Clinical characteristics of occult macular dystrophy in family with mutation of RP1l1 gene.

PubMed

Tsunoda, Kazushige; Usui, Tomoaki; Hatase, Tetsuhisa; Yamai, Satoshi; Fujinami, Kaoru; Hanazono, Gen; Shinoda, Kei; Ohde, Hisao; Akahori, Masakazu; Iwata, Takeshi; Miyake, Yozo

2012-06-01

To report the clinical characteristics of occult macular dystrophy (OMD) in members of one family with a mutation of the RP1L1 gene. Fourteen members with a p.Arg45Trp mutation in the RP1L1 gene were examined. The visual acuity, visual fields, fundus photographs, fluorescein angiograms, full-field electroretinograms, multifocal electroretinograms, and optical coherence tomographic images were examined. The clinical symptoms and signs and course of the disease were documented. All the members with the RP1L1 mutation except one woman had ocular symptoms and signs of OMD. The fundus was normal in all the patients during the entire follow-up period except in one patient with diabetic retinopathy. Optical coherence tomography detected the early morphologic abnormalities both in the photoreceptor inner/outer segment line and cone outer segment tip line. However, the multifocal electroretinograms were more reliable in detecting minimal macular dysfunction at an early stage of OMD. The abnormalities in the multifocal electroretinograms and optical coherence tomography observed in the OMD patients of different durations strongly support the contribution of RP1L1 mutation to the presence of this disease.

Genetic variations of the NPC1L1 gene associated with hepatitis C virus (HCV) infection and biochemical characteristics of HCV patients in China.

PubMed

Zhang, A-Mei; Zhang, Cheng-Lin; Song, Yuzhu; Zhao, Ping; Feng, Yue; Wang, Binghui; Li, Zheng; Liu, Li; Xia, Xueshan

2016-12-01

About 2% of the world population is infected with hepatitis C virus (HCV), a leading cause of hepatic cirrhosis and hepatocellular carcinoma. The Niemann-Pick C1-like 1 cholesterol absorption receptor (NPC1L1) was recently identified to be an important factor for HCV entry into host cells. Whether genetic variations of the NPC1L1 gene are associated with HCV infection is unknown. In this study, five single nucleotide polymorphisms (SNPs) of the NPC1L1 gene were analyzed in 261 HCV-infected individuals and 265 general controls from Yunnan Province, China. No significant differences were identified in genotypes or alleles of the SNPs between the two groups. After constructing haplotypes based on the five SNPs, a significant difference between HCV-infected individuals and general controls was shown for two haplotypes. Haplotype GCCTT appeared to be a protective factor and haplotype GCCCT was a risk factor for HCV-infected individuals. Genotypes of four SNPs correlated with biochemical characteristics of HCV-infected persons. Genotypes of SNPs rs799444 and rs2070607 were correlated with total bilirubin. Genotype TT of rs917098 was a risk factor for the gamma-glutamyltransferase level. Furthermore, HCV-infected individuals carrying genotype GG of rs41279633 showed statistically higher gamma-glutamyltransferase levels than HCV-infected persons with GT and TT. The results of this study identified the association between genetic susceptibility of the NPC1L1 gene and HCV infection, as well as biochemical characteristics of HCV-infected persons in Yunnan, China. Copyright © 2016 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Fine mapping of the genic male-sterile ms 1 gene in Capsicum annuum L.

PubMed

Jeong, Kyumi; Choi, Doil; Lee, Jundae

2018-01-01

The genomic region cosegregating with the genic male-sterile ms 1 gene of Capsicum annuum L. was delimited to a region of 869.9 kb on chromosome 5 through fine mapping analysis. A strong candidate gene, CA05g06780, a homolog of the Arabidopsis MALE STERILITY 1 gene that controls pollen development, was identified in this region. Genic male sterility caused by the ms 1 gene has been used for the economically efficient production of massive hybrid seeds in paprika (Capsicum annuum L.), a colored bell-type sweet pepper. Previously, a CAPS marker, PmsM1-CAPS, located about 2-3 cM from the ms 1 locus, was reported. In this study, we constructed a fine map near the ms 1 locus using high-resolution melting (HRM) markers in an F 2 population consisting of 1118 individual plants, which segregated into 867 male-fertile and 251 male-sterile plants. A total of 12 HRM markers linked to the ms 1 locus were developed from 53 primer sets targeting intraspecific SNPs derived by comparing genome-wide sequences obtained by next-generation resequencing analysis. Using this approach, we narrowed down the region cosegregating with the ms 1 gene to 869.9 kb of sequence. Gene prediction analysis revealed 11 open reading frames in this region. A strong candidate gene, CA05g06780, was identified; this gene is a homolog of the Arabidopsis MALE STERILITY 1 (MS1) gene, which encodes a PHD-type transcription factor that regulates pollen and tapetum development. Sequence comparison analysis suggested that the CA05g06780 gene is the strongest candidate for the ms 1 gene of paprika. To summarize, we developed a cosegregated marker, 32187928-HRM, for marker-assisted selection and identified a strong candidate for the ms 1 gene.

Early-onset obesity and paternal 2pter deletion encompassing the ACP1, TMEM18, and MYT1L genes.

PubMed

Doco-Fenzy, Martine; Leroy, Camille; Schneider, Anouck; Petit, Florence; Delrue, Marie-Ange; Andrieux, Joris; Perrin-Sabourin, Laurence; Landais, Emilie; Aboura, Azzedine; Puechberty, Jacques; Girard, Manon; Tournaire, Magali; Sanchez, Elodie; Rooryck, Caroline; Ameil, Agnès; Goossens, Michel; Jonveaux, Philippe; Lefort, Geneviève; Taine, Laurence; Cailley, Dorothée; Gaillard, Dominique; Leheup, Bruno; Sarda, Pierre; Geneviève, David

2014-04-01

Obesity is a common but highly, clinically, and genetically heterogeneous disease. Deletion of the terminal region of the short arm of chromosome 2 is rare and has been reported in about 13 patients in the literature often associated with a Prader-Willi-like phenotype. We report on five unrelated patients with 2p25 deletion of paternal origin presenting with early-onset obesity, hyperphagia, intellectual deficiency, and behavioural difficulties. Among these patients, three had de novo pure 2pter deletions, one presented with a paternal derivative der(2)t(2;15)(p25.3;q26) with deletion in the 2pter region and the last patient presented with an interstitial 2p25 deletion. The size of the deletions was characterized by SNP array or array-CGH and was confirmed by fluorescence in situ hybridization (FISH) studies. Four patients shared a 2p25.3 deletion with a minimal critical region estimated at 1.97 Mb and encompassing seven genes, namely SH3HYL1, ACP1, TMEMI8, SNTG2, TPO, PXDN, and MYT1L genes. The fifth patient had a smaller interstitial deletion encompassing the TPO, PXDN, and MYT1L genes. Paternal origin of the deletion was determined by genotyping using microsatellite markers. Analysis of the genes encompassed in the deleted region led us to speculate that the ACP1, TMEM18, and/or MYT1L genes might be involved in early-onset obesity. In addition, intellectual deficiency and behavioural troubles can be explained by the heterozygous loss of the SNTG2 and MYT1L genes. Finally, we discuss the parent-of-origin of the deletion.

Genetic variation at the NPC1L1 gene locus, plasma lipoproteins, and heart disease risk in the elderly

USDA-ARS?s Scientific Manuscript database

Niemann-Pick C1-like 1 protein (NPC1L1) plays a critical role in intestinal cholesterol absorption. Our objective was to examine whether five variants (-133A>G, -18A>C, L272L, V1296V, and U3_28650A>G) at the NPC1L1 gene have effects on lipid levels, prevalence, and incidence of coronary heart diseas...

Angioimmunoblastic T-cell lymphoma and hypereosinophilic syndrome with FIP1L1/PDGFRA fusion gene effectively treated with imatinib: A case report.

PubMed

Yamamoto, Masayo; Ikuta, Katsuya; Toki, Yasumichi; Hatayama, Mayumi; Shindo, Motohiro; Torimoto, Yoshihiro; Okumura, Toshikatsu

2017-09-01

Hypereosinophilic syndrome (HES) is a rare disorder characterized by hypereosinophilia and organ damage. Some cases of HES are caused by the FIP1L1/PDGFRA fusion gene and respond to imatinib. FIP1L1/PDGFRA-positive HES occasionally evolves into chronic eosinophilic leukemia or into another form of myeloproliferative neoplasm; however, the development of a malignant lymphoma is very rare. We present a rare case of angioimmunoblastic T-cell lymphoma (AITL) and HES with the FIP1L1/PDGFRA gene rearrangement. A man in his 30s presented to our hospital with fever, hypereosinophilia, widespread lymphadenopathy, and splenomegaly. Laboratory tests showed hypereosinophilia, increased soluble interleukin-2 receptor, and increased vitamin B12. Positron-emission tomography with F fluorodeoxyglucose (FDG) showed positive FDG uptake in multiple enlarged lymph nodes throughout the body and the red bone marrow. A bone-marrow biopsy showed hypereosinophilia without dysplasia and an increased number of blasts. The FIP1L1/PDGFRA fusion gene was positive upon fluorescence in situ hybridization (FISH) analysis of the peripheral blood. Furthermore, biopsy of a lymph node from the neck revealed restiform hyperplasia of capillary vessels, with small lymphoma cells arranged around the capillaries. Lymphoma cells were positive for CD3, CD4, and CD10, and negative for CD20. Lymphoma cells were also positive for the FIP1L1/PDGFRA fusion gene by FISH analysis. From these findings, the patient was diagnosed with HES and AITL with FIP1L1/PDGFRA. After the diagnosis, corticosteroid was administered but was ineffective. Imatinib was then administered. Imatinib was very effective for treating HES and AITL, and complete remission was achieved in both. This report presents the first case in which the FIP1L1/PDGFRA fusion gene was positive both in peripheral blood and lymph nodes, implying the possibility that the tumor cells acquired the FIP1L1/PDGFRA fusion gene in the early stage of hematopoietic

In planta Transformed Cumin (Cuminum cyminum L.) Plants, Overexpressing the SbNHX1 Gene Showed Enhanced Salt Endurance

PubMed Central

Pandey, Sonika; Patel, Manish Kumar; Jha, Bhavanath

2016-01-01

Cumin is an annual, herbaceous, medicinal, aromatic, spice glycophyte that contains diverse applications as a food and flavoring additive, and therapeutic agents. An efficient, less time consuming, Agrobacterium-mediated, a tissue culture-independent in planta genetic transformation method was established for the first time using cumin seeds. The SbNHX1 gene, cloned from an extreme halophyte Salicornia brachiata was transformed in cumin using optimized in planta transformation method. The SbNHX1 gene encodes a vacuolar Na+/H+ antiporter and is involved in the compartmentalization of excess Na+ ions into the vacuole and maintenance of ion homeostasis Transgenic cumin plants were confirmed by PCR using gene (SbNHX1, uidA and hptII) specific primers. The single gene integration event and overexpression of the gene were confirmed by Southern hybridization and competitive RT-PCR, respectively. Transgenic lines L3 and L13 showed high expression of the SbNHX1 gene compared to L6 whereas moderate expression was detected in L5 and L10 transgenic lines. Transgenic lines (L3, L5, L10 and L13), overexpressing the SbNHX1 gene, showed higher photosynthetic pigments (chlorophyll a, b and carotenoid), and lower electrolytic leakage, lipid peroxidation (MDA content) and proline content as compared to wild type plants under salinity stress. Though transgenic lines were also affected by salinity stress but performed better compared to WT plants. The ectopic expression of the SbNHX1 gene confirmed enhanced salinity stress tolerance in cumin as compared to wild type plants under stress condition. The present study is the first report of engineering salt tolerance in cumin, so far and the plant may be utilized for the cultivation in saline areas. PMID:27411057

In planta Transformed Cumin (Cuminum cyminum L.) Plants, Overexpressing the SbNHX1 Gene Showed Enhanced Salt Endurance.

PubMed

Pandey, Sonika; Patel, Manish Kumar; Mishra, Avinash; Jha, Bhavanath

2016-01-01

Cumin is an annual, herbaceous, medicinal, aromatic, spice glycophyte that contains diverse applications as a food and flavoring additive, and therapeutic agents. An efficient, less time consuming, Agrobacterium-mediated, a tissue culture-independent in planta genetic transformation method was established for the first time using cumin seeds. The SbNHX1 gene, cloned from an extreme halophyte Salicornia brachiata was transformed in cumin using optimized in planta transformation method. The SbNHX1 gene encodes a vacuolar Na+/H+ antiporter and is involved in the compartmentalization of excess Na+ ions into the vacuole and maintenance of ion homeostasis Transgenic cumin plants were confirmed by PCR using gene (SbNHX1, uidA and hptII) specific primers. The single gene integration event and overexpression of the gene were confirmed by Southern hybridization and competitive RT-PCR, respectively. Transgenic lines L3 and L13 showed high expression of the SbNHX1 gene compared to L6 whereas moderate expression was detected in L5 and L10 transgenic lines. Transgenic lines (L3, L5, L10 and L13), overexpressing the SbNHX1 gene, showed higher photosynthetic pigments (chlorophyll a, b and carotenoid), and lower electrolytic leakage, lipid peroxidation (MDA content) and proline content as compared to wild type plants under salinity stress. Though transgenic lines were also affected by salinity stress but performed better compared to WT plants. The ectopic expression of the SbNHX1 gene confirmed enhanced salinity stress tolerance in cumin as compared to wild type plants under stress condition. The present study is the first report of engineering salt tolerance in cumin, so far and the plant may be utilized for the cultivation in saline areas.

Constitutively overexpressing a tomato fructokinase gene (lefrk1) in cotton (Gossypium hirsutum L. cv. coker 312) positively affects plant vegetative growth, boll number and seed cotton yield.

USDA-ARS?s Scientific Manuscript database

Increasing fructokinase (FRK) activity in cotton (Gossypium hirsutum L.) plants may reduce fructose inhibition of sucrose synthase (Sus) and lead to improved fibre yield and quality. Cotton was transformed with a tomato (Solanum lycopersicum L.) fructokinase gene (LeFRK1) under the control of the C...

Possible roles of the transcription factor Nrf1 (NFE2L1) in neural homeostasis by regulating the gene expression of deubiquitinating enzymes.

PubMed

Taniguchi, Hiroaki; Okamuro, Shota; Koji, Misaki; Waku, Tsuyoshi; Kubo, Kaori; Hatanaka, Atsushi; Sun, Yimeng; Chowdhury, A M Masudul Azad; Fukamizu, Akiyoshi; Kobayashi, Akira

2017-02-26

The transcription factor Nrf1 (NFE2L1) maintains protein homeostasis (proteostasis) by regulating the gene expression of proteasome subunits in response to proteasome inhibition. The deletion of the Nrf1 gene in neural stem/progenitor cells causes severe neurodegeneration due to the accumulation of ubiquitinated proteins in Purkinje cells and motor neurons (Nrf1 NKO mice). However, the molecular mechanisms governing this neurodegenerative process remain unclear. We demonstrate herein that the loss of Nrf1 leads to the reduced gene expression of the deubiquitinating enzymes (DUBs) but not proteasome subunits in Nrf1 NKO mice between P7 and P18. First, we show that K48-linked polyubiquitinated proteins accumulate in Nrf1-deficient Purkinje cells and cerebral cortex neurons. Nevertheless, loss of Nrf1 does not alter the expression and proteolytic activity of proteasome. A significantly reduced expression of deubiquitinating enzymes was also demonstrated in Nrf1-deficient cerebellar tissue using microarray analysis. The genome database further reveals species-conserved ARE, a Nrf1 recognition element, in the regulatory region of certain DUB genes. Furthermore, we show that Nrf1 can activate Usp9x gene expression related to neurodegeneration. Altogether these findings suggest that neurodegeneration in Nrf1 NKO mice may stem from the dysfunction of the ubiquitin-mediated regulation of neuronal proteins. Copyright © 2017 Elsevier Inc. All rights reserved.

MASA syndrome is caused by mutations in the neural cell adhesion gene, L1CAM

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

Schwartz, C.E.; Wang, Y.; Schroer, R.J.

1994-09-01

The MASA syndrome is a recessive X-linked disorder characterized by Mental retardation, Adducted thumbs, Shuffling gait and Aphasia. Recently we found that MASA in one family was likely caused by a point mutation in exon 6 of the L1CAM gene. This gene has also been shown to be involved in X-linked hydrocephalus (HSAS). We have screened 60 patients with either sporadic HSAS or MASA as well as two additional families with MASA. For the screening, we initially utilized 3 cDNA probes for the L1CAM gene. In one of the MASA families, K8310, two affected males were found to have anmore » altered BglII band. The band was present in their carrier mother but not in their normal brothers. This band was detected by the entire cDNA probe as well as the cDNA probe for 3{prime} end of the gene. Analysis of the L1CAM sequence indicated the altered BglII site is distal to the exon 28 but proximal to the punative poly A signal site. It is hypothesized that this point mutation alters the stability of the L1CAM mRNA. This is being tested using cell lines established from the two affected males.« less

Evaluation of the effect of divalent metal transporter 1 gene polymorphism on blood iron, lead and cadmium levels

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

Kayaaltı, Zeliha, E-mail: kayaalti@ankara.edu.tr; Akyüzlü, Dilek Kaya; Söylemezoğlu, Tülin

Divalent metal transporter 1 (DMT1), a member of the proton-coupled metal ion transporter family, mediates transport of ferrous iron from the lumen of the intestine into the enterocyte and export of iron from endocytic vesicles. It has an affinity not only for iron but also for other divalent cations including manganese, cobalt, nickel, cadmium, lead, copper, and zinc. DMT1 is encoded by the SLC11a2 gene that is located on chromosome 12q13 in humans and express four major mammalian isoforms (1A/+IRE, 1A/-IRE, 2/+IRE and 2/-IRE). Mutations or polymorphisms of DMT1 gene may have an impact on human health by disturbing metalmore » trafficking. To study the possible association of DMT1 gene with the blood levels of some divalent cations such as iron, lead and cadmium, a single nucleotide polymorphism (SNP) (IVS4+44C/A) in DMT1 gene was investigated in 486 unrelated and healthy individuals in a Turkish population by method of polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP). The genotype frequencies were found as 49.8% homozygote typical (CC), 38.3% heterozygote (CA) and 11.9% homozygote atypical (AA). Metal levels were analyzed by dual atomic absorption spectrometer system and the average levels of iron, lead and cadmium in the blood samples were 446.01±81.87 ppm, 35.59±17.72 ppb and 1.25±0.87 ppb, respectively. Individuals with the CC genotype had higher blood iron, lead and cadmium levels than those with AA and CA genotypes. Highly statistically significant associations were detected between IVS4+44 C/A polymorphism in the DMT1 gene and iron and lead levels (p=0.001 and p=0.036, respectively), but no association was found with cadmium level (p=0.344). This study suggested that DMT1 IVS4+44 C/A polymorphism is associated with inter-individual variations in blood iron, lead and cadmium levels. - Highlights: • DMT1 IVS4+44 C/A polymorphism is associated with inter-individual variations in blood iron, cadmium and lead levels. �

Characterization of a Mobile clpL Gene from Lactobacillus rhamnosus

PubMed Central

Suokko, Aki; Savijoki, Kirsi; Malinen, Erja; Palva, Airi; Varmanen, Pekka

2005-01-01

Two genes encoding ClpL ATPase proteins were identified in a probiotic Lactobacillus rhamnosus strain, E-97800. Sequence analyses revealed that the genes, designated clpL1 and clpL2, share 80% identity. The clpL2 gene showed the highest degree of identity (98.5%) to a clpL gene from Lactobacillus plantarum WCFSI, while it was not detected in three other L. rhamnosus strains studied. According to Northern analyses, the expression of clpL1 and the clpL2 were induced during heat shock by >20- and 3-fold, respectively. The functional promoter regions were determined by primer extension analyses, and the clpL1 promoter was found to be overlapped by an inverted repeat structure identical to the conserved CIRCE element, indicating that clpL1 belongs to the HrcA regulon in L. rhamnosus. No consensus binding sites for HrcA or CtsR could be identified in the clpL2 promoter region. Interestingly, the clpL2 gene was found to be surrounded by truncated transposase genes and flanked by inverted repeat structures nearly identical to the terminal repeats of the ISLpl1 from L. plantarum HN38. Furthermore, clpL2 was shown to be mobilized during prolonged cultivation at elevated temperature. The presence of a gene almost identical to clpL2 in L. plantarum and its absence in other L. rhamnosus strains suggest that the L. rhamnosus E-97800 has acquired the clpL2 gene via horizontal transfer. No change in the stress tolerance of the ClpL2-deficient derivative of E-97800 compared to the parental strain was observed. PMID:15812039

AtFXG1, an Arabidopsis gene encoding alpha-L-fucosidase active against fucosylated xyloglucan oligosaccharides.

PubMed

de La Torre, Francisco; Sampedro, Javier; Zarra, Ignacio; Revilla, Gloria

2002-01-01

An alpha-L-fucosidase (EC 3.2.1.51) able to release the t-fucosyl residue from the side chain of xyloglucan oligosaccharides has been detected in the leaves of Arabidopsis plants. Moreover, an alpha-L-fucosidase with similar substrate specificity was purified from cabbage (Brassica oleracea) leaves to render a single band on SDS-PAGE. Two peptide sequences were obtained from this protein band, and they were used to identify an Arabidopsis gene coding for an alpha-fucosidase that we propose to call AtFXG1. In addition, an Arabidopsis gene with homology with known alpha-L-fucosidases has been also found, and we proposed to name it as AtFUC1. Both AtFXG1 and ATFUC1 were heterologously expressed in Pichia pastoris cells and the alpha-L-fucosidase activities secreted to the culture medium. The alpha-L-fucosidase encoded by AtFXG1 was active against the oligosaccharides from xyloglucan XXFG as well as against 2'-fucosyl-lactitol but not against p-nitrophenyl-alpha-L-fucopyranoside. However, the AtFUC1 heterologously expressed was active only against 2'-fucosyl-lactitol. Thus, the former must be related to xyloglucan metabolism.

Intronic L1 Retrotransposons and Nested Genes Cause Transcriptional Interference by Inducing Intron Retention, Exonization and Cryptic Polyadenylation

PubMed Central

Kaer, Kristel; Branovets, Jelena; Hallikma, Anni; Nigumann, Pilvi; Speek, Mart

2011-01-01

Background Transcriptional interference has been recently recognized as an unexpectedly complex and mostly negative regulation of genes. Despite a relatively few studies that emerged in recent years, it has been demonstrated that a readthrough transcription derived from one gene can influence the transcription of another overlapping or nested gene. However, the molecular effects resulting from this interaction are largely unknown. Methodology/Principal Findings Using in silico chromosome walking, we searched for prematurely terminated transcripts bearing signatures of intron retention or exonization of intronic sequence at their 3′ ends upstream to human L1 retrotransposons, protein-coding and noncoding nested genes. We demonstrate that transcriptional interference induced by intronic L1s (or other repeated DNAs) and nested genes could be characterized by intron retention, forced exonization and cryptic polyadenylation. These molecular effects were revealed from the analysis of endogenous transcripts derived from different cell lines and tissues and confirmed by the expression of three minigenes in cell culture. While intron retention and exonization were comparably observed in introns upstream to L1s, forced exonization was preferentially detected in nested genes. Transcriptional interference induced by L1 or nested genes was dependent on the presence or absence of cryptic splice sites, affected the inclusion or exclusion of the upstream exon and the use of cryptic polyadenylation signals. Conclusions/Significance Our results suggest that transcriptional interference induced by intronic L1s and nested genes could influence the transcription of the large number of genes in normal as well as in tumor tissues. Therefore, this type of interference could have a major impact on the regulation of the host gene expression. PMID:22022525

A possible regulatory link between Twist 1 and PPARγ gene regulation in 3T3-L1 adipocytes.

PubMed

Ren, Rui; Chen, Zhufeng; Zhao, Xia; Sun, Tao; Zhang, Yuchao; Chen, Jie; Lu, Sumei; Ma, Wanshan

2016-11-08

Peroxisome proliferator-activated receptor γ (PPARγ) is a critical gene that regulates the function of adipocytes. Therefore, studies on the molecular regulation mechanism of PPARγ are important to understand the function of adipose tissue. Twist 1 is another important functional gene in adipose tissue, and hundreds of genes are regulated by Twist 1. The aim of this study was to investigate the regulation of Twist 1 and PPARγ expression in 3T3-L1 mature adipocytes. We induced differentiation in 3T3-L1 preadipocytes and examined alterations in Twist 1 and PPARγ expression. We used the PPARγ agonist pioglitazone and the PPARγ antagonist T0070907 to investigate the effect of PPARγ on Twist 1 expression. In addition, we utilized retroviral interference and overexpression of Twist 1 to determine the effects of Twist 1 on PPARγ expression. The expression levels of Twist 1 and PPARγ were induced during differentiation in 3T3-L1 adipocytes. Application of either a PPARγ agonist (pioglitazone) or antagonist (T0070907) influenced Twist 1 expression, with up-regulation of Twist 1 under pioglitazone (1 μM, 24 h) and down-regulation of Twist 1 under T0070907 (100 μM, 24 h) exposure. Furthermore, the retroviral interference of Twist 1 decreased the protein and mRNA expression of PPARγ, while Twist 1 overexpression had the opposite effect. There was a possible regulatory link between Twist 1 and PPARγ in 3T3-L1 mature adipocytes. This regulatory link enhanced the regulation of PPARγ and may be a functional mechanism of Twist 1 regulation of adipocyte physiology and pathology.

Association between PD-L1 expression and driven gene status in NSCLC: A meta-analysis.

PubMed

Li, D; Zhu, X; Wang, H; Li, N

2017-07-01

We explored the potential clinical association between programmed death-ligand 1 (PD-L1) expression and driven gene status in non-small cell lung cancer (NSCLC). We systemically searched through October 2015. Odd ratios (ORs) with 95% CIs were calculated to examine the association of PD-L1 expression with driven gene status. A random- or fixed-effects model was used. Nine studies were identified. KRAS-mutant tumors were more likely to be PD-L1 positive than KRAS-wild type tumors (51% vs 36%; OR 1.69; 95% CI 1.01-2.84; p = 0.045). In contrast, PD-L1 expression did not differ by EGFR (OR 0.86; 95% CI 0.43-1.73; p = 0.675) or ALK (OR 1.02; 95% CI 0.44-2.37; p = 0.954) status. In subgroup analysis, there was also no significant association between PD-L1 expression and EGFR status in term of the cut-offs or ethnicity. In conclusion, NSCLC with KRAS mutations showed a trend for higher frequency of positive PD-L1 expression. Copyright © 2017 Elsevier Ltd, BASO ~ The Association for Cancer Surgery, and the European Society of Surgical Oncology. All rights reserved.

An NPC1L1 gene promoter variant is associated with autosomal dominant hypercholesterolemia.

PubMed

Martín, B; Solanas-Barca, M; García-Otín, A-L; Pampín, S; Cofán, M; Ros, E; Rodríguez-Rey, J-C; Pocoví, M; Civeira, F

2010-05-01

A substantial number of subjects with autosomal dominant hypercholesterolemia (ADH) do not have LDL receptor (LDLR) or apolipoprotein B (APOB) mutations. Some ADH subjects appear to hyperabsorb sterols from the intestine, thus we hypothesized that they could have variants of the Niemann-Pick C1-Like 1 gene (NPC1L1). NPC1L1 encodes a crucial protein involved in intestinal sterol absorption. Four NPC1L1 variants (-133A>G, -18C>A, 1679C>G, 28650A>G) were analyzed in 271 (155 women and 116 men) ADH bearers without mutations in LDLR or APOB aged 30-70years and 274 (180 women and 94 men) control subjects aged 25-65years. The AC haplotype determined by the -133A>G and -18C>A variants was underrepresented in ADH subjects compared to controls (p=0.01). In the ADH group, cholesterol absorption/synthesis markers were significantly lower in AC homozygotes that in all others haplotypes. Electrophoretic mobility shift assay (EMSA) results revealed that the -133A-specific oligonucleotide produced a retarded band stronger than the -133G allele. Luciferase activity with NPC1L1 -133G variant was 2.5-fold higher than with the -133A variant. The -133A>G polymorphism exerts a significant effect on NPC1L1 promoter activity. NPC1L1 promoter variants might explain in part the hypercholesterolemic phenotype of some subjects with nonLDLR/nonAPOB ADH. Copyright 2009 Elsevier B.V. All rights reserved.

Genetic variability in E6, E7 and L1 genes of Human Papillomavirus 62 and its prevalence in Mexico.

PubMed

Artaza-Irigaray, Cristina; Flores-Miramontes, María Guadalupe; Olszewski, Dominik; Magaña-Torres, María Teresa; López-Cardona, María Guadalupe; Leal-Herrera, Yelda Aurora; Piña-Sánchez, Patricia; Jave-Suárez, Luis Felipe; Aguilar-Lemarroy, Adriana

2017-01-01

population and in women with CIN1, and in 90.5% of the cases it was found in coinfection with other HPVs. Novel nucleotide changes in its L1, E6 and E7 genes were detected, some of them lead to changes in the protein sequence.

Sex-based differences in gene expression in hippocampus following postnatal lead exposure

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

Schneider, J.S., E-mail: jay.schneider@jefferson.edu; Anderson, D.W.; Sonnenahalli, H.

The influence of sex as an effect modifier of childhood lead poisoning has received little systematic attention. Considering the paucity of information available concerning the interactive effects of lead and sex on the brain, the current study examined the interactive effects of lead and sex on gene expression patterns in the hippocampus, a structure involved in learning and memory. Male or female rats were fed either 1500 ppm lead-containing chow or control chow for 30 days beginning at weaning.Blood lead levels were 26.7 {+-} 2.1 {mu}g/dl and 27.1 {+-} 1.7 {mu}g/dl for females and males, respectively. The expression of 175more » unique genes was differentially regulated between control male and female rats. A total of 167 unique genes were differentially expressed in response to lead in either males or females. Lead exposure had a significant effect without a significant difference between male and female responses in 77 of these genes. In another set of 71 genes, there were significant differences in male vs. female response. A third set of 30 genes was differentially expressed in opposite directions in males vs. females, with the majority of genes expressed at a lower level in females than in males. Highly differentially expressed genes in males and females following lead exposure were associated with diverse biological pathways and functions. These results show that a brief exposure to lead produced significant changes in expression of a variety of genes in the hippocampus and that the response of the brain to a given lead exposure may vary depending on sex. - Highlights: > Postnatal lead exposure has a significant effect on hippocampal gene expression patterns. > At least one set of genes was affected in opposite directions in males and females. > Differentially expressed genes were associated with diverse biological pathways.« less

Blood lead levels, iron metabolism gene polymorphisms and homocysteine: a gene-environment interaction study.

PubMed

Kim, Kyoung-Nam; Lee, Mee-Ri; Lim, Youn-Hee; Hong, Yun-Chul

2017-12-01

Homocysteine has been causally associated with various adverse health outcomes. Evidence supporting the relationship between lead and homocysteine levels has been accumulating, but most prior studies have not focused on the interaction with genetic polymorphisms. From a community-based prospective cohort, we analysed 386 participants (aged 41-71 years) with information regarding blood lead and plasma homocysteine levels. Blood lead levels were measured between 2001 and 2003, and plasma homocysteine levels were measured in 2007. Interactions of lead levels with 42 genotyped single-nucleotide polymorphisms (SNPs) in five genes ( TF , HFE , CBS , BHMT and MTR ) were assessed via a 2-degree of freedom (df) joint test and a 1-df interaction test. In secondary analyses using imputation, we further assessed 58 imputed SNPs in the TF and MTHFR genes. Blood lead concentrations were positively associated with plasma homocysteine levels (p=0.0276). Six SNPs in the TF and MTR genes were screened using the 2-df joint test, and among them, three SNPs in the TF gene showed interactions with lead with respect to homocysteine levels through the 1-df interaction test (p<0.0083). Seven SNPs in the MTHFR gene were associated with homocysteine levels at an α-level of 0.05, but the associations did not persist after Bonferroni correction. These SNPs did not show interactions with lead levels. Blood lead levels were positively associated with plasma homocysteine levels measured 4-6 years later, and three SNPs in the TF gene modified the association. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

A new insight to adsorption and accumulation of high lead concentration by exopolymer and whole cells of lead-resistant bacterium Acinetobacter junii L. Pb1 isolated from coal mine dump.

PubMed

Kushwaha, Anamika; Rani, Radha; Kumar, Sanjay; Thomas, Tarence; David, Arun Alfred; Ahmed, Meraz

2017-04-01

A lead-resistant bacterial strain was isolated from coal mine dump and identified as Acinetobacter junii Pb1 on basis of 16S rRNA (ribosomal ribonucleic acid) gene sequencing. The minimum inhibitory concentration of lead for the strain was 16,000 mg l -1 and it showed antibiotic and multi metal resistance. In aqueous culture, at an initial lead (Pb(II)) concentration of 100 and 500 mg l -1 , lead adsorption and accumulation by the isolate was 100 and 60%, at pH 7 at 30 °C after 48 and 120 h, respectively. The two fractions of exopolysaccharide (EPS), loosely associated EPS (laEPS) and bound EPS (bEPS), and whole cells (devoid of EPS) showed high binding affinity towards Pb(II). The binding affinity of laEPS towards Pb(II) (1071 mg Pb g -1 ) was three times higher than that of bEPS (321.5 mg Pb g -1 ) and 6.5 times higher than that of whole cells (165 mg Pb g -1 ). The binding affinity of EPS and whole cells with Pb(II), reported in the current study, is considerably higher as compared to that reported in the literature, till date. SEM analysis, showed an increase in thickness of cells on exposure to Pb(II) and TEM analysis, revealed its accumulation (interior of cell) and its adsorption (with the external cell surface). The isolate was also found to be positive for indole acetic acid (IAA) and 1-aminocyclopropane-1-carboxylate (ACC) deaminase production which helps in promoting plant growth. Thus, this study provides a new understanding towards Pb(II) uptake by A. junii Pb1, highlighting its potential on the restoration of Pb(II) contaminated repositories.

Gene expression profiling of 3T3-L1 adipocytes exposed to phloretin.

PubMed

Hassan, Meryl; El Yazidi, Claire; Malezet-Desmoulins, Christiane; Amiot, Marie-Josèphe; Margotat, Alain

2010-07-01

Adipocyte dysfunction plays a major role in the outcome of obesity, insulin resistance and related cardiovascular complications. Thus, considerable efforts are underway in the pharmaceutical industry to find molecules that target the now well-documented pleiotropic functions of adipocyte. We previously reported that the dietary flavonoid phloretin enhances 3T3-L1 adipocyte differentiation and adiponectin expression at least in part through PPAR gamma activation. The present study was designed to further characterize the molecular mechanisms underlying the phloretin-mediated effects on 3T3-L1 adipocytes using microarray technology. We show that phloretin positively regulates the expression of numerous genes involved in lipogenesis and triglyceride storage, including GLUT4, ACSL1, PEPCK1, lipin-1 and perilipin (more than twofold). The expression of several genes encoding adipokines, in addition to adiponectin and its receptor, is positively or negatively regulated in a way that suggests a possible reduction in systemic insulin resistance and obesity-associated inflammation. Improvement of insulin sensitivity is also suggested by the overexpression of genes associated with insulin signal transduction, such as CAP, PDK1 and Akt2. Many of these genes are PPAR gamma targets, confirming the involvement of PPAR gamma pathway in the phloretin effects on adipocytes. In light of these microarray data, it is reasonable to assume that phloretin may be beneficial for reducing insulin resistance, in a similar way to the thiazolidinedione class of antidiabetic drugs. (c) 2010 Elsevier Inc. All rights reserved.

High-throughput detection of human papillomavirus-18 L1 gene methylation, a candidate biomarker for the progression of cervical neoplasia.

PubMed

Turan, Tolga; Kalantari, Mina; Cuschieri, Kate; Cubie, Heather A; Skomedal, Hanne; Bernard, Hans-Ulrich

2007-04-25

The L1 gene of human papillomavirus-18 (HPV-18) is consistently hypermethylated in cervical carcinomas, but frequently hypo- or unmethylated in exfoliated cells from asymptomatic patients. In precancerous lesions, L1 is sporadically hypermethylated, correlating with the severity of the neoplasia. In order to explore the potential of using L1 methylation as a workable biomarker for carcinogenic progression of HPV-18 infections in routinely taken samples, our aim was to develop methylation-detection techniques that were sensitive and rapid without being overly complex technically. Therein, we developed a methylation-specific PCR (MSP) through the design of primer sets that specifically amplify either methylated or unmethylated HPV-18 L1 DNA within bisulfite-modified sample DNA. Amplification of unmethylated and in vitro methylated HPV-18 DNA by MSP resulted in 2500 copies of either of the two L1 DNA species being detected, a satisfactory sensitivity considering that bisulfite treatment leads to the fragmentation of about 99% of sample DNA. The primers proved specific and did not generate false positive results at concentrations exceeding the lowest limit of detection by a factor of 400. DNA from carcinomas yielded PCR signals only with the methylation-specific primers, and not with primers specific for unmethylated L1 genes. The inverse result was obtained with DNA from precursor lesions that contained only hypomethylated DNA. High-grade precursor lesions and carcinomas that contained hyper- as well as hypomethylated L1 DNA yielded PCR signals with both primers. By developing a fluorescence based real-time PCR, we quantitatively analyzed samples with in vitro methylated and unmethylated L1 DNA, and could distinguish clinical samples with hyper- and hypomethylated DNA or mixtures of both DNAs. The methylation-specific and real-time PCR techniques permitted efficient HPV-18 L1 methylation analyses and open the door for larger-scale clinical studies where the utility of

QCD on the BlueGene/L Supercomputer

NASA Astrophysics Data System (ADS)

Bhanot, G.; Chen, D.; Gara, A.; Sexton, J.; Vranas, P.

2005-03-01

In June 2004 QCD was simulated for the first time at sustained speed exceeding 1 TeraFlops in the BlueGene/L supercomputer at the IBM T.J. Watson Research Lab. The implementation and performance of QCD in the BlueGene/L is presented.

Ascorbic acid deficiency leads to increased grain chalkiness in transgenic rice for suppressed of L-GalLDH.

PubMed

Yu, Le; Liu, Yonghai; Lu, Lina; Zhang, Qilei; Chen, Yezheng; Zhou, Liping; Chen, Hua; Peng, Changlian

2017-04-01

The grain chalkiness of rice (Oryza sativa L.), which determines the rice quality and price, is a major concern in rice breeding. Reactive oxygen species (ROS) plays a critical role in regulating rice endosperm chalkiness. Ascorbic acid (Asc) is a major plant antioxidant, which strictly regulates the levels of ROS. l-galactono-1, 4-lactone dehydrogenase (L-GalLDH, EC 1.3.2.3) is an enzyme that catalyzes the last step of Asc biosynthesis in higher plants. Here we show that the L-GalLDH-suppressed transgenic rice, GI-1 and GI-2, which have constitutively low (between 30% and 50%) leaf and grain Asc content compared with the wild-type (WT), exhibit significantly increased grain chalkiness. Further examination showed that the deficiency of Asc resulted in a higher lipid peroxidation and H 2 O 2 content, accompanied by a lower hydroxyl radical scavenging rate, total antioxidant capacity and photosynthetic ability. In addition, changes of the enzyme activities and gene transcript abundances related to starch synthesis were also observed in GI-1 and GI-2 grains. The results we presented here suggest a close correlation between Asc deficiency and grain chalkiness in the L-GalLDH-suppressed transgenics. Asc deficiency leads to the accumulation of H 2 O 2 , affecting antioxidant capacity and photosynthetic function, changing enzyme activities and gene transcript abundances related to starch synthesis, finally leading to the increased grain chalkiness. Copyright © 2017 Elsevier GmbH. All rights reserved.

E3L and F1L Gene Functions Modulate the Protective Capacity of Modified Vaccinia Virus Ankara Immunization in Murine Model of Human Smallpox.

PubMed

Volz, Asisa; Jany, Sylvia; Freudenstein, Astrid; Lantermann, Markus; Ludwig, Holger; Sutter, Gerd

2018-01-04

The highly attenuated Modified Vaccinia virus Ankara (MVA) lacks most of the known vaccinia virus (VACV) virulence and immune evasion genes. Today MVA can serve as a safety-tested next-generation smallpox vaccine. Yet, we still need to learn about regulatory gene functions preserved in the MVA genome, such as the apoptosis inhibitor genes F1L and E3L . Here, we tested MVA vaccine preparations on the basis of the deletion mutant viruses MVA-ΔF1L and MVA-ΔE3L for efficacy against ectromelia virus (ECTV) challenge infections in mice. In non-permissive human tissue culture the MVA deletion mutant viruses produced reduced levels of the VACV envelope antigen B5. Upon mousepox challenge at three weeks after vaccination, MVA-ΔF1L and MVA-ΔE3L exhibited reduced protective capacity in comparison to wildtype MVA. Surprisingly, however, all vaccines proved equally protective against a lethal ECTV infection at two days after vaccination. Accordingly, the deletion mutant MVA vaccines induced high levels of virus-specific CD8+ T cells previously shown to be essential for rapidly protective MVA vaccination. These results suggest that inactivation of the anti-apoptotic genes F1L or E3L modulates the protective capacity of MVA vaccination most likely through the induction of distinct orthopoxvirus specific immunity in the absence of these viral regulatory proteins.

Polymorphisms of vitamin K-related genes (EPHX1 and VKORC1L1) and stable warfarin doses.

PubMed

Chung, Jee-Eun; Lee, Kyung Eun; Chang, Byung Chul; Gwak, Hye Sun

2018-01-30

The aim of this study was to investigate the possible effects of EPHX1 and VKORC1L1 polymorphisms on variability of responses to warfarin. Sixteen single nucleotide polymorphisms (SNPs) in 201 patients with stable warfarin doses were analyzed including genes of VKORC1, CYP2C9, CYP4F2, GGCX, EPHX1 and VKORC1L1. Univariate analysis was conducted for the association of genotypes with stable warfarin doses. Multiple linear regression analysis was used to investigate factors that independently affected the inter-individual variability of warfarin dose requirements. The rs4072879 of VKORC1L1 (A>G) was significantly associated with stable warfarin doses; wild homozygote carriers (AA) required significantly lower stable warfarin doses than those with the variant G allele (5.02±1.56 vs. 5.96±2.01mg; p=0.001). Multivariate analysis showed that EPHX1 rs1877724 and VKORC1L1 rs4072879 accounted for 1.5% and 1.3% of the warfarin dose variability. Adding EPHX1 and VKORC1L1 SNPs to the base model including non-genetic variables (operation age, body weight and the therapy of ACEI or ARB) and genetic variables (VKORC1 rs9934438, CYP2C9 rs1057910, and CYP4F2 rs2108622) gave a number needed to genotype of 34. This study showed that polymorphisms of EPHX1 and VKORC1L1 could be determinants of stable warfarin doses. Copyright © 2017. Published by Elsevier B.V.

GhL1L1 affects cell fate specification by regulating GhPIN1-mediated auxin distribution.

PubMed

Xu, Jiao; Yang, Xiyan; Li, Baoqi; Chen, Lin; Min, Ling; Zhang, Xianlong

2018-05-13

Auxin is as an efficient initiator and regulator of cell fate during somatic embryogenesis (SE), but the molecular mechanisms and regulating networks of this process are not well understood. In this report, we analysed SE process induced by Leafy cotyledon1-like 1 (GhL1L1), a NF-YB subfamily gene specifically expressed in embryonic tissues in cotton. We also identified the target gene of GhL1L1, and its role in auxin distribution and cell fate specification during embryonic development was analysed. Overexpression of GhL1L1 accelerated embryonic cell formation, associated with an increased concentration of IAA in embryogenic calluses (ECs) and in the shoot apical meristem (SAM), corresponding to altered expression of the auxin transport gene GhPIN1. By contrast, GhL1L1-deficient explants showed retarded embryonic cell formation, and the concentration of IAA was decreased in GhL1L1-deficient ECs. Disruption of auxin distribution accelerated the specification of embryonic cell fate together with regulation of GhPIN1. Furthermore, we showed that PHOSPHATASE 2AA2 (GhPP2AA2) was activated by GhL1L1 through targeting the G-box of its promoter, hence regulating the activity of GhPIN1 protein. Our results indicate that GhL1L1 functions as a key regulator in auxin distribution to regulate cell fate specification in cotton and contribute to the understanding of the complex process of SE in plant species. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Expression and regulation of glycoprotein C gene of herpes simplex virus 1 resident in a clonal L-cell line.

PubMed Central

Arsenakis, M; Tomasi, L F; Speziali, V; Roizman, B; Campadelli-Fiume, G

1986-01-01

Ltk- cells were transfected with a plasmid containing the entire domain of glycoprotein C (gC), a true gamma or gamma 2 gene of herpes simplex virus 1 (HSV-1) and the methotrexate-resistant mouse dihydrofolate reductase mutant gene. The resulting methotrexate-resistant cell line was cloned; of the 39 clonal lines tested only 1, L3153(28), expressed gC after infection with HSV-1(MP), a gC- mutant, and none expressed gC constitutively. The induction of gC was optimal at multiplicities ranging between 0.5 and 2 PFU per cell, and the quantities produced were equivalent to or higher than those made by methotrexate-resistant gC- L cells infected with wild-type (gC+) virus. The gC gene resident in the L3153(28) cells was regulated as a beta gene inasmuch as the amounts of gC made in infected L3153(28) cells exposed to concentrations of phosphonoacetate that inhibited viral DNA synthesis were higher than those made in the absence of the drug, gC was induced at both permissive and nonpermissive temperatures by the DNA- mutant tsHA1 carrying a lesion in the gene specifying the major DNA-binding protein and which does not express gamma 2 genes at the nonpermissive temperature, and gC was induced only at the permissive temperature in cells infected with ts502 containing a mutation in the alpha 4 gene. The gC induced in L3153(28) cells was made earlier and processed faster to the mature form than that induced in a gC- clone of methotrexate-resistant cells infected with wild-type virus. Unlike virus stocks made in gC- cells, HSV-1(MP) made in L3153(28) cells was susceptible to neutralization by anti-gC monoclonal antibody. Images PMID:3009854

Genetic polymorphism in ATG16L1 gene influences the response to adalimumab in Crohn's disease patients.

PubMed

Koder, Silvo; Repnik, Katja; Ferkolj, Ivan; Pernat, Cvetka; Skok, Pavel; Weersma, Rinse K; Potočnik, Uroš

2015-01-01

To see if SNPs could help predict response to biological therapy using adalimumab (ADA) in Crohn's disease (CD). IBDQ index and CRP levels were used to monitor therapy response. We genotyped 31 CD-associated genes in 102 Slovenian CD patients. The strongest association for treatment response defined as decrease in CRP levels was found for ATG16L1 SNP rs10210302. Additional SNPs in 7 out of 31 tested CD-associated genes (PTGER4, CASP9, IL27, C11orf30, CCNY, IL13, NR1I2) showed suggestive association with ADA response. Our results suggest ADA response in CD patients is genetically predisposed by SNPs in CD risk genes and suggest ATG16L1 as most promising candidate gene for drug response in ADA treatment. Original submitted 24 September 2014; Revision submitted 1 December 2014.

Virus-Plus-Susceptibility Gene Interaction Determines Crohn’s Disease Gene Atg16L1 Phenotypes in Intestine

PubMed Central

Cadwell, Ken; Patel, Khushbu K.; Maloney, Nicole S.; Liu, Ta-Chiang; Ng, Aylwin C.Y.; Storer, Chad E.; Head, Richard D.; Xavier, Ramnik; Stappenbeck, Thaddeus S.; Virgin, Herbert W.

2010-01-01

SUMMARY It is unclear why disease occurs in only a small proportion of persons carrying common risk alleles of disease susceptibility genes. Here we demonstrate that an interaction between a specific virus infection and a mutation in the Crohn’s disease susceptibility gene Atg16L1 induces intestinal pathologies in mice. This virus-plus-susceptibility gene interaction generated abnormalities in granule packaging and unique patterns of gene expression in Paneth cells. Further, the response to injury induced by the toxic substance dextran sodium sulfate was fundamentally altered to include pathologies resembling aspects of Crohn’s disease. These pathologies triggered by virus-plus-susceptibility gene interaction were dependent on TNFα and IFNγ and were prevented by treatment with broad spectrum antibiotics. Thus, we provide a specific example of how a virus-plus-susceptibility gene interaction can, in combination with additional environmental factors and commensal bacteria, determine the phenotype of hosts carrying common risk alleles for inflammatory disease. PMID:20602997

A novel zinc finger protein 219-like (ZNF219L) is involved in the regulation of collagen type 2 alpha 1a (col2a1a) gene expression in zebrafish notochord.

PubMed

Lien, Huang-Wei; Yang, Chung-Hsiang; Cheng, Chia-Hsiung; Hung, Chin-Chun; Liao, Wei-Hao; Hwang, Pung-Pung; Han, Yu-San; Huang, Chang-Jen

2013-01-01

The notochord is required for body plan patterning in vertebrates, and defects in notochord development during embryogenesis can lead to diseases affecting the adult. It is therefore important to elucidate the gene regulatory mechanism underlying notochord formation. In this study, we cloned the zebrafish zinc finger 219-like (ZNF219L) based on mammalian ZNF219, which contains nine C2H2-type zinc finger domains. Through whole-mount in situ hybridization, we found that znf219L mRNA is mainly expressed in the zebrafish midbrain-hindbrain boundary, hindbrain, and notochord during development. The znf219L morpholino knockdown caused partial abnormal notochord phenotype and reduced expression of endogenous col2a1a in the notochord specifically. In addition, ZNF219L could recognize binding sites with GGGGG motifs and trigger augmented activity of the col2a1a promoter in a luciferase assay. Furthermore, in vitro binding experiments revealed that ZNF219L recognizes the GGGGG motifs in the promoter region of the zebrafish col2a1a gene through its sixth and ninth zinc finger domains. Taken together, our results reveal that ZNF219L is involved in regulating the expression of col2a1a in zebrafish notochord specifically.

A Novel Zinc Finger Protein 219-like (ZNF219L) is Involved in the Regulation of Collagen Type 2 Alpha 1a (col2a1a) Gene Expression in Zebrafish Notochord

PubMed Central

Lien, Huang-Wei; Yang, Chung-Hsiang; Cheng, Chia-Hsiung; Hung, Chin-Chun; Liao, Wei-Hao; Hwang, Pung-Pung; Han, Yu-San; Huang, Chang-Jen

2013-01-01

The notochord is required for body plan patterning in vertebrates, and defects in notochord development during embryogenesis can lead to diseases affecting the adult. It is therefore important to elucidate the gene regulatory mechanism underlying notochord formation. In this study, we cloned the zebrafish zinc finger 219-like (ZNF219L) based on mammalian ZNF219, which contains nine C2H2-type zinc finger domains. Through whole-mount in situ hybridization, we found that znf219L mRNA is mainly expressed in the zebrafish midbrain-hindbrain boundary, hindbrain, and notochord during development. The znf219L morpholino knockdown caused partial abnormal notochord phenotype and reduced expression of endogenous col2a1a in the notochord specifically. In addition, ZNF219L could recognize binding sites with GGGGG motifs and trigger augmented activity of the col2a1a promoter in a luciferase assay. Furthermore, in vitro binding experiments revealed that ZNF219L recognizes the GGGGG motifs in the promoter region of the zebrafish col2a1a gene through its sixth and ninth zinc finger domains. Taken together, our results reveal that ZNF219L is involved in regulating the expression of col2a1a in zebrafish notochord specifically. PMID:24155663

Biological and Clinical Significance of MAD2L1 and BUB1, Genes Frequently Appearing in Expression Signatures for Breast Cancer Prognosis

PubMed Central

Wang, Zhanwei; Katsaros, Dionyssios; Shen, Yi; Fu, Yuanyuan; Canuto, Emilie Marion; Benedetto, Chiara; Lu, Lingeng; Chu, Wen-Ming; Risch, Harvey A.; Yu, Herbert

2015-01-01

To investigate the biologic relevance and clinical implication of genes involved in multiple gene expression signatures for breast cancer prognosis, we identified 16 published gene expression signatures, and selected two genes, MAD2L1 and BUB1. These genes appeared in 5 signatures and were involved in cell-cycle regulation. We analyzed the expression of these genes in relation to tumor features and disease outcomes. In vitro experiments were also performed in two breast cancer cell lines, MDA-MB-231 and MDA-MB-468, to assess cell proliferation, migration and invasion after knocking down the expression of these genes. High expression of these genes was found to be associated with aggressive tumors and poor disease-free survival of 203 breast cancer patients in our study, and the association with survival was confirmed in an online database consisting of 914 patients. In vitro experiments demonstrated that lowering the expression of these genes by siRNAs reduced tumor cell growth and inhibited cell migration and invasion. Our investigation suggests that MAD2L1 and BUB1 may play important roles in breast cancer progression, and measuring the expression of these genes may assist the prediction of breast cancer prognosis. PMID:26287798

[Relationship between XRCC3 gene polymorphism and susceptibility to lead poisoning in male lead-exposed workers].

PubMed

Liu, Xiang-quan; Zhang, Zhong

2013-06-01

To investigate the relationship between genetic polymorphism of X-ray repair cross-complementing gene 3 (XRCC3) and susceptibility to lead poisoning in male lead-exposed workers. Peripheral venous blood and morning urine samples were collected from 326 male lead-exposed workers in a storage battery factory in Fuzhou. Blood lead, urine lead, blood zinc protoporphyrin (ZPP), blood calcium, and blood iron were measured. The genotype of XRCC3 was determined by polymerase chain reaction-restriction fragment length polymorphism method. The relationship between XRCC3 gene polymorphism and susceptibility to lead poisoning in male lead-exposed workers was analyzed. Genetic polymorphism of XRCC3 was seen in the 326 subjects. The frequency distribution of XRCC3 genotypes, XRCC3-241CC (wild type), XRCC3-241CT (heterozygous mutation), and XRCC3-241TT (homozygous mutation), was in accordance with the Hardy-Weinberg equilibrium (P > 0.05). There were no significant differences in urine lead, blood ZPP, blood calcium, and blood iron between the lead-exposed workers with different XRCC3 genotypes (P > 0.05). The workers with XRCC3-241CT/TT had a significantly higher mean blood lead level than those with XRCC3-241CC (P < 0.05). With a blood lead level of 1.90 µmol/L as the cutoff value, the chi-square test and logistic regression analysis showed that the proportion of workers with XRCC3-241CT/TT was significantly higher than that of workers with XRCC3-241CC in the subjects with high blood leads (P < 0.05) and that the risk of high blood lead was significantly higher in the workers with XRCC3-241CT/TT than in those with XRCC3-241CC (OR = 2.34, 95%CI = 1.61 ∼ 5.13); the multivariate linear regression analysis showed that the workers with XRCC3-241CT/TT had high blood lead levels (β = 0.116, P < 0.05), the workers with smoking habit demonstrated marked lead absorption (β = 0.188, P < 0.05), good individual protection could reduce lead absorption (β = -0.247, P < 0.05), and the

Specific interaction between hnRNP H and HPV16 L1 proteins: Implications for late gene auto-regulation enabling rapid viral capsid protein production

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

Zheng, Zi-Zheng; Sun, Yuan-Yuan; Zhao, Min

2013-01-18

Highlights: ► The RNA-binding hnRNP H regulates late viral gene expression. ► hnRNP H activity was inhibited by a late viral protein. ► Specific interaction between HPV L1 and hnRNP H was demonstrated. ► Co-localization of HPV L1 and hnRNP H inside cells was observed. ► Viral capsid protein production, enabling rapid capsid assembly, was implicated. -- Abstract: Heterogeneous nuclear ribonucleoproteins (hnRNPs), including hnRNP H, are RNA-binding proteins that function as splicing factors and are involved in downstream gene regulation. hnRNP H, which binds to G triplet regions in RNA, has been shown to play an important role in regulatingmore » the staged expression of late proteins in viral systems. Here, we report that the specific association between hnRNP H and a late viral capsid protein, human papillomavirus (HPV) L1 protein, leads to the suppressed function of hnRNP H in the presence of the L1 protein. The direct interaction between the L1 protein and hnRNP H was demonstrated by complex formation in solution and intracellularly using a variety of biochemical and immunochemical methods, including peptide mapping, specific co-immunoprecipitation and confocal fluorescence microscopy. These results support a working hypothesis that a late viral protein HPV16 L1, which is down regulated by hnRNP H early in the viral life cycle may provide an auto-regulatory positive feedback loop that allows the rapid production of HPV capsid proteins through suppression of the function of hnRNP H at the late stage of the viral life cycle. In this positive feedback loop, the late viral gene products that were down regulated earlier themselves disable their suppressors, and this feedback mechanism could facilitate the rapid production of capsid proteins, allowing staged and efficient viral capsid assembly.« less

Heading date QTL in winter wheat (Triticum aestivum L.) coincide with major developmental genes Vernalization-1 and Photoperiod-1

USDA-ARS?s Scientific Manuscript database

In wheat (Triticum aestivum L.), time from planting to spike emergence is influenced by genes controlling vernalization requirement and photoperiod response. Characterizing the available genetic diversity of known and novel alleles of Vernalization-1 (Vrn-1) and Photoperiod-1 (Ppd-1) in winter wheat...

Standardization of PCR-RFLP analysis of nsSNP rs1468384 of NPC1L1 gene

PubMed Central

Balgir, Praveen P.; Khanna, Divya; Kaur, Gurlovleen

2008-01-01

Niemann-Pick C1-like 1 (NPC1L1) protein, a newly identified sterol influx transporter, located at the apical membrane of the enterocyte, which may actively facilitate the uptake of cholesterol by promoting the passage of sterols across the brush border membrane of the enterocyte. It effects intestinal cholesterol absorption and intracellular transport and as such is an integral part of complex process of cholesterol homeostasis. The study of population data for the distribution of these single nucleotide polymorphisms (SNP) of NPC1L1 has lead to the identification of six non-synonymous single nucleotide polymorphisms (nsSNP). The in vitro analysis using the software MuPro and StructureSNP shows that nsSNP M510I (rs1468384), which involves A→G base pair change leads to decrease in the stability of the protein. A reproducible and a cost-effective PCR-RFLP based assay was developed to screen for the SNP among population data. This SNP has been studied in Caucasian, Asian, and African American populations. Till date, no data is available on Indian population. The distribution of M510I NPC1L1 genotype was estimated in the North Western Indian Population as a test case. The allele distribution in Indian Population differs significantly from that of other populations. The methodology thus proved to be robust enough to bring out these differences. PMID:20300301

Iron metabolism mutant hbd mice have a deletion in Sec15l1, which has homology to a yeast gene for vesicle docking.

PubMed

White, Robert A; Boydston, Leigh A; Brookshier, Terri R; McNulty, Steven G; Nsumu, Ndona N; Brewer, Brandon P; Blackmore, Krista

2005-12-01

Defects in iron absorption and utilization lead to iron deficiency and anemia. While iron transport by transferrin receptor-mediated endocytosis is well understood, it is not completely clear how iron is transported from the endosome to the mitochondria where heme is synthesized. We undertook a positional cloning project to identify the causative mutation for the hemoglobin-deficit (hbd) mouse mutant, which suffers from a microcytic, hypochromic anemia apparently due to defective iron transport in the endocytosis cycle. As shown by previous studies, reticulocyte iron accumulation in homozygous hbd/hbd mice is deficient despite normal binding of transferrin to its receptor and normal transferrin uptake in the cell. We have identified a strong candidate gene for hbd, Sec15l1, a homologue to yeast SEC15, which encodes a key protein in vesicle docking. The hbd mice have an exon deletion in Sec15l1, which is the first known mutation of a SEC gene homologue in mammals.

l-Proline Accumulation and Freeze Tolerance in Saccharomyces cerevisiae Are Caused by a Mutation in the PRO1 Gene Encoding γ-Glutamyl Kinase

PubMed Central

Morita, Yuko; Nakamori, Shigeru; Takagi, Hiroshi

2003-01-01

We previously isolated a mutant which showed a high tolerance to freezing that correlated with higher levels of intracellular l-proline derived from l-proline analogue-resistant mutants. The mutation responsible for the analogue resistance and l-proline accumulation was a single nuclear dominant mutation. By introducing the mutant-derived genomic library into a non-l-proline-utilizing strain, the mutant was found to carry an allele of the wild-type PRO1 gene encoding γ-glutamyl kinase, which resulted in a single amino acid replacement; Asp (GAC) at position 154 was replaced by Asn (AAC). Interestingly, the allele of PRO1 was shown to enhance the activities of γ-glutamyl kinase and γ-glutamyl phosphate reductase, both of which catalyze the first two steps of l-proline synthesis from l-glutamate and which together may form a complex in vivo. When cultured in liquid minimal medium, yeast cells expressing the mutated γ-glutamyl kinase were found to accumulate intracellular l-proline and showed a prominent increase in cell viability after freezing at −20°C compared to the viability of cells harboring the wild-type PRO1 gene. These results suggest that the altered γ-glutamyl kinase results in stabilization of the complex or has an indirect effect on γ-glutamyl phosphate reductase activity, which leads to an increase in l-proline production in Saccharomyces cerevisiae. The approach described in this paper could be a practical method for breeding novel freeze-tolerant yeast strains. PMID:12513997

Analysis of the L1 gene product of human papillomavirus type 16 by expression in a vaccinia virus recombinant.

PubMed

Browne, H M; Churcher, M J; Stanley, M A; Smith, G L; Minson, A C

1988-06-01

The L1 open reading frame of human papillomavirus type 16 (HPV16) has been expressed in vaccinia virus under the control of both the 7.5K early and late promoter, and the 4b major late promoter. Antibodies to a beta-galactosidase fusion protein containing a C-terminal portion of the HPV16 L1 gene product were used to compare the levels of L1 expression in the two recombinants, and showed that greater levels of expression were obtained when the gene was placed under the control of the 4b late promoter. Immunofluorescence studies revealed a nuclear location of the L1 gene product when expressed in vaccinia virus. Antibodies to the beta-galactosidase fusion protein detected a major polypeptide species of 57K and a minor species of 64K in Western blots of recombinant-infected cell lysates. The 64K species was not detected when cells were infected in the presence of tunicamycin, indicating that the primary translation product of the HPV16 L1 open reading frame is modified by N-linked glycosylation when expressed in vaccinia virus. Whereas antibodies to HPV16 L1 fusion proteins and to a peptide containing amino acids from the C terminus of HPV16 L1 reacted well in Western blots with the HPV16 L1 target expressed in vaccinia virus, no reactivity was observed with antibodies to bovine papillomavirus type 1 particles or to a HPV6b fusion protein.

Ectopic expression of class 1 KNOX genes induce and adventitious shoot regeneration and alter growth and development of tobacco (Nicotiana tabacum L) and European plum (Prunus domestica L)

USDA-ARS?s Scientific Manuscript database

Transgenic plants of tobacco (Nicotiana tabacum L) and plum (Prunus domestica L) were produced by transforming with apple class 1 KNOX genes (MdKN1 and MdKN2) or corn KN1 gene. Transgenic tobacco plants were regenerated in vitro from transformed leaf discs cultured in a tissue medium lacking cytoki...

The midgut cadherin-like gene is not associated with resistance to Bacillus thuringiensis toxin Cry1Ac in Plutella xylostella (L.).

PubMed

Guo, Zhaojiang; Kang, Shi; Zhu, Xun; Wu, Qingjun; Wang, Shaoli; Xie, Wen; Zhang, Youjun

2015-03-01

The Gram-positive bacterium Bacillus thuringiensis (Bt) produces Cry toxins that have been used to control important agricultural pests. Evolution of resistance in target pests threatens the effectiveness of these toxins when used either in sprayed biopesticides or in Bt transgenic crops. Although alterations of the midgut cadherin-like receptor can lead to Bt Cry toxin resistance in many insects, whether the cadherin gene is involved in Cry1Ac resistance of Plutella xylostella (L.) remains unclear. Here, we present experimental evidence that resistance to Cry1Ac or Bt var. kurstaki (Btk) in P. xylostella is not due to alterations of the cadherin gene. The bona fide P. xylostella cadherin cDNA sequence was cloned and analyzed, and comparisons of the cadherin cDNA sequence among susceptible and resistant P. xylostella strains confirmed that Cry1Ac resistance was independent of mutations in this gene. In addition, real-time quantitative PCR (qPCR) indicated that cadherin transcript levels did not significantly differ among susceptible and resistant P. xylostella strains. RNA interference (RNAi)-mediated suppression of cadherin gene expression did not affect larval susceptibility to Cry1Ac toxin. Furthermore, genetic linkage assays using four cadherin gDNA allelic biomarkers confirmed that the cadherin gene is not linked to resistance against Cry1Ac in P. xylostella. Taken together, our findings demonstrate that Cry1Ac resistance of P. xylostella is independent of the cadherin gene. Copyright © 2015 Elsevier Inc. All rights reserved.

Gene within gene configuration and expression of the Drosophila melanogaster genes lethal(2) neighbour of tid [l(2)not] and lethal(2) relative of tid[l(2)rot].

PubMed

Kurzik-Dumke, U; Kaymer, M; Gundacker, D; Debes, A; Labitzke, K

1997-10-24

In this paper, we describe the structure and temporal expression pattern of the Drosophila melanogaster genes l(2)not and l(2)rot located at locus 59F5 vis à vis the tumor suppressor gene l(2)tid described previously and exhibiting a gene within gene configuration. The l(2)not protein coding region, 1530 nt, is divided into two exons by an intron, 2645 nt, harboring the genes l(2)rot, co-transcribed from the same DNA strand, and l(2)tid, co-transcribed from the opposite DNA strand, located vis à vis. To determine proteins encoded by the genes described in this study polyclonal rabbit antibodies (Ab), anti-Not and anti-Rot, were generated. Immunostaining of developmental Western blots with the anti-Not Ab resulted in the identification of a 45-kDa protein, Not45, which is smaller than the Not56 protein predicted from the sequence. Its localization in endoplasmic reticulum (ER) was established by immunoelectron microscopy of Drosophila melanogaster Schneider 2 cells. Not45 shows significant homology to yeast ALG3 protein acting as a dolichol mannosyltransferase in the asparagine-linked glycosylation. It is synthesized ubiquitously throughout embryonic life. The protein predicted from the l(2)rot sequence, Rot57, shows a homology to the NS2B protein of the yellow fever virus1 (yefv1). The results of l(2)rot RNA analysis by developmental Northern blot and by in situ RNA localization, as well as the results of the protein analysis via Western blot and immunohistochemistry suggest that l(2)rot is transcribed but not translated. Since RNAs encoded by the genes l(2)tid and l(2)rot are complementary and l(2)rot is presumably not translated we performed preliminary experiments on the function of the l(2)rot RNA as a natural antisense RNA (asRNA) regulator of l(2)tid expression, expressed in the same temporal and spatial manner as the l(2)tid- and l(2)not RNA. l(2)tid knock-out by antisense RNA yielded late embryonic lethality resulting from multiple morphogenetic defects.

Environmental Lead Exposure, Catalase Gene, and Markers of Antioxidant and Oxidative Stress Relation to Hypertension: An Analysis Based on the EGAT Study

PubMed Central

Kaojarern, Sukhumpun; Chanprasertyothin, Suwannee; Panpunuan, Pachara; Petchpoung, Krittaya; Tatsaneeyapant, Aninthita; Yoovathaworn, Krongtong; Sura, Thunyachai; Kaojarern, Sming; Sritara, Piyamit

2015-01-01

Lead has been linked to the development of hypertension via oxidative stress. Catalase plays an important role in the disposal of hydrogen peroxide in erythrocyte and its activity was determined by CAT gene. The aims of this study were to investigate (1) the association between blood levels of antioxidant markers such as catalase, superoxide dismutase, glutathione, glutathione peroxidase, oxidative stress-marker (malondialdehyde), and blood lead level and (2) the influence of genetic polymorphism of CAT gene (rs769217) on change in blood pressure in general population of EGAT study project. This is a cross-sectional study of 332 normotensive, 432 prehypertensive, and 222 hypertensive male subjects. Hypertensive subjects had significantly higher blood lead level (5.28 μg/dL) compared to normotensive (4.41 μg/dL) and prehypertensive (4.55 μg/dL) subjects (P < 0.05). These significant findings are also found in MDA levels. Moreover, individuals with TT genotype in hypertensive group had significantly higher blood lead and MDA levels (6.06 μg/dL and 9.67 μmol/L) than those with CC genotype (5.32 μg/dL and 8.31 μmol/L, P < 0.05). Our findings suggested that decreased blood catalase activity in this polymorphism together with low level lead exposure induced lipid peroxidation may be responsible for hypertension. PMID:25793211

Enhancement of ginsenoside Rg(1) in Panax ginseng hairy root by overexpressing the α-L-rhamnosidase gene from Bifidobacterium breve.

PubMed

Zhang, Ru; Zhang, Bian-Ling; Li, Gu-Cai; Xie, Tao; Hu, Teng; Luo, Zhi-Yong

2015-10-01

To improve the production of ginsenoside Rg1 in Panax ginseng. The α-L-rhamnosidase gene from Bifidobacterium breve (BbRha) was overexpressed into hairy root culture system using Agrobacterium rhizogenes A4. Ginsenoside Rg1 in hairy roots was obtained following transformation via overexpressed gene representing 2.2-fold higher than those of control lines. Several overexpression transgenic hairy root lines were obtained exhibiting markedly increased levels of the corresponding α-L-rhamnosidase enzymatic activity relative to control. Ginsenoside Rg1 levels in the transgenic lines were higher (2.2-fold) than those of control after following 30 days culturing, while ginsenoside Re contents in tested transgenic lines were found to be lower. The transgenic hairy roots harboring α-L-rhamnosidase gene improved the accumulation of ginsenoside Rg1 up to 3.6 mg g(-1) dry weight. BbRha gene selectively enhances the production of ginsenoside Rg1 in P. ginseng hairy roots.

Cloning and Expression Analysis of Genes Encoding Lytic Endopeptidases L1 and L5 from Lysobacter sp. Strain XL1

PubMed Central

Lapteva, Y. S.; Zolova, O. E.; Shlyapnikov, M. G.; Tsfasman, I. M.; Muranova, T. A.; Stepnaya, O. A.; Kulaev, I. S.

2012-01-01

Lytic enzymes are the group of hydrolases that break down structural polymers of the cell walls of various microorganisms. In this work, we determined the nucleotide sequences of the Lysobacter sp. strain XL1 alpA and alpB genes, which code for, respectively, secreted lytic endopeptidases L1 (AlpA) and L5 (AlpB). In silico analysis of their amino acid sequences showed these endopeptidases to be homologous proteins synthesized as precursors similar in structural organization: the mature enzyme sequence is preceded by an N-terminal signal peptide and a pro region. On the basis of phylogenetic analysis, endopeptidases AlpA and AlpB were assigned to the S1E family [clan PA(S)] of serine peptidases. Expression of the alpA and alpB open reading frames (ORFs) in Escherichia coli confirmed that they code for functionally active lytic enzymes. Each ORF was predicted to have the Shine-Dalgarno sequence located at a canonical distance from the start codon and a potential Rho-independent transcription terminator immediately after the stop codon. The alpA and alpB mRNAs were experimentally found to be monocistronic; transcription start points were determined for both mRNAs. The synthesis of the alpA and alpB mRNAs was shown to occur predominantly in the late logarithmic growth phase. The amount of alpA mRNA in cells of Lysobacter sp. strain XL1 was much higher, which correlates with greater production of endopeptidase L1 than of L5. PMID:22865082

Cloning and expression analysis of genes encoding lytic endopeptidases L1 and L5 from Lysobacter sp. strain XL1.

PubMed

Lapteva, Y S; Zolova, O E; Shlyapnikov, M G; Tsfasman, I M; Muranova, T A; Stepnaya, O A; Kulaev, I S; Granovsky, I E

2012-10-01

Lytic enzymes are the group of hydrolases that break down structural polymers of the cell walls of various microorganisms. In this work, we determined the nucleotide sequences of the Lysobacter sp. strain XL1 alpA and alpB genes, which code for, respectively, secreted lytic endopeptidases L1 (AlpA) and L5 (AlpB). In silico analysis of their amino acid sequences showed these endopeptidases to be homologous proteins synthesized as precursors similar in structural organization: the mature enzyme sequence is preceded by an N-terminal signal peptide and a pro region. On the basis of phylogenetic analysis, endopeptidases AlpA and AlpB were assigned to the S1E family [clan PA(S)] of serine peptidases. Expression of the alpA and alpB open reading frames (ORFs) in Escherichia coli confirmed that they code for functionally active lytic enzymes. Each ORF was predicted to have the Shine-Dalgarno sequence located at a canonical distance from the start codon and a potential Rho-independent transcription terminator immediately after the stop codon. The alpA and alpB mRNAs were experimentally found to be monocistronic; transcription start points were determined for both mRNAs. The synthesis of the alpA and alpB mRNAs was shown to occur predominantly in the late logarithmic growth phase. The amount of alpA mRNA in cells of Lysobacter sp. strain XL1 was much higher, which correlates with greater production of endopeptidase L1 than of L5.

Polymorphisms of CCL3L1/CCR5 genes and recurrence of hepatitis B in liver transplant recipients.

PubMed

Li, Hong; Xie, Hai-Yang; Zhou, Lin; Wang, Wei-Lin; Liang, Ting-Bo; Zhang, Min; Zheng, Shu-Sen

2011-12-01

The genetic diversity of chemokines and chemokine receptors has been associated with the outcome of hepatitis B virus infection. The aim of this study was to evaluate whether the copy number variation in the CCL3L1 gene and the polymorphisms of CCR5Δ32 and CCR5-2459A→G (rs1799987) are associated with recurrent hepatitis B in liver transplantation for hepatitis B virus infection-related end-stage liver disease. A total of 185 transplant recipients were enrolled in this study. The genomic DNA was extracted from whole blood, the copy number of the CCL3L1 gene was determined by a quantitative real-time PCR based assay, CCR5Δ32 was detected by a sizing PCR method, and a single-nucleotide polymorphism in CCR5-2459 was detected by restriction fragment length polymorphism PCR. No CCR5Δ32 mutation was detected in any of the individuals from China. Neither copy number variation nor polymorphism in CCR5-2459 was associated with post-transplant re-infection with hepatitis B virus. However, patients with fewer copies (<4) of the CCL3L1 gene compared with the population median in combination with the CCR5G allele had a significantly higher risk for recurrent hepatitis B (odds ratio=1.93, 95% CI: 1.00-3.69; P=0.047). Patients possessing the compound decreased functional genotype of both CCL3L1 and CCR5 genes might be more likely to have recurrence of hepatitis B after transplantation.

The Psen1-L166P-knock-in mutation leads to amyloid deposition in human wild-type amyloid precursor protein YAC transgenic mice

PubMed Central

Vidal, Ruben; Sammeta, Neeraja; Garringer, Holly J.; Sambamurti, Kumar; Miravalle, Leticia; Lamb, Bruce T.; Ghetti, Bernardino

2012-01-01

Genetically engineered mice have been generated to model cerebral β-amyloidosis, one of the hallmarks of Alzheimer disease (AD) pathology, based on the overexpression of a mutated cDNA of the amyloid-β precursor protein (AβPP) or by knock-in of the murine Aβpp gene alone or with presenilin1 mutations. Here we describe the generation and initial characterization of a new mouse line based on the presence of 2 copies of the human genomic region encoding the wild-type AβPP and the L166P presenilin 1 mutation. At ∼6 mo of age, double-mutant mice develop amyloid pathology, with signs of neuritic dystrophy, intracellular Aβ accumulation, and glial inflammation, an increase in AβPP C-terminal fragments, and an 8 times increase in Aβ42 levels with a 40% decrease in Aβ40 levels, leading to a significant increase (14 times) of Aβ42/Aβ40 ratios, with minimal effects on presenilin or the Notch1 pathway in the brain. We conclude that in mice, neither mutations in AβPP nor overexpression of an AβPP isoform are a prerequisite for Aβ pathology. This model will allow the study of AD pathogenesis and testing of therapeutic strategies in a more relevant environment without experimental artifacts due to the overexpression of a single-mutant AβPP isoform using exogenous promoters.—Vidal, R., Sammeta, N., Garringer, H. J., Sambamurti, K., Miravalle, L., Lamb B. T., Ghetti, B. The Psen1-L166P-knock-in mutation leads to amyloid deposition in human wild-type amyloid precursor protein YAC transgenic mice. PMID:22459153

Effects of L-cysteine on lead acetate induced neurotoxicity in albino mice.

PubMed

Mahmoud, Y I; Sayed, S S

2016-07-01

Lead is a toxic heavy metal that adversely affects nervous tissues; it often occurs as an environmental pollutant. We investigated histological changes in the cerebral cortex, hippocampus and cerebellum of adult albino mice following exposure to lead acetate. We also studied the possible ameliorative effect of the chelating agent, L-cysteine, on lead-induced neurotoxicity. We divided albino mice into six groups: 1) vehicle-only control, 2) L-cysteine control, 3 and 4) treated for 7 days with 20 and 40 mg/kg lead acetate, respectively, and 5 and 6) treated for 7 days with 20 and 40 mg/kg lead acetate, respectively, followed by 50 mg/kg L-cysteine for 7 days. Lead acetate administration caused disorganization of cell layers, neuronal loss and degeneration, and neuropil vacuolization. Brain sections from lead-intoxicated mice treated with L-cysteine showed fewer pathological changes; the neuropil showed less vacuolization and the neurons appeared less damaged. L-cysteine at the dose we used only marginally alleviated lead-induced toxicity.

Role of plnB gene in the regulation of bacteriocin production in Lactobacillus paraplantarum L-XM1.

PubMed

Zhang, Xiangmei; Shang, Nan; Zhang, Xu; Gui, Meng; Li, Pinglan

2013-06-12

Homologues of plnB gene have been shown to participate in regulation of bacteriocin production through quorum sensing system in other organisms, to investigate the possible role of plnB gene in Lactobacillus paraplantarum L-XM1, we cloned and insertionally inactivated the plnB gene. The plnB knockout mutant ΔplnB21 showed loss of bacteriocin production, its Bac⁺ phenotype could not be restored even after the addition of PlnA. Furthermore, reverse transcription-PCR analysis from total RNA preparations showed that the bacteriocin structural genes of the plnEF and plnJK were not transcribed in the plnB knockout mutant compared with the wild-type strain. It was therefore concluded that plnB is invovled in a quorum sensing based bacteriocin production. This is the first demonstration of a role for plnB by gene knockout in L. paraplantarum. Copyright © 2012 Elsevier GmbH. All rights reserved.

Differential expression of immune-regulatory genes associated with PD-L1 display in melanoma: implications for PD-1 pathway blockade

PubMed Central

Taube, Janis M.; Young, Geoffrey D.; McMiller, Tracee L.; Chen, Shuming; Salas, January T.; Pritchard, Theresa S.; Xu, Haiying; Meeker, Alan K.; Fan, Jinshui; Cheadle, Chris; Berger, Alan E.; Pardoll, Drew M.; Topalian, Suzanne L.

2015-01-01

Purpose Blocking the immunosuppressive PD-1/PD-L1 pathway has anti-tumor activity in multiple cancer types, and PD-L1 expression on tumor cells and infiltrating myeloid cells correlates with the likelihood of response. We previously found that IFNG (interferon-gamma) was over-expressed by TILs in PD-L1+ vs. PD-L1(−) melanomas, creating adaptive immune resistance by promoting PD-L1 display. The current study was undertaken to identify additional factors in the PD-L1+ melanoma microenvironment coordinately contributing to immunosuppression. Experimental design Archived, formalin-fixed paraffin-embedded melanoma specimens were assessed for PD-L1 protein expression at the tumor cell surface with immunohistochemistry (IHC). Whole genome expression analysis, quantitative (q)RT-PCR, immunohistochemistry, and functional in vitro validation studies were employed to assess factors differentially expressed in PD-L1+ versus PD-L1(−) melanomas. Results Functional annotation clustering based on whole genome expression profiling revealed pathways up-regulated in PD-L1+ melanomas, involving immune cell activation, inflammation, and antigen processing and presentation. Analysis by qRT-PCR demonstrated over-expression of functionally related genes in PD-L1+ melanomas, involved in CD8+ T cell activation (CD8A, IFNG, PRF1, CCL5), antigen presentation (CD163, TLR3, CXCL1, LYZ), and immunosuppression [PDCD1 (PD-1), CD274(PD-L1), LAG3, IL10]. Functional studies demonstrated that some factors, including IL-10 and IL-32-gamma, induced PD-L1 expression on monocytes but not tumor cells. Conclusions These studies elucidate the complexity of immune checkpoint regulation in the tumor microenvironment, identifying multiple factors likely contributing to coordinated immunosuppression. These factors may provide tumor escape mechanisms from anti-PD-1/PD-L1 therapy, and should be considered for co-targeting in combinatorial immunomodulation treatment strategies. PMID:25944800

Multiple Genes Repress Motility in Uropathogenic Escherichia coli Constitutively Expressing Type 1 Fimbriae▿ †

PubMed Central

Simms, Amy N.; Mobley, Harry L. T.

2008-01-01

Two surface organelles of uropathogenic Escherichia coli (UPEC), flagella and type 1 fimbriae, are critical for colonization of the urinary tract but mediate opposite actions. Flagella propel bacteria through urine and along mucus layers, while type 1 fimbriae allow bacteria to adhere to specific receptors present on uroepithelial cells. Constitutive expression of type 1 fimbriae leads to repression of motility and chemotaxis in UPEC strain CFT073, suggesting that UPEC may coordinately regulate motility and adherence. To identify genes involved in this regulation of motility by type 1 fimbriae, transposon mutagenesis was performed on a phase-locked type 1 fimbrial ON variant of strain CFT073 (CFT073 fim L-ON), followed by a screen for restoration of motility in soft agar. Functions of the genes identified included attachment, metabolism, transport, DNA mismatch repair, and transcriptional regulation, and a number of genes had hypothetical function. Isogenic deletion mutants of these genes were also constructed in CFT073 fim L-ON. Motility was partially restored in six of these mutants, including complementable mutations in four genes encoding known transcriptional regulators, lrhA, lrp, slyA, and papX; a mismatch repair gene, mutS; and one hypothetical gene, ydiV. Type 1 fimbrial expression in these mutants was unaltered, and the majority of these mutants expressed larger amounts of flagellin than the fim L-ON parental strain. Our results indicate that repression of motility in CFT073 fim L-ON is not solely due to the constitutive expression of type 1 fimbriae on the surfaces of the bacteria and that multiple genes may contribute to this repression. PMID:18359812

Aberrant PD-L1 expression through 3'-UTR disruption in multiple cancers.

PubMed

Kataoka, Keisuke; Shiraishi, Yuichi; Takeda, Yohei; Sakata, Seiji; Matsumoto, Misako; Nagano, Seiji; Maeda, Takuya; Nagata, Yasunobu; Kitanaka, Akira; Mizuno, Seiya; Tanaka, Hiroko; Chiba, Kenichi; Ito, Satoshi; Watatani, Yosaku; Kakiuchi, Nobuyuki; Suzuki, Hiromichi; Yoshizato, Tetsuichi; Yoshida, Kenichi; Sanada, Masashi; Itonaga, Hidehiro; Imaizumi, Yoshitaka; Totoki, Yasushi; Munakata, Wataru; Nakamura, Hiromi; Hama, Natsuko; Shide, Kotaro; Kubuki, Yoko; Hidaka, Tomonori; Kameda, Takuro; Masuda, Kyoko; Minato, Nagahiro; Kashiwase, Koichi; Izutsu, Koji; Takaori-Kondo, Akifumi; Miyazaki, Yasushi; Takahashi, Satoru; Shibata, Tatsuhiro; Kawamoto, Hiroshi; Akatsuka, Yoshiki; Shimoda, Kazuya; Takeuchi, Kengo; Seya, Tsukasa; Miyano, Satoru; Ogawa, Seishi

2016-06-16

Successful treatment of many patients with advanced cancer using antibodies against programmed cell death 1 (PD-1; also known as PDCD1) and its ligand (PD-L1; also known as CD274) has highlighted the critical importance of PD-1/PD-L1-mediated immune escape in cancer development. However, the genetic basis for the immune escape has not been fully elucidated, with the exception of elevated PD-L1 expression by gene amplification and utilization of an ectopic promoter by translocation, as reported in Hodgkin and other B-cell lymphomas, as well as stomach adenocarcinoma. Here we show a unique genetic mechanism of immune escape caused by structural variations (SVs) commonly disrupting the 3' region of the PD-L1 gene. Widely affecting multiple common human cancer types, including adult T-cell leukaemia/lymphoma (27%), diffuse large B-cell lymphoma (8%), and stomach adenocarcinoma (2%), these SVs invariably lead to a marked elevation of aberrant PD-L1 transcripts that are stabilized by truncation of the 3'-untranslated region (UTR). Disruption of the Pd-l1 3'-UTR in mice enables immune evasion of EG7-OVA tumour cells with elevated Pd-l1 expression in vivo, which is effectively inhibited by Pd-1/Pd-l1 blockade, supporting the role of relevant SVs in clonal selection through immune evasion. Our findings not only unmask a novel regulatory mechanism of PD-L1 expression, but also suggest that PD-L1 3'-UTR disruption could serve as a genetic marker to identify cancers that actively evade anti-tumour immunity through PD-L1 overexpression.

A radiosensitivity gene signature and PD-L1 status predict clinical outcome of patients with invasive breast carcinoma in The Cancer Genome Atlas (TCGA) dataset.

PubMed

Jang, Bum-Sup; Kim, In Ah

2017-09-01

We investigated the link between the radiosensitivity gene signature and programmed cell death ligand 1 (PD-L1) status and clinical outcome in order to identify a group of patients that would possibly receive clinical benefit of radiotherapy (RT) combined with anti-PD1/PD-L1 therapy. We validated the identified gene signature related to radiosensitivity and analyzed the PD-L1 status of invasive breast cancer in The Cancer Genome Atlas (TCGA) dataset. To validate the gene signature, 1045 patients were selected and divided into two clusters using a consensus clustering algorithm based on their radiosensitive (RS) or radioresistant (RR) designation according to their prognosis. Patients were also stratified as PD-L1-high or PD-L1-low based on the median value of CD274 mRNA expression level as surrogates of PD-L1. Patents assigned to the RS group had decreased risk of recurrence-free survival (RFS) rate than patients in the RR group by univariate analysis (HR 0.45, 95% CI 0.25-0.81, p=0.008) only when treated with RT. The RS group was independently associated with the PD-L1-high group, and CD274 mRNA expression was significantly higher in the RS group (p<0.001) than the RR group. In the PD-L1-high group, the RS group was associated with better RFS compared to the RR group (HR 0.37, 95% CI 0.16-0.87, p=0.022) in multivariate analysis. The level of PD-L1 expression may represent the immunogenicity of tumors, and thus, we speculated that the PD-L1-high group had more immunogenic tumors, which could be more sensitive to radiation-induced immunologic cell death. We first evaluated the predictive value of the radiosensitivity gene signature and described a relationship with this radiosensitivity gene signature and PD-L1. The radiosensitivity gene signature and PD-L1 status were important factors for prediction of the clinical outcome of RT in patients with invasive breast cancer and may be used for selecting patients who will benefit from RT combined with anti-PD1/PDL1

Deficiency of Suppressor Enhancer Lin12 1 Like (SEL1L) in Mice Leads to Systemic Endoplasmic Reticulum Stress and Embryonic Lethality*

PubMed Central

Francisco, Adam B.; Singh, Rajni; Li, Shuai; Vani, Anish K.; Yang, Liu; Munroe, Robert J.; Diaferia, Giuseppe; Cardano, Marina; Biunno, Ida; Qi, Ling; Schimenti, John C.; Long, Qiaoming

2010-01-01

Stress in the endoplasmic reticulum (ER) plays an important causal role in the pathogenesis of several chronic diseases such as Alzheimer, Parkinson, and diabetes mellitus. Insight into the genetic determinants responsible for ER homeostasis will greatly facilitate the development of therapeutic strategies for the treatment of these debilitating diseases. Suppressor enhancer Lin12 1 like (SEL1L) is an ER membrane protein and was thought to be involved in the quality control of secreted proteins. Here we show that the mice homozygous mutant for SEL1L were embryonic lethal. Electron microscopy studies revealed a severely dilated ER in the fetal liver of mutant embryos, indicative of alteration in ER homeostasis. Consistent with this, several ER stress responsive genes were significantly up-regulated in the mutant embryos. Mouse embryonic fibroblast cells deficient in SEL1L exhibited activated unfolded protein response at the basal state, impaired ER-associated protein degradation, and reduced protein secretion. Furthermore, markedly increased apoptosis was observed in the forebrain and dorsal root ganglions of mutant embryos. Taken together, our results demonstrate an essential role for SEL1L in protein quality control during mouse embryonic development. PMID:20197277

Deficiency of suppressor enhancer Lin12 1 like (SEL1L) in mice leads to systemic endoplasmic reticulum stress and embryonic lethality.

PubMed

Francisco, Adam B; Singh, Rajni; Li, Shuai; Vani, Anish K; Yang, Liu; Munroe, Robert J; Diaferia, Giuseppe; Cardano, Marina; Biunno, Ida; Qi, Ling; Schimenti, John C; Long, Qiaoming

2010-04-30

Stress in the endoplasmic reticulum (ER) plays an important causal role in the pathogenesis of several chronic diseases such as Alzheimer, Parkinson, and diabetes mellitus. Insight into the genetic determinants responsible for ER homeostasis will greatly facilitate the development of therapeutic strategies for the treatment of these debilitating diseases. Suppressor enhancer Lin12 1 like (SEL1L) is an ER membrane protein and was thought to be involved in the quality control of secreted proteins. Here we show that the mice homozygous mutant for SEL1L were embryonic lethal. Electron microscopy studies revealed a severely dilated ER in the fetal liver of mutant embryos, indicative of alteration in ER homeostasis. Consistent with this, several ER stress responsive genes were significantly up-regulated in the mutant embryos. Mouse embryonic fibroblast cells deficient in SEL1L exhibited activated unfolded protein response at the basal state, impaired ER-associated protein degradation, and reduced protein secretion. Furthermore, markedly increased apoptosis was observed in the forebrain and dorsal root ganglions of mutant embryos. Taken together, our results demonstrate an essential role for SEL1L in protein quality control during mouse embryonic development.

Genetics Home Reference: L1 syndrome

MedlinePlus

... X-linked hydrocephalus: evidence for closely related clinical entities of unknown molecular bases. Acta Neuropathol. 2013 Sep; ... F. Three cases with L1 syndrome and two novel mutations in the L1CAM gene. Eur J Pediatr. ...

RNA interference of 1-aminocyclopropane-1-carboxylic acid oxidase (ACO1 and ACO2) genes expression prolongs the shelf life of Eksotika (Carica papaya L.) papaya fruit.

PubMed

Sekeli, Rogayah; Abdullah, Janna Ong; Namasivayam, Parameswari; Muda, Pauziah; Abu Bakar, Umi Kalsom; Yeong, Wee Chien; Pillai, Vilasini

2014-06-19

The purpose of this study was to evaluate the effectiveness of using RNA interference in down regulating the expression of 1-aminocyclopropane-1-carboxylic acid oxidase gene in Eksotika papaya. One-month old embryogenic calli were separately transformed with Agrobacterium strain LBA 4404 harbouring the three different RNAi pOpOff2 constructs bearing the 1-aminocyclopropane-1-carboxylic acid oxidase gene. A total of 176 putative transformed lines were produced from 15,000 calli transformed, selected, then regenerated on medium supplemented with kanamycin. Integration and expression of the targeted gene in putatively transformed lines were verified by PCR and real-time RT-PCR. Confined field evaluation of a total of 31 putative transgenic lines planted showed a knockdown expression of the targeted ACO1 and ACO2 genes in 13 lines, which required more than 8 days to achieve the full yellow colour (Index 6). Fruits harvested from lines pRNAiACO2 L2-9 and pRNAiACO1 L2 exhibited about 20 and 14 days extended post-harvest shelf life to reach Index 6, respectively. The total soluble solids contents of the fruits ranged from 11 to 14° Brix, a range similar to fruits from non-transformed, wild type seed-derived plants.

[Blue-light induced expression of S-adenosy-L-homocysteine hydrolase-like gene in Mucor amphibiorum RCS1].

PubMed

Gao, Ya; Wang, Shu; Fu, Mingjia; Zhong, Guolin

2013-09-04

To determine blue-light induced expression of S-adenosyl-L-homocysteine hydrolase-like (sahhl) gene in fungus Mucor amphibiorum RCS1. In the random process of PCR, a sequence of 555 bp was obtained from M. amphibiorum RCS1. The 555 bp sequence was labeled with digoxin to prepare the probe for northern hybridization. By northern hybridization, the transcription of sahhl gene was analyzed in M. amphibiorum RCS1 mycelia culture process from darkness to blue light to darkness. Simultaneously real-time PCR method was used to the sahhl gene expression analysis. Compared with the sequence of sahh gene from Homo sapiens, Mus musculus and some fungi species, a high homology of the 555 bp sequence was confirmed. Therefore, the preliminary confirmation has supported that the 555 bp sequence should be sahhl gene from M. amphibiorum RCS1. Under the dark pre-culture in 24 h, a large amounts of transcript of sahhl gene in the mycelia can be detected by northern hybridization and real-time PCR in the condition of 24 h blue light. But a large amounts of transcript of sahhl gene were not found in other detection for the dark pre-culture of 48 h, even though M. amphibiorum RCS1 mycelia were induced by blue light. Blue light can induce the expression of sahhl gene in the vigorous growth of M. amphibiorum RCS1 mycelia.

Ectopic expression of class 1 KNOX genes induce adventitious shoot regeneration and alter growth and development of tobacco (Nicotiana tabacum L) and European plum (Prunus domestica L).

PubMed

Srinivasan, C; Liu, Zongrang; Scorza, Ralph

2011-04-01

Transgenic plants of tobacco (Nicotiana tabacum L) and European plum (Prunus domestica L) were produced by transforming with the apple class 1 KNOX genes (MdKN1 and MdKN2) or corn KNOX1 gene. Transgenic tobacco plants were regenerated in vitro from transformed leaf discs cultured in a medium lacking cytokinin. Ectopic expression of KNOX genes retarded shoot growth by suppressing elongation of internodes in transgenic tobacco plants. Expression of each of the three KNOX1 genes induced malformation and extensive lobbing in tobacco leaves. In situ regeneration of adventitious shoots was observed from leaves and roots of transgenic tobacco plants expressing each of the three KNOX genes. In vitro culture of leaf explants and internode sections excised from in vitro grown MdKN1 expressing tobacco shoots regenerated adventitious shoots on MS (Murashige and Skoog 1962) basal medium in the absence of exogenous cytokinin. Transgenic plum plants that expressed the MdKN2 or corn KNOX1 gene grew normally but MdKN1 caused a significant reduction in plant height, leaf shape and size and produced malformed curly leaves. A high frequency of adventitious shoot regeneration (96%) was observed in cultures of leaf explants excised from corn KNOX1-expressing transgenic plum shoots. In contrast to KNOX1-expressing tobacco, leaf and internode explants of corn KNOX1-expressing plum required synthetic cytokinin (thidiazuron) in the culture medium to induce adventitious shoot regeneration. The induction of high-frequency regeneration of adventitious shoots in vitro from leaves and stem internodal sections of plum through the ectopic expression of a KNOX1 gene is the first such report for a woody perennial fruit trees.

PPARγ partial agonist GQ-16 strongly represses a subset of genes in 3T3-L1 adipocytes

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

Milton, Flora Aparecida; Genomic Medicine, Houston Methodist Research Institute, Houston, TX; Cvoro, Aleksandra

Thiazolidinediones (TZDs) are peroxisome proliferator-activated receptor gamma (PPARγ) agonists that improve insulin resistance but trigger side effects such as weight gain, edema, congestive heart failure and bone loss. GQ-16 is a PPARγ partial agonist that improves glucose tolerance and insulin sensitivity in mouse models of obesity and diabetes without inducing weight gain or edema. It is not clear whether GQ-16 acts as a partial agonist at all PPARγ target genes, or whether it displays gene-selective actions. To determine how GQ-16 influences PPARγ activity on a gene by gene basis, we compared effects of rosiglitazone (Rosi) and GQ-16 in mature 3T3-L1more » adipocytes using microarray and qRT-PCR. Rosi changed expression of 1156 genes in 3T3-L1, but GQ-16 only changed 89 genes. GQ-16 generally showed weak effects upon Rosi induced genes, consistent with partial agonist actions, but a subset of modestly Rosi induced and strongly repressed genes displayed disproportionately strong GQ-16 responses. PPARγ partial agonists MLR24 and SR1664 also exhibit disproportionately strong effects on transcriptional repression. We conclude that GQ-16 displays a continuum of weak partial agonist effects but efficiently represses some negatively regulated PPARγ responsive genes. Strong repressive effects could contribute to physiologic actions of GQ-16. - Highlights: • GQ-16 is an insulin sensitizing PPARγ ligand with reduced harmful side effects. • GQ-16 displays a continuum of weak partial agonist activities at PPARγ-induced genes. • GQ-16 exerts strong repressive effects at a subset of genes. • These inhibitor actions should be evaluated in models of adipose tissue inflammation.« less

Cloning, expression, and characterization of bacterial L-arabinose 1-dehydrogenase involved in an alternative pathway of L-arabinose metabolism.

PubMed

Watanabe, Seiya; Kodaki, Tsutomu; Kodak, Tsutomu; Makino, Keisuke

2006-02-03

Azospirillum brasiliense converts L-arabinose to alpha-ketoglutarate via five hypothetical enzymatic steps. We purified and characterized L-arabinose 1-dehydrogenase (EC 1.1.1.46), catalyzing the conversion of L-arabinose to L-arabino-gamma-lactone as an enzyme responsible for the first step of this alternative pathway of L-arabinose metabolism. The purified enzyme preferred NADP+ to NAD+ as a coenzyme. Kinetic analysis revealed that the enzyme had high catalytic efficiency for both L-arabinose and D-galactose. The gene encoding L-arabinose 1-dehydrogenase was cloned using a partial peptide sequence of the purified enzyme and was overexpressed in Escherichia coli as a fully active enzyme. The enzyme consists of 308 amino acids and has a calculated molecular mass of 33,663.92 Da. The deduced amino acid sequence had some similarity to glucose-fructose oxidoreductase, D-xylose 1-dehydrogenase, and D-galactose 1-dehydrogenase. Site-directed mutagenesis revealed that the enzyme possesses unique catalytic amino acid residues. Northern blot analysis showed that this gene was induced by L-arabinose but not by D-galactose. Furthermore, a disruptant of the L-arabinose 1-dehydrogenase gene did not grow on L-arabinose but grew on D-galactose at the same growth rate as the wild-type strain. There was a partial gene for L-arabinose transport in the flanking region of the L-arabinose 1-dehydrogenase gene. These results indicated that the enzyme is involved in the metabolism of L-arabinose but not D-galactose. This is the first identification of a gene involved in an alternative pathway of L-arabinose metabolism in bacterium.

[Establishment of L-periaxin gene knock-out RSC96 cell line].

PubMed

Liang, Min; Peng, Tingting; Shi, Yawei

2016-12-25

Periaxin, a protein of noncompact myelin, is specifically expressed in the peripheral nervous system (PNS). There are two protein isoform L-periaxin and S-Periaxin by alternative splicing of periaxin gene, playing an important role in the initiation of myelin formation. So far, 18 different mutation sites in L-periaxin gene have been found to induce the peripheral demyelinating neurological charcot-marie-tooth diseases subtype 4F (CMT4F). The technique of activation of transcription activator-like effector nucleases (TALENS) was used to knock out the L-periaxin gene in RSC 96 cell line of Rattus. According to the design principle, the knock-out site of L-periaxin was assured to NLS domain of L-periaxin, which is target sequence of left and right arms of TALEN. The knock-out vectors of TALEN-L and TALEN-R were established and transfected into RSC96 cell. After puromycin screening, L-periaxin was knocked out successfully in RSC96 cell, which is confirmed by DNA sequence. The mutation efficiency is 21.6%. S-periaxin, not L-periaxin can be detected by Western blotting in L-periaxin gene knock-out RSC96 cell. The cell growth rate was decreased and the number of cells in G1 increased and decreased in S phase in L-periaxin gene knock-out RSC96 cell by flow cytometry and MTT assay.

Evaluation of cellular uptake and gene transfer efficiency of pegylated poly-L-lysine compacted DNA: implications for cancer gene therapy.

PubMed

Walsh, M; Tangney, M; O'Neill, M J; Larkin, J O; Soden, D M; McKenna, S L; Darcy, R; O'Sullivan, G C; O'Driscoll, C M

2006-01-01

Recent success in phase I/II clinical trials (Konstan, M. W.; Davis, P. B.; Wagener, J. S.; Hilliard, K. A.; Stern, R. C.; Milgram, L. J.; Kowalczyk, T. H.; Hyatt, S. L.; Fink, T. L.; Gedeon, C. R.; Oette, S. M.; Payne, J. M.; Muhammad, O.; Ziady, A. G.; Moen, R. C.; Cooper, M. J. Hum. Gene Ther. 2004, 15 (12), 1255-69) has highlighted pegylated poly-L-lysine (C1K30-PEG) as a nonviral gene delivery agent capable of achieving clinically significant gene transfer levels in vivo. This study investigates the potential of a C1K30-PEG gene delivery system for cancer gene therapy and evaluates its mode of cellular entry with the purpose of developing an optimally formulated prototype for tumor cell transfection. C1K30-PEG complexes have a neutral charge and form rod-like and toroid-like nanoparticles. Comparison of the transfection efficiency achieved by C1K30-PEG with other cationic lipid and polymeric vectors demonstrates that C1K30-PEG transfects cells more efficiently than unpegylated poly-L-lysine and compares well to commercially available vectors. In vivo gene delivery by C1K30-PEG nanoparticles to a growing subcutaneous murine tumor was also demonstrated. To determine potential barriers to C1K30-PEG gene delivery, the entry mechanism and intracellular fate of rhodamine labeled complexes were investigated. Using cellular markers to delineate the pathway taken by the complexes upon cellular entry, only minor colocalization was observed with EEA-1, a marker of early endosomes. No colocalization was observed between the complexes and the transferrin receptor, which is a marker for clathrin-coated pits. In addition, complexes were not observed to enter late endosomes/lysosomes. Cellular entry of the complexes was completely inhibited by the macropinocytosis inhibitor, amiloride, indicating that the complexes enter cells via macropinosomes. Such mechanistic studies are an essential step to support future rational design of pegylated poly-L-lysine vectors to improve the

Differential expression of a novel gene during seed triacylglycerol accumulation in lupin species ( Lupinus angustifolius L. and L. mutabilis L.).

PubMed

Francki, Michael G; Whitaker, Peta; Smith, Penelope M; Atkins, Craig A

2002-11-01

Seed triacylglycerols (TAGs) are stored as energy reserves and extracted for various end-product uses. In lupins, seed oil content varies from 16% in Lupinus mutabilisto 8% in L. angustifolius. We have shown that TAGs rapidly accumulate during mid-stages of seed development in L. mutabilis compared to the lower seed oil species, L. angustifolius. In this study, we have targeted the key enzymes of the lipid biosynthetic pathway, acetyl-CoA carboxylase (ACCase) and diacylglycerol acyltransferase (DAGAT), to determine factors regulating TAG accumulation between two lupin species. A twofold increase in ACCase activity was observed in L. mutabilis relative to L. angustifolius and correlated with rapid TAG accumulation. No difference in DAGAT activity was detected. We have identified, cloned and partially characterised a novel gene differentially expressed during TAG accumulation between L. angustifolius and L. mutabilis. The gene has some identity to the glucose dehydrogenase family previously described in barley and bacteria and the significance of its expression levels during seed development in relation to TAG accumulation is discussed. DNA sequence analysis of the promoter in both L. angustifolius and L. mutabilis identified putative matrix attachment regions and recognition sequences for transcription binding sites similar to those found in the Adh1 gene from Arabidopsis. The identical promoter regions between species indicate that differential gene expression is controlled by alternative transcription factors, accessibility to binding sites or a combination of both.

Gene Expression ‏‏‏‏Profiles of BAD and Bcl-xL in the CA1 Region of the Hippocampus Following Global Ischemic/Reperfusion and FK-506 Administration.

PubMed

Badr, Ramak; Hashemi, Mehrdad; Javadi, Gholamreza; Movafagh, Abolfazl; Mahdian, Reza

2015-12-01

The hippocampus is a tiny nub in the mammalian brain that is involved in forming, organizing, and storing memories. Global cerebral ischemia (GCI) and reperfusion induced apoptosis lead to cell injury and death. FK-506 is a strong immunosuppressant drug that has neuroprotective effects on the hypoxic-ischemic effects of brain damage. BAD and Bcl-xL are pro-apoptotic and anti-apoptotic genes, respectively. These genes belong to The B-cell lymphoma-2 (Bcl-2) family. In this study, we assessed the neurotrophic properties of FK-506 on expression of the BAD and Bcl-xL genes in the hippocampus following global ischemia and reperfusion. In the present experimental study, adult male Wistar rats were obtained and housed under standard conditions in the Tehran University of Medical Science in Iran. Rats were equally distributed in groups of three among the following groups: normal control, treated-1 (ischemia/reperfusion), and treated-2 (ischemia/reperfusion followed by FK-506). Global ischemia was induced for animals in the treated-1 and treated-2 groups. In treated-2, two doses of FK-506 were injected: one dose as an IV injection immediately after reperfusion and another as an intra-peritoneal (IP) injection after 48 hours. Then, the hippocampus tissue was removed after anaesthetizing the rats. RNA was isolated, cDNA was synthesized, and real-time PCR was performed. Finally, the obtained data were analyzed statistically (P value ˂ 0.05). The quantitative results of real-time PCR show that the mRNA expression ratio of Bcl-xL down-regulated was 0.75 ± 0.06 in the ischemia/reperfusion group versus 1.57 ± 0.09 in the control group (P value < 0.001), whereas Bcl-xL gene expression was greater in the ischemia/reperfusion +FK506 group (1.93 ± 0.15) than in the ischemia/reperfusion group. Moreover, the mRNA expression ratio of BAD up-regulated in the ischemia/reperfusion + FK506 group was 3.65 ± 0.49 compared to Normal control (1.39 ± 0.09) and Ischemia/reperfusion + FK506

Bi-allelic Mutations in PKD1L1 Are Associated with Laterality Defects in Humans.

PubMed

Vetrini, Francesco; D'Alessandro, Lisa C A; Akdemir, Zeynep C; Braxton, Alicia; Azamian, Mahshid S; Eldomery, Mohammad K; Miller, Kathryn; Kois, Chelsea; Sack, Virginia; Shur, Natasha; Rijhsinghani, Asha; Chandarana, Jignesh; Ding, Yan; Holtzman, Judy; Jhangiani, Shalini N; Muzny, Donna M; Gibbs, Richard A; Eng, Christine M; Hanchard, Neil A; Harel, Tamar; Rosenfeld, Jill A; Belmont, John W; Lupski, James R; Yang, Yaping

2016-10-06

Disruption of the establishment of left-right (L-R) asymmetry leads to situs anomalies ranging from situs inversus totalis (SIT) to situs ambiguus (heterotaxy). The genetic causes of laterality defects in humans are highly heterogeneous. Via whole-exome sequencing (WES), we identified homozygous mutations in PKD1L1 from three affected individuals in two unrelated families. PKD1L1 encodes a polycystin-1-like protein and its loss of function is known to cause laterality defects in mouse and medaka fish models. Family 1 had one fetus and one deceased child with heterotaxy and complex congenital heart malformations. WES identified a homozygous splicing mutation, c.6473+2_6473+3delTG, which disrupts the invariant splice donor site in intron 42, in both affected individuals. In the second family, a homozygous c.5072G>C (p.Cys1691Ser) missense mutation was detected in an individual with SIT and congenital heart disease. The p.Cys1691Ser substitution affects a highly conserved cysteine residue and is predicted by molecular modeling to disrupt a disulfide bridge essential for the proper folding of the G protein-coupled receptor proteolytic site (GPS) motif. Damaging effects associated with substitutions of this conserved cysteine residue in the GPS motif have also been reported in other genes, namely GPR56, BAI3, and PKD1 in human and lat-1 in C. elegans, further supporting the likely pathogenicity of p.Cys1691Ser in PKD1L1. The identification of bi-allelic PKD1L1 mutations recapitulates previous findings regarding phenotypic consequences of loss of function of the orthologous genes in mice and medaka fish and further expands our understanding of genetic contributions to laterality defects in humans. Copyright © 2016 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

FUN-L: gene prioritization for RNAi screens.

PubMed

Lees, Jonathan G; Hériché, Jean-Karim; Morilla, Ian; Fernández, José M; Adler, Priit; Krallinger, Martin; Vilo, Jaak; Valencia, Alfonso; Ellenberg, Jan; Ranea, Juan A; Orengo, Christine

2015-06-15

Most biological processes remain only partially characterized with many components still to be identified. Given that a whole genome can usually not be tested in a functional assay, identifying the genes most likely to be of interest is of critical importance to avoid wasting resources. Given a set of known functionally related genes and using a state-of-the-art approach to data integration and mining, our Functional Lists (FUN-L) method provides a ranked list of candidate genes for testing. Validation of predictions from FUN-L with independent RNAi screens confirms that FUN-L-produced lists are enriched in genes with the expected phenotypes. In this article, we describe a website front end to FUN-L. The website is freely available to use at http://funl.org © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Modifications to a LATE MERISTEM IDENTITY1 gene are responsible for the major leaf shapes of Upland cotton (Gossypium hirsutum L.).

PubMed

Andres, Ryan J; Coneva, Viktoriya; Frank, Margaret H; Tuttle, John R; Samayoa, Luis Fernando; Han, Sang-Won; Kaur, Baljinder; Zhu, Linglong; Fang, Hui; Bowman, Daryl T; Rojas-Pierce, Marcela; Haigler, Candace H; Jones, Don C; Holland, James B; Chitwood, Daniel H; Kuraparthy, Vasu

2017-01-03

Leaf shape varies spectacularly among plants. Leaves are the primary source of photoassimilate in crop plants, and understanding the genetic basis of variation in leaf morphology is critical to improving agricultural productivity. Leaf shape played a unique role in cotton improvement, as breeders have selected for entire and lobed leaf morphs resulting from a single locus, okra (l-D 1 ), which is responsible for the major leaf shapes in cotton. The l-D 1 locus is not only of agricultural importance in cotton, but through pioneering chimeric and morphometric studies, it has contributed to fundamental knowledge about leaf development. Here we show that an HD-Zip transcription factor homologous to the LATE MERISTEM IDENTITY1 (LMI1) gene of Arabidopsis is the causal gene underlying the l-D 1 locus. The classical okra leaf shape allele has a 133-bp tandem duplication in the promoter, correlated with elevated expression, whereas an 8-bp deletion in the third exon of the presumed wild-type normal allele causes a frame-shifted and truncated coding sequence. Our results indicate that subokra is the ancestral leaf shape of tetraploid cotton that gave rise to the okra allele and that normal is a derived mutant allele that came to predominate and define the leaf shape of cultivated cotton. Virus-induced gene silencing (VIGS) of the LMI1-like gene in an okra variety was sufficient to induce normal leaf formation. The developmental changes in leaves conferred by this gene are associated with a photosynthetic transcriptomic signature, substantiating its use by breeders to produce a superior cotton ideotype.

Effects of MicroRNA-23a on Differentiation and Gene Expression Profiles in 3T3-L1 Adipocytes

PubMed Central

Huang, Yong; Huang, Jinxiu; Qi, Renli; Wang, Qi; Wu, Yongjiang; Wang, Jing

2016-01-01

MicroRNAs (miRNAs) are small non-coding RNA molecules that regulate growth, development, and programmed death of cells. A newly-published study has shown that miRNA-23a could regulate 3T3-L1 adipocyte differentiation. Here, we identified miRNA-23a as a negative regulator of 3T3-L1 adipocyte differentiation again. Over-expression of miRNA-23a inhibited differentiation and decreased lipogenesis as well as down-regulated mRNA and protein expression of both peroxisome proliferator-activated receptor (PPAR) γ and fatty acid binding protein (FABP) 4, whereas knock down of miRNA-23a showed the opposite effects on differentiation as well as increasing the number of apoptotic cells. Additionally, digital gene expression profiling sequencing (DGE-Seq) was used to assay changes in gene expression profiles following alterations in the level of miR-23a. In total, over-expression or knock down of miRNA-23a significantly changed the expression of 313 and 425 genes, respectively. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses indicated that these genes were mainly involved in the stress response, immune system, metabolism, cell cycle, among other pathways. Additionally, the signal transducer and activator of transcription 1 (Stat1) was shown to be a target of miRNA-23a by computational and dual-luciferase reporter assays that indicated Janus Kinase (Jak)-Stat signal pathway was implicated in regulating adipogenesis mediated by miRNA-23a in adipocytes. PMID:27783036

Quantitative analyses of ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO) large-subunit genes (cbbL) in typical paddy soils.

PubMed

Xiao, Ke-Qing; Bao, Peng; Bao, Qiong-Li; Jia, Yan; Huang, Fu-Yi; Su, Jian-Qiang; Zhu, Yong-Guan

2014-01-01

The Calvin cycle is known to be the major pathway for CO2 fixation, but our current understanding of its occurrence and importance in paddy soils is poor. In this study, the diversity of three ribulose-1,5-bisphosphate carboxylase/oxygenase large-subunit genes (cbbLG, cbbLR, cbbM) was investigated by clone library, T-RFLP, qPCR, and enzyme assay in five paddy soils in China. The cbbLG sequences revealed a relatively low level of diversity and were mostly related to the sequences of species from Thiobacillus. In contrast, highly diverse cbbLR and cbbM sequences were dispersed on the phylogenetic trees, and most of them were distantly related to known sequences, even forming separate clusters. Abundances of three cbbL genes ranged from 10(6) to 10(9) copies g(-1) soil, and cbbLR outnumbered cbbM and cbbLG in all soil samples, indicating that cbbLR may play a more important role than other two cbbL genes. Soil properties significantly influenced cbbL diversity in five paddy soils, of which clay content, C/N ratio, CEC, pH, and SOC correlated well with variations in microbial composition and abundance. In summary, this study provided a comparison of three cbbL genes, advancing our understanding of their role in carbon sequestration and nutrient turnover in the paddy soil. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Lead facilitates foci formation in a Balb/c-3T3 two-step cell transformation model: role of Ape1 function.

PubMed

Hernández-Franco, Pablo; Silva, Martín; Franco, Rodrigo; Valverde, Mahara; Rojas, Emilio

2018-04-01

Several possible mechanisms have been examined to gain an understanding on the carcinogenic properties of lead, which include among others, mitogenesis, alteration of gene expression, oxidative damage, and inhibition of DNA repair. The aim of the present study was to explore if low concentrations of lead, relevant for human exposure, interfere with Ape1 function, a base excision repair enzyme, and its role in cell transformation in Balb/c-3T3. Lead acetate 5 and 30 μM induced APE1 mRNA and upregulation of protein expression. This increase in mRNA expression is consistent throughout the chronic exposure. Additionally, we also found an impaired function of Ape1 through molecular beacon-based assay. To evaluate the impact of lead on foci formation, a Balb/c-3T3 two-step transformation model was used. Balb/c-3T3 cells were pretreated 1 week with low concentrations of lead before induction of transformation with n-methyl-n-nitrosoguanidine (MNNG) (0.5 μg/mL) and 12-O-tetradecanoylphorbol-13-acetate (TPA) (0.1 μg/mL) (a classical two-step protocol). Morphological cell transformation increased in response to lead pretreatment that was paralleled with an increase in Ape1 mRNA and protein overexpression and an impairment of Ape1 activity and correlating with foci number. In addition, we found that lead pretreatment and MNNG (transformation initiator) increased DNA damage, determined by comet assay. Our data suggest that low lead concentrations (5, 30 μM) could play a facilitating role in cellular transformation, probably through the impaired function of housekeeping genes such as Ape1, leading to DNA damage accumulation and chromosomal instability, one of the most important hallmarks of cancer induced by chronic exposures.

PD-1/PD-L1 in disease.

PubMed

Kuol, Nyanbol; Stojanovska, Lily; Nurgali, Kulmira; Apostolopoulos, Vasso

2018-02-01

Expression of PD-1 on T/B cells regulates peripheral tolerance and autoimmunity. Binding of PD-1 to its ligand, PD-L1, leads to protection against self-reactivity. In contrary, tumor cells have evolved immune escape mechanisms whereby overexpression of PD-L1 induces anergy and/or apoptosis of PD-1 positive T cells by interfering with T cell receptor signal transduction. PD-L1 and PD-1 blockade using antibodies are in human clinical trials as an alternative cancer treatment modality. Areas covered: We describe the role of PD-1/PD-L1 in disease in the context of autoimmunity, neurological disorders, stroke and cancer. For immunotherapy/vaccines to be successful, the expression of PD-L1/PD-1 on immune cells should be considered, and the combination of checkpoint inhibitors and vaccines may pave the way for successful outcomes to disease.

Cloning and expression of L-asparaginase gene in Escherichia coli.

PubMed

Wang, Y; Qian, S; Meng, G; Zhang, S

2001-08-01

The L-asparaginase (ASN) from Escherichia coli AS1.357 was cloned as a DNA fragment generated using polymerase chain reaction technology and primers derived from conserved regions of published ASN gene sequences. Recombinant plasmid pASN containing ASN gene and expression vector pBV220 was transformed in different E. coli host strains. The activity and expression level of ASN in the engineering strains could reach 228 IU/mL of culture fluid and about 50% of the total soluble cell protein respectively, more than 40-fold the enzyme activity of the wild strain. The recombinant plasmid in E. coli AS1.357 remained stable after 72 h of cultivation and 5 h of heat induction without selective pressure. The ASN gene of E. coli AS1.357 was sequenced and had high homology compared to the reported data.

Modifications to a LATE MERISTEM IDENTITY1 gene are responsible for the major leaf shapes of Upland cotton (Gossypium hirsutum L.)

PubMed Central

Andres, Ryan J.; Coneva, Viktoriya; Frank, Margaret H.; Tuttle, John R.; Samayoa, Luis Fernando; Han, Sang-Won; Kaur, Baljinder; Zhu, Linglong; Fang, Hui; Bowman, Daryl T.; Rojas-Pierce, Marcela; Haigler, Candace H.; Jones, Don C.; Holland, James B.; Chitwood, Daniel H.; Kuraparthy, Vasu

2017-01-01

Leaf shape varies spectacularly among plants. Leaves are the primary source of photoassimilate in crop plants, and understanding the genetic basis of variation in leaf morphology is critical to improving agricultural productivity. Leaf shape played a unique role in cotton improvement, as breeders have selected for entire and lobed leaf morphs resulting from a single locus, okra (l-D1), which is responsible for the major leaf shapes in cotton. The l-D1 locus is not only of agricultural importance in cotton, but through pioneering chimeric and morphometric studies, it has contributed to fundamental knowledge about leaf development. Here we show that an HD-Zip transcription factor homologous to the LATE MERISTEM IDENTITY1 (LMI1) gene of Arabidopsis is the causal gene underlying the l-D1 locus. The classical okra leaf shape allele has a 133-bp tandem duplication in the promoter, correlated with elevated expression, whereas an 8-bp deletion in the third exon of the presumed wild-type normal allele causes a frame-shifted and truncated coding sequence. Our results indicate that subokra is the ancestral leaf shape of tetraploid cotton that gave rise to the okra allele and that normal is a derived mutant allele that came to predominate and define the leaf shape of cultivated cotton. Virus-induced gene silencing (VIGS) of the LMI1-like gene in an okra variety was sufficient to induce normal leaf formation. The developmental changes in leaves conferred by this gene are associated with a photosynthetic transcriptomic signature, substantiating its use by breeders to produce a superior cotton ideotype. PMID:27999177

Correlation between lower balance of Th2 helper T-cells and expression of PD-L1/PD-1 axis genes enables prognostic prediction in patients with glioblastoma.

PubMed

Takashima, Yasuo; Kawaguchi, Atsushi; Kanayama, Tomohiko; Hayano, Azusa; Yamanaka, Ryuya

2018-04-10

Common cancer treatments include radiation therapy, chemotherapy including molecular targeted drugs and anticancer drugs, and surgical treatment. Recent studies have focused on investigating the mechanisms by which immune cells attack cancer cells and produce immune tolerance-suppressing cytokines, as well as on their potential application in cancer immunotherapy. We conducted expression profiling of CD274 ( PD-L1 ), GATA3, IFNG, IL12R, IL12RB2, IL4, PDCD1 ( PD-1 ), PDCD1LG2 ( PD-L2 ), and TBX21 ( T-bet ) using data of 158 glioblastoma multiforme (GBM) patients with clinical information available at The Cancer Genome Atlas. Principal component analysis of the expression profiling data was used to derive an equation for evaluating the status of Th1 and Th2 cells. GBM specimens were divided based on the median of the Th scores. The results revealed that Th1 High Th2 Low and Th1 Low Th2 Low statuses indicated better prognosis than Th1 High Th2 High , and were evaluated based on the downregulation of PD-L1, PD-L2, and PD-1. Furthermore, Th2 Low divided based on the threshold, as well as CD274 Low and PDCD1 Low , were associated with good prognosis. In the Th2 Low subgroup, 14 genes were identified as potential prognostic markers. Of these, SLC11A1 Low , TNFRSF1B Low , and LTBR Low also indicated good prognosis. These results suggest that low Th2 balance and low activity of the PD-L1/PD-1 axis predict good prognosis in GBM. The set of genes identified in the present study could reliably predict survival in GBM patients and serve as useful molecular markers. Furthermore, this set of genes could prove to be novel targets for cancer immunotherapy.

Genetic variability in E6, E7, and L1 genes of human papillomavirus genotype 52 from Southwest China.

PubMed

Zhang, Yiwen; Cao, Man; Wang, Mengting; Ding, Xianping; Jing, Yaling; Chen, Zuyi; Ma, Tengjiao; Chen, Honghan

2016-07-01

Human papillomavirus (HPV) is the major causative agent of cervical cancer, which accounts for the second highest cancer burden in women worldwide. HPV-52, the prevalent subtype in Asia, especially in southwest China, was analyzed in this study. To analyze polymorphisms, intratypic variants, and genetic variability in the E6-E7 (n=26) and L1 (n=53) genes of HPV-52, these genes were sequenced and the sequences were submitted to GenBank. Phylogenetic trees were constructed using the neighbor-joining and Kimura 2-parameters methods, followed by analysis of the diversity of secondary structure. Finally, we estimated the selection pressures acting on the E6-E7 and L1 genes. Fifty-one novel variants of HPV-52 L1, and two novel variants of HPV-52 E6-E7 were identified in this study. Thirty single nucleotide changes were observed in HPV-52 E6-E7 sequences with 19/30 non-synonymous mutations and 11/30 synonymous mutations (five in the alpha helix and five in the beta sheet). Fifty-five single nucleotide changes were observed in HPV-52 L1 sequences with 17/55 non-synonymous mutations (seven in the alpha helix and fourteen in the beta sheet) and 38/55 synonymous mutations. Selective pressure analysis predicted that most of these mutations reflect positive selection. Identifying new variants in HPV-52 may inform the rational design of new vaccines specifically for women in southwest China. Knowledge of genetic variation in HPV may be useful as an epidemiologic correlate of cervical cancer risk, or may even provide critical information for developing diagnostic probes. Copyright © 2016 Elsevier B.V. All rights reserved.

Saccharomyces cerevisiae sigma 1278b has novel genes of the N-acetyltransferase gene superfamily required for L-proline analogue resistance.

PubMed

Takagi, H; Shichiri, M; Takemura, M; Mohri, M; Nakamori, S

2000-08-01

We discovered on the chromosome of Saccharomyces cerevisiae Sigma 1278b novel genes involved in L-proline analogue L-azetidine-2-carboxylic acid resistance which are not present in the standard laboratory strains. The 5.4 kb-DNA fragment was cloned from the genomic library of the L-azetidine-2-carboxylic acid-resistant mutant derived from a cross between S. cerevisiae strains S288C and Sigma 1278b. The nucleotide sequence of a 4.5-kb segment exhibited no identity with the sequence in the genome project involving strain S288C. Deletion analysis indicated that one open reading frame encoding a predicted protein of 229 amino acids is indispensable for L-azetidine-2-carboxylic acid resistance. The protein sequence was found to be a member of the N-acetyltransferase superfamily. Genomic Southern analysis and gene disruption showed that two copies of the novel gene with one amino acid change at position 85 required for L-azetidine-2-carboxylic acid resistance were present on chromosomes X and XIV of Sigma 1278b background strains. When this novel MPR1 or MPR2 gene (sigma 1278b gene for L-proline analogue resistance) was introduced into the other S. cerevisiae strains, all of the recombinants were resistant to L-azetidine-2-carboxylic acid, indicating that both MPR1 and MPR2 are expressed and have a global function in S. cerevisiae.

Transcriptional Repression of the Dspp Gene Leads to Dentinogenesis Imperfecta Phenotype in Col1a1-Trps1 Transgenic Mice

PubMed Central

Napierala, Dobrawa; Sun, Yao; Maciejewska, Izabela; Bertin, Terry K; Dawson, Brian; D'Souza, Rena; Qin, Chunlin; Lee, Brendan

2012-01-01

Dentinogenesis imperfecta (DGI) is a hereditary defect of dentin, a calcified tissue that is the most abundant component of teeth. Most commonly, DGI is manifested as a part of osteogenesis imperfecta (OI) or the phenotype is restricted to dental findings only. In the latter case, DGI is caused by mutations in the DSPP gene, which codes for dentin sialoprotein (DSP) and dentin phosphoprotein (DPP). Although these two proteins together constitute the majority of noncollagenous proteins of the dentin, little is known about their transcriptional regulation. Here we demonstrate that mice overexpressing the Trps1 transcription factor (Col1a1-Trps1 mice) in dentin-producing cells, odontoblasts, present with severe defects of dentin formation that resemble DGI. Combined micro–computed tomography (µCT) and histological analyses revealed tooth fragility due to severe hypomineralization of dentin and a diminished dentin layer with irregular mineralization in Col1a1-Trps1 mice. Biochemical analyses of noncollagenous dentin matrix proteins demonstrated decreased levels of both DSP and DPP proteins in Col1a1-Trps1 mice. On the molecular level, we demonstrated that sustained high levels of Trps1 in odontoblasts lead to dramatic decrease of Dspp expression as a result of direct inhibition of the Dspp promoter by Trps1. During tooth development Trps1 is highly expressed in preodontoblasts, but in mature odontoblasts secreting matrix its expression significantly decreases, which suggests a Trps1 role in odontoblast development. In these studies we identified Trps1 as a potent inhibitor of Dspp expression and the subsequent mineralization of dentin. Thus, we provide novel insights into mechanisms of transcriptional dysregulation that leads to DGI. © 2012 American Society for Bone and Mineral Research. PMID:22508542

Marker-assisted pyramiding of brown planthopper (Nilaparvata lugens Stål) resistance genes Bph1 and Bph2 on rice chromosome 12.

PubMed

Sharma, Prem N; Torii, Akihide; Takumi, Shigeo; Mori, Naoki; Nakamura, Chiharu

2004-01-01

Brown planthopper (BPH) (Nilaparvata lugens Stål) is a significant insect pest of rice (Oryza sativa L.). We constructed a gene-pyramided japonica line, in which two BPH resistance genes Bph1 and Bph2 on the long arm of chromosome 12 independently derived from two indica resistance lines were combined through the recombinant selection. The gene-pyramiding was achieved based on the previously constructed high-resolution linkage maps of the two genes. Two co-dominant and four dominant PCR-based markers flanking the loci were used to select for a homozygous recombinant line in a segregating population that was derived from a cross between the parental homozygous single-gene introgression lines. BPH bioassay showed that the resistance level of the pyramided line was equivalent to that of the Bph1-single introgression line, which showed a higher level of resistance than the Bph2-single introgression line. The pyramid line should provide a useful experimental means for studying the fine structure of the chromosomal region covering these two major BPH resistance genes.

Chronic exposure of μg/L range Bisphenol A to adult zebrafish (Danio rerio) leading to adipogenesis

NASA Astrophysics Data System (ADS)

Ngo, Mai Thi; Doan, Thao Thi-Phuong; Nguyen, Cong Thanh; Vo, Diem Thi-Ngoc; Do, Chi Hong-Lan; Le, Nga Phi

2017-09-01

Bisphenol A (BPA) is known as an endocrine disruptor compound and commonly found in food-packaging plastics and in aquatic environment. It is scientifically concerned that BPA may causes significant health risks to human and aquatic animals. In fact, most of scientific studies have been conducted with BPA-acute toxicity in early stages of animal development, while far lesser researches have been focused on BPA-chronic toxicity in adults. This study was to investigate the chronic effects of environmental 1, 10 and 100 µg/L BPA to four-week-old zebrafishes (Danio rerio) after 60 days of exposure in terms of changing of body morphology, hepatocyte morphology and the transcriptional expression of biomarker gene for lipid metabolism. As a result, significant effects were found in: 1/ the increase of body weight, but not body length; 2/ the increase in the number of vacuolated hepatocytes corresponding to relatively higher glycogen and lipid content; 3/ shifting hepatocyte morphology to basophilic cytoplasm and cytoplasmic and/or nuclear enlargement, but not inflammation signs (macrophages, granuloma, fibrosis/cirrhosis); 4/ the decrease of mRNA level of PPARγ and C/EBPα genes in liver. Our study here indicates that the exposure to μg/L range concentration of BPA leads to adipogensis in adult zebrafishes.

Gene mapping and functional analysis of the novel leaf color gene SiYGL1 in foxtail millet [Setaria italica (L.) P. Beauv].

PubMed

Li, Wen; Tang, Sha; Zhang, Shuo; Shan, Jianguo; Tang, Chanjuan; Chen, Qiannan; Jia, Guanqing; Han, Yuanhuai; Zhi, Hui; Diao, Xianmin

2016-05-01

Setaria italica and its wild ancestor Setaria viridis are emerging as model systems for genetics and functional genomics research. However, few systematic gene mapping or functional analyses have been reported in these promising C4 models. We herein isolated the yellow-green leaf mutant (siygl1) in S. italica using forward genetics approaches. Map-based cloning revealed that SiYGL1, which is a recessive nuclear gene encoding a magnesium-chelatase D subunit (CHLD), is responsible for the mutant phenotype. A single Phe to Leu amino acid change occurring near the ATPase-conserved domain resulted in decreased chlorophyll (Chl) accumulation and modified chloroplast ultrastructure. However, the mutation enhanced the light-use efficiency of the siygl1 mutant, suggesting that the mutated CHLD protein does not completely lose its original activity, but instead, gains novel features. A transcriptional analysis of Chl a oxygenase revealed that there is a strong negative feedback control of Chl b biosynthesis in S. italica. The SiYGL1 mRNA was expressed in all examined tissues, with higher expression observed in the leaves. Comparison of gene expression profiles in wild-type and siygl1 mutant plants indicated that SiYGL1 regulates a subset of genes involved in photosynthesis (rbcL and LHCB1), thylakoid development (DEG2) and chloroplast signaling (SRP54CP). These results provide information regarding the mutant phenotype at the transcriptional level. This study demonstrated that the genetic material of a Setaria species could be ideal for gene discovery investigations using forward genetics approaches and may help to explain the molecular mechanisms associated with leaf color variation. © 2015 Scandinavian Plant Physiology Society.

Overexpression of the Transcription Factor Sp1 Activates the OAS-RNAse L-RIG-I Pathway

PubMed Central

Dupuis-Maurin, Valéryane; Brinza, Lilia; Baguet, Joël; Plantamura, Emilie; Schicklin, Stéphane; Chambion, Solène; Macari, Claire; Tomkowiak, Martine; Deniaud, Emmanuelle; Leverrier, Yann

2015-01-01

Deregulated expression of oncogenes or transcription factors such as specificity protein 1 (Sp1) is observed in many human cancers and plays a role in tumor maintenance. Paradoxically in untransformed cells, Sp1 overexpression induces late apoptosis but the early intrinsic response is poorly characterized. In the present work, we studied increased Sp1 level consequences in untransformed cells and showed that it turns on an early innate immune transcriptome. Sp1 overexpression does not activate known cellular stress pathways such as DNA damage response or endoplasmic reticulum stress, but induces the activation of the OAS-RNase L pathway and the generation of small self-RNAs, leading to the upregulation of genes of the antiviral RIG-I pathway at the transcriptional and translational levels. Finally, Sp1-induced intrinsic innate immune response leads to the production of the chemokine CXCL4 and to the recruitment of inflammatory cells in vitro and in vivo. Altogether our results showed that increased Sp1 level in untransformed cells constitutes a novel danger signal sensed by the OAS-RNase L axis leading to the activation of the RIG-I pathway. These results suggested that the OAS-RNase L-RIG-I pathway may be activated in sterile condition in absence of pathogen. PMID:25738304

Mutations in GREB1L Cause Bilateral Kidney Agenesis in Humans and Mice.

PubMed

De Tomasi, Lara; David, Pierre; Humbert, Camille; Silbermann, Flora; Arrondel, Christelle; Tores, Frédéric; Fouquet, Stéphane; Desgrange, Audrey; Niel, Olivier; Bole-Feysot, Christine; Nitschké, Patrick; Roume, Joëlle; Cordier, Marie-Pierre; Pietrement, Christine; Isidor, Bertrand; Khau Van Kien, Philippe; Gonzales, Marie; Saint-Frison, Marie-Hélène; Martinovic, Jelena; Novo, Robert; Piard, Juliette; Cabrol, Christelle; Verma, Ishwar C; Puri, Ratna; Journel, Hubert; Aziza, Jacqueline; Gavard, Laurent; Said-Menthon, Marie-Hélène; Heidet, Laurence; Saunier, Sophie; Jeanpierre, Cécile

2017-11-02

Congenital anomalies of the kidney and urinary tract (CAKUT) constitute a major cause of chronic kidney disease in children and 20% of prenatally detected anomalies. CAKUT encompass a spectrum of developmental kidney defects, including renal agenesis, hypoplasia, and cystic and non-cystic dysplasia. More than 50 genes have been reported as mutated in CAKUT-affected case subjects. However, the pathophysiological mechanisms leading to bilateral kidney agenesis (BKA) remain largely elusive. Whole-exome or targeted exome sequencing of 183 unrelated familial and/or severe CAKUT-affected case subjects, including 54 fetuses with BKA, led to the identification of 16 heterozygous variants in GREB1L (growth regulation by estrogen in breast cancer 1-like), a gene reported as a target of retinoic acid signaling. Four loss-of-function and 12 damaging missense variants, 14 being absent from GnomAD, were identified. Twelve of them were present in familial or simplex BKA-affected case subjects. Female BKA-affected fetuses also displayed uterus agenesis. We demonstrated a significant association between GREB1L variants and BKA. By in situ hybridization, we showed expression of Greb1l in the nephrogenic zone in developing mouse kidney. We generated a Greb1l knock-out mouse model by CRISPR-Cas9. Analysis at E13.5 revealed lack of kidneys and genital tract anomalies in male and female Greb1l -/- embryos and a slight decrease in ureteric bud branching in Greb1l +/- embryos. We showed that Greb1l invalidation in mIMCD3 cells affected tubulomorphogenesis in 3D-collagen culture, a phenotype rescued by expression of the wild-type human protein. This demonstrates that GREB1L plays a major role in early metanephros and genital development in mice and humans. Copyright © 2017 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

Identification and fine mapping of a stay-green gene (Brnye1) in pakchoi (Brassica campestris L. ssp. chinensis).

PubMed

Wang, Nan; Liu, Zhiyong; Zhang, Yun; Li, Chengyu; Feng, Hui

2018-03-01

Using bulked segregant analysis combined with next-generation sequencing, we delimited the Brnye1 gene responsible for the stay-green trait of nye in pakchoi. Sequence analysis identified Bra019346 as the candidate gene. "Stay-green" refers to a plant trait whereby leaves remain green during senescence. This trait is useful in the cultivation of pakchoi (Brassica campestris L. ssp. chinensis), which is marketed as a green leaf product. This study aimed to identify the gene responsible for the stay-green trait in pakchoi. We identified a stay-green mutant in pakchoi, which we termed "nye". Genetic analysis revealed that the stay-green trait is controlled by a single recessive gene, Brnye1. Using the BSA-seq method, a 3.0-Mb candidate region was mapped on chromosome A03, which helped us localize Brnye1 to an 81.01-kb interval between SSR markers SSRWN27 and SSRWN30 via linkage analysis in an F 2 population. We identified 12 genes in this region, 11 of which were annotated based on the Brassica rapa annotation database, and one was a functionally unknown gene. An orthologous gene of the Arabidopsis gene AtNYE1, Bra019346, was identified as the potential candidate for Brnye1. Sequence analysis revealed a 40-bp insertion in the second exon of Bra019346 in nye, which generated the TAA stop codon. A candidate gene-specific Indel marker in 1561 F 2 individuals showed perfect cosegregation with Brnye1 in the nye mutant. These results provide a foundation for uncovering the molecular mechanism of the stay-green trait in pakchoi.

Robust Gaussian Graphical Modeling via l1 Penalization

PubMed Central

Sun, Hokeun; Li, Hongzhe

2012-01-01

Summary Gaussian graphical models have been widely used as an effective method for studying the conditional independency structure among genes and for constructing genetic networks. However, gene expression data typically have heavier tails or more outlying observations than the standard Gaussian distribution. Such outliers in gene expression data can lead to wrong inference on the dependency structure among the genes. We propose a l1 penalized estimation procedure for the sparse Gaussian graphical models that is robustified against possible outliers. The likelihood function is weighted according to how the observation is deviated, where the deviation of the observation is measured based on its own likelihood. An efficient computational algorithm based on the coordinate gradient descent method is developed to obtain the minimizer of the negative penalized robustified-likelihood, where nonzero elements of the concentration matrix represents the graphical links among the genes. After the graphical structure is obtained, we re-estimate the positive definite concentration matrix using an iterative proportional fitting algorithm. Through simulations, we demonstrate that the proposed robust method performs much better than the graphical Lasso for the Gaussian graphical models in terms of both graph structure selection and estimation when outliers are present. We apply the robust estimation procedure to an analysis of yeast gene expression data and show that the resulting graph has better biological interpretation than that obtained from the graphical Lasso. PMID:23020775

Leigh syndrome associated with mitochondrial complex I deficiency due to a novel mutation in the NDUFS1 gene.

PubMed

Martín, Miguel A; Blázquez, Alberto; Gutierrez-Solana, Luis G; Fernández-Moreira, Daniel; Briones, Paz; Andreu, Antoni L; Garesse, Rafael; Campos, Yolanda; Arenas, Joaquín

2005-04-01

Mutations in the nuclear-encoded subunits of complex I of the mitochondrial respiratory chain are a recognized cause of Leigh syndrome (LS). Recently, 6 mutations in the NDUFS1 gene were identified in 3 families. To describe a Spanish family with LS, complex I deficiency in muscle, and a novel mutation in the NDUFS1 gene. Using molecular genetic approaches, we identified the underlying molecular defect in a patient with LS with a complex I defect. The proband was a child who displayed the clinical features of LS. Muscle biochemistry results showed a complex I defect of the mitochondrial respiratory chain. Sequencing analysis of the mitochondrial DNA-encoded ND genes, the nuclear DNA-encoded NDUFV1, NDUFS1, NDUFS2, NDUFS4, NDUFS6, NDUFS7, NDUFS8, and NDUFAB1 genes, and the complex I assembly factor CIA30 gene revealed a novel homozygous L231V mutation (c.691C-->G) in the NDUFS1 gene. The parents were heterozygous carriers of the L231V mutation. Identifying nuclear mutations as a cause of respiratory chain disorders will enhance the possibility of prenatal diagnosis and help us understand how molecular defects can lead to complex I deficiency.

BCoR-L1 variation and breast cancer.

PubMed

Lose, Felicity; Arnold, Jeremy; Young, David B; Brown, Carolyn J; Mann, Graham J; Pupo, Gulietta M; Khanna, Kum Kum; Chenevix-Trench, Georgia; Spurdle, Amanda B

2007-01-01

BRCA1 is involved in numerous essential processes in the cell, and the effects of BRCA1 dysfunction in breast cancer carcinogenesis are well described. Many of the breast cancer susceptibility genes such as BRCA2, p53, ATM, CHEK2, and BRIP1 encode proteins that interact with BRCA1. BCL6 corepressor-like 1 (BCoR-L1) is a newly described BRCA1-interacting protein that displays high homology to several proteins known to be involved in the fundamental processes of DNA damage repair and transcription regulation. BCoR-L1 has been shown to play a role in transcription corepression, and expression of the X-linked BCoR-L1 gene has been reported to be dysregulated in breast cancer subjects. BCoR-L1 is located on the X chromosome and is subject to X inactivation. We performed mutation analysis of 38 BRCA1/2 mutation-negative breast cancer families with male breast cancer, prostate cancer, and/or haplotype sharing around BCoR-L1 to determine whether there is a role for BCoR-L1 as a high-risk breast cancer predisposition gene. In addition, we conducted quantitative real-time PCR (qRT-PCR) on lymphoblastoid cell lines (LCLs) from the index cases from these families and a number of cancer cell lines to assess the role of BCoR-L1 dysregulation in cancer and cancer families. Very little variation was detected in the coding region, and qRT-PCR analysis revealed that BCoR-L1 expression is highly variable in cancer-free subjects, high-risk breast cancer patients, and cancer cell lines. We also report the investigation of a new expression control, DIDO1 (death inducer-obliterator 1), that is superior to GAPDH (glyceraldehyde-3-phosphate dehydrogenase) and UBC (ubiquitin C) for analysis of expression in LCLs. Our results suggest that BCoR-L1 expression does not play a large role in predisposition to familial breast cancer.

Potential role of TCF7L2 gene variants on cardiac sympathetic/parasympathetic activity.

PubMed

Boccardi, Virginia; Ambrosino, Immacolata; Papa, Michela; Fiore, Daniela; Rizzo, Maria Rosaria; Paolisso, Giuseppe; Barbieri, Michelangela

2010-12-01

Variants in transcription factor 7-like 2 (266096218TCF7L2266096218USuser266096218Gene names have been italicized per house style. Please check and confirm whether there are other instances that need to be italicized or instances where italics have been inappropriately applied.) gene have been found strongly associated with an increased risk of type 2 diabetes, as well as with an impairment of glucagon-like peptide-1 (GLP-1) signalling chain. In rats, stimulation of central GLP-1 receptors increases heart rate and activates autonomic regulatory neurons. We aimed to evaluate the potential role of TCF7L2 gene polymorphisms on sympathovagal response in relation to changes in plasma insulin and/or GLP-1 concentration after glucose ingestion. Genotyping was performed for rs12255372 and rs7903146 TCF7L2 gene variants in 250 non-related healthy volunteers (mean age 27±3 years). Consistent with previous reports, both single-nucleotide polymorphisms were in strong linkage disequilibrium (D'=0.87, r(2)=0.76). A subset of 167 patients underwent an oral glucose tolerance test while a continuous recording of heart rate variability was performed. At baseline, no differences in fasting plasma insulin, in GLP-1 levels and in LF/HF (low frequency/high frequency) ratio between the three genotypes were found. Along with glucose ingestion TT subjects had lower INS(AUC) (insulin area under curve), as well as higher LF/HF(AUC) (LF/HF area under curve) values. No difference in GLP-1(AUC) (GLP-1 area under curve) between TCF7L2 gene variants was found. A multivariate analysis including multiple covariates showed that only INS(AUC,) GLP-1(AUC) and TCF7L2 gene variants were independently associated with LF/HF(AUC). In conclusion, TT genotype of rs12255372 and rs7903146 TCF7L2 gene variants is associated with lower insulin secretion and higher cardiosympathetic activity. Moreover, such effect is independent of GLP-1 and insulin plasma concentrations suggesting a potential role of such gene

GhNAC12, a neutral candidate gene, leads to early aging in cotton (Gossypium hirsutum L).

PubMed

Zhao, Fengli; Ma, Jianhui; Li, Libei; Fan, Shuli; Guo, Yaning; Song, Meizhen; Wei, Hengling; Pang, Chaoyou; Yu, Shuxun

2016-01-15

NAC (NAM, ATAF, and CUC) is one of the largest transcription factor families in plants, and its members play various roles in plant growth, development, and the response to biotic and abiotic stresses. Currently, 77 NAC genes have been reported in cotton (Gossypium hirsutum L.). And GhNAC12 showed up-regulation during leaf senescence, but its role in this process is poorly understood. In the present study, a preliminary function analysis of GhNAC12 was performed during leaf senescence. qRT-PCR analysis indicated that GhNAC12 expression increased during the early-aging process and the aging of cotyledons. Additionally, we observed that overexpression of GhNAC12 in Arabidopsis led to early senescence (early aging). Our findings suggest that GhNAC12 is a candidate gene for early aging in upland cotton cultivars. Neutrality tests suggested that there was no selection pressure imposed on GhNAC12 during the domestication of upland cotton. Copyright © 2015 Elsevier B.V. All rights reserved.

Specific immune response genes of the guinea pig. II. Relationship between the poly-L-lysine gene and the genes controlling immune responsiveness to copolymers of L-glutamic acid and L-alanine and L-glutamic acid and L-tyrosine in random-bred Hartley guinea pigs.

PubMed

Bluestein, H G; Green, I; Benacerraf, B

1971-08-01

The ability of guinea pigs to make immune responses to GA, a linear random copolymer of L-glutamic acid and L-alanine, GT, a random linear copolymer of L-glutamic acid and L-tyrosine, and PLL, a linear homopolymer of L-lysine, is controlled by different autosomal dominant genes specific for each of those polymers. We have investigated the relationship between the PLL gene and the GA and GT immune response genes by simultaneously immunizing random-bred Hartley strain guinea pigs with GA and PLL, GT and PLL, or GA and GT. In most Hartley guinea pigs the ability to respond immunologically to GA and to PLL is inherited together; that is, most animals responding to GA respond to PLL and vice versa. However, a few animals respond to either GA or to PLL but not both, demonstrating that the GA and PLL immune response genes are not identical but linked in most Hartley animals. Conversely, when simultaneously immunized with GT and PLL, most Hartley guinea pigs respond to either PLL or GT but not both, indicating that GT and PLL responsiveness tends to segregate away from each other. Thus, the GT and PLL immune response genes also are not inherited independently but, rather, behave as alleles or pseudoalleles. Similar results are observed when Hartley guinea pigs are simultaneously immunized with GA and GT. The ability to respond to GA segregates away from the ability to respond to GT. Our studies demonstrated that the specific immune response genes thus far identified in guinea pigs controlling the ability to respond to GA, GT, and PLL, respectively, are found on the same chromosome. In most Hartley animals, the GA and PLL immune response genes are often linked, i.e. occur on the same chromosome strand, and tend to behave as alleles or pseudoalleles to the GT immune response gene.

Involvement of interleukin-1 in lead nitrate-induced hypercholesterolemia in mice.

PubMed

Kojima, Misaki; Ashino, Takashi; Yoshida, Takemi; Iwakura, Yoichiro; Degawa, Masakuni

2012-01-01

Hepatic 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) and cholesterol 7α-hydroxylase (Cyp7a1) are rate-limiting enzymes for cholesterol biosynthesis and catabolism, respectively. Involvement of inflammatory cytokines, particularly interleukin-1 (IL-1), in alterations of HMGR and Cyp7a1 gene expression during development of lead nitrate (LN)-induced hypercholesterolemia was examined in IL-1α/β-knockout (IL-1-KO) and wild-type (WT) mice. Lead nitrate treatment of WT mice led to not only a marked downregulation of the Cyp7a1 gene at 6-12 h, but also a significant upregulation of the HMGR gene at 12 h. However, such changes were not observed at significant levels in IL-1-KO mice, although a slight, transient downregulation of the Cyp7a1 gene and a minimal upregulation of the HMGR gene occurred at 6 h and 24 h, respectively. Consequently, LN treatment led to development of hypercholesterolemia at 24 h in WT mice, but not in IL-1-KO mice. Furthermore, in WT mice, significant LN-mediated increases were observed at 3-6 h in hepatic IL-1 levels, which can modulate gene expression of Cyp7a1 and HMGR. These findings indicate that, in mice, LN-mediated increases in hepatic IL-1 levels contribute, at least in part, to altered expressions of Cyp7a1 and HMGR genes, and eventually to hypercholesterolemia development.

L1-CAM in cancerous tissues.

PubMed

Gavert, Nancy; Ben-Shmuel, Amir; Raveh, Shani; Ben-Ze'ev, Avri

2008-11-01

L1-cell adhesion molecule (L1-CAM) is a cell adhesion receptor of the immunoglobulin superfamily, known for its roles in nerve cell function. While originally believed to be present only in brain cells, in recent years L1-CAM has been detected in other tissues, and in a variety of cancer cells, including some common types of human cancer. We review the prevalence of L1-CAM in human cancer, the possible mechanisms involved in L1-CAM-mediated tumorigenesis, and cancer therapies based upon L1-CAM antibody treatment. In colon cancer cells, the L1-CAM gene was identified as a target of the Wnt/beta-catenin-TCF signaling pathway, and L1-CAM was exclusively detected at the invasive front of colon and ovarian cancer tissue. The expression of L1-CAM in normal and cancer cells enhanced tumorigenesis and conferred metastasis in colon cancer cells. Antibodies against the L1-CAM ectodomain severely inhibited the proliferation of a variety of cancer cells in culture and reduced tumor burden when injected into mice harboring cancer cells expressing L1-CAM. These results, in addition to the presence of L1-CAM on the cell surface and its restricted distribution in normal tissues, make it an ideal target for tumor therapy.

Dopamine Signaling Leads to Loss of Polycomb Repression and Aberrant Gene Activation in Experimental Parkinsonism

PubMed Central

Lerdrup, Mads; Gomes, Ana-Luisa; Kryh, Hanna; Spigolon, Giada; Caboche, Jocelyne; Fisone, Gilberto; Hansen, Klaus

2014-01-01

Polycomb group (PcG) proteins bind to and repress genes in embryonic stem cells through lineage commitment to the terminal differentiated state. PcG repressed genes are commonly characterized by the presence of the epigenetic histone mark H3K27me3, catalyzed by the Polycomb repressive complex 2. Here, we present in vivo evidence for a previously unrecognized plasticity of PcG-repressed genes in terminally differentiated brain neurons of parkisonian mice. We show that acute administration of the dopamine precursor, L-DOPA, induces a remarkable increase in H3K27me3S28 phosphorylation. The induction of the H3K27me3S28p histone mark specifically occurs in medium spiny neurons expressing dopamine D1 receptors and is dependent on Msk1 kinase activity and DARPP-32-mediated inhibition of protein phosphatase-1. Chromatin immunoprecipitation (ChIP) experiments showed that increased H3K27me3S28p was accompanied by reduced PcG binding to regulatory regions of genes. An analysis of the genome wide distribution of L-DOPA-induced H3K27me3S28 phosphorylation by ChIP sequencing (ChIP-seq) in combination with expression analysis by RNA-sequencing (RNA-seq) showed that the induction of H3K27me3S28p correlated with increased expression of a subset of PcG repressed genes. We found that induction of H3K27me3S28p persisted during chronic L-DOPA administration to parkisonian mice and correlated with aberrant gene expression. We propose that dopaminergic transmission can activate PcG repressed genes in the adult brain and thereby contribute to long-term maladaptive responses including the motor complications, or dyskinesia, caused by prolonged administration of L-DOPA in Parkinson's disease. PMID:25254549

Clinical differences between patients with MODY-3, MODY-2 and type 2 diabetes mellitus with I27L polymorphism in the HNF1alpha gene.

PubMed

Pinés Corrales, Pedro José; López Garrido, María P; Aznar Rodríguez, Silvia; Louhibi Rubio, Lynda; López Jiménez, Luz M; Lamas Oliveira, Cristina; Alfaro Martínez, Jose J; Lozano García, Jose J; Hernández López, Antonio; Requejo Castillo, Ramón; Escribano Martínez, Julio; Botella Romero, Francisco

2010-01-01

The aim of our study was to describe and evaluate the clinical and metabolic characteristics of patients with MODY-3, MODY-2 or type 2 diabetes who presented I27L polymorphism in the HNF1alpha gene. The study included 31 previously diagnosed subjects under follow-up for MODY-3 (10 subjects from 5 families), MODY-2 (15 subjects from 9 families), or type 2 diabetes (6 subjects) with I27L polymorphism in the HNF1alpha gene. The demographic, clinical, metabolic, and genetic characteristics of all patients were analyzed. No differences were observed in distribution according to sex, age of onset, or form of diagnosis. All patients with MODY-2 or MODY-3 had a family history of diabetes. In contrast, 33.3% of patients with type 2 diabetes mellitus and I27L polymorphism in the HNF1alpha gene had no family history of diabetes (p < 0.05). No differences were observed in body mass index, prevalence of hypertension, or microvascular or macrovascular complications. Drug therapy was required by 100% of MODY-3 patients, but not required by 100% of MODY-2 patients or 16.7% of patients with type 2 diabetes mellitus and I27L polymorphism in the HNF1alpha gene (p < 0.05). Occasional difficulties may be encountered when classifying patients with MODY-2, MODY-3 or type 2 diabetes of atypical characteristics, in this case patients who present I27L polymorphism in the HNF1alpha gene. Copyright 2010 Sociedad Española de Endocrinología y Nutrición. Published by Elsevier Espana. All rights reserved.

Transcriptome-Wide Identification of Differentially Expressed Genes in Solanum lycopersicon L. in Response to an Alfalfa-Protein Hydrolysate Using Microarrays.

PubMed

Ertani, Andrea; Schiavon, Michela; Nardi, Serenella

2017-01-01

An alfalfa -based protein hydrolysate (EM) has been tested in tomato ( Solanum lycopersicon L.) plants at two different concentrations (0.1 and 1 mL L -1 ) to get insight on its efficacy as biostimulant in this species and to unravel possible metabolic targets and molecular mechanisms that may shed light on its mode of action. EM was efficient in promoting the fresh biomass and content in chlorophyll and soluble sugars of tomato plants, especially when it was applied at the concentration of 1 mL L -1 . This effect on plant productivity was likely related to the EM-dependent up-regulation of genes identified via microarray and involved in primary carbon and nitrogen metabolism, photosynthesis, nutrient uptake and developmental processes. EM also up-regulated a number of genes implied in the secondary metabolism that leads to the synthesis of compounds (phenols and terpenes) functioning in plant development and interaction with the environment. Concomitantly, phenol content was enhanced in EM-treated plants. Several new genes have been identified in tomato as potential targets of EM action, like those involved in detoxification processes from reactive oxygen species and xenobiotic (particularly glutathione/ascorbate cycle-related and ABC transporters), and defense against abiotic and biotic stress. The model hypothesized is that elicitors present in the EM formulation like auxins, phenolics, and amino acids, may trigger a signal transduction pathway via modulation of the intracellular levels of the hormones ethylene, jasmonic acid and abscissic acid, which then further prompt the activation of a cascade events requiring the presence and activity of many kinases and transcription factors to activate stress-related genes. The genes identified suggest these kinases and transcription factors as players involved in a complex crosstalk between biotic and abiotic stress signaling pathways. We conclude that EM acts as a biostimulant in tomato due to its capacity to stimulate

Transcriptome-Wide Identification of Differentially Expressed Genes in Solanum lycopersicon L. in Response to an Alfalfa-Protein Hydrolysate Using Microarrays

PubMed Central

Ertani, Andrea; Schiavon, Michela; Nardi, Serenella

2017-01-01

An alfalfa-based protein hydrolysate (EM) has been tested in tomato (Solanum lycopersicon L.) plants at two different concentrations (0.1 and 1 mL L-1) to get insight on its efficacy as biostimulant in this species and to unravel possible metabolic targets and molecular mechanisms that may shed light on its mode of action. EM was efficient in promoting the fresh biomass and content in chlorophyll and soluble sugars of tomato plants, especially when it was applied at the concentration of 1 mL L-1. This effect on plant productivity was likely related to the EM-dependent up-regulation of genes identified via microarray and involved in primary carbon and nitrogen metabolism, photosynthesis, nutrient uptake and developmental processes. EM also up-regulated a number of genes implied in the secondary metabolism that leads to the synthesis of compounds (phenols and terpenes) functioning in plant development and interaction with the environment. Concomitantly, phenol content was enhanced in EM-treated plants. Several new genes have been identified in tomato as potential targets of EM action, like those involved in detoxification processes from reactive oxygen species and xenobiotic (particularly glutathione/ascorbate cycle-related and ABC transporters), and defense against abiotic and biotic stress. The model hypothesized is that elicitors present in the EM formulation like auxins, phenolics, and amino acids, may trigger a signal transduction pathway via modulation of the intracellular levels of the hormones ethylene, jasmonic acid and abscissic acid, which then further prompt the activation of a cascade events requiring the presence and activity of many kinases and transcription factors to activate stress-related genes. The genes identified suggest these kinases and transcription factors as players involved in a complex crosstalk between biotic and abiotic stress signaling pathways. We conclude that EM acts as a biostimulant in tomato due to its capacity to stimulate plant

Insulin-like growth factor-1 enhances rat skeletal muscle charge movement and L-type Ca2+ channel gene expression

PubMed Central

Wang, Zhong-Min; Laura Messi, María; Renganathan, Muthukrishnan; Delbono, Osvaldo

1999-01-01

We investigated whether insulin-like growth factor-1 (IGF-1), an endogenous potent activator of skeletal muscle proliferation and differentiation, enhances L-type Ca2+ channel gene expression resulting in increased functional voltage sensors in single skeletal muscle cells. Charge movement and inward Ca2+ current were recorded in primary cultured rat myoballs using the whole-cell configuration of the patch-clamp technique. Ca2+ current and maximum charge movement (Qmax) were potentiated in cells treated with IGF-1 without significant changes in their voltage dependence. Peak Ca2+ current in control and IGF-1-treated cells was -7·8 ± 0·44 and -10·5 ± 0·37 pA pF−1, respectively (P < 0·01), whilst Qmax was 12·9 ± 0·4 and 22·0 ± 0·3 nC μF−1, respectively (P < 0·01). The number of L-type Ca2+ channels was found to increase in the same preparation. The maximum binding capacity (Bmax) of the high-affinity radioligand [3H]PN200-110 in control and IGF-1-treated cells was 1·21 ± 0·25 and 3·15 ± 0·5 pmol (mg protein)−1, respectively (P < 0·01). No significant change in the dissociation constant for [3H]PN200-110 was found. Antisense RNA amplification showed a significant increase in the level of mRNA encoding the L-type Ca2+ channel α1-subunit in IGF-1-treated cells. This study demonstrates that IGF-1 regulates charge movement and the level of L-type Ca2+ channel α1-subunits through activation of gene expression in skeletal muscle cells. PMID:10087334

Overexpression of Transcription Factor Sp1 Leads to Gene Expression Perturbations and Cell Cycle Inhibition

PubMed Central

Deniaud, Emmanuelle; Baguet, Joël; Chalard, Roxane; Blanquier, Bariza; Brinza, Lilia; Meunier, Julien; Michallet, Marie-Cécile; Laugraud, Aurélie; Ah-Soon, Claudette; Wierinckx, Anne; Castellazzi, Marc; Lachuer, Joël; Gautier, Christian

2009-01-01

Background The ubiquitous transcription factor Sp1 regulates the expression of a vast number of genes involved in many cellular functions ranging from differentiation to proliferation and apoptosis. Sp1 expression levels show a dramatic increase during transformation and this could play a critical role for tumour development or maintenance. Although Sp1 deregulation might be beneficial for tumour cells, its overexpression induces apoptosis of untransformed cells. Here we further characterised the functional and transcriptional responses of untransformed cells following Sp1 overexpression. Methodology and Principal Findings We made use of wild-type and DNA-binding-deficient Sp1 to demonstrate that the induction of apoptosis by Sp1 is dependent on its capacity to bind DNA. Genome-wide expression profiling identified genes involved in cancer, cell death and cell cycle as being enriched among differentially expressed genes following Sp1 overexpression. In silico search to determine the presence of Sp1 binding sites in the promoter region of modulated genes was conducted. Genes that contained Sp1 binding sites in their promoters were enriched among down-regulated genes. The endogenous sp1 gene is one of the most down-regulated suggesting a negative feedback loop induced by overexpressed Sp1. In contrast, genes containing Sp1 binding sites in their promoters were not enriched among up-regulated genes. These results suggest that the transcriptional response involves both direct Sp1-driven transcription and indirect mechanisms. Finally, we show that Sp1 overexpression led to a modified expression of G1/S transition regulatory genes such as the down-regulation of cyclin D2 and the up-regulation of cyclin G2 and cdkn2c/p18 expression. The biological significance of these modifications was confirmed by showing that the cells accumulated in the G1 phase of the cell cycle before the onset of apoptosis. Conclusion This study shows that the binding to DNA of overexpressed Sp1

Characterization of the NPC1L1 gene and proteome from an exceptional responder to ezetimibe.

PubMed

Schweitzer, Morris; Makhoul, Sandra; Paliouras, Miltiadis; Beitel, Lenore K; Gottlieb, Bruce; Trifiro, Mark; Chowdhury, Shafinaz F; Zaman, Naif M; Wang, Edwin; Davis, Harry; Chalifour, Lorraine E

2016-03-01

Strategies to reduce LDL-cholesterol involve reductions in cholesterol synthesis or absorption. We identified a familial hypercholesterolemia patient with an exceptional response to the cholesterol absorption inhibitor, ezetimibe. Niemann-Pick C 1-like 1 (NPC1L1) is the molecular target of ezetimibe. Sequencing identified nucleotide changes predicted to change amino acids 52 (L52P), 300 (I300T) and 489 (S489G) in exceptional NPC1L1. In silico analyses identified increased stability and cholesterol binding affinity in L52P-NPC1L1 versus WT-NPC1L1. HEK293 cells overexpressing WT-NPC1L1 or NPC1L1 harboring amino acid changes singly or in combination (Comb-NPC1L1) had reduced cholesterol uptake in Comb-NPC1L1 when ezetimibe was present. Cholesterol uptake was reduced by ezetimibe in L52P-NPC1L1, I300T-NPC1L1, but increased in S489G-NPC1L1 overexpressing cells. Immunolocalization studies found preferential plasma membrane localization of mutant NPC1L1 independent of ezetimibe. Flotillin 1 and 2 expression was reduced and binding to Comb-NPC1L1 was reduced independent of ezetimibe exposure. Proteomic analyses identified increased association with proteins that modulate intermediate filament proteins in Comb-NPC1L1 versus WT-NPC1L1 treated with ezetimibe. This is the first detailed analysis of the role of NPC1L1 mutations in an exceptional responder to ezetimibe. The results point to a complex set of events in which the combined mutations were shown to affect cholesterol uptake in the presence of ezetimibe. Proteomic analysis suggests that the exceptional response may also lie in the nature of interactions with cytosolic proteins. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Molecular characterization and expression analysis of Zar1 and Zar1-like genes in rainbow trout

USDA-ARS?s Scientific Manuscript database

Zygote arrest 1 (Zar1) is a maternal effect gene that is essential for early embryonic development. Recently, a novel gene called Zar1-like (Zar1l) was discovered. Functional studies showed that ZAR1L plays an important role in regulating oocyte-to-embryo transition in mouse. The objectives of this ...

Expression of L1-CAM and ADAM10 in human colon cancer cells induces metastasis.

PubMed

Gavert, Nancy; Sheffer, Michal; Raveh, Shani; Spaderna, Simone; Shtutman, Michael; Brabletz, Thomas; Barany, Francis; Paty, Phillip; Notterman, Daniel; Domany, Eytan; Ben-Ze'ev, Avri

2007-08-15

L1-CAM, a neuronal cell adhesion receptor, is also expressed in a variety of cancer cells. Recent studies identified L1-CAM as a target gene of beta-catenin-T-cell factor (TCF) signaling expressed at the invasive front of human colon cancer tissue. We found that L1-CAM expression in colon cancer cells lacking L1-CAM confers metastatic capacity, and mice injected in their spleen with such cells form liver metastases. We identified ADAM10, a metalloproteinase that cleaves the L1-CAM extracellular domain, as a novel target gene of beta-catenin-TCF signaling. ADAM10 overexpression in colon cancer cells displaying endogenous L1-CAM enhanced L1-CAM cleavage and induced liver metastasis, and ADAM10 also enhanced metastasis in colon cancer cells stably transfected with L1-CAM. DNA microarray analysis of genes induced by L1-CAM in colon cancer cells identified a cluster of genes also elevated in a large set of human colon carcinoma tissue samples. Expression of these genes in normal colon epithelium was low. These results indicate that there is a gene program induced by L1-CAM in colon cancer cells that is also present in colorectal cancer tissue and suggest that L1-CAM can serve as target for colon cancer therapy.

Antiproliferative activity of aqueous leaf extract of Annona muricata L. on the prostate, BPH-1 cells, and some target genes.

PubMed

Asare, George Awuku; Afriyie, Dan; Ngala, Robert A; Abutiate, Harry; Doku, Derek; Mahmood, Seidu A; Rahman, Habibur

2015-01-01

Annona muricata L. has been reported to possess antitumor and antiproliferative properties. Not much work has been done on its effect on BPH-1 cell lines, and no in vivo studies targeting the prostate organ exist. The study determined the effect of A muricata on human BPH-1 cells and prostate organ. The MTT assay was performed on BPH-1 cells using the aqueous leaf extract of A muricata. Cells (1 × 10(5) per well) were challenged with 0.5, 1.0, and 1.5 mg/mL extract for 24, 48, and 72 hours. Cell proliferation and morphology were examined microscopically. BPH-1 cells (1 × 10(4) per well) were seeded into 6-well plates and incubated for 48 hours with 0.5, 1.0, and 1.5 mg/mL A muricata extract. Reverse transcriptase polymerase chain reaction was performed using mRNA extracted from the cells. Possible target genes, Bax and Bcl-2, were examined. Twenty F344 male rats (≈200 g) were gavaged 30 mg/mL (10 rats) and 300 mg/mL (10 rats) and fed ad libitum alongside 10 control rats. Rats were sacrificed after 60 days. The prostate, seminal vesicles, and testes were harvested for histological examination. Annona muricata demonstrated antiproliferative effects with an IC50 of 1.36 mg/mL. Best results were obtained after 48 hours, with near cell extinction at 72 hours. Bax gene was upregulated, while Bcl-2 was downregulated. Normal histological architecture was observed for all testes. Seminal vesicle was significantly reduced in test groups (P < .05) and demonstrated marked atrophy with increased cellularity and the acinii, empty of secretion. Prostate of test groups were reduced with epithelial lining showing pyknotic nucleus, condensation, and marginalization of the nuclear material, characteristic of apoptosis of the glandular epithelium. Furthermore, scanty prostatic secretion with flattening of acinar epithelial lining occurred. Annona muricata has antiproliferative effects on BPH-1 cells and reduces prostate size, possibly through apoptosis. © The Author(s) 2014.

Expression pattern of L-FABP gene in different tissues and its regulation of fat metabolism-related genes in duck.

PubMed

He, Jun; Tian, Yong; Li, Jinjun; Shen, Junda; Tao, Zhengrong; Fu, Yan; Niu, Dong; Lu, Lizhi

2013-01-01

Liver fatty acid binding protein (L-FABP) is a member of intracellular lipid-binding proteins responsible for the transportation of fatty acids. The expression pattern of duck L-FABP mRNA was examined in this study by quantitative RT-PCR. The results showed that duck L-FABP gene was expressed in many tissues, including heart, lung, kidney, muscle, ovary, brain, intestine, stomach and adipocyte tissues, and highly expressed in liver. Several lipid metabolism-related genes were selected to detect the regulation of L-FABP in duck. The expression of L-FABP and lipoprotein lipase was promoted by oleic acid. The L-FABP knockdown decreased the expression levels of peroxisome proliferator-activated receptor α (PPARα), fatty acid synthase and lipoprotein lipase by 61.1, 42.3 and 53.7 %, respectively (P < 0.05), but had no influences on the mRNA levels of PPARγ and leptin receptor. L-FABP might function through the PPARα to regulate the fat metabolism-related gene expression and play important roles in lipid metabolism in duck hepatocytes.

Validation of reference genes for quantitative RT-PCR studies of gene expression in perennial ryegrass (Lolium perenne L.)

PubMed Central

2010-01-01

Background Perennial ryegrass (Lolium perenne L.) is an important pasture and turf crop. Biotechniques such as gene expression studies are being employed to improve traits in this temperate grass. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) is among the best methods available for determining changes in gene expression. Before analysis of target gene expression, it is essential to select an appropriate normalisation strategy to control for non-specific variation between samples. Reference genes that have stable expression at different biological and physiological states can be effectively used for normalisation; however, their expression stability must be validated before use. Results Existing Serial Analysis of Gene Expression data were queried to identify six moderately expressed genes that had relatively stable gene expression throughout the year. These six candidate reference genes (eukaryotic elongation factor 1 alpha, eEF1A; TAT-binding protein homolog 1, TBP-1; eukaryotic translation initiation factor 4 alpha, eIF4A; YT521-B-like protein family protein, YT521-B; histone 3, H3; ubiquitin-conjugating enzyme, E2) were validated for qRT-PCR normalisation in 442 diverse perennial ryegrass (Lolium perenne L.) samples sourced from field- and laboratory-grown plants under a wide range of experimental conditions. Eukaryotic EF1A is encoded by members of a multigene family exhibiting differential expression and necessitated the expression analysis of different eEF1A encoding genes; a highly expressed eEF1A (h), a moderately, but stably expressed eEF1A (s), and combined expression of multigene eEF1A (m). NormFinder identified eEF1A (s) and YT521-B as the best combination of two genes for normalisation of gene expression data in perennial ryegrass following different defoliation management in the field. Conclusions This study is unique in the magnitude of samples tested with the inclusion of numerous field-grown samples, helping pave the way to

Novel roles for LIX1L in promoting cancer cell proliferation through ROS1-mediated LIX1L phosphorylation

PubMed Central

Nakamura, Satoki; Kahyo, Tomoaki; Tao, Hong; Shibata, Kiyoshi; Kurabe, Nobuya; Yamada, Hidetaka; Shinmura, Kazuya; Ohnishi, Kazunori; Sugimura, Haruhiko

2015-01-01

Herein, we report the characterization of Limb expression 1-like, (LIX1L), a putative RNA-binding protein (RBP) containing a double-stranded RNA binding motif, which is highly expressed in various cancer tissues. Analysis of MALDI-TOF/TOF mass spectrometry and RNA immunoprecipitation-sequencing of interacting proteins and the microRNAs (miRNAs) bound to LIX1L revealed that LIX1L interacts with proteins (RIOK1, nucleolin and PABPC4) and miRNAs (has-miRNA-520a-5p, −300, −216b, −326, −190a, −548b-3p, −7–5p and −1296) in HEK-293 cells. Moreover, the reduction of phosphorylated Tyr136 (pTyr136) in LIX1L through the homeodomain peptide, PY136, inhibited LIX1L-induced cell proliferation in vitro, and PY136 inhibited MKN45 cell proliferation in vivo. We also determined the miRNA-targeted genes and showed that was apoptosis induced through the reduction of pTyr136. Moreover, ROS1, HCK, ABL1, ABL2, JAK3, LCK and TYR03 were identified as candidate kinases responsible for the phosphorylation of Tyr136 of LIX1L. These data provide novel insights into the biological significance of LIX1L, suggesting that this protein might be an RBP, with implications for therapeutic approaches for targeting LIX1L in LIX1L-expressing cancer cells. PMID:26310847

Lead phytoextraction from printed circuit computer boards by Lolium perenne L. and Medicago sativa L.

PubMed

Díaz Martínez, María Esther; Argumedo-Delira, Rosalba; Sánchez Viveros, Gabriela; Alarcón, Alejandro; Trejo-Téllez, Libia Iris

2018-04-16

This work assessed the ability of Lolium perenne and Medicago sativa for extracting lead (Pb) from particulate printed circuit computer boards (PCB) mixed in sand with the following concentrations: 0.5, 1.0 and 1.5 g of PCB, and including a control treatment without PCB. The PCB were obtained from computers, and grinded in two particle sizes: 0.0594 mm (PCB1) and 0.0706 mm (PCB2). The PCB particle sizes at their corresponding concentrations were applied to L. perenne and M. sativa by using three experimental assays. In assay II, PCB2 affected the biomass production for both plants. For assay III, the PCB1 increased the biomass of M. sativa (236.5%) and L. perenne (142.2%) when applying either 0.5 or 1.0 g, respectively. In regards to phytoextraction, assay I showed the highest Pb-extraction by roots of L. perenne (4.7%) when exposed to 1.5 g of PCB1. At assay I, L. perenne showed a Pb-bioconcentration factor higher than 1.0 when growing at 0.5 g of PCB1, and when HNO 3 was used as digestion solution; moreover, in assay III both plants showed a Pb-translocation factor higher than 1.0. Therefore, Lolium perenne and Medicago sativa are able to recover Pb from electronic wastes (PCB).

A guanine insert in OsBBS1 leads to early leaf senescence and salt stress sensitivity in rice (Oryza sativa L.).

PubMed

Zeng, Dong-Dong; Yang, Cheng-Cong; Qin, Ran; Alamin, Md; Yue, Er-Kui; Jin, Xiao-Li; Shi, Chun-Hai

2018-06-01

A rice receptor-like kinase gene OSBBS1/OsRLCK109 was identified; this gene played vital roles in leaf senescence and the salt stress response. Early leaf senescence can cause negative effects on rice yield, but the underlying molecular regulation is not fully understood. bilateral blade senescence 1 (bbs1), an early leaf senescence mutant with a premature senescence phenotype that occurs mainly performing at the leaf margins, was isolated from a rice mutant population generated by ethylmethane sulfonate (EMS) treatment. The mutant showed premature leaf senescence beginning at the tillering stage and exhibited severe symptoms at the late grain-filling stage. bbs1 showed accelerated dark-induced leaf senescence. The OsBBS1 gene was cloned by a map-based cloning strategy, and a guanine (G) insertion was found in the first exon of LOC_Os03g24930. This gene encodes a receptor-like cytoplasmic kinase and was named OsRLCK109 in a previous study. Transgenic LOC_Os03g24930 knockout plants generated by a CRISPR/Cas9 strategy exhibited similar early leaf senescence phenotypes as did the bbs1 mutant, which confirmed that LOC_Os03g24930 was the OsBBS1 gene. OsBBS1/OsRLCK109 was expressed in all detected tissues and was predominantly expressed in the main vein region of mature leaves. The expression of OsBBS1 could be greatly induced by salt stress, and the bbs1 mutant exhibited hypersensitivity to salt stress. In conclusion, this is the first identification of OsRLCKs participating in leaf senescence and playing critical roles in the salt stress response in rice (Oryza sativa L.).

Overexpression of Arabidopsis Molybdenum Cofactor Sulfurase Gene Confers Drought Tolerance in Maize (Zea mays L.)

PubMed Central

Zhang, Jiachang; Xiao, Yitao; Yue, Yuesen; Duan, Liusheng; Zhang, Mingcai; Li, Zhaohu

2013-01-01

Abscisic acid (ABA) is a key component of the signaling system that integrates plant adaptive responses to abiotic stress. Overexpression of Arabidopsis molybdenum cofactor sulfurase gene (LOS5) in maize markedly enhanced the expression of ZmAO and aldehyde oxidase (AO) activity, leading to ABA accumulation and increased drought tolerance. Transgenic maize (Zea mays L.) exhibited the expected reductions in stomatal aperture, which led to decreased water loss and maintenance of higher relative water content (RWC) and leaf water potential. Also, transgenic maize subjected to drought treatment exhibited lower leaf wilting, electrolyte leakage, malondialdehyde (MDA) and H2O2 content, and higher activities of antioxidative enzymes and proline content compared to wild-type (WT) maize. Moreover, overexpression of LOS5 enhanced the expression of stress-regulated genes such as Rad 17, NCED1, CAT1, and ZmP5CS1 under drought stress conditions, and increased root system development and biomass yield after re-watering. The increased drought tolerance in transgenic plants was associated with ABA accumulation via activated AO and expression of stress-related gene via ABA induction, which sequentially induced a set of favorable stress-related physiological and biochemical responses. PMID:23326325

Overexpression of Arabidopsis molybdenum cofactor sulfurase gene confers drought tolerance in maize (Zea mays L.).

PubMed

Lu, Yao; Li, Yajun; Zhang, Jiachang; Xiao, Yitao; Yue, Yuesen; Duan, Liusheng; Zhang, Mingcai; Li, Zhaohu

2013-01-01

Abscisic acid (ABA) is a key component of the signaling system that integrates plant adaptive responses to abiotic stress. Overexpression of Arabidopsis molybdenum cofactor sulfurase gene (LOS5) in maize markedly enhanced the expression of ZmAO and aldehyde oxidase (AO) activity, leading to ABA accumulation and increased drought tolerance. Transgenic maize (Zea mays L.) exhibited the expected reductions in stomatal aperture, which led to decreased water loss and maintenance of higher relative water content (RWC) and leaf water potential. Also, transgenic maize subjected to drought treatment exhibited lower leaf wilting, electrolyte leakage, malondialdehyde (MDA) and H(2)O(2) content, and higher activities of antioxidative enzymes and proline content compared to wild-type (WT) maize. Moreover, overexpression of LOS5 enhanced the expression of stress-regulated genes such as Rad 17, NCED1, CAT1, and ZmP5CS1 under drought stress conditions, and increased root system development and biomass yield after re-watering. The increased drought tolerance in transgenic plants was associated with ABA accumulation via activated AO and expression of stress-related gene via ABA induction, which sequentially induced a set of favorable stress-related physiological and biochemical responses.

Two Novel Mutations in the GDAP1 and PRX Genes in Early Onset Charcot-Marie-Tooth Syndrome

PubMed Central

Auer-Grumbach, M.; Fischer, C.; Papić, L.; John, E.; Plecko, B.; Bittner, R. E.; Bernert, G.; Pieber, T. R.; Miltenberger, G.; Schwarz, R.; Windpassinger, C.; Grill, F.; Timmerman, V.; Speicher, M. R.; Janecke, A. R.

2011-01-01

Autosomal recessive Charcot-Marie-Tooth syndrome (AR-CMT) is often characterised by an infantile disease onset and a severe phenotype. Mutations in the ganglioside-induced differentiation-associated protein 1 (GDAP1) gene are thought to be a common cause of AR-CMT. Mutations in the periaxin (PRX) gene are rare. They are associated with severe demyelination of the peripheral nerves and sometimes lead to prominent sensory disturbances. To evaluate the frequency of GDAP1 and PRX mutations in early onset CMT, we examined seven AR-CMT families and 12 sporadic CMT patients, all presenting with progressive distal muscle weakness and wasting. In one family also prominent sensory abnormalities and sensory ataxia were apparent from early childhood. In three families we detected four GDAP1 mutations (L58LfsX4, R191X, L239F and P153L), one of which is novel and is predicted to cause a loss of protein function. In one additional family with prominent sensory abnormalities a novel homozygous PRX mutation was found (A700PfsX17). No mutations were identified in 12 sporadic cases. This study suggests that mutations in the GDAP1 gene are a common cause of early-onset AR-CMT. In patients with early-onset demyelinating AR-CMT and severe sensory loss PRX is one of the genes to be tested. PMID:18504680

PiiL: visualization of DNA methylation and gene expression data in gene pathways.

PubMed

Moghadam, Behrooz Torabi; Zamani, Neda; Komorowski, Jan; Grabherr, Manfred

2017-08-02

DNA methylation is a major mechanism involved in the epigenetic state of a cell. It has been observed that the methylation status of certain CpG sites close to or within a gene can directly affect its expression, either by silencing or, in some cases, up-regulating transcription. However, a vertebrate genome contains millions of CpG sites, all of which are potential targets for methylation, and the specific effects of most sites have not been characterized to date. To study the complex interplay between methylation status, cellular programs, and the resulting phenotypes, we present PiiL, an interactive gene expression pathway browser, facilitating analyses through an integrated view of methylation and expression on multiple levels. PiiL allows for specific hypothesis testing by quickly assessing pathways or gene networks, where the data is projected onto pathways that can be downloaded directly from the online KEGG database. PiiL provides a comprehensive set of analysis features that allow for quick and specific pattern searches. Individual CpG sites and their impact on host gene expression, as well as the impact on other genes present in the regulatory network, can be examined. To exemplify the power of this approach, we analyzed two types of brain tumors, Glioblastoma multiform and lower grade gliomas. At a glance, we could confirm earlier findings that the predominant methylation and expression patterns separate perfectly by mutations in the IDH genes, rather than by histology. We could also infer the IDH mutation status for samples for which the genotype was not known. By applying different filtering methods, we show that a subset of CpG sites exhibits consistent methylation patterns, and that the status of sites affect the expression of key regulator genes, as well as other genes located downstream in the same pathways. PiiL is implemented in Java with focus on a user-friendly graphical interface. The source code is available under the GPL license from https://github.com/behroozt/PiiL.git .

Acetate alters expression of genes involved in beige adipogenesis in 3T3-L1 cells and obese KK-Ay mice

PubMed Central

Hanatani, Satoko; Motoshima, Hiroyuki; Takaki, Yuki; Kawasaki, Shuji; Igata, Motoyuki; Matsumura, Takeshi; Kondo, Tatsuya; Senokuchi, Takafumi; Ishii, Norio; Kawashima, Junji; Kukidome, Daisuke; Shimoda, Seiya; Nishikawa, Takeshi; Araki, Eiichi

2016-01-01

The induction of beige adipogenesis within white adipose tissue, known as “browning”, has received attention as a novel potential anti-obesity strategy. The expression of some characteristic genes including PR domain containing 16 is induced during the browning process. Although acetate has been reported to suppress weight gain in both rodents and humans, its potential effects on beige adipogenesis in white adipose tissue have not been fully characterized. We examined the effects of acetate treatment on 3T3-L1 cells and in obese diabetic KK-Ay mice. The mRNA expression levels of genes involved in beige adipocyte differentiation and genes selectively expressed in beige adipocytes were significantly elevated in both 3T3-L1 cells incubated with 1.0 mM acetate and the visceral white adipose tissue from mice treated with 0.6% acetate for 16 weeks. In KK-Ay mice, acetate reduced the food efficiency ratio and increased the whole-body oxygen consumption rate. Additionally, reduction of adipocyte size and uncoupling protein 1-positive adipocytes and interstitial areas with multilocular adipocytes appeared in the visceral white adipose tissue of acetate-treated mice, suggesting that acetate induced initial changes of “browning”. In conclusion, acetate alters the expression of genes involved in beige adipogenesis and might represent a potential therapeutic agent to combat obesity. PMID:27895388

Genes encoding Xenopus laevis Ig L chains: Implications for the evolution of [kappa] and [lambda] chains

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

Zezza, D.J.; Stewart, S.E.; Steiner, L.A.

1992-12-15

Xenopus laevis Ig contain two distinct types of L chains, designated [rho] or L1 and [sigma] or L2. The authors have analyzed Xenopus genomic DNA by Southern blotting with cDNA probes specific for L1 V and C regions. Many fragments hybridized to the V probe, but only one or two fragments hybridized to the C probe. Corresponding C, J, and V gene segments were identified on clones isolated from a genomic library prepared from the same DNA. One clone contains a C gene segment separated from a J gene segment by an intron of 3.4 kb. The J and Cmore » gene segments are nearly identical in sequence to cDNA clones analyzed previously. The C segment is somewhat more similar and the J segment considerably more similar in sequence to the corresponding segments of mammalian [kappa] chains than to those of mammalian [lambda] chains. Upstream of the J segment is a typical recombination signal sequence with a spacer of 23 bp, as in J[kappa]. A second clone from the library contains four V gene segments, separated by 2.1 to 3.6 kb. Two of these, V1 and V3, have the expected structural and regulatory features of V genes, and are very similar in sequence to each other and to mammalian V[kappa]. A third gene segment, V2, resembles V1 and V3 in its coding region and nearby 5[prime]-flanking region, but diverges in sequence 5[prime] to position [minus]95 with loss of the octamer promoter element. The fourth V-like segment is similar to the others at the 3[prime]-end, but upstream of codon 64 bears no resemblance in sequence to any Ig V region. All four V segments have typical recombination signal sequences with 12-bp spacers at their 3[prime]-ends, as in V[kappa]. Taken together, the data suggest that Xenopus L1 L chain genes are members of the [kappa] gene family. 80 refs., 9 figs.« less

[Reconstruction of Leptospira interrogans lipL21 gene and characteristics of its expression product].

PubMed

Luo, Dong-jiao; Hu, Ye; Dennin, R H; Yan, Jie

2007-09-01

To reconstruct the nucleotide sequence of Leptospira interrogans lipL21 gene for increasing the output of prokaryotic expression and to understand the changes on immunogenicity of the expression products before and after reconstruction, and to determine the position of envelope lipoprotein LipL21 on the surface of leptospiral body. According to the preferred codons of E.coli, the nucleotide sequence of lipL21 gene was designed and synthesized, and then its prokaryotic expression system was constructed. By using SDS-PAGE plus BioRad agarose image analysor, the expression level changes of lipL21 genes before and after reconstruction were measured. A Western blot assay using rabbit anti-TR/Patoc I serum as the first antibody was performed to identify the immunoreactivity of the two target recombinant proteins rLipL21s before and after reconstruction. The changes of cross agglutination titers of antisera against two rLipL21s before and after reconstruction to the different leptospiral serogroups were demonstrated using microscope agglutination test (MAT). Immuno-electronmicroscopy was applied to confirm the location of LipL21s. The expression outputs of original and reconstructed lipL21 genes were 8.5 % and 46.5 % of the total bacterial proteins, respectively. Both the two rLipL21s could take place immune conjugation reaction with TR/Patoc I antiserum. After immunization with each of the two rLipL21s in rabbits, the animals could produce specific antibody. Similar MAT titers with 1:80 - 1:320 of the two antisera against rLipL21s were present. LipL21 was confirmed to locate on the surface of leptospiral envelope. LipL21 is a superficial antigen of Leptospira interrogans. The expression output of the reconstructed lipL21 gene is remarkably increased. The expression rLipL21 maintains fine antigenicity and immunoreactivity and its antibody still shows an extensive cross immunoagglutination activity. The high expression of the reconstructed lipL21 gene will offer a

EBV+ and MSI Gastric Cancers Harbor High PD-L1/PD-1 Expression and High CD8+ Intratumoral Lymphocytes

PubMed Central

De Rosa, Simona; Tibiletti, Maria Grazia; Magnoli, Francesca; Vanoli, Alessandro; Sessa, Fausto; Chiaravalli, Anna Maria

2018-01-01

Both EBV+ and MSI gastric cancers (GCs) have high lymphoid infiltration which is rare in MSS/EBV− cancers. PD-L1/PD-1 interaction leads to a down-regulated immune response and it is one of the most promising targets for gastric cancer immunotherapy. PD-L1/PD-1 and CD8 expression were immunohistochemically investigated in a series of 169 FFPE GCs, including 33 EBV+, 59 MSI and 77 MSS/EBV− cases. PD-L1 membrane immunoreactivity in more than 5% of tumor cells was present in 31/169 GCs and was associated with high levels of CD8 intraepithelial lymphocytes (TILs; p < 0.001). PD-L1+ cases were mainly poorly differentiated (71%), intestinal type (85%) and high lymphoid response (HLR; 90%) tumors. PD-L1 expression was only present in EBV⁺ (46%), MSI (24%) and rare MSS/EBV− (3%) GCs with high CD8+ TILs (p < 0.001). Despite being associated with a better prognosis both in the whole series (p < 0.05) and in the MSI subset, PD-L1 is not an independent prognostic factor. PD-L1 gene amplification was detected in 3/17 cases, including 2/7 EBV+ and 1/8 MSI GC. PD-1⁺ TILs were significantly higher in EBV⁺ than MSI and MSS/EBV− cases. PD-L1/PD-1 pathway is selectively activated in HLR GCs and could be considered an emerging therapeutic target, particularly for EBV and MSI GCs. PMID:29614789

Ectopic overexpression of the cell wall invertase gene CIN1 leads to dehydration avoidance in tomato.

PubMed

Albacete, Alfonso; Cantero-Navarro, Elena; Großkinsky, Dominik K; Arias, Cintia L; Balibrea, María Encarnación; Bru, Roque; Fragner, Lena; Ghanem, Michel E; González, María de la Cruz; Hernández, Jose A; Martínez-Andújar, Cristina; van der Graaff, Eric; Weckwerth, Wolfram; Zellnig, Günther; Pérez-Alfocea, Francisco; Roitsch, Thomas

2015-02-01

Drought stress conditions modify source-sink relations, thereby influencing plant growth, adaptive responses, and consequently crop yield. Invertases are key metabolic enzymes regulating sink activity through the hydrolytic cleavage of sucrose into hexose monomers, thus playing a crucial role in plant growth and development. However, the physiological role of invertases during adaptation to abiotic stress conditions is not yet fully understood. Here it is shown that plant adaptation to drought stress can be markedly improved in tomato (Solanum lycopersicum L.) by overexpression of the cell wall invertase (cwInv) gene CIN1 from Chenopodium rubrum. CIN1 overexpression limited stomatal conductance under normal watering regimes, leading to reduced water consumption during the drought period, while photosynthetic activity was maintained. This caused a strong increase in water use efficiency (up to 50%), markedly improving water stress adaptation through an efficient physiological strategy of dehydration avoidance. Drought stress strongly reduced cwInv activity and induced its proteinaceous inhibitor in the leaves of the wild-type plants. However, the CIN1-overexpressing plants registered 3- to 6-fold higher cwInv activity in all analysed conditions. Surprisingly, the enhanced invertase activity did not result in increased hexose concentrations due to the activation of the metabolic carbohydrate fluxes, as reflected by the maintenance of the activity of key enzymes of primary metabolism and increased levels of sugar-phosphate intermediates under water deprivation. The induced sink metabolism in the leaves explained the maintenance of photosynthetic activity, delayed senescence, and increased source activity under drought stress. Moreover, CIN1 plants also presented a better control of production of reactive oxygen species and sustained membrane protection. Those metabolic changes conferred by CIN1 overexpression were accompanied by increases in the concentrations of the

A Crohn's disease variant in Atg16l1 enhances its degradation by caspase 3

NASA Astrophysics Data System (ADS)

Murthy, Aditya; Li, Yun; Peng, Ivan; Reichelt, Mike; Katakam, Anand Kumar; Noubade, Rajkumar; Roose-Girma, Merone; Devoss, Jason; Diehl, Lauri; Graham, Robert R.; van Lookeren Campagne, Menno

2014-02-01

Crohn's disease is a debilitating inflammatory bowel disease (IBD) that can involve the entire digestive tract. A single-nucleotide polymorphism (SNP) encoding a missense variant in the autophagy gene ATG16L1 (rs2241880, Thr300Ala) is strongly associated with the incidence of Crohn's disease. Numerous studies have demonstrated the effect of ATG16L1 deletion or deficiency; however, the molecular consequences of the Thr300Ala (T300A) variant remains unknown. Here we show that amino acids 296-299 constitute a caspase cleavage motif in ATG16L1 and that the T300A variant (T316A in mice) significantly increases ATG16L1 sensitization to caspase-3-mediated processing. We observed that death-receptor activation or starvation-induced metabolic stress in human and murine macrophages increased degradation of the T300A or T316A variants of ATG16L1, respectively, resulting in diminished autophagy. Knock-in mice harbouring the T316A variant showed defective clearance of the ileal pathogen Yersinia enterocolitica and an elevated inflammatory cytokine response. In turn, deletion of the caspase-3-encoding gene, Casp3, or elimination of the caspase cleavage site by site-directed mutagenesis rescued starvation-induced autophagy and pathogen clearance, respectively. These findings demonstrate that caspase 3 activation in the presence of a common risk allele leads to accelerated degradation of ATG16L1, placing cellular stress, apoptotic stimuli and impaired autophagy in a unified pathway that predisposes to Crohn's disease.

Isolation of the endosperm-specific LPAAT gene promoter from coconut (Cocos nucifera L.) and its functional analysis in transgenic rice plants.

PubMed

Xu, Li; Ye, Rongjian; Zheng, Yusheng; Wang, Zhekui; Zhou, Peng; Lin, Yongjun; Li, Dongdong

2010-09-01

As one of the key tropical crops, coconut (Cocos nucifera L.) is a member of the monocotyledonous family Aracaceae (Palmaceae). In this study, we amplified the upstream region of an endosperm-specific expression gene, Lysophosphatidyl acyltransferase (LPAAT), from the coconut genomic DNA by chromosome walking. In this sequence, we found several types of promoter-related elements including TATA-box, CAAT-box and Skn1-motif. In order to further examine its function, three different 5'-deletion fragments were inserted into pBI101.3, a plant expression vector harboring the LPAAT upstream sequence, leading to pBI101.3-L1, pBI101.3-L2 and pBI101.3-L3, respectively. We obtained transgenic plants of rice by Agrobacterium-mediated callus transformation and plant regeneration and detected the expression of gus gene by histochemical staining and fluorometric determination. We found that gus gene driven by the three deletion fragments was specifically expressed in the endosperm of rice seeds, but not in the empty vector of pBI101.3 and other tissues. The highest expression level of GUS was at 15 DAF in pBI101.3-L3 and pBI101.3-L2 transgenic lines, while the same level was detected at 10 DAF in pBI101.3-L1. The expression driven by the whole fragment was up to 1.76- and 2.8-fold higher than those driven by the -817 bp and -453 bp upstream fragments, and 10.7-fold higher than that driven by the vector without the promoter. Taken together, our results strongly suggest that these promoter fragments from coconut have a significant potential in genetically improving endosperm in main crops.

A novel Zea mays ssp. mexicana L. MYC-type ICE-like transcription factor gene ZmmICE1, enhances freezing tolerance in transgenic Arabidopsis thaliana.

PubMed

Lu, Xiang; Yang, Lei; Yu, Mengyuan; Lai, Jianbin; Wang, Chao; McNeil, David; Zhou, Meixue; Yang, Chengwei

2017-04-01

The annual Zea mays ssp. mexicana L., a member of the teosinte group, is a close wild relative of maize and thus can be effectively used in maize improvement. In this study, an ICE-like gene, ZmmICE1, was isolated from a cDNA library of RNA-Seq from cold-treated seedling tissues of Zea mays ssp. mexicana L. The deduced protein of ZmmICE1 contains a highly conserved basic helix-loop-helix (bHLH) domain and C-terminal region of ICE-like proteins. The ZmmICE1 protein localizes to the nucleus and shows sumoylation when expressed in an Escherichia coli reconstitution system. In addition, yeast one hybrid assays indicated that ZmmICE1 has transactivation activities. Moreover, ectopic expression of ZmmICE1 in the Arabidopsis ice1-2 mutant increased freezing tolerance. The ZmmICE1 overexpressed plants showed lower electrolyte leakage (EL), reduced contents of malondialdehyde (MDA). The expression of downstream cold related genes of Arabidopsis C-repeat-binding factors (AtCBF1, AtCBF2 and AtCBF3), cold-responsive genes (AtCOR15A and AtCOR47), kinesin-1 member gene (AtKIN1) and responsive to desiccation gene (AtRD29A) was significantly induced when compared with wild type under low temperature treatment. Taken together, these results indicated that ZmmICE1 is the homolog of Arabidopsis inducer of CBF expression genes (AtICE1/2) and plays an important role in the regulation of freezing stress response. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

The analysis of blood lead levels changeability over the 5-year observation in workers occupationally exposed to lead.

PubMed

Dobrakowski, Michał; Boroń, Marta; Kasperczyk, Sławomir; Kozłowska, Agnieszka; Kasperczyk, Aleksandra; Płachetka, Anna; Pawlas, Natalia

2017-06-01

The aim of the present study was to compare a group of workers with stable lead levels with a group of workers with fluctuating lead levels in terms of selected hematological, biochemical, and immunological parameters. The examined group included male workers occupationally exposed to lead. Blood lead (PbB) levels were measured every 3 months during the 5-year observation. Based on standard deviation of mean PbB levels, the examined population was divided into two groups: low level of fluctuation (L-SD) and high level of fluctuation (H-SD) groups. The mean and maximal PbB levels were significantly higher in the H-SD group than in the L-SD group by 9 and 22%, respectively. At the same time, the maximal level of zinc protoporphyrin (ZPP) and standard deviation of mean ZPP level were higher in the H-SD group by 29 and 55%, respectively. The maximal level of hemoglobin and white blood cell (WBC) count as well as standard deviation of the mean hemoglobin level and WBC count were higher in the H-SD group by 2, 8, 58, and 24%, respectively. The expression of nuclear factor kappa-B1 gene and telomerase reverse transcriptase gene was significantly greater in the H-SD group than in the L-SD group by 11 and 28%, respectively. Workers occupationally exposed to lead do not represent a homogenous population. Some present stable lead levels, whereas others have fluctuating lead levels. These fluctuations are related to secondary changes in ZPP and hemoglobin levels as well as WBC count.

De novo and rare mutations in the HSPA1L heat shock gene associated with inflammatory bowel disease.

PubMed

Takahashi, Shinichi; Andreoletti, Gaia; Chen, Rui; Munehira, Yoichi; Batra, Akshay; Afzal, Nadeem A; Beattie, R Mark; Bernstein, Jonathan A; Ennis, Sarah; Snyder, Michael

2017-01-26

Inflammatory bowel disease (IBD) is a chronic, relapsing inflammatory disease of the gastrointestinal tract which includes ulcerative colitis and Crohn's disease. Genetic risk factors for IBD are not well understood. We performed a family-based whole exome sequencing (WES) analysis on a core family (Family A) to identify potential causal mutations and then analyzed exome data from a Caucasian pediatric cohort (136 patients and 106 controls) to validate the presence of mutations in the candidate gene, heat shock 70 kDa protein 1-like (HSPA1L). Biochemical assays of the de novo and rare (minor allele frequency, MAF < 0.01) mutation variant proteins further validated the predicted deleterious effects of the identified alleles. In the proband of Family A, we found a heterozygous de novo mutation (c.830C > T; p.Ser277Leu) in HSPA1L. Through analysis of WES data of 136 patients, we identified five additional rare HSPA1L mutations (p.Gly77Ser, p.Leu172del, p.Thr267Ile, p.Ala268Thr, p.Glu558Asp) in six patients. In contrast, rare HSPA1L mutations were not observed in controls, and were significantly enriched in patients (P = 0.02). Interestingly, we did not find non-synonymous rare mutations in the HSP70 isoforms HSPA1A and HSPA1B. Biochemical assays revealed that all six rare HSPA1L variant proteins showed decreased chaperone activity in vitro. Moreover, three variants demonstrated dominant negative effects on HSPA1L and HSPA1A protein activity. Our results indicate that de novo and rare mutations in HSPA1L are associated with IBD and provide insights into the pathogenesis of IBD, and also expand our understanding of the roles of HSP70s in human disease.

Isolation and characterization of cbbL and cbbS genes encoding form I ribulose-1,5-bisphosphate carboxylase/oxygenase large and small subunits in Nitrosomonas sp. strain ENI-11.

PubMed

Hirota, Ryuichi; Kato, Junichi; Morita, Hiromu; Kuroda, Akio; Ikeda, Tsukasa; Takiguchi, Noboru; Ohtake, Hisao

2002-03-01

The cbbL and cbbS genes encoding form I ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO) large and small subunits in the ammonia-oxidizing bacterium Nitrosomonas sp. strain ENI-11 were cloned and sequenced. The deduced gene products, CbbL and CbbS, had 93 and 87% identity with Thiobacillus intermedius CbbL and Nitrobacter winogradskyi CbbS, respectively. Expression of cbbL and cbbS in Escherichia coli led to the detection of RubisCO activity in the presence of 0.1 mM isopropyl-beta-D-thiogalactopyranoside (IPTG). To our knowledge, this is the first paper to report the genes involved in the carbon fixation reaction in chemolithotrophic ammonia-oxidizing bacteria.

Transcriptome Profiling of Radish (Raphanus sativus L.) Root and Identification of Genes Involved in Response to Lead (Pb) Stress with Next Generation Sequencing

PubMed Central

Wang, Yan; Xu, Liang; Chen, Yinglong; Shen, Hong; Gong, Yiqin; Limera, Cecilia; Liu, Liwang

2013-01-01

Lead (Pb), one of the most toxic heavy metals, can be absorbed and accumulated by plant roots and then enter the food chain resulting in potential health risks for human beings. The radish (Raphanus sativus L.) is an important root vegetable crop with fleshy taproots as the edible parts. Little is known about the mechanism by which radishes respond to Pb stress at the molecular level. In this study, Next Generation Sequencing (NGS)–based RNA-seq technology was employed to characterize the de novo transcriptome of radish roots and identify differentially expressed genes (DEGs) during Pb stress. A total of 68,940 assembled unique transcripts including 33,337 unigenes were obtained from radish root cDNA samples. Based on the assembled de novo transcriptome, 4,614 DEGs were detected between the two libraries of untreated (CK) and Pb-treated (Pb1000) roots. Gene Ontology (GO) and pathway enrichment analysis revealed that upregulated DEGs under Pb stress are predominately involved in defense responses in cell walls and glutathione metabolism-related processes, while downregulated DEGs were mainly involved in carbohydrate metabolism-related pathways. The expression patterns of 22 selected genes were validated by quantitative real-time PCR, and the results were highly accordant with the Solexa analysis. Furthermore, many candidate genes, which were involved in defense and detoxification mechanisms including signaling protein kinases, transcription factors, metal transporters and chelate compound biosynthesis related enzymes, were successfully identified in response to heavy metal Pb. Identification of potential DEGs involved in responses to Pb stress significantly reflected alterations in major biological processes and metabolic pathways. The molecular basis of the response to Pb stress in radishes was comprehensively characterized. Useful information and new insights were provided for investigating the molecular regulation mechanism of heavy metal Pb accumulation and

Nuclear factor I-A represses expression of the cell adhesion molecule L1

PubMed Central

2009-01-01

Background The neural cell adhesion molecule L1 plays a crucial role in development and plasticity of the nervous system. Neural cells thus require precise control of L1 expression. Results We identified a full binding site for nuclear factor I (NFI) transcription factors in the regulatory region of the mouse L1 gene. Electrophoretic mobility shift assay (EMSA) showed binding of nuclear factor I-A (NFI-A) to this site. Moreover, for a brain-specific isoform of NFI-A (NFI-A bs), we confirmed the interaction in vivo using chromatin immunoprecipitation (ChIP). Reporter gene assays showed that in neuroblastoma cells, overexpression of NFI-A bs repressed L1 expression threefold. Conclusion Our findings suggest that NFI-A, in particular its brain-specific isoform, represses L1 gene expression, and might act as a second silencer of L1 in addition to the neural restrictive silencer factor (NRSF). PMID:20003413

Interferon regulatory factor 1 and a variant of heterogeneous nuclear ribonucleoprotein L coordinately silence the gene for adhesion protein CEACAM1.

PubMed

Dery, Kenneth J; Silver, Craig; Yang, Lu; Shively, John E

2018-06-15

The adhesion protein carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) is widely expressed in epithelial cells as a short cytoplasmic isoform (S-iso) and in leukocytes as a long cytoplasmic isoform (L-iso) and is frequently silenced in cancer by unknown mechanisms. Previously, we reported that interferon response factor 1 (IRF1) biases alternative splicing (AS) to include the variable exon 7 (E7) in CEACAM1, generating long cytoplasmic isoforms. We now show that IRF1 and a variant of heterogeneous nuclear ribonucleoprotein L (Lv1) coordinately silence the CEACAM1 gene. RNAi-mediated Lv1 depletion in IRF1-treated HeLa and melanoma cells induced significant CEACAM1 protein expression, reversed by ectopic Lv1 expression. The Lv1-mediated CEACAM1 repression resided in residues Gly 71 -Gly 89 and Ala 38 -Gly 89 in Lv1's N-terminal extension. ChIP analysis of IRF1- and FLAG-tagged Lv1-treated HeLa cells and global treatment with the global epigenetic modifiers 5-aza-2'-deoxycytidine and trichostatin A indicated that IRF1 and Lv1 together induce chromatin remodeling, restricting IRF1 access to the CEACAM1 promoter. In interferon γ-treated HeLa cells, the transcription factor SP1 did not associate with the CEACAM1 promoter, but binding by upstream transcription factor 1 (USF1), a known CEACAM1 regulator, was greatly enhanced. ChIP-sequencing revealed that Lv1 overexpression in IRF1-treated cells induces transcriptional silencing across many genes, including DCC ( d eleted in c olorectal c arcinoma), associated with CEACAM5 in colon cancer. Notably, IRF1, but not IRF3 and IRF7, affected CEACAM1 expression via translational repression. We conclude that IRF1 and Lv1 coordinately regulate CEACAM1 transcription, alternative splicing, and translation and may significantly contribute to CEACAM1 silencing in cancer. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

The GhTT2_A07 gene is linked to the brown colour and natural flame retardancy phenotypes of Lc1 cotton (Gossypium hirsutum L.) fibres

PubMed Central

Hinchliffe, Doug J.; Condon, Brian D.; Thyssen, Gregory; Naoumkina, Marina; Madison, Crista A.; Reynolds, Michael; Delhom, Christopher D.; Fang, David D.; Li, Ping; McCarty, Jack

2016-01-01

Some naturally coloured brown cotton fibres from accessions of Gossypium hirsutum L. can be used to make textiles with enhanced flame retardancy (FR). Several independent brown fibre loci have been identified and mapped to chromosomes, but the underlying genes have not yet been identified, and the mechanism of lint fibre FR is not yet fully understood. In this study, we show that both the brown colour and enhanced FR of the Lc1 lint colour locus are linked to a 1.4Mb inversion on chromosome A07 that is immediately upstream of a gene with similarity to Arabidopsis TRANSPARENT TESTA 2 (TT2). As a result of the alternative upstream sequence, the transcription factor GhTT2_A07 is highly up-regulated in developing fibres. In turn, genes in the phenylpropanoid metabolic pathway are activated, leