently, research has shown that these LPs also contribute to the phenotypic features of Bacillus strains, such as hemolytic activity, swarming motility, biofilm formation, and colony morphology. Bacillus subtilis 916 not only coproduces the three families of well-known LPs, i.e., surfactins, bacillomycin Ls (iturin family), and fengycins, but also produces a new family of LP called locillomycins. The genome of B. subtilis 916 contains four nonribosomal peptide synthase (NRPS) gene clusters, srf, bmy, fen, and loc, which are responsible for the biosynthesis of surfactins, bacillomycin Ls, fengycins, and locillomycins, respectively. By studying B. subtilis 916 mutants lacking production of one, two, or three LPs, we attempted to unveil the connections between LPs and phenotypic features. We demonstrated that bacillomycin Ls and fengycins contribute mainly to antifungal activity. Although surfactins have weak antifungal activity in vitro, the strain mutated in srfAA had significantly decreased antifungal activity. This may be due to the impaired productions of fengycins and bacillomycin Ls. We also found that the disruption of any LP gene cluster other than fen resulted in a change in colony morphology. While surfactins and bacillomycin Ls play very important roles in hemolytic activity, swarming motility, and biofilm formation, the fengycins and locillomycins had little influence on these phenotypic features. In conclusion, B. subtilis 916 coproduces four families of LPs which contribute to the phenotypic features of B. subtilis 916 in an intricate way. PMID:25362061

  13. The gene encoding the iron regulatory peptide hepcidin is regulated by anemia, hypoxia, and inflammation.

    PubMed

    Nicolas, Gaël; Chauvet, Caroline; Viatte, Lydie; Danan, Jean Louis; Bigard, Xavier; Devaux, Isabelle; Beaumont, Carole; Kahn, Axel; Vaulont, Sophie

    2002-10-01

    The present study was aimed at determining whether hepcidin, a recently identified peptide involved in iron metabolism, plays a role in conditions associated with both iron overload and iron deficiency. Hepcidin mRNA levels were assessed in two models of anemia, acute hemolysis provoked by phenylhydrazine and bleeding provoked by repeated phlebotomies. Hepcidin response to hypoxia was also studied, both ex vivo, in human hepatoma cells, and in vivo. Anemia and hypoxia were associated with a dramatic decrease in liver hepcidin gene expression, which may account for the increase in iron release from reticuloendothelial cells and increase in iron absorption frequently observed in these situations. A single injection of turpentine for 16 hours induced a sixfold increase in liver hepcidin mRNA levels and a twofold decrease in serum iron. The hyposideremic effect of turpentine was completely blunted in hepcidin-deficient mice, revealing hepcidin participation in anemia of inflammatory states. These modifications of hepcidin gene expression further suggest a key role for hepcidin in iron homeostasis under various pathophysiological conditions, which may support the pharmaceutical use of hepcidin agonists and antagonists in various iron homeostasis disorders. PMID:12370282

  14. The gene encoding the iron regulatory peptide hepcidin is regulated by anemia, hypoxia, and inflammation

    PubMed Central

    Nicolas, Gaël; Chauvet, Caroline; Viatte, Lydie; Danan, Jean Louis; Bigard, Xavier; Devaux, Isabelle; Beaumont, Carole; Kahn, Axel; Vaulont, Sophie

    2002-01-01

    The present study was aimed at determining whether hepcidin, a recently identified peptide involved in iron metabolism, plays a role in conditions associated with both iron overload and iron deficiency. Hepcidin mRNA levels were assessed in two models of anemia, acute hemolysis provoked by phenylhydrazine and bleeding provoked by repeated phlebotomies. Hepcidin response to hypoxia was also studied, both ex vivo, in human hepatoma cells, and in vivo. Anemia and hypoxia were associated with a dramatic decrease in liver hepcidin gene expression, which may account for the increase in iron release from reticuloendothelial cells and increase in iron absorption frequently observed in these situations. A single injection of turpentine for 16 hours induced a sixfold increase in liver hepcidin mRNA levels and a twofold decrease in serum iron. The hyposideremic effect of turpentine was completely blunted in hepcidin-deficient mice, revealing hepcidin participation in anemia of inflammatory states. These modifications of hepcidin gene expression further suggest a key role for hepcidin in iron homeostasis under various pathophysiological conditions, which may support the pharmaceutical use of hepcidin agonists and antagonists in various iron homeostasis disorders. PMID:12370282

  15. A novel peptide-modified and gene-activated biomimetic bone matrix accelerating bone regeneration.

    PubMed

    Pan, Haitao; Zheng, Qixin; Yang, Shuhua; Guo, Xiaodong; Wu, Bin; Zou, Zhenwei; Duan, Zhixia

    2014-08-01

    The osteogenic differentiation of bone marrow stromal cells (BMSCs) can be regulated by systemic or local growth factor, especially by transforming growth factor beta 1 (TGF-β1). However, how to maintain the bioactivity of exogenous TGF-β1 is a great challenge due to its short half-life time. The most promising solution is to transfer TGF-β1 gene into seed cells through transgenic technology and then transgenic cells to continuously secret endogenous TGF-β1 protein via gene expression. In this study, a novel non-viral vector (K)16GRGDSPC was chemically linked to bioactive bone matrices PLGA-[ASP-PEG]n using cross-linker to construct a novel non-viral gene transfer system. TGF-β1 gene was incubated with this system and subsequently rabbit-derived BMSCs were co-cultured with this gene-activated PLGA-[ASP-PEG]n, while co-cultured with PLGA-[ASP-PEG]n modified with (K)16GRGDSPC only and original PLGA-[ASP-PEG]n as control. Thus we fabricated three kinds of composites: Group A (BMSCs-TGF-β1DNA-(K)16GRGDSPC-PLGA-[ASP-PEG]n composite); Group B (BMSCs-(K)16GRGDSPC-PLGA-[ASP-PEG]n composite); and Group C (BMSCs-PLGA-[ASP-PEG]n composite). TGF-β1 and other osteogenic phenotype markers of alkaline phosphatase, osteocalcin, osteopontin and type I collagen in Group A were all significantly higher than the other two groups ex vivo. In vivo, 15-mm long segmental rabbit bone defects were created and randomly implanted the aforementioned composites separately, and then fixed with plate-screws. The results demonstrated that the implants in Group A significantly accelerated bone regeneration compared with the other implants based on X-rays, histological and biomechanical examinations. Therefore, we conclude this novel peptide-modified and gene-activated biomimetic bone matrix of TGF-β1DNA-(K)16GRGDSPC-PLGA-[ASP-PEG]n is a very promising scaffold biomaterial for accelerating bone regeneration. PMID:24115366

  16. Direct comparison between genomic constitution and flavonoid contents in Allium multiple alien addition lines reveals chromosomal locations of genes related to biosynthesis from dihydrokaempferol to quercetin glucosides in scaly leaf of shallot (Allium cepa L.).

    PubMed

    Masuzaki, S; Shigyo, M; Yamauchi, N

    2006-02-01

    The extrachromosome 5A of shallot (Allium cepa L., genomes AA) has an important role in flavonoid biosynthesis in the scaly leaf of Allium fistulosum-shallot monosomic addition lines (FF+nA). This study deals with the production and biochemical characterisation of A. fistulosum-shallot multiple alien addition lines carrying at least 5A to determine the chromosomal locations of genes for quercetin formation. The multiple alien additions were selected from the crossing between allotriploid FFA (female symbol) and A. fistulosum (male symbol). The 113 plants obtained from this cross were analysed by a chromosome 5A-specific PGI isozyme marker of shallot. Thirty plants were preliminarily selected for an alien addition carrying 5A. The chromosome numbers of the 30 plants varied from 18 to 23. The other extrachromosomes in 19 plants were completely identified by using seven other chromosome markers of shallot. High-performance liquid chromatography analyses of the 19 multiple additions were conducted to identify the flavonoid compounds produced in the scaly leaves. Direct comparisons between the chromosomal constitution and the flavonoid contents of the multiple alien additions revealed that a flavonoid 3'-hydroxylase (F3'H) gene for the synthesis of quercetin from kaempferol was located on 7A and that an anonymous gene involved in the glucosidation of quercetin was on 3A or 4A. As a result of supplemental SCAR analyses by using genomic DNAs from two complete sets of A. fistulosum-shallot monosomic additions, we have assigned F3'H to 7A and flavonol synthase to 4A. PMID:16411131

  17. Degradable polyethylenimine derivate coupled to a bifunctional peptide R13 as a new gene-delivery vector

    PubMed Central

    Liu, Kehai; Wang, Xiaoyu; Fan, Wei; Zhu, Qing; Yang, Jingya; Gao, Jing; Gao, Shen

    2012-01-01

    Background To solve the efficiency versus cytotoxicity and tumor-targeting problems of polyethylenimine (PEI) used as a nonviral gene delivery vector, a degradable PEI derivate coupled to a bifunctional peptide R13 was developed. Methods First, we synthesized a degradable PEI derivate by crosslinking low-molecular-weight PEI with pluronic P123, then used tumor-targeting peptide arginine-glycine-aspartate-cysteine (RGDC), in conjunction with the cell-penetrating peptide Tat (49–57), to yield a bifunctional peptide RGDC-Tat (49–57) named R13, which can improve cell selection and increase cellular uptake, and, lastly, adopted R13 to modify the PEI derivates so as to prepare a new polymeric gene vector (P123-PEI-R13). The new gene vector was characterized in terms of its chemical structure and biophysical parameters. We also investigated the specificity, cytotoxicity, and gene transfection efficiency of this vector in αvβ3-positive human cervical carcinoma Hela cells and murine melanoma B16 cells in vitro. Results The vector showed controlled degradation, strong targeting specificity to αvβ3 receptor, and noncytotoxicity in Hela cells and B16 cells at higher doses, in contrast to PEI 25 KDa. The particle size of P123-PEI-R13/DNA complexes was around 100–250 nm, with proper zeta potential. The nanoparticles can protect plasmid DNA from being digested by DNase I at a concentration of 6 U DNase I/μg DNA. The nanoparticles were resistant to dissociation induced by 50% fetal bovine serum and 600 μg/mL sodium heparin. P123-PEI-R13 also revealed higher transfection efficiency in two cell lines as compared with PEI 25 KDa. Conclusion P123-PEI-R13 is a potential candidate as a safe and efficient gene-delivery carrier for gene therapy. PMID:22412301

  18. De-novo assembly and characterization of Chlorella minutissima UTEX2341 transcriptome by paired-end sequencing and the identification of genes related to the biosynthesis of lipids for biodiesel.

    PubMed

    Yu, Mingjia; Yang, Shanjun; Lin, Xiangzhi

    2016-02-01

    Chlorella minutissima is considered to be one of the promising feedstocks for biofuels in the future. In this study, the transcriptome from the oil-rich strain UTEX2341 of C. minutissima was generated based on Illumina paired-end sequencing. Through de-novo assembly, a total of 14,905 isogenes were obtained and compacted into 6216 unigenes. A total of 80% of the unigenes were assigned with GO terms and were further subdivided into 55 sub-categories. KEGG analysis demonstrated that 37.2% of the unigenes could be accessed and mapped into 278 pathways. Interestingly, the genes that encoded key enzymes that are involved in the biosynthesis, elongation, and metabolism of fatty acids were identified, including malonyl-CoA-ACP transacylase, 3-ketoacyl-ACP synthase, 3-ketoacyl-ACP reductase, and others. Moreover, the genes that are involved in triacylglycerol (TAG) biosynthesis and metabolism were also observed. Therefore, the transcriptome analysis of C. minutissima UTEX2341 not only supplies comprehensive insight into the molecular pathway that is involved in the biosynthesis of biofuel precursors but also provides substantial valuable genomic resources to accelerate the further development and utilization of biofuels. PMID:26590019

  19. The myeloid-binding peptide adenoviral vector enables multi-organ vascular endothelial gene targeting.

    PubMed

    Lu, Zhi Hong; Kaliberov, Sergey; Zhang, Jingzhu; Muz, Barbara; Azab, Abdel K; Sohn, Rebecca E; Kaliberova, Lyudmila; Du, Yingqiu; Curiel, David T; Arbeit, Jeffrey M

    2014-08-01

    Vascular endothelial cells (ECs) are ideal gene therapy targets as they provide widespread tissue access and are the first contact surfaces following intravenous vector administration. Human recombinant adenovirus serotype 5 (Ad5) is the most frequently used gene transfer system because of its appreciable transgene payload capacity and lack of somatic mutation risk. However, standard Ad5 vectors predominantly transduce liver but not the vasculature following intravenous administration. We recently developed an Ad5 vector with a myeloid cell-binding peptide (MBP) incorporated into the knob-deleted, T4 fibritin chimeric fiber (Ad.MBP). This vector was shown to transduce pulmonary ECs presumably via a vector handoff mechanism. Here we tested the body-wide tropism of the Ad.MBP vector, its myeloid cell necessity, and vector-EC expression dose response. Using comprehensive multi-organ co-immunofluorescence analysis, we discovered that Ad.MBP produced widespread EC transduction in the lung, heart, kidney, skeletal muscle, pancreas, small bowel, and brain. Surprisingly, Ad.MBP retained hepatocyte tropism albeit at a reduced frequency compared with the standard Ad5. While binding specifically to myeloid cells ex vivo, multi-organ Ad.MBP expression was not dependent on circulating monocytes or macrophages. Ad.MBP dose de-escalation maintained full lung-targeting capacity but drastically reduced transgene expression in other organs. Swapping the EC-specific ROBO4 for the CMV promoter/enhancer abrogated hepatocyte expression but also reduced gene expression in other organs. Collectively, our multilevel targeting strategy could enable therapeutic biological production in previously inaccessible organs that pertain to the most debilitating or lethal human diseases. PMID:24955893

  20. Leucocyte expression of genes implicated in the plasminogen activation cascade is modulated by yoghurt peptides.

    PubMed

    Theodorou, Georgios; Politis, Ioannis

    2016-08-01

    The urokinase-plasminogen activator (u-PA), its receptor (u-PAR) and the inhibitors of u-PA (PAI-1 and PAI-2) provide a multi-molecular system in leucocytes that exerts pleiotropic functions influencing the development of inflammatory and immune responses. The objective of the present study was to examine the ability of water soluble extracts (WSE) obtained from traditional Greek yoghurt made from bovine or ovine milk to modulate the expression of u-PA, u-PAR, PAI-1 and PAI-2 in ovine monocytes and neutrophils. WSE were obtained from 8 commercial traditional type Greek yoghurts made from ovine or bovine milk. WSE upregulated the expression of all 4 u-PA related genes in monocytes but the upregulation was much higher in the PAI-1 (10-fold) than in u-PA and u-PAR (3-4 fold) thus, shifting the system towards inhibition. In line with this observation, WSE reduced total and membrane-bound u-PA activity in monocytes. In neutrophils, WSE caused small (50-60%) but significant (P < 0·05) reductions in expression of u-PAR and PAI-2 but had no effect on expression of u-PA, PAI-1 and on total cell-associated and membrane-bound u-PA activity. WSE from yoghurts made from bovine or ovine milk were essentially equally effective in affecting the u-PA system except for the u-PAR gene in ovine neutrophils that was affected (reduced) by the ovine and not the bovine WSE. In conclusion, peptides present in WSE modulated the expression of u-PA related genes but the effect was much more prominent in monocytes than in neutrophils. PMID:27600972

  1. The Novel Gene CRNDE Encodes a Nuclear Peptide (CRNDEP) Which Is Overexpressed in Highly Proliferating Tissues

    PubMed Central

    Szafron, Lukasz Michal; Balcerak, Anna; Grzybowska, Ewa Anna; Pienkowska-Grela, Barbara; Felisiak-Golabek, Anna; Podgorska, Agnieszka; Kulesza, Magdalena; Nowak, Natalia; Pomorski, Pawel; Wysocki, Juliusz; Rubel, Tymon; Dansonka-Mieszkowska, Agnieszka; Konopka, Bozena; Lukasik, Martyna; Kupryjanczyk, Jolanta

    2015-01-01

    CRNDE, recently described as the lncRNA-coding gene, is overexpressed at RNA level in human malignancies. Its role in gametogenesis, cellular differentiation and pluripotency has been suggested as well. Herein, we aimed to verify our hypothesis that the CRNDE gene may encode a protein product, CRNDEP. By using bioinformatics methods, we identified the 84-amino acid ORF encoded by one of two CRNDE transcripts, previously described by our research team. This ORF was cloned into two expression vectors, subsequently utilized in localization studies in HeLa cells. We also developed a polyclonal antibody against CRNDEP. Its specificity was confirmed in immunohistochemical, cellular localization, Western blot and immunoprecipitation experiments, as well as by showing a statistically significant decrease of endogenous CRNDEP expression in the cells with transient shRNA-mediated knockdown of CRNDE. Endogenous CRNDEP localizes predominantly to the nucleus and its expression seems to be elevated in highly proliferating tissues, like the parabasal layer of the squamous epithelium, intestinal crypts or spermatocytes. After its artificial overexpression in HeLa cells, in a fusion with either the EGFP or DsRed Monomer fluorescent tag, CRNDEP seems to stimulate the formation of stress granules and localize to them. Although the exact role of CRNDEP is unknown, our preliminary results suggest that it may be involved in the regulation of the cell proliferation. Possibly, CRNDEP also participates in oxygen metabolism, considering our in silico results, and the correlation between its enforced overexpression and the formation of stress granules. This is the first report showing the existence of a peptide encoded by the CRNDE gene. PMID:25978564

  2. Construction of a novel peptide nucleic acid piezoelectric gene sensor microarray detection system.

    PubMed

    Chen, Ming; Liu, Minghua; Yu, Lili; Cai, Guoru; Chen, Qinghai; Wu, Rong; Wang, Feng; Zhang, Bo; Jiang, Tianlun; Fu, Welling

    2005-08-01

    A novel 2 x 5 clamped style piezoelectric gene sensor microarray has been successfully constructed. Every crystal unit of the fabricated gene sensor can oscillate independently without interfering with each other. The bis-peptide nucleic acid (bis-PNA) probe, which can combine with target DNA or RNA sequences more effectively and specifically than a DNA probe, was designed and immobilized on the surface of the gene sensor microarray to substitute the conventional DNA probe for direct detection of the hepatitis B virus (HBV) genomic DNA. Detection conditions were then explored and optimized. Results showed that PBS buffer of pH 6.8, an ion concentration of 20 mmol/liter, and a probe concentration of 1.5 micromol/liter were optimal for the detection system. Under such optimized experimental conditions, the specificity of bis-PNA was proved much higher than that of DNA probe. The relationship between quantity of target and decrease of frequency showed a typical saturation curve when concentrations of target HBV DNA varied from 10 pg/liter to 100 microg/liter, and 10 microg/liter was the watershed, with a statistic linear regression equation of I gC = -2.7455 + 0.0691 deltaF and the correlating coefficient of 0.9923. Fortunately, this is exactly the most ordinary variant range of the HBV virus concentration in clinical hepatitis samples. So, a good technical platform is successfully constructed and it will be applied to detect HBV quantitatively in clinical samples. PMID:16193990

  3. A Peptide Derived from G0/G1 Switch Gene 2 Acts as Noncompetitive Inhibitor of Adipose Triglyceride Lipase*

    PubMed Central

    Cerk, Ines K.; Salzburger, Barbara; Boeszoermenyi, Andras; Heier, Christoph; Pillip, Christoph; Romauch, Matthias; Schweiger, Martina; Cornaciu, Irina; Lass, Achim; Zimmermann, Robert; Zechner, Rudolf; Oberer, Monika

    2014-01-01

    The protein G0/G1 switch gene 2 (G0S2) is a small basic protein that functions as an endogenous inhibitor of adipose triglyceride lipase (ATGL), a key enzyme in intracellular lipolysis. In this study, we identified a short sequence covering residues Lys-20 to Ala-52 in G0S2 that is still fully capable of inhibiting mouse and human ATGL. We found that a synthetic peptide corresponding to this region inhibits ATGL in a noncompetitive manner in the nanomolar range. This peptide is highly selective for ATGL and does not inhibit other lipases, including hormone-sensitive lipase, monoacylglycerol lipase, lipoprotein lipase, and patatin domain-containing phospholipases 6 and 7. Because increased lipolysis is linked to the development of metabolic disorders, the inhibition of ATGL by G0S2-derived peptides may represent a novel therapeutic tool to modulate lipolysis. PMID:25258314

  4. Salmonella enterica Serovar Enteritidis Antimicrobial Peptide Resistance Genes Aid in Defense against Chicken Innate Immunity, Fecal Shedding, and Egg Deposition

    PubMed Central

    McKelvey, Jessica A.; Yang, Ming; Jiang, Yanhua

    2014-01-01

    Salmonella enterica serovar Enteritidis (S. Enteritidis) is a major etiologic agent of nontyphoid salmonellosis in the United States. S. Enteritidis persistently and silently colonizes the intestinal and reproductive tract of laying hens, resulting in contaminated poultry products. The consumption of contaminated poultry products has been identified as a significant risk factor for human salmonellosis. To understand the mechanisms S. Enteritidis utilizes to colonize and persist in laying hens, we used selective capture of transcribed sequences to identify genes overexpressed in the HD11 chicken macrophage cell line and in primary chicken oviduct epithelial cells. From the 15 genes found to be overexpressed in both cell types, we characterized the antimicrobial peptide resistance (AMPR) genes, virK and ybjX, in vitro and in vivo. In vitro, AMPR genes were required for natural morphology, motility, secretion, defense against detergents such as EDTA and bile salts, and resistance to antimicrobial peptides polymyxin B and avian β-defensins. From this, we inferred the AMPR genes play a role in outer membrane stability and/or modulation. In the intestinal tract, AMPR genes were involved in early intestinal colonization and fecal shedding. In the reproductive tract, virK was required in early colonization whereas a deletion of ybjX caused prolonged ovary colonization and egg deposition. Data from the present study indicate that AMPR genes are differentially utilized in various host environments, which may ultimately assist S. Enteritidis in persistent and silent colonization of chickens. PMID:25267840

  5. Cloning, characterization, and embryonic expression analysis of the Drosophila melanogaster gene encoding insulin/relaxin-like peptide.

    PubMed

    Nasonkin, Igor O; Alikasifoglu, Ayfer; Barrette, Terry; Cheng, Michael M; Thomas, Pamela M; Nikitin, Alexey G

    2002-07-12

    Insulin is one of the key peptide hormones that regulates growth and metabolism in vertebrates. Evolutionary conservation of many elements of the insulin/IGF signaling network makes it possible to study the basic genetic function of this pathway in lower metazoan models such as Drosophila. Here we report the cloning and characterization of the gene for Drosophila insulin/relaxin-like peptide (DIRLP). The predicted protein structure of DIRLP greatly resembles typical insulin structure and contains features that differentiate it from the Drosophila juvenile hormone, another member of the insulin family. The Dirlp gene is represented as a single copy in the Drosophila melanogaster genome (compared to multiple copies for Drosophila juvenile hormone) and shows evolutionary conservation of genetic structure. The gene was mapped to the Drosophila chromosome 3, region 67D2. In situ hybridization of whole-mount Drosophila embryos with Dirlp antisense RNA probe reveals early embryonic mesodermal/ventral furrow expression pattern, consistent with earlier observation of the insulin protein immunoreactivity in Drosophila embryos. The in situ hybridization pattern was found to be identical to that obtained during immunohistochemistry analysis of the Drosophila embryos using various insulin monoclonal and polyclonal antibodies that do not recognize Drosophila juvenile hormone, supporting the idea that Dirlp is a possible Drosophila insulin ortholog. Identification of the gene for DIRLP provides a new approach for study of the regulatory pathway of the insulin family of peptides. PMID:12150949

  6. In Vitro Efficient Transfection by CM18-Tat11 Hybrid Peptide: A New Tool for Gene-Delivery Applications

    PubMed Central

    Salomone, Fabrizio; Cardarelli, Francesco; Signore, Giovanni; Boccardi, Claudia; Beltram, Fabio

    2013-01-01

    Cell penetrating peptides (CPPs) are actively researched as non-viral molecular carriers for the controlled delivery of nucleic acids into cells, but widespread application is severely hampered by their trapping into endosomes. Here we show that the recently introduced endosomolytic CM18-Tat11 hybrid peptide (KWKLFKKIGAVLKVLTTG-YGRKKRRQRRR, residues 1-7 of Cecropin-A, 2-12 of Melittin, and 47-57 of HIV-1 Tat protein) can be exploited to obtain a self-assembled peptide-DNA vector which maintains the CM18-Tat11 ability to enter cells and destabilize vesicular membranes, concomitantly yielding high DNA transfection efficiency with no detectable cytotoxic effects. Different peptide-DNA stoichiometric ratios were tested to optimize vector size, charge, and stability characteristics. The transfection efficiency of selected candidates is quantitatively investigated by the luciferase-reporter assay. Vector intracellular trafficking is monitored in real time and in live cells by confocal microscopy. In particular, fluorescence resonant energy transfer (FRET) between suitably-labeled peptide and DNA modules was exploited to monitor complex disassembly during endocytosis, and this process is correlated to transfection timing and efficiency. We argue that these results can open the way to the rational design and application of CM18-Tat11–based systems for gene-delivery purposes. PMID:23922923

  7. Cell penetrating peptide-based polyplexes shelled with polysaccharide to improve stability and gene transfection

    NASA Astrophysics Data System (ADS)

    Li, Wenyu; Liu, Yajie; Du, Jianwei; Ren, Kefeng; Wang, Youxiang

    2015-04-01

    Cell-penetrating peptides (CPP) have been widely developed as a strategy to enhance cell penetrating ability and transfection. In this work, octa-arginine modified dextran gene vector with pH-sensitivity was developed via host-guest interactions. α-Cyclodextrin was modified with octa-arginine (CDR), which had excellent cell penetrating ability. Dextran was selected as a backbone and modified with azobenzene as guest units by acid-labile imine bonds (Az-I-Dex). The supramolecular polymer CDR/Az-I-Dex with high a C/A molar ratio (molar ratio of CD on CDR to Az on Az-I-Dex) was unfavorable for DNA condensation. The dextran shell of CDR/Az-I-Dex/DNA polyplexes improved the stability under physiological conditions. However, once treated with acetate buffer (pH 5.4) for 3 h, large aggregates formed rapidly due to the cleavage of the dextran shell. As expected, the vector had cell viability of 80% even when the CDR concentration increased to 100 μg mL-1. Moreover, due to the effective cellular uptake efficiency, CDR/Az-I-Dex/DNA polyplexes had 6-300 times higher transfection efficiency than CDR/DNA polyplexes. It was even higher than high molecular weight PLL-based polyplexes of HEK293 T cells. Importantly, chloroquine as an endosomal escape agent could not improve the transfection of CDR/Az-I-Dex/DNA polyplexes, which indicated that the CDR/Az-I-Dex supramolecular polymer had its own ability for endosomal escape. These results suggested that the CPP-based polyplexes shelled with polysaccharide can be promising non-viral gene delivery carriers.Cell-penetrating peptides (CPP) have been widely developed as a strategy to enhance cell penetrating ability and transfection. In this work, octa-arginine modified dextran gene vector with pH-sensitivity was developed via host-guest interactions. α-Cyclodextrin was modified with octa-arginine (CDR), which had excellent cell penetrating ability. Dextran was selected as a backbone and modified with azobenzene as guest units by acid

  8. Maternal dietary vitamin D carry-over alters offspring growth, skeletal mineralisation and tissue mRNA expressions of genes related to vitamin D, calcium and phosphorus homoeostasis in swine.

    PubMed

    Amundson, Laura A; Hernandez, Laura L; Laporta, Jimena; Crenshaw, Thomas D

    2016-09-01

    Maternal dietary vitamin D carry-over effects were assessed in young pigs to characterise skeletal abnormalities in a diet-induced model of kyphosis. Bone abnormalities were previously induced and bone mineral density (BMD) reduced in offspring from sows fed diets with inadequate vitamin D3. In a nested design, pigs from sows (n 23) fed diets with 0 (-D), 8·125 (+D) or 43·750 (++D) µg D3/kg from breeding through lactation were weaned and, within litter, fed nursery diets arranged as a 2×2 factorial design with 0 (-D) or 7·0 (+D) µg D3/kg, each with 95 % (95P) or 120 % (120P) of P requirements. Selected pigs were euthanised before colostrum consumption at birth (0 weeks, n 23), weaning (3 weeks, n 22) and after a growth period (8 weeks, n 185) for BMD, bone mechanical tests and tissue mRNA analysis. Pigs produced by +D or ++D sows had increased gain at 3 weeks (P<0·05), and at 8 weeks had increased BMD and improved femur mechanical properties. However, responses to nursery diets depended on maternal diets (P<0·05). Relative mRNA expressions of genes revealed a maternal dietary influence at birth in bone osteocalcin and at weaning in kidney 24-hydroxylase (P<0·05). Nursery treatments affected mRNA expressions at 8 weeks. Detection of a maternal and nursery diet interaction (P<0·05) provided insights into the long-term effects of maternal nutritional inputs. Characterising early stages of bone abnormalities provided inferences for humans and animals about maternal dietary influence on offspring skeletal health. PMID:27480125

  9. Expression of antimicrobial peptide genes in Bombyx mori gut modulated by oral bacterial infection and development.

    PubMed

    Wu, Shan; Zhang, Xiaofeng; He, Yongqiang; Shuai, Jiangbing; Chen, Xiaomei; Ling, Erjun

    2010-11-01

    Although Bombyx mori systematic immunity is extensively studied, little is known about the silkworm's intestine-specific responses to bacterial infection. Antimicrobial peptides (AMPs) gene expression analysis of B. mori intestinal tissue to oral infection with the Gram-positive (Staphylococcus aureus) and -negative (Escherichia coli) bacteria revealed that there is specificity in the interaction between host immune responses and parasite types. Neither Att1 nor Leb could be stimulated by S. aureus and E. coli. However, CecA1, Glo1, Glo2, Glo3, Glo4 and Lys, could only be trigged by S. aureus. On the contrary, E. coli stimulation caused the decrease in the expression of CecA1, Glo3 and Glo4 in some time points. Interestingly, there is regional specificity in the silkworm local gut immunity. During the immune response, the increase in Def, Hem and LLP3 was only detected in the foregut and midgut. For CecB1, CecD, LLP2 and Mor, after orally administered with E. coli, the up-regulation was only limited in the midgut and hindgut. CecE was the only AMP that positively responses to the both bacteria in all the testing situations. With development, the expression levels of the AMPs were also changed dramatically. That is, at spinning and prepupa stages, a large increase in the expression of CecA1, CecB1, CecD, CecE, Glo1, Glo2, Glo3, Glo4, Leb, Def, Hem, Mor and Lys was detected in the gut. Unexpectedly, in addition to the IMD pathway genes, the Toll and JAK/STAT pathway genes in the silkworm gut can also be activated by microbial oral infection. But in the developmental course, corresponding to the increase in expression of AMPs at spinning and prepupa stages, only the Toll pathway genes in the gut exhibit the similar increasing trend. Our results imply that the immune responses in the silkworm gut are synergistically regulated by the Toll, JAK/STAT and IMD pathways. However, as the time for approaching pupation, the Toll pathway may play a role in the AMPs expression

  10. A cyclic peptide synthetase gene required for pathogenicity of the fungus Cochliobolus carbonum on maize.

    PubMed Central

    Panaccione, D G; Scott-Craig, J S; Pocard, J A; Walton, J D

    1992-01-01

    Specificity in many plant-pathogen interactions is determined by single genes in pathogen and host. The single locus for host-selective pathogenicity (TOX2) in the fungus Cochliobolus carbonum governs production of a cyclic tetrapeptide named HC-toxin. We have isolated a chromosomal region, 22 kilobases (kb) long, that contains a 15.7-kb open reading frame (HTS1) encoding a multifunctional cyclic peptide synthetase. The 22-kb chromosomal region is duplicated in toxin-producing isolates of the fungus but is completely absent from the genomes of toxin-nonproducing isolates. Mutants of the fungus with disruptions in both copies of HTS1, at either of two different sites within HTS1, were engineered by DNA-mediated transformation. Disruption of both copies at either site resulted in loss of ability to produce HC-toxin and loss of host-selective pathogenicity, but the mutants displayed different biochemical phenotypes depending on the site of disruption. The results demonstrate that TOX2 encodes, at least in part, a large, multifunctional biosynthetic enzyme and that the evolution of host range in C. carbonum involved the insertion or deletion of a large piece of chromosomal DNA. Images PMID:11607305

  11. Involvement of Relish gene from Macrobrachium rosenbergii in the expression of anti-microbial peptides.

    PubMed

    Shi, Yan-Ru; Jin, Min; Ma, Fu-Tong; Huang, Ying; Huang, Xin; Feng, Jin-Ling; Zhao, Ling-Ling; Chen, Yi-Hong; Ren, Qian

    2015-10-01

    Relish is an NF-kB transcription factor involved in immune-deficiency (IMD) signal pathway. In this study, a Relish gene (MrRelish) was identified from Macrobrachium rosenbergii. The full length of MrRelish comprises 5072 bp, including a 3510 bp open reading frame encoding a 1169 bp amino acid protein. MrRelish contains a Rel homology domain (RHD), a nucleus localization signal, an IκB-like domain (6 ankyrin repeats), and a death domain. Phylogenetic analysis showed that MrRelish and other Relish from crustaceans belong to one group. MrRelish was expressed in all detected tissues, with the highest expression level in hemocytes and intestines. MrRelish was also upregulated in hepatopancreas at 6 h after Vibrio anguillarum challenge. The over-expression of MrRelish could induce the expression of antimicrobial peptides (AMPs), such as Drosophila Metchnikowin (Mtk), Attacin (Atta), Drosomycin (Drs), and Cecropin (CecA) and shrimp Penaeidin (Pen4). The RNAi of MrRelish in gills showed that the expression of crustin (cru) 2, Cru5, Cru8, lysozyme (Lyso) 1, and Lyso2 was inhibited. However, the expression of anti-lipopolysaccharide factor (ALF) 1 and ALF3 did not change when MrRelish was knocked down. These results indicate that MrRelish may play an important role in innate immune defense against V. anguillarum in M. rosenbergii. PMID:26026243

  12. Cell penetrating peptide-based polyplexes shelled with polysaccharide to improve stability and gene transfection.

    PubMed

    Li, Wenyu; Liu, Yajie; Du, Jianwei; Ren, Kefeng; Wang, Youxiang

    2015-05-14

    Cell-penetrating peptides (CPP) have been widely developed as a strategy to enhance cell penetrating ability and transfection. In this work, octa-arginine modified dextran gene vector with pH-sensitivity was developed via host-guest interactions. α-Cyclodextrin was modified with octa-arginine (CDR), which had excellent cell penetrating ability. Dextran was selected as a backbone and modified with azobenzene as guest units by acid-labile imine bonds (Az-I-Dex). The supramolecular polymer CDR/Az-I-Dex with high a C/A molar ratio (molar ratio of CD on CDR to Az on Az-I-Dex) was unfavorable for DNA condensation. The dextran shell of CDR/Az-I-Dex/DNA polyplexes improved the stability under physiological conditions. However, once treated with acetate buffer (pH 5.4) for 3 h, large aggregates formed rapidly due to the cleavage of the dextran shell. As expected, the vector had cell viability of 80% even when the CDR concentration increased to 100 μg mL(-1). Moreover, due to the effective cellular uptake efficiency, CDR/Az-I-Dex/DNA polyplexes had 6-300 times higher transfection efficiency than CDR/DNA polyplexes. It was even higher than high molecular weight PLL-based polyplexes of HEK293 T cells. Importantly, chloroquine as an endosomal escape agent could not improve the transfection of CDR/Az-I-Dex/DNA polyplexes, which indicated that the CDR/Az-I-Dex supramolecular polymer had its own ability for endosomal escape. These results suggested that the CPP-based polyplexes shelled with polysaccharide can be promising non-viral gene delivery carriers. PMID:25893559

  13. Amelogenin signal peptide mutation: Correlation between mutations in the amelogenin gene (AMGX) and manifestations of X-linked amelogenesis imperfecta

    SciTech Connect

    Lagerstroem-Fermer, M.; Nilsson, M.; Pettersson, U.

    1995-03-01

    Formation of tooth enamel is a poorly understood biological process. In this study the authors describe a 9-bp deletion in exon 2 of the amelogenin gene (AMGX) causing X-linked hypoplastic amelogenesis imperfecta, a disease characterized by defective enamel. The mutation results in the loss of 3 amino acids and exchange of 1 in the signal peptide of the amelogenin protein. This deletion in the signal peptide probably interferes with translocation of the amelogenin protein during synthesis, resulting in the thin enamel observed in affected members of the family. The authors compare this mutation to a previously reported mutation in the amelogenin gene that causes a different disease phenotype. The study illustrates that molecular analysis can help explain the various manifestations of a tooth disorder and thereby provide insights into the mechanisms of tooth enamel formation. 16 refs., 2 figs., 1 tab.

  14. Identification, expression, and innate immune responses of two insulin-like peptide genes in the razor clam Sinonovacula constricta.

    PubMed

    Niu, Donghong; Wang, Fei; Zhao, Honggang; Wang, Ze; Xie, Shumei; Li, Jiale

    2016-04-01

    Insulin-like peptide (ILP) has emerged as a cell regulatory factor with multiple functions in vertebrates and invertebrates. In the present study, we identified and characterized two ILP genes, ILP1 and ILP2, in the razor clam Sinonovacula constricta. Both ILPs have a signal peptide and a mature domain consisting of six strictly conserved cysteines. The tertiary structure is divided into three main α-helices with a C-domain loop that separates helix 1 from helix 2. Both of ILPs were found to be regulated according to tissue type and developmental stage. After challenge with Vibrio anguillarum, Vibrio parahaemolyticus and Micrococcus lysodeikticus, the expression of two ILP genes was significantly up-regulated in the liver, hemocytes and mantle tissues, suggesting that the ILPs may play roles in the innate immunity in the razor clam Sinonovacula constricta. PMID:26980611

  15. Enhanced blood-brain barrier penetration and glioma therapy mediated by a new peptide modified gene delivery system.

    PubMed

    Yao, Hui; Wang, Kaiyuan; Wang, Yi; Wang, Shanshan; Li, Jianfeng; Lou, Jinning; Ye, Liya; Yan, Xueying; Lu, Weiyue; Huang, Rongqin

    2015-01-01

    Successful glioma gene therapy lays on two important factors, the therapeutic genes and efficient delivery vehicles to cross the blood-brain barrier (BBB) and reach gliomas. In this work, a new gene vector was constructed based on dendrigraft poly-l-lysines (DGL) and polyethyleneglycol (PEG), conjugated with a cell-penetrating peptide, the nucleolar translocation signal (NoLS) sequence of the LIM Kinase 2 (LIMK2) protein (LIMK2 NoLS peptide, LNP), yielding DGL-PEG-LNP. Plasmid DNA encoding inhibitor of growth 4 (ING4) was applied as the therapeutic gene. DGL-PEG-LNP/DNA nanoparticles (NPs) were monodispersed, with a mean diameter of 90.6 ± 8.9 nm. The conjugation of LNP significantly enhanced the BBB-crossing efficiency, cellular uptake and gene expression within tumor cells. Mechanism studies suggested the involvement of energy, caveolae-mediated endocytosis and macropinocytosis in cellular uptake of LNP-modified NPs. MTT results showed that no apparent cytotoxicity was observed when cells were treated with synthesized vectors. Furthermore, LNP-modified NPs mediated strongest and most intensive apoptosis on the tumor site, and the longest median survival time of glioma-bearing mice. All the results demonstrated that LNP is a kind of efficient CPPs especially for BBB-crossing application, and DGL-PEG-LNP/DNA is a potential non-viral platform for glioma gene therapy via intravenous administration. PMID:25453963

  16. Integration of antimicrobial peptides with gold nanoparticles as unique non-viral vectors for gene delivery to mesenchymal stem cells with antibacterial activity.

    PubMed

    Peng, Li-Hua; Huang, Yan-Fen; Zhang, Chen-Zhen; Niu, Jie; Chen, Ying; Chu, Yang; Jiang, Zhi-Hong; Gao, Jian-Qing; Mao, Zheng-Wei

    2016-10-01

    Gold nanoparticles (AuNPs) have emerged as attractive non-viral gene vectors. However their application in regenerative medicine is still limited partially due to a lack of an intrinsic capacity to transfect difficult-to-transfect cells such as primary cells or stem cells. In current study, we report the synthesis of antimicrobial peptide conjugated cationic AuNPs (AuNPs@PEP) as highly efficient carriers for gene delivery to stem cells with antibacterial ability. The AuNPs@PEP integrate the advantages of cationic AuNPs and antibacterial peptides: the presence of cationic AuNPs can effectively condense DNA and the antimicrobial peptides are essential for the cellular & nucleus entry enhancement to achieve high transfection efficiency and antibacterial ability. As a result, antimicrobial peptides conjugated AuNPs significantly promoted the gene transfection efficiency in rat mesenchymal stem cells than pristine AuNPs, with a similar extent to those expressed by TAT (a well-known cell-penetrating peptide) modified AuNPs. More interestingly, the combinational system has better antibacterial ability than free antimicrobial peptides in vitro and in vivo, possibly due to the high density of peptides on the surface of AuNPs. Finally we present the concept-proving results that AuPs@PEP can be used as a carrier for in vivo gene activation in tissue regeneration, suggesting its potential as a multifunctional system with both gene delivery and antibacterial abilities in clinic. PMID:27376562

  17. Genomic Organization, Molecular Diversification, and Evolution of Antimicrobial Peptide Myticin-C Genes in the Mussel (Mytilus galloprovincialis)

    PubMed Central

    Vera, Manuel; Martínez, Paulino; Poisa-Beiro, Laura; Figueras, Antonio; Novoa, Beatriz

    2011-01-01

    Myticin-C is a highly variable antimicrobial peptide associated to immune response in Mediterranean mussel (Mytilus galloprovincialis). In this study, we tried to ascertain the genetic organization and the mechanisms underlying myticin-C variation and evolution of this gene family. We took advantage of the large intron size variation to find out the number of myticin-C genes. Using fragment analysis a maximum of four alleles was detected per individual at both introns in a large mussel sample suggesting a minimum of two myticin-C genes. The transmission pattern of size variants in two full-sib families was also used to ascertain the number of myticin-C genes underlying the variability observed. Results in both families were in accordance with two myticin-C genes organized in tandem. A more detailed analysis of myticin-C variation was carried out by sequencing a large sample of complementary (cDNA) and genomic DNA (gDNA) in 10 individuals. Two basic sequences were detected at most individuals and several sequences were constituted by combination of two different basic sequences, strongly suggesting somatic recombination or gene conversion. Slight within-basic sequence variation detected in all individuals was attributed to somatic mutation. Such mutations were more frequently at the C-terminal domain and mostly determined non-synonymous substitutions. The mature peptide domain showed the highest variation both in the whole cDNA and in the basic-sequence samples, which is in accordance with the pathogen recognition function associated to this domain. Although most tests suggested neutrality for myticin-C variation, evidence indicated positive selection in the mature peptide and C-terminal region. Three main highly supported clusters were observed when reconstructing phylogeny on basic sequences, meiotic recombination playing a relevant role on myticin-C evolution. This study demonstrates that mechanisms to generate molecular variation similar to that observed in

  18. Expression pattern of peptide and amino acid genes in digestive tract of transporter juvenile turbot ( Scophthalmus maximus L.)

    NASA Astrophysics Data System (ADS)

    Xu, Dandan; He, Gen; Mai, Kangsen; Zhou, Huihui; Xu, Wei; Song, Fei

    2016-04-01

    Turbot ( Scophthalmus maximus L.), a carnivorous fish species with high dietary protein requirement, was chosen to examine the expression pattern of peptide and amino acid transporter genes along its digestive tract which was divided into six segments including stomach, pyloric caeca, rectum, and three equal parts of the remainder of the intestine. The results showed that the expression of two peptide and eleven amino acid transporters genes exhibited distinct patterns. Peptide transporter 1 (PepT1) was rich in proximal intestine while peptide transporter 2 (PepT2) was abundant in distal intestine. A number of neutral and cationic amino acid transporters expressed richly in whole intestine including B0-type amino acid transporter 1 (B0AT1), L-type amino acid transporter 2 (LAT2), T-type amino acid transporter 1 (TAT1), proton-coupled amino acid transporter 1 (PAT1), y+L-type amino acid transporter 1 (y+LAT1), and cationic amino acid transporter 2 (CAT2) while ASC amino acid transporter 2 (ASCT2), sodium-coupled neutral amino acid transporter 2 (SNAT2), and y+L-type amino acid transporter 2 (y+LAT2) abundantly expressed in stomach. In addition, system b0,+ transporters (rBAT and b0,+AT) existed richly in distal intestine. These findings comprehensively characterized the distribution of solute carrier family proteins, which revealed the relative importance of peptide and amino acid absorption through luminal membrane. Our findings are helpful to understand the mechanism of the utilization of dietary protein in fish with a short digestive tract.

  19. Tissue-specific expression of the human brain natriuretic peptide gene in cardiac myocytes.

    PubMed

    LaPointe, M C; Wu, G; Garami, M; Yang, X P; Gardner, D G

    1996-03-01

    Brain natriuretic peptide (BNP) is a cardiac hormone constitutively expressed in the adult heart. To identify the cis-acting elements involved in regulation of the human BNP gene, we subcloned the full-length promoter (-1818 to +100) and deletions thereof upstream from a luciferase reporter gene and transiently transfected them into primary cultures of neonatal rat atrial and ventricular myocytes and myocardial fibroblasts. Luciferase activity of the full-length construct was higher in ventricular (39064 +/- 8488 relative light units, N=11) and atrial (11225 +/- 1907, N=17) myocytes than myocardial fibroblasts (329 +/- 113, n=5). Maximal promoter activity in ventricular and atrial myocytes was maintained by sequences positioned between -1818 and -1283 relative to the transcription start site. Deletion to -1175 resulted in a decrease, whereas further deletion to -500 effected an increase in reporter activity in both cell types. In ventricular and atrial myocytes, deletion from -500 to -40 reduced luciferase activity 20-fold and 2-fold, respectively, whereas in myocardial fibroblasts, deletion to -40 upregulated the BNP promoter 2-fold. Of note, deleting 16 bp between -127 and -111 reduced luciferase activity 7-fold and 4-fold in ventricular and atrial myocytes, respectively, but had essentially no effect on luciferase activity in fibroblasts. Placement of sequences lying between -127 and -40 upstream from a heterologous thymidine kinase promoter resulted in reporter expression that was 7.4-fold greater than the vector alone in ventricular myocytes, approximately 2-fold greater in atrial myocytes, and equivalent to the vector alone in fibroblasts. For study of activity of the human BNP promoter in adult myocytes, either 408 or 97 bp of 5' flanking sequence coupled to the luciferase reporter gene was injected into the apex of adult male Sprague-Dawley rat hearts. After 7 days, luciferase activity in the injected myocardium was 9.8-fold higher for the longer construct

  20. Cloning, sequencing and overexpression of the gene for prokaryotic factor EF-P involved in peptide bond synthesis.

    PubMed Central

    Aoki, H; Adams, S L; Chung, D G; Yaguchi, M; Chuang, S E; Ganoza, M C

    1991-01-01

    A soluble protein EF-P (elongation factor P) from Escherichia coli has been purified and shown to stimulate efficient translation and peptide-bond synthesis on native or reconstituted 70S ribosomes in vitro. Based on the partial amino acid sequence of EF-P, 18- and 24-nucleotide DNA probes were synthesized and used to screen lambda phage clones from the Kohara Gene Bank. The entire EF-P gene was detected on lambda clone #650 which contains sequences from the 94 minute region of the E.coli genome. Two DNA fragments, 3.0 and 0.78 kilobases in length encompassing the gene, were isolated and cloned into pUC18 and pUC19. Partially purified extracts from cells transformed with these plasmids overrepresented a protein which co-migrates with EF-P upon SDS polyacrylamide gel electrophoresis, and also exhibited increased EF-P mediated peptide-bond synthetic activity. Based on DNA sequence analysis of this gene, the EF-P protein consists of 187 amino acids with a calculated molecular weight of 20,447. The sequence and chromosomal location of EF-P establishes it as a unique gene product. Images PMID:1956781

  1. Structural Mechanisms of Peptide Recognition and Allosteric Modulation of Gene Regulation by the RRNPP Family of Quorum-Sensing Regulators.

    PubMed

    Do, Hackwon; Kumaraswami, Muthiah

    2016-07-17

    The members of RRNPP family of bacterial regulators sense population density-specific secreted oligopeptides and modulate the expression of genes involved in cellular processes, such as sporulation, competence, virulence, biofilm formation, conjugative plasmid transfer and antibiotic resistance. Signaling by RRNPP regulators include several steps: generation and secretion of the signaling oligopeptides, re-internalization of the signaling molecules into the cytoplasm, signal sensing by the cytosolic RRNPP regulators, signal-specific allosteric structural changes in the regulators, and interaction of the regulators with their respective regulatory target and gene regulation. The recently determined structures of the RRNPP regulators provide insight into the mechanistic aspects for several steps in this signaling circuit. In this review, we discuss the structural principles underlying peptide specificity, regulatory target recognition, and ligand-induced allostery in RRNPP regulators and its impact on gene regulation. Despite the conserved tertiary structure of these regulators, structural analyses revealed unexpected diversity in the mechanism of activation and molecular strategies that couple the peptide-induced allostery to gene regulation. Although these structural studies provide a sophisticated understanding of gene regulation by RRNPP regulators, much needs to be learned regarding the target DNA binding by yet-to-be characterized RNPP regulators and the several aspects of signaling by Rgg regulators. PMID:27283781

  2. Detection of nonribosomal peptide synthetase genes in Xylaria sp. BCC1067 and cloning of XyNRPSA.

    PubMed

    Paungmoung, Porntip; Punya, Jantira; Pongpattanakitshote, Somchai; Jeamton, Wattana; Vichisoonthonkul, Taweerat; Bhumiratana, Sakarindr; Tanticharoen, Morakot; Linne, Uwe; Marahiel, Mohamed A; Cheevadhanarak, Supapon

    2007-09-01

    Nonribosomal peptides, synthesized by nonribosomal peptide synthetases (NRPS), are an important group of diverse bioactive fungal metabolites. Xylaria sp. BCC1067, which is known to produce a variety of biologically active metabolites, was studied for gene encoding NRPS by two different PCR-based methods and seven different NRPS fragments were obtained. In addition, screening a genomic library with an amplified NRPS fragment as a probe identified a putative NRPS gene named XyNRPSA. The functionality of XyNRPSA for the production of a corresponding metabolite was probed by gene insertion inactivation. Comparing the disrupting metabolite profile with that of the wild type led to the identification of a speculated metabolite. The crude extract of Xylaria sp. BCC1067 also exhibits antifungal activity against the human pathogens Candida albicans and Trichophyton mentagrophytes. However, the evaluation of biological activity of the XyNRPSA product suggests that it is neither a compound with antifungal activity nor a siderophore. In the vicinity of XyNRPSA, a second gene (named XyPtB) was identified. Its localization and homology to orfB of the ergot alkaloid biosynthetic gene cluster suggests that XyPtB may be involved in XyNRPSA product biosynthesis. PMID:17623029

  3. Genes encoding 4-Cys antimicrobial peptides in wheat Triticum kiharae Dorof. et Migush.: multimodular structural organization, instraspecific variability, distribution and role in defence.

    PubMed

    Utkina, Lyubov L; Andreev, Yaroslav A; Rogozhin, Eugene A; Korostyleva, Tatyana V; Slavokhotova, Anna A; Oparin, Peter B; Vassilevski, Alexander A; Grishin, Eugene V; Egorov, Tsezi A; Odintsova, Tatyana I

    2013-08-01

    A novel family of antifungal peptides was discovered in the wheat Triticum kiharae Dorof. et Migusch. Two members of the family, designated Tk-AMP-X1 and Tk-AMP-X2, were completely sequenced and shown to belong to the α-hairpinin structural family of plant peptides with a characteristic C1XXXC2-X(n)-C3XXXC4 motif. The peptides inhibit the spore germination of several fungal pathogens in vitro. cDNA and gene cloning disclosed unique structure of genes encoding Tk-AMP-X peptides. They code for precursor proteins of unusual multimodular structure, consisting of a signal peptide, several α-hairpinin (4-Cys) peptide domains with a characteristic cysteine pattern separated by linkers and a C-terminal prodomain. Three types of precursor proteins, with five, six or seven 4-Cys peptide modules, were found in wheat. Among the predicted family members, several peptides previously isolated from T. kiharae seeds were identified. Genes encoding Tk-AMP-X precursors have no introns in the protein-coding regions and are upregulated by fungal pathogens and abiotic stress, providing conclusive evidence for their role in stress response. A combined PCR-based and bioinformatics approach was used to search for related genes in the plant kingdom. Homologous genes differing in the number of peptide modules were discovered in phylogenetically-related Triticum and Aegilops species, including polyploid wheat genome donors. Association of the Tk-AMP-X genes with A, B/G or D genomes of hexaploid wheat was demonstrated. Furthermore, Tk-AMP-X-related sequences were shown to be widespread in the Poaceae family among economically important crops, such as barley, rice and maize. PMID:23702306

  4. Use of the "blue halo" assay in the identification of genes encoding exported proteins with cleavable signal peptides: cloning of a Borrelia burgdorferi plasmid gene with a signal peptide.

    PubMed

    Giladi, M; Champion, C I; Haake, D A; Blanco, D R; Miller, J F; Miller, J N; Lovett, M A

    1993-07-01

    We have recently reported a phoA expression vector, termed pMG, which, like TnphoA, is useful in identifying genes encoding membrane-spanning sequences or signal peptides. This cloning system has been modified to facilitate the distinction of outer membrane and periplasmic alkaline phosphatase (AP) fusion proteins from inner membrane AP fusion proteins by transforming pMG recombinants into Escherichia coli KS330, the strain utilized in the "blue halo" assay first described by Strauch and Beckwith (Proc. Natl. Acad. Sci. USA 85:1576-1580, 1988). The lipoprotein mutation lpp-5508 of KS330 results in an outer membrane that is leaky to macromolecules, and its degP4 mutation greatly reduces periplasmic proteolytic degradation of AP fusion proteins. pMG AP fusions containing cleavable signal peptides, including the E. coli periplasmic protein beta-lactamase, the E. coli and Chlamydia trachomatis outer membrane proteins OmpA and MOMP, respectively, and Tp 9, a Treponema pallidum AP recombinant, diffused through the leaky outer membrane of KS330 and resulted in blue colonies with blue halos. In contrast, inner membrane AP fusions derived from E. coli proteins, including leader peptidase, SecY, and the tetracycline resistance gene product, as well as Tp 70, a T. pallidum AP recombinant which does not contain a signal peptide, resulted in blue colonies without blue halos. Lipoprotein-AP fusions, including the Borrelia burgdorferi OspA and T. pallidum Tp 75 and TmpA showed halo formation, although there was significantly less halo formation than that produced by either periplasmic or outer membrane AP fusions. In addition, we applied this approach to screen recombinants constructed from a 9.0-kb plasmid isolated from the B31 virulent strain of B. burgdorferi. One of the blue halo colonies identified produced an AP fusion protein which contained a signal peptide with a leader peptidase I cleavage recognition site. The pMG/KS330r- cloning and screening approach can identify

  5. Effects of vaspin, chemerin and omentin-1 on feeding behavior and hypothalamic peptide gene expression in the rat.

    PubMed

    Brunetti, Luigi; Di Nisio, Chiara; Recinella, Lucia; Chiavaroli, Annalisa; Leone, Sheila; Ferrante, Claudio; Orlando, Giustino; Vacca, Michele

    2011-09-01

    Visceral adipose tissue-derived serpin (vaspin) improves glucose tolerance and insulin sensitivity in diet-induced obese mice. Chemerin may increase insulin sensitivity in adipose tissue and seems to be associated with several key aspects of metabolic syndrome. Decreased levels of omentin-1 are associated with increasing obesity and insulin resistance. Our study aimed to investigate the effects of vaspin, chemerin and omentin-1 acute administration on feeding and hypothalamic gene expression of peptides which play a key role in feeding regulation. 35 rats were injected into the arcuate nucleus (ARC) of the hypothalamus with either saline (n=8), vaspin (1μg/kg; n=9), chemerin (8μg/kg; n=9), or omentin-1 (8μg/kg; n=9). Food intake in the following 24h was recorded, thereafter rats were sacrificed. Total RNA was extracted from hypothalami and reverse transcribed to evaluate hypothalamic gene expression of agouti-related peptide (AgRP), neuropeptide Y (NPY), orexin-A, cocaine- and amphetamine-regulated transcript (CART), corticotrophin releasing hormone (CRH) and proopiomelanocortin (POMC), by real-time reverse transcription polymerase chain reaction. Compared to vehicle, vaspin injection significantly decreased feeding, while chemerin and omentin-1 had no effect in the tested dose. Vaspin treatment significantly decreased NPY and increased POMC gene expression. Chemerin treatment led to a significant increase of both AgRP and POMC gene expression. Omentin-1 treatment did not modify gene expression of the investigated peptides. Therefore, vaspin is an adipokine triggering anorectic pathways in the hypothalamus, where reduction of NPY and increase of POMC mRNA levels mediate feeding inhibition. Chemerin and omentin-1 have no effect on feeding in the tested dose. PMID:21855588

  6. Effect of corticosterone on gene expression of feed intake regulatory peptides in laying hens.

    PubMed

    Liu, Lei; Song, Zhigang; Sheikhahmadi, Ardashir; Jiao, Hongchao; Lin, Hai

    2012-08-01

    The present study was conducted to explore the effects of corticosterone (CORT) on the regulation of appetite-associated genes in laying hens. Forty eight laying hens were randomly divided into two groups: one received subcutaneous injection of CORT (2mg/kg body weight, CORT-exposed) and the other received sham-treatment (Control). Treatment of hens with CORT stimulated an increase (P<0.05) in plasma CORT, glucose, uric acid (UA), insulin, cholesterol (Chol) and triiodothyronine (T(3)), but the concentrations of plasma non-esterified fatty acids (NEFA) and triacylglycerol (TG) were decreased (P<0.05). CORT treatment had no significant effect (P>0.05) on the mRNA levels of neuropeptide Y (NPY), corticotropin-releasing hormone (CRH), melanocortin receptor 4 and 5 (MCR-4 and MCR-5) and cholecystokinin (CCK) in the hypothalamus when compared with control hens. However, the expression of pro-opiomelanocortin (POMC), agouti-related protein (AgRP) and melanocortin recepter 1 (MCR-1) were significantly (P<0.05) suppressed while the mRNA levels of ghrelin and cocaine-and amphetamine-regulated transcript (CART) were significantly upregulated (P<0.05) in CORT-treated hens. Treatment of laying hens with CORT had no significant (P>0.05) effect on the mRNA levels of CCK in the glandular stomach and the duodenum, and those of ghrelin in the glandular stomach, the duodenum and the jejunum. However, the mRNA levels of CCK in the jejunum and the ileum, and those of ghrelin in the ileum were significantly (P<0.05) suppressed by CORT treatment. In conclusion, these results suggest that CORT plays a unique role in some special neuropeptides (e.g., ghrelin, CART, POMC, CCK and MCRs) and a dynamic balance between these appetite-associated peptides in the hypothalamus and the gastrointestinal tract defines the feeding status of CORT-exposed laying hens. PMID:22554475

  7. Development of self-assembling peptide nanovesicle with bilayers for enhanced EGFR-targeted drug and gene delivery.

    PubMed

    Liang, Xiaofei; Shi, Bizhi; Wang, Kai; Fan, Mingliang; Jiao, Dejin; Ao, Junping; Song, Na; Wang, Chun; Gu, Jianren; Li, Zonghai

    2016-03-01

    Development of rational vectors for efficient drug and gene delivery is crucial for cancer treatment. In this study, epidermal growth factor receptor (EGFR)-binding peptide amphiphile (PA) were used as the primary bilayer skeleton material to construct ultra-stable self-assembling peptide nanovesicle (SPV). The resulted EGFR-targeted SPV (ESPV) could efficiently encapsulate therapeutic cargos (drugs or small interfering RNAs [siRNAs]) or labelled fluorescent cargo (quantum dots [QDs]) and exhibited excellent affinity for EGFR-positive cancer cells. Moreover, ESPV could deliver more drug or plasmid DNA to tumour sites and promote gene expression (a three-fold ratio of ESPVs vs cationic liposomes). Notably, the individual delivery or co-delivery of doxorubicin (DOX) and the acetylcholinesterase (AChE) gene via the ESPVs resulted in excellent drug/gene delivery both in vitro and in vivo and exerted a significant growth-suppressing effect on a liver cancer xenograft. This nanoscale, targeted cargo-packaging technology may provide a new strategy for the design of highly targeted cancer therapy vectors. PMID:26763734

  8. Peptide-Mediated Tumor Targeting by a Degradable Nano Gene Delivery Vector Based on Pluronic-Modified Polyethylenimine

    NASA Astrophysics Data System (ADS)

    Wu, Zhaoyong; Zhan, Shuyu; Fan, Wei; Ding, Xueying; Wu, Xin; Zhang, Wei; Fu, Yinghua; Huang, Yueyan; Huang, Xuan; Chen, Rubing; Li, Mingjuan; Xu, Ningyin; Zheng, Yongxia; Ding, Baoyue

    2016-03-01

    Polyethylenimine (PEI) is considered to be a promising non-viral gene delivery vector. To solve the toxicity versus efficacy and tumor-targeting challenges of PEI used as gene delivery vector, we constructed a novel non-viral vector DR5-TAT-modified Pluronic-PEI (Pluronic-PEI-DR5-TAT), which was based on the attachment of low-molecular-weight polyethylenimine (LMW-PEI) to the amphiphilic polymer Pluronic to prepare Pluronic-modified LMW-PEI (Pluronic-PEI). This was then conjugated to a multifunctional peptide containing a cell-penetrating peptide (TAT) and a synthetic peptide that would bind to DR5—a receptor that is overexpressed in cancer cells. The vector showed controlled degradation, favorable DNA condensation and protection performance. The Pluronic-PEI-DR5-TAT/DNA complexes at an N/P ratio of 15:1 were spherical nanoparticles of 122 ± 11.6 nm and a zeta potential of about 22 ± 2.8 mV. In vitro biological characterization results indicated that Pluronic-PEI-DR5-TAT/DNA complexes had a higher specificity for the DR5 receptor and were taken up more efficiently by tumor cells than normal cells, compared to complexes formed with PEI 25 kDa or Pluronic-PEI. Thus, the novel complexes showed much lower cytotoxicity to normal cells and higher gene transfection efficiency in tumor cells than that exhibited by PEI 25 kDa and Pluronic-PEI. In summary, our novel, degradable non-viral tumor-targeting vector is a promising candidate for use in gene therapy.

  9. Peptide-Mediated Tumor Targeting by a Degradable Nano Gene Delivery Vector Based on Pluronic-Modified Polyethylenimine.

    PubMed

    Wu, Zhaoyong; Zhan, Shuyu; Fan, Wei; Ding, Xueying; Wu, Xin; Zhang, Wei; Fu, Yinghua; Huang, Yueyan; Huang, Xuan; Chen, Rubing; Li, Mingjuan; Xu, Ningyin; Zheng, Yongxia; Ding, Baoyue

    2016-12-01

    Polyethylenimine (PEI) is considered to be a promising non-viral gene delivery vector. To solve the toxicity versus efficacy and tumor-targeting challenges of PEI used as gene delivery vector, we constructed a novel non-viral vector DR5-TAT-modified Pluronic-PEI (Pluronic-PEI-DR5-TAT), which was based on the attachment of low-molecular-weight polyethylenimine (LMW-PEI) to the amphiphilic polymer Pluronic to prepare Pluronic-modified LMW-PEI (Pluronic-PEI). This was then conjugated to a multifunctional peptide containing a cell-penetrating peptide (TAT) and a synthetic peptide that would bind to DR5-a receptor that is overexpressed in cancer cells. The vector showed controlled degradation, favorable DNA condensation and protection performance. The Pluronic-PEI-DR5-TAT/DNA complexes at an N/P ratio of 15:1 were spherical nanoparticles of 122 ± 11.6 nm and a zeta potential of about 22 ± 2.8 mV. In vitro biological characterization results indicated that Pluronic-PEI-DR5-TAT/DNA complexes had a higher specificity for the DR5 receptor and were taken up more efficiently by tumor cells than normal cells, compared to complexes formed with PEI 25 kDa or Pluronic-PEI. Thus, the novel complexes showed much lower cytotoxicity to normal cells and higher gene transfection efficiency in tumor cells than that exhibited by PEI 25 kDa and Pluronic-PEI. In summary, our novel, degradable non-viral tumor-targeting vector is a promising candidate for use in gene therapy. PMID:26932761

  10. Expression and characterization of Drosophila signal peptide peptidase-like (sppL), a gene that encodes an intramembrane protease.

    PubMed

    Casso, David J; Liu, Songmei; Biehs, Brian; Kornberg, Thomas B

    2012-01-01

    Intramembrane proteases of the Signal Peptide Peptidase (SPP) family play important roles in developmental, metabolic and signaling pathways. Although vertebrates have one SPP and four SPP-like (SPPL) genes, we found that insect genomes encode one Spp and one SppL. Characterization of the Drosophila sppL gene revealed that the predicted SppL protein is a highly conserved structural homolog of the vertebrate SPPL3 proteases, with a predicted nine-transmembrane topology, an active site containing aspartyl residues within a transmembrane region, and a carboxy-terminal PAL domain. SppL protein localized to both the Golgi and ER. Whereas spp is an essential gene that is required during early larval stages and whereas spp loss-of-function reduced the unfolded protein response (UPR), sppL loss of function had no apparent phenotype. This was unexpected given that genetic knockdown phenotypes in other organisms suggested significant roles for Spp-related proteases. PMID:22439002

  11. The permease homologue Ssy1p controls the expression of amino acid and peptide transporter genes in Saccharomyces cerevisiae.

    PubMed

    Didion, T; Regenberg, B; Jørgensen, M U; Kielland-Brandt, M C; Andersen, H A

    1998-02-01

    Amino acid transporters of the yeast plasma membrane (permeases) belong to a family of integral membrane proteins with pronounced structural similarity. We present evidence that a member of this family, encoded by the open reading frame (ORF) YDR160w (SSY1), is required for the expression of a set of transporter genes. Thus, deletion of the SSY1 gene causes loss of leucine-inducible transcription of the amino acid permease genes BAP2, TAT1 and BAP3 (ORF YDR046c) and the peptide transporter, PTR2. D-leucine can generate the signal without entering the cell. We propose that Ssy1p is situated in the plasma membrane and is involved in sensing leucine in the medium. PMID:9489675

  12. Bioinformatic and phylogenetic analysis of the CLAVATA3/EMBRYO-SURROUNDING REGION (CLE) and the CLE-LIKE signal peptide genes in the Pinophyta

    PubMed Central

    2014-01-01

    Background There is a rapidly growing awareness that plant peptide signalling molecules are numerous and varied and they are known to play fundamental roles in angiosperm plant growth and development. Two closely related peptide signalling molecule families are the CLAVATA3-EMBRYO-SURROUNDING REGION (CLE) and CLE-LIKE (CLEL) genes, which encode precursors of secreted peptide ligands that have roles in meristem maintenance and root gravitropism. Progress in peptide signalling molecule research in gymnosperms has lagged behind that of angiosperms. We therefore sought to identify CLE and CLEL genes in gymnosperms and conduct a comparative analysis of these gene families with angiosperms. Results We undertook a meta-analysis of the GenBank/EMBL/DDBJ gymnosperm EST database and the Picea abies and P. glauca genomes and identified 93 putative CLE genes and 11 CLEL genes among eight Pinophyta species, in the genera Cryptomeria, Pinus and Picea. The predicted conifer CLE and CLEL protein sequences had close phylogenetic relationships with their homologues in Arabidopsis. Notably, perfect conservation of the active CLE dodecapeptide in presumed orthologues of the Arabidopsis CLE41/44-TRACHEARY ELEMENT DIFFERENTIATION (TDIF) protein, an inhibitor of tracheary element (xylem) differentiation, was seen in all eight conifer species. We cloned the Pinus radiata CLE41/44-TDIF orthologues. These genes were preferentially expressed in phloem in planta as expected, but unexpectedly, also in differentiating tracheary element (TE) cultures. Surprisingly, transcript abundances of these TE differentiation-inhibitors sharply increased during early TE differentiation, suggesting that some cells differentiate into phloem cells in addition to TEs in these cultures. Applied CLE13 and CLE41/44 peptides inhibited root elongation in Pinus radiata seedlings. We show evidence that two CLEL genes are alternatively spliced via 3′-terminal acceptor exons encoding separate CLEL peptides

  13. Mining for Nonribosomal Peptide Synthetase and Polyketide Synthase Genes Revealed a High Level of Diversity in the Sphagnum Bog Metagenome

    PubMed Central

    Müller, Christina A.; Oberauner-Wappis, Lisa; Peyman, Armin; Amos, Gregory C. A.; Wellington, Elizabeth M. H.

    2015-01-01

    Sphagnum bog ecosystems are among the oldest vegetation forms harboring a specific microbial community and are known to produce an exceptionally wide variety of bioactive substances. Although the Sphagnum metagenome shows a rich secondary metabolism, the genes have not yet been explored. To analyze nonribosomal peptide synthetases (NRPSs) and polyketide synthases (PKSs), the diversity of NRPS and PKS genes in Sphagnum-associated metagenomes was investigated by in silico data mining and sequence-based screening (PCR amplification of 9,500 fosmid clones). The in silico Illumina-based metagenomic approach resulted in the identification of 279 NRPSs and 346 PKSs, as well as 40 PKS-NRPS hybrid gene sequences. The occurrence of NRPS sequences was strongly dominated by the members of the Protebacteria phylum, especially by species of the Burkholderia genus, while PKS sequences were mainly affiliated with Actinobacteria. Thirteen novel NRPS-related sequences were identified by PCR amplification screening, displaying amino acid identities of 48% to 91% to annotated sequences of members of the phyla Proteobacteria, Actinobacteria, and Cyanobacteria. Some of the identified metagenomic clones showed the closest similarity to peptide synthases from Burkholderia or Lysobacter, which are emerging bacterial sources of as-yet-undescribed bioactive metabolites. This report highlights the role of the extreme natural ecosystems as a promising source for detection of secondary compounds and enzymes, serving as a source for biotechnological applications. PMID:26002894

  14. Expression of 2A peptide mediated tri-fluorescent protein genes were regulated by epigenetics in transgenic sheep.

    PubMed

    Tian, Yongzhi; Li, Wenrong; Wang, Liqin; Liu, Chenxi; Lin, Jiapeng; Zhang, Xuemei; Zhang, Ning; He, Sangang; Huang, Juncheng; Jia, Bin; Liu, Mingjun

    2013-05-10

    A number of gene therapy applications and basic research would benefit from vectors expressing multiple genes. In this study, we constructed 2A peptide based tricistronic lentiviral vector and generated transgenic lambs by injecting lentivirus carrying the tricistronic vector into perivitelline space of zygotes. Of 7 lambs born, 2 lambs (#6 and #7) carried the transgene. However, no fluorescent proteins were identified in transgenic sheep. To investigate why the transgene was silenced in transgenic sheep, we analyzed the methylation status of transgene. The methylation level of CMV promoter was 76.25% in #6, and 64.7% in #7. In the coding region of three fluorescent protein genes, methylation levels were extremely high, with the average level of 98.3% in #6 and 98.4% in #7 respectively. Furthermore, the ratio of GFP(+) cells were increased significantly when the fibroblasts derived from the transgenic sheep were treated with 5-azaC and/ or TSA. Our results showed that 2A peptide based tricistronic construct was subjected to hypermethylation in transgenic sheep. Moreover, the silencing could be relieved by treating with methytransferase inhibitor and/or deacetylase inhibitor. PMID:23603255

  15. Mining for Nonribosomal Peptide Synthetase and Polyketide Synthase Genes Revealed a High Level of Diversity in the Sphagnum Bog Metagenome.

    PubMed

    Müller, Christina A; Oberauner-Wappis, Lisa; Peyman, Armin; Amos, Gregory C A; Wellington, Elizabeth M H; Berg, Gabriele

    2015-08-01

    Sphagnum bog ecosystems are among the oldest vegetation forms harboring a specific microbial community and are known to produce an exceptionally wide variety of bioactive substances. Although the Sphagnum metagenome shows a rich secondary metabolism, the genes have not yet been explored. To analyze nonribosomal peptide synthetases (NRPSs) and polyketide synthases (PKSs), the diversity of NRPS and PKS genes in Sphagnum-associated metagenomes was investigated by in silico data mining and sequence-based screening (PCR amplification of 9,500 fosmid clones). The in silico Illumina-based metagenomic approach resulted in the identification of 279 NRPSs and 346 PKSs, as well as 40 PKS-NRPS hybrid gene sequences. The occurrence of NRPS sequences was strongly dominated by the members of the Protebacteria phylum, especially by species of the Burkholderia genus, while PKS sequences were mainly affiliated with Actinobacteria. Thirteen novel NRPS-related sequences were identified by PCR amplification screening, displaying amino acid identities of 48% to 91% to annotated sequences of members of the phyla Proteobacteria, Actinobacteria, and Cyanobacteria. Some of the identified metagenomic clones showed the closest similarity to peptide synthases from Burkholderia or Lysobacter, which are emerging bacterial sources of as-yet-undescribed bioactive metabolites. This report highlights the role of the extreme natural ecosystems as a promising source for detection of secondary compounds and enzymes, serving as a source for biotechnological applications. PMID:26002894

  16. Genetic Diversity of Polyketide Synthase/Nonribosomal Peptide Synthetase Genes in Isolates of the Barley Net Blotch Fungus Pyrenophora teres f. teres

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Polyketide synthases (PKSs) and nonribosomal peptide synthetases (NRPSs) are multifunctional enzymes responsible for biosynthesis of diverse small molecules (e.g., mycotoxins and phytotoxins) in filamentous ascomycetes. Both PKS and NRPS genes are present in fungal genomes as large gene families but...

  17. 2A self-cleaving peptide-based multi-gene expression system in the silkworm Bombyx mori

    PubMed Central

    Wang, Yuancheng; Wang, Feng; Wang, Riyuan; Zhao, Ping; Xia, Qingyou

    2015-01-01

    Fundamental and applied studies of silkworms have entered the functional genomics era. Here, we report a multi-gene expression system (MGES) based on 2A self-cleaving peptide (2A), which regulates the simultaneous expression and cleavage of multiple gene targets in the silk gland of transgenic silkworms. First, a glycine-serine-glycine spacer (GSG) was found to significantly improve the cleavage efficiency of 2A. Then, the cleavage efficiency of six types of 2As with GSG was analyzed. The shortest porcine teschovirus-1 2A (P2A-GSG) exhibited the highest cleavage efficiency in all insect cell lines that we tested. Next, P2A-GSG successfully cleaved the artificial human serum albumin (66 kDa) linked with human acidic fibroblast growth factor (20.2 kDa) fusion genes and vitellogenin receptor fragment (196 kD) of silkworm linked with EGFP fusion genes, importantly, vitellogenin receptor protein was secreted to the outside of cells. Furthermore, P2A-GSG successfully mediated the simultaneous expression and cleavage of a DsRed and EGFP fusion gene in silk glands and caused secretion into the cocoon of transgenic silkworms using our sericin1 expression system. We predicted that the MGES would be an efficient tool for gene function research and innovative research on various functional silk materials in medicine, cosmetics, and other biomedical areas. PMID:26537835

  18. Therapeutic targets for olive pollen allergy defined by gene markers modulated by Ole e 1-derived peptides.

    PubMed

    Calzada, David; Aguerri, Miriam; Baos, Selene; Montaner, David; Mata, Manuel; Dopazo, Joaquín; Quiralte, Joaquín; Florido, Fernando; Lahoz, Carlos; Cárdaba, Blanca

    2015-04-01

    Two regions of Ole e 1, the major olive-pollen allergen, have been characterized as T-cell epitopes, one as immunodominant region (aa91-130) and the other, as mainly recognized by non-allergic subjects (aa10-31). This report tries to characterize the specific relevance of these epitopes in the allergic response to olive pollen by analyzing the secreted cytokines and the gene expression profiles induced after specific stimulation of peripheral blood mononuclear cells (PBMCs). PBMCs from olive pollen-allergic and non-allergic control subjects were stimulated with olive-pollen extract and Ole e 1 dodecapeptides containing relevant T-cell epitopes. Levels of cytokines were measured in cellular supernatants and gene expression was determined by microarrays, on the RNAs extracted from PBMCs. One hundred eighty-nine differential genes (fold change >2 or <-2, P<0.05) were validated by qRT-PCR in a large population. It was not possible to define a pattern of response according the overall cytokine results but interesting differences were observed, mainly in the regulatory cytokines. Principal component (PCA) gene-expression analysis defined clusters that correlated with the experimental conditions in the group of allergic subjects. Gene expression and functional analyses revealed differential genes and pathways among the experimental conditions. A set of 51 genes (many essential to T-cell tolerance and homeostasis) correlated with the response to aa10-31 of Ole e 1. In conclusion, two peptides derived from Ole e 1 could regulate the immune response in allergic patients, by gene-expression modification of several regulation-related genes. These results open new research ways to the regulation of allergy by Oleaceae family members. PMID:25553522

  19. Isolation, characterization, and expression analyses of plant elicitor peptides (Pep) genes in maize

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Insect-induced defenses occur in nearly all plants and are regulated by conserved signaling pathways. In plant families, peptides with analogous activity have remained elusive. Peps are conserved signals across diverse plant families regulating antiherbivore defenses and are likely to be the missing...

  20. Butyrate enhances disease resistance of chickens by inducing antimicrobial host defense peptide gene expression

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Host defense peptides (HDPs) constitute a large group of natural broad-spectrum antimicrobials and an important first line of immunity in virtually all forms of life. Specific augmentation of synthesis of endogenous HDPs may represent a promising antibiotic-alternative approach to disease control. I...

  1. Immunologically active peptides capable of inducing immunization against malaria and genes encoding therefor

    SciTech Connect

    Dame, J.B.; Williams, J.L.; McCutchan, T.F.; Schneider, I.

    1987-11-17

    An antimalarial immunogenic stimulant is described comprising an immunogenic carrier and a peptide sequence of between 2 and 1000 consecutive repeats of a sequence Asn-X-Y-Pro, wherein X is Ala or Val and Y is Asn or Asp.

  2. Putative Nonribosomal Peptide Synthetase and Cytochrome P450 Genes Responsible for Tentoxin Biosynthesis in Alternaria alternata ZJ33

    PubMed Central

    Li, You-Hai; Han, Wen-Jin; Gui, Xi-Wu; Wei, Tao; Tang, Shuang-Yan; Jin, Jian-Ming

    2016-01-01

    Tentoxin, a cyclic tetrapeptide produced by several Alternaria species, inhibits the F1-ATPase activity of chloroplasts, resulting in chlorosis in sensitive plants. In this study, we report two clustered genes, encoding a putative non-ribosome peptide synthetase (NRPS) TES and a cytochrome P450 protein TES1, that are required for tentoxin biosynthesis in Alternaria alternata strain ZJ33, which was isolated from blighted leaves of Eupatorium adenophorum. Using a pair of primers designed according to the consensus sequences of the adenylation domain of NRPSs, two fragments containing putative adenylation domains were amplified from A. alternata ZJ33, and subsequent PCR analyses demonstrated that these fragments belonged to the same NRPS coding sequence. With no introns, TES consists of a single 15,486 base pair open reading frame encoding a predicted 5161 amino acid protein. Meanwhile, the TES1 gene is predicted to contain five introns and encode a 506 amino acid protein. The TES protein is predicted to be comprised of four peptide synthase modules with two additional N-methylation domains, and the number and arrangement of the modules in TES were consistent with the number and arrangement of the amino acid residues of tentoxin, respectively. Notably, both TES and TES1 null mutants generated via homologous recombination failed to produce tentoxin. This study provides the first evidence concerning the biosynthesis of tentoxin in A. alternata. PMID:27490569

  3. Putative Nonribosomal Peptide Synthetase and Cytochrome P450 Genes Responsible for Tentoxin Biosynthesis in Alternaria alternata ZJ33.

    PubMed

    Li, You-Hai; Han, Wen-Jin; Gui, Xi-Wu; Wei, Tao; Tang, Shuang-Yan; Jin, Jian-Ming

    2016-01-01

    Tentoxin, a cyclic tetrapeptide produced by several Alternaria species, inhibits the F₁-ATPase activity of chloroplasts, resulting in chlorosis in sensitive plants. In this study, we report two clustered genes, encoding a putative non-ribosome peptide synthetase (NRPS) TES and a cytochrome P450 protein TES1, that are required for tentoxin biosynthesis in Alternaria alternata strain ZJ33, which was isolated from blighted leaves of Eupatorium adenophorum. Using a pair of primers designed according to the consensus sequences of the adenylation domain of NRPSs, two fragments containing putative adenylation domains were amplified from A. alternata ZJ33, and subsequent PCR analyses demonstrated that these fragments belonged to the same NRPS coding sequence. With no introns, TES consists of a single 15,486 base pair open reading frame encoding a predicted 5161 amino acid protein. Meanwhile, the TES1 gene is predicted to contain five introns and encode a 506 amino acid protein. The TES protein is predicted to be comprised of four peptide synthase modules with two additional N-methylation domains, and the number and arrangement of the modules in TES were consistent with the number and arrangement of the amino acid residues of tentoxin, respectively. Notably, both TES and TES1 null mutants generated via homologous recombination failed to produce tentoxin. This study provides the first evidence concerning the biosynthesis of tentoxin in A. alternata. PMID:27490569

  4. Two roles for the Drosophila IKK complex in the activation of Relish and the induction of antimicrobial peptide genes

    PubMed Central

    Ertürk-Hasdemir, Deniz; Broemer, Meike; Leulier, François; Lane, William S.; Paquette, Nicholas; Hwang, Daye; Kim, Chan-Hee; Stöven, Svenja; Meier, Pascal; Silverman, Neal

    2009-01-01

    The Drosophila NF-κB transcription factor Relish is an essential regulator of antimicrobial peptide gene induction after Gram-negative bacterial infection. Relish is a bipartite NF-κB precursor protein, with an N-terminal Rel homology domain and a C-terminal IκB-like domain, similar to mammalian p100 and p105. Unlike these mammalian homologs, Relish is endoproteolytically cleaved after infection, allowing the N-terminal NF-κB module to translocate to the nucleus. Signal-dependent activation of Relish, including cleavage, requires both the Drosophila IκB kinase (IKK) and death-related ced-3/Nedd2-like protein (DREDD), the Drosophila caspase-8 like protease. In this report, we show that the IKK complex controls Relish by direct phosphorylation on serines 528 and 529. Surprisingly, these phosphorylation sites are not required for Relish cleavage, nuclear translocation, or DNA binding. Instead they are critical for recruitment of RNA polymerase II and antimicrobial peptide gene induction, whereas IKK functions noncatalytically to support Dredd-mediated cleavage of Relish. PMID:19497884

  5. Characterization of Fpr-rs8, an atypical member of the mouse formyl peptide receptor gene family.

    PubMed

    Tiffany, H Lee; Gao, Ji-Liang; Roffe, Ester; Sechler, Joan M G; Murphy, Philip M

    2011-01-01

    The formyl peptide receptor gene family encodes G protein-coupled receptors for phagocyte chemoattractants, including bacteria- and mitochondria-derived N-formylpeptides. The human family has 3 functional genes, whereas the mouse family has 7 functional genes and 2 possible pseudogenes (ΨFpr-rs2 and ΨFpr-rs3). Here we characterize ΨFpr-rs2, a duplication of Fpr-rs2. Compared to Fpr-rs2, the ΨFpr-rs2 ORF is 186 nucleotides shorter but 98% identical. Due to a deletion and frame shift, the sequences lack homology from amino acid 219-289. Both transcripts were detected constitutively in multiple immune organs; however, ΨFpr-rs2 was consistently less abundant than Fpr-rs2. LPS induced expression of ΨFpr-rs2, but not Fpr-rs2, in spleen and bone marrow. Both transcripts were detected constitutively in thioglycollate-elicited peritoneal neutrophils, whereas only Fpr-rs2 was detected in thioglycollate-elicited peritoneal macrophages. Both transcripts were induced in LPS-stimulated macrophages. ΨFpr-rs2-GFP fusion protein appeared in cytoplasm but not plasma membrane of transfected HEK 293 cells, whereas Fpr-rs2-GFP labeled only plasma membrane. Survival of ΨFpr-rs2(-/-) mice was 33% shorter than that of wild-type and heterozygous littermates (p < 0.05), but no signature pathology was identified. Since ΨFpr-rs2 is expressed in phagocytes and regulated by bacterial products, and may affect longevity, we propose renaming it Fpr-rs8, an atypical member of the formyl peptide receptor gene family. PMID:21691049

  6. Characterization of Fpr-rs8, an Atypical Member of the Mouse Formyl Peptide Receptor Gene Family

    PubMed Central

    Tiffany, H. Lee; Gao, Ji-Liang; Roffe, Ester; Sechler, Joan M.G.; Murphy, Philip M.

    2011-01-01

    The formyl peptide receptor gene family encodes G protein-coupled receptors for phagocyte chemoattractants, including bacteria- and mitochondria-derived N-formylpeptides. The human family has 3 functional genes, whereas the mouse family has 7 functional genes and 2 possible pseudogenes (ΨFpr-rs2 and ΨFpr-rs3). Here we characterize ΨFpr-rs2, a duplication of Fpr-rs2. Compared to Fpr-rs2, the ΨFpr-rs2 ORF is 186 nucleotides shorter but 98% identical. Due to a deletion and frame shift, the sequences lack homology from amino acid 219–289. Both transcripts were detected constitutively in multiple immune organs; however, ΨFpr-rs2 was consistently less abundant than Fpr-rs2. LPS induced expression of ΨFpr-rs2, but not Fpr-rs2, in spleen and bone marrow. Both transcripts were detected constitutively in thioglycollate-elicited peritoneal neutrophils, whereas only Fpr-rs2 was detected in thioglycollate-elicited peritoneal macrophages. Both transcripts were induced in LPS-stimulated macrophages. ΨFpr-rs2-GFP fusion protein appeared in cytoplasm but not plasma membrane of transfected HEK 293 cells, whereas Fpr-rs2-GFP labeled only plasma membrane. Survival of ΨFpr-rs2–/– mice was 33% shorter than that of wild-type and heterozygous littermates (p < 0.05), but no signature pathology was identified. Since ΨFpr-rs2 is expressed in phagocytes and regulated by bacterial products, and may affect longevity, we propose renaming it Fpr-rs8, an atypical member of the formyl peptide receptor gene family. Copyright © 2011 S. Karger AG, Basel PMID:21691049

  7. Inefficient constitutive inhibition of P2X3 receptors by brain natriuretic peptide system contributes to sensitization of trigeminal sensory neurons in a genetic mouse model of familial hemiplegic migraine

    PubMed Central

    Marchenkova, Anna; Vilotti, Sandra; Ntamati, Niels; van den Maagdenberg, Arn MJM

    2016-01-01

    Background On trigeminal ganglion neurons, pain-sensing P2X3 receptors are constitutively inhibited by brain natriuretic peptide via its natriuretic peptide receptor-A. This inhibition is associated with increased P2X3 serine phosphorylation and receptor redistribution to non-lipid raft membrane compartments. The natriuretic peptide receptor-A antagonist anantin reverses these effects. We studied whether P2X3 inhibition is dysfunctional in a genetic familial hemiplegic migraine type-1 model produced by introduction of the human pathogenic R192Q missense mutation into the mouse CACNA1A gene (knock-in phenotype). This model faithfully replicates several properties of familial hemiplegic migraine type-1, with gain-of-function of CaV2.1 Ca2+ channels, raised levels of the algogenic peptide calcitonin gene-related peptide, and enhanced activity of P2X3 receptors in trigeminal ganglia. Results In knock-in neurons, anantin did not affect P2X3 receptor activity, membrane distribution, or serine phosphorylation level, implying ineffective inhibition by the constitutive brain natriuretic peptide/natriuretic peptide receptor-A pathway. However, expression and functional properties of this pathway remained intact together with its ability to downregulate TRPV1 channels. Reversing the familial hemiplegic migraine type-1 phenotype with the CaV2.1-specific antagonist, ω-agatoxin IVA restored P2X3 activity to wild-type level and enabled the potentiating effects of anantin again. After blocking calcitonin gene-related peptide receptors, P2X3 receptors exhibited wild-type properties and were again potentiated by anantin. Conclusions P2X3 receptors on mouse trigeminal ganglion neurons are subjected to contrasting modulation by inhibitory brain natriuretic peptide and facilitatory calcitonin gene-related peptide that both operate via complex intracellular signaling. In the familial hemiplegic migraine type-1 migraine model, the action of calcitonin gene-related peptide appears to

  8. RGD peptide-modified dendrimer-entrapped gold nanoparticles enable highly efficient and specific gene delivery to stem cells.

    PubMed

    Kong, Lingdan; Alves, Carla S; Hou, Wenxiu; Qiu, Jieru; Möhwald, Helmuth; Tomás, Helena; Shi, Xiangyang

    2015-03-01

    We report the use of arginine-glycine-aspartic (Arg-Gly-Asp, RGD) peptide-modified dendrimer-entrapped gold nanoparticles (Au DENPs) for highly efficient and specific gene delivery to stem cells. In this study, generation 5 poly(amidoamine) dendrimers modified with RGD via a poly(ethylene glycol) (PEG) spacer and with PEG monomethyl ether were used as templates to entrap gold nanoparticles (AuNPs). The native and the RGD-modified PEGylated dendrimers and the respective well characterized Au DENPs were used as vectors to transfect human mesenchymal stem cells (hMSCs) with plasmid DNA (pDNA) carrying both the enhanced green fluorescent protein and the luciferase (pEGFPLuc) reporter genes, as well as pDNA encoding the human bone morphogenetic protein-2 (hBMP-2) gene. We show that all vectors are capable of transfecting the hMSCs with both pDNAs. Gene transfection using pEGFPLuc was demonstrated by quantitative Luc activity assay and qualitative evaluation by fluorescence microscopy. For the transfection with hBMP-2, the gene delivery efficiency was evaluated by monitoring the hBMP-2 concentration and the level of osteogenic differentiation of the hMSCs via alkaline phosphatase activity, osteocalcin secretion, calcium deposition, and von Kossa staining assays. Our results reveal that the stem cell gene delivery efficiency is largely dependent on the composition and the surface functionality of the dendrimer-based vectors. The coexistence of RGD and AuNPs rendered the designed dendrimeric vector with specific stem cell binding ability likely via binding of integrin receptor on the cell surface and improved three-dimensional conformation of dendrimers, which is beneficial for highly efficient and specific stem cell gene delivery applications. PMID:25658033

  9. Targeted disruption of the CCR5 gene in human hematopoietic stem cells stimulated by peptide nucleic acids

    PubMed Central

    Schleifman, Erica B.; Bindra, Ranjit; Leif, Jean; Campo, Jacob del; Rogers, Faye A.; Uchil, Pradeep; Kutsch, Olaf; Shultz, Leonard D.; Kumar, Priti; Greiner, Dale L.; Glazer, Peter M.

    2011-01-01

    SUMMARY Peptide nucleic acids (PNAs) bind duplex DNA in a sequence-specific manner, creating triplex structures that can provoke DNA repair and produce genome modification. CCR5 encodes a chemokine receptor required for HIV-1 entry into human cells and individuals carrying mutations in this gene are resistant to HIV-1 infection. Transfection of human cells with PNAs targeted to the CCR5 gene, plus donor DNAs designed to introduce stop codons mimicking the naturally occurring CCR5-delta32 mutation, produced 2.46% targeted gene modification. CCR5 modification was confirmed at the DNA, RNA and protein levels and was shown to confer resistance to infection with HIV-1. Targeting of CCR5 was achieved in human CD34+ hematopoietic stem cells (HSCs) with subsequent engraftment into mice and persistence of the gene modification more than four months post-transplantation. This work suggests a therapeutic strategy for CCR5 knockout in HSCs from HIV-1-infected individuals, rendering cells resistant to HIV-1 and preserving immune system function. PMID:21944757

  10. Site-directed gene mutation at mixed sequence targets by psoralen-conjugated pseudo-complementary peptide nucleic acids.

    PubMed

    Kim, Ki-Hyun; Nielsen, Peter E; Glazer, Peter M

    2007-01-01

    Sequence-specific DNA-binding molecules such as triple helix-forming oligonucleotides (TFOs) provide a means for inducing site-specific mutagenesis and recombination at chromosomal sites in mammalian cells. However, the utility of TFOs is limited by the requirement for homopurine stretches in the target duplex DNA. Here, we report the use of pseudo-complementary peptide nucleic acids (pcPNAs) for intracellular gene targeting at mixed sequence sites. Due to steric hindrance, pcPNAs are unable to form pcPNA-pcPNA duplexes but can bind to complementary DNA sequences by Watson-Crick pairing via double duplex-invasion complex formation. We show that psoralen-conjugated pcPNAs can deliver site-specific photoadducts and mediate targeted gene modification within both episomal and chromosomal DNA in mammalian cells without detectable off-target effects. Most of the induced psoralen-pcPNA mutations were single-base substitutions and deletions at the predicted pcPNA-binding sites. The pcPNA-directed mutagenesis was found to be dependent on PNA concentration and UVA dose and required matched pairs of pcPNAs. Neither of the individual pcPNAs alone had any effect nor did complementary PNA pairs of the same sequence. These results identify pcPNAs as new tools for site-specific gene modification in mammalian cells without purine sequence restriction, thereby providing a general strategy for designing gene targeting molecules. PMID:17977869

  11. Effective gene delivery into human stem cells with a cell-targeting Peptide-modified bioreducible polymer.

    PubMed

    Beloor, Jagadish; Ramakrishna, Suresh; Nam, Kihoon; Seon Choi, Chang; Kim, Jongkil; Kim, Sung Hwa; Cho, Hyong Jin; Shin, HeungSoo; Kim, Hyongbum; Kim, Sung Wan; Lee, Sang-Kyung; Kumar, Priti

    2015-05-01

    Stem cells are poorly permissive to non-viral gene transfection reagents. In this study, we explored the possibility of improving gene delivery into human embryonic (hESC) and mesenchymal (hMSC) stem cells by synergizing the activity of a cell-binding ligand with a polymer that releases nucleic acids in a cytoplasm-responsive manner. A 29 amino acid long peptide, RVG, targeting the nicotinic acetylcholine receptor (nAchR) was identified to bind both hMSC and H9-derived hESC. Conjugating RVG to a redox-sensitive biodegradable dendrimer-type arginine-grafted polymer (PAM-ABP) enabled nanoparticle formation with plasmid DNA without altering the environment-sensitive DNA release property and favorable toxicity profile of the parent polymer. Importantly, RVG-PAM-ABP quantitatively enhanced transfection into both hMSC and hESC compared to commercial transfection reagents like Lipofectamine 2000 and Fugene. ∼60% and 50% of hMSC and hESC were respectively transfected, and at increased levels on a per cell basis, without affecting pluripotency marker expression. RVG-PAM-ABP is thus a novel bioreducible, biocompatible, non-toxic, synthetic gene delivery system for nAchR-expressing stem cells. Our data also demonstrates that a cell-binding ligand like RVG can cooperate with a gene delivery system like PAM-ABP to enable transfection of poorly-permissive cells. PMID:25515928

  12. High-resolution structure of HLA-A*0201 in complex with a tumour-specific antigenic peptide encoded by the MAGE-A4 gene.

    PubMed

    Hillig, R C; Coulie, P G; Stroobant, V; Saenger, W; Ziegler, A; Hülsmeyer, M

    2001-07-27

    The heterotrimeric complex of the human major histocompatibity complex (MHC) molecule HLA-A*0201, beta2-microglobulin and the decameric peptide GVYDGREHTV derived from the melanoma antigen (MAGE-A4 protein has been determined by X-ray crystallography at 1.4 A resolution. MAGE-A4 belongs to a family of genes that are specifically expressed in a variety of tumours. MAGE-A4-derived peptides are presented by MHC molecules at the cell surface to cytotoxic T-lymphocytes. As the HLA-A*0201:MAGE-A4 complex occurs only on tumour cells, it is considered to be an appropriate target for immunotherapy. The structure presented here reveals potential epitopes specific to the complex and indicates which peptide residues could be recognised by T-cell receptors. In addition, as the structure could be refined anisotropically, it was possible to describe the movements of the bound peptide in more detail. PMID:11502003

  13. A relationship between antimicrobial peptide gene expression and capacity of a selected shrimp line to survive a Vibrio infection.

    PubMed

    de Lorgeril, Julien; Gueguen, Yannick; Goarant, Cyrille; Goyard, Emmanuel; Mugnier, Chantal; Fievet, Julie; Piquemal, David; Bachère, Evelyne

    2008-07-01

    Understanding of antimicrobial defence mechanisms of penaeid shrimp should help in the design of efficient strategies for the management and disease control in aquaculture. In this study, we have specifically analysed the expression in circulating hemocytes of antimicrobial peptides (AMPs) encoding genes, such as PEN2 and PEN3, ALF, crustin, lysozyme and a putative cysteine-rich peptide. We evidenced a relationship between the level of expression of some AMPs and the successful response of the shrimp, Litopenaeus stylirostris, to circumvent a pathogenic Vibrio penaeicida infection. Additionally, significant differences in some AMP transcript amounts are evidenced between control, non-selected shrimp line and the third generation breeding of shrimp selected for their survival to natural V. penaeicida infections. On the basis of these results, it will now be of great interest to determine if these AMPs are directly involved in the resistance of shrimp to infection or if they only reflect other acquired defence mechanisms which can confer a resistance. PMID:18486974

  14. Cathelicidin LL-37 and HSV-1 Corneal Infection: Peptide Versus Gene Therapy

    PubMed Central

    Lee, Chyan-Jang; Buznyk, Oleksiy; Kuffova, Lucia; Rajendran, Vijayalakshmi; Forrester, John V.; Phopase, Jaywant; Islam, Mohammad M.; Skog, Mårten; Ahlqvist, Jenny; Griffith, May

    2014-01-01

    Purpose To evaluate the potential utility of collagen-based corneal implants with anti–Herpes Simplex Virus (HSV)-1 activity achieved through sustained release of LL-37, from incorporated nanoparticles, as compared with cell-based delivery from model human corneal epithelial cells (HCECs) transfected to produce endogenous LL-37. Methods We tested the ability of collagen-phosphorylcholine implants to tolerate the adverse microenvironment of herpetic murine corneas. Then, we investigated the efficacy of LL-37 peptides delivered through nanoparticles incorporated within the corneal implants to block HSV-1 viral activity. In addition, LL-37 complementary DNA (cDNA) was transferred into HCECs to confer viral resistance, and their response to HSV-1 infection was examined. Results Our implants remained in herpetic murine corneas 7 days longer than allografts. LL-37 released from the implants blocked HSV-1 infection of HCECs by interfering with viral binding. However, in pre-infected HCECs, LL-37 delayed but could not prevent viral spreading nor clear viruses from the infected cells. HCECs transfected with the LL-37 expressed and secreted the peptide. Secreted LL-37 inhibited viral binding in vitro but was insufficient to protect cells completely from HSV-1 infection. Nevertheless, secreted LL-37 reduced both the incidence of plaque formation and plaque size. Conclusion LL-37 released from composite nanoparticle-hydrogel corneal implants and HCEC-produced peptide, both showed anti–HSV-1 activity by blocking binding. However, while both slowed down virus spread, neither was able on its own to completely inhibit the viruses. Translational Relevance LL-37 releasing hydrogels may have potential utility as corneal substitutes for grafting in HSV-1 infected corneas, possibly in combination with LL-37 producing therapeutic cells. PMID:24932432

  15. Expression of proglucagon and proglucagon-derived peptide hormone receptor genes in the chicken

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To better understand how the glucagon system functions in birds, we utilized a molecular cloning strategy to sequence and characterize the chicken proglucagon gene. This gene has seven exons and six introns with evidence for an additional (alternate) first exon and two promoter regions. Two classes ...

  16. Synthesis and characterization of a PAMAM dendrimer nanocarrier functionalized by SRL peptide for targeted gene delivery to the brain.

    PubMed

    Zarebkohan, Amir; Najafi, Farhood; Moghimi, Hamid Reza; Hemmati, Mohammad; Deevband, Mohammad Reza; Kazemi, Bahram

    2015-10-12

    Blood-brain barrier inhibits most of drugs and genetic materials from reaching the brain. So, developing high efficiency carriers for gene and drug delivery to the brain, is the challenging area in pharmaceutical sciences. This investigation aimed to target DNA to brain using Serine-Arginine-Leucine (SRL) functionalized PAMAM dendrimers as a novel gene delivery system. The SRL peptide was linked on G4 PAMAM dendrimers using bifunctional PEG. DNA was then loaded in these functionalized nanoparticles and their physicochemical properties and cellular uptake/distribution evaluated by AFM, NMR, FTIR and fluorescence and confocal microscopy. Also, biodistribution and brain localization of nanoparticles were studied after IV injection of nanoparticles into rat tail. Unmodified nanoparticles were used as control in all evaluations. In vitro studies showed that SRL-modified nanoparticles have good transfection efficacy and low toxicity. Results also showed that SRL is a LRP ligand and SRL-modified nanoparticles internalized by clathrin/caveolin energy-dependent endocytosis to brain capillary endothelial cells. After intravenous administration, the SRL-modified nanoparticles were able to cross the blood-brain barrier and enter the brain parenchyma. Our result showed that, SRL-modified nanoparticles provide a safe and effective nanocarrier for brain gene delivery. PMID:26118442

  17. New Insight into the Ochratoxin A Biosynthetic Pathway through Deletion of a Nonribosomal Peptide Synthetase Gene in Aspergillus carbonarius

    SciTech Connect

    Gallo, A.; Bruno, K. S.; Solfrizzo, M.; Perrone, G.; Mule, G.; Visconti, A.; Baker, S. E.

    2012-09-14

    Ochratoxin A (OTA), a mycotoxin produced by Aspergillus and Penicillium species, is composed of a dihydroisocoumarin ring linked to phenylalanine and its biosynthetic pathway has not yet been completely elucidated. Most of the knowledge regarding the genetic and enzymatic aspects of OTA biosynthesis has been obtained in Penicillium species. In Aspergillus species only pks genes involved in the initial steps of the pathway have been partially characterized. In our study, the inactivation of a gene encoding a nonribosomal peptide synthetase in OTA producing A. carbonarius ITEM 5010 has removed the ability of the fungus to produce OTA. This is the first report on the involvement of an nrps gene product in OTA biosynthetic pathway in Aspergillus species. The absence of OTA and ochratoxin α-the isocoumaric derivative of OTA, and the concomitant increase of ochratoxin β- the dechloro analog of ochratoxin α- were observed in the liquid culture of transformed strain. The data provide the first evidence that the enzymatic step adding phenylalanine to polyketide dihydroisocoumarin precedes the chlorination step to form OTA in A. carbonarius, and that ochratoxin α is a product of hydrolysis of OTA, giving an interesting new insight in the biosynthetic pathway of the toxin.

  18. Mass spectrometric identification, sequence evolution, and intraspecific variability of dimeric peptides encoded by cockroach akh genes.

    PubMed

    Sturm, Sebastian; Predel, Reinhard

    2015-02-01

    Neuropeptides are structurally the most diverse group of messenger molecules of the nervous system. Regarding neuropeptide identification, distribution, function, and evolution, insects are among the best studied invertebrates. Indeed, more than 100 neuropeptides are known from single species. Most of these peptides can easily be identified by direct tissue or cell profiling using MALDI-TOF MS. In these experiments, protein hormones with extensive post-translational modifications such as inter- and intramolecular disulfides are usually missed. It is evident that an exclusion of these bioactive molecules hinders the utilization of direct profiling methods in comprehensive peptidomic analyses. In the current study, we focus on the detection and structural elucidation of homo- and heterodimeric adipokinetic hormone precursor-related peptides (APRPs) of cockroaches. The physiological relevance of these molecules with highly conserved sequences in insects is still uncertain. Sequence similarities with vertebrate growth hormone-releasing factors have been reported, but remarkably, few data regarding APRP processing exist and these data are restricted to locusts. Here, we elucidated sequences of carbamidomethylated APRP monomers of different cockroaches by means of MALDI-TOF MS(2), and we were able to identify a surprisingly large number of APRP sequences, resulting either from intraspecific amino acid substitutions within the APRP sequences or C-terminal truncated APRPs. PMID:25524231

  19. The IDA/IDA-LIKE and PIP/PIP-LIKE gene families in Arabidopsis: phylogenetic relationship, expression patterns, and transcriptional effect of the PIPL3 peptide

    PubMed Central

    Vie, Ane Kjersti; Najafi, Javad; Liu, Bin; Winge, Per; Butenko, Melinka A.; Hornslien, Karina S.; Kumpf, Robert; Aalen, Reidunn B.; Bones, Atle M.; Brembu, Tore

    2015-01-01

    Peptide ligands play crucial roles in the life cycle of plants by modulating the innate immunity against pathogens and regulating growth and developmental processes. One well-studied example is INFLORESCENCE DEFICIENT IN ABSCISSION (IDA), which controls floral organ abscission and lateral root emergence in Arabidopsis thaliana. IDA belongs to a family of five additional IDA-LIKE (IDL) members that have all been suggested to be involved in regulation of Arabidopsis development. Here we present three novel members of the IDL subfamily and show that two of them are strongly and rapidly induced by different biotic and abiotic stresses. Furthermore, we provide data that the recently identified PAMP-INDUCED SECRETED PEPTIDE (PIP) and PIP-LIKE (PIPL) peptides, which show similarity to the IDL and C-TERMINALLY ENCODED PEPTIDE (CEP) peptides, are not only involved in innate immune response in Arabidopsis but are also induced by abiotic stress. Expression patterns of the IDA/IDL and PIP/PIPL genes were analysed using in silico data, qRT-PCR and GUS promoter lines. Transcriptomic responses to PIPL3 peptide treatment suggested a role in regulation of biotic stress responses and cell wall modification. PMID:26062745

  20. The IDA/IDA-LIKE and PIP/PIP-LIKE gene families in Arabidopsis: phylogenetic relationship, expression patterns, and transcriptional effect of the PIPL3 peptide.

    PubMed

    Vie, Ane Kjersti; Najafi, Javad; Liu, Bin; Winge, Per; Butenko, Melinka A; Hornslien, Karina S; Kumpf, Robert; Aalen, Reidunn B; Bones, Atle M; Brembu, Tore

    2015-08-01

    Peptide ligands play crucial roles in the life cycle of plants by modulating the innate immunity against pathogens and regulating growth and developmental processes. One well-studied example is INFLORESCENCE DEFICIENT IN ABSCISSION (IDA), which controls floral organ abscission and lateral root emergence in Arabidopsis thaliana. IDA belongs to a family of five additional IDA-LIKE (IDL) members that have all been suggested to be involved in regulation of Arabidopsis development. Here we present three novel members of the IDL subfamily and show that two of them are strongly and rapidly induced by different biotic and abiotic stresses. Furthermore, we provide data that the recently identified PAMP-INDUCED SECRETED PEPTIDE (PIP) and PIP-LIKE (PIPL) peptides, which show similarity to the IDL and C-TERMINALLY ENCODED PEPTIDE (CEP) peptides, are not only involved in innate immune response in Arabidopsis but are also induced by abiotic stress. Expression patterns of the IDA/IDL and PIP/PIPL genes were analysed using in silico data, qRT-PCR and GUS promoter lines. Transcriptomic responses to PIPL3 peptide treatment suggested a role in regulation of biotic stress responses and cell wall modification. PMID:26062745

  1. The gene expression of the hypothalamic feeding-regulating peptides in cisplatin-induced anorexic rats.

    PubMed

    Yoshimura, Mitsuhiro; Matsuura, Takanori; Ohkubo, Junichi; Ohno, Motoko; Maruyama, Takashi; Ishikura, Toru; Hashimoto, Hirofumi; Kakuma, Tetsuya; Yoshimatsu, Hironobu; Terawaki, Kiyoshi; Uezono, Yasuhito; Ueta, Yoichi

    2013-08-01

    Cisplatin has been widely used; however, various disadvantageous side effects afflict patients. Rikkunshito (RKT), a traditional Japanese herbal medicine, has been widely prescribed in Japan to improve anorexia; but the mechanisms are unknown. Here we studied whether RKT could improve anorexia induced by cisplatin and changes in feeding-regulating peptides in the hypothalamus in rats. Adult male rats were divided into 4 groups: water+saline (WS), water+cisplatin (WC), RKT+saline (RS), and RKT+cisplatin (RC) groups. Water or RKT (1g/kg) was intragastrically administered for 4 days, from day -1 to day 2, and saline or cisplatin (6mg/kg) was intraperitoneally (i.p.) administered at day 0. After i.p. administration, cumulative food intake, water intake, urine volume and body weight were measured. The rats were then decapitated, followed by removal of the brain, and feeding-regulating peptides in the hypothalamus were measured by in situ hybridization histochemistry. In the three-day measurements, there were no significant changes in cumulative water intake and urine volume. The body weight and cumulative food intake in WC significantly decreased compared to WS, whereas these were not observed in RC. Pro-opiomelanocortin (POMC) and cocaine and amphetamine-regulated transcript (CART) in the arcuate nucleus (ARC) in WC significantly increased, and neuropeptide Y (NPY) in the ARC decreased compared to WS, whereas those in RS and RC were comparable to WS. These results suggest that RKT may have therapeutic potential for anorexia induced by cisplatin. PMID:23684922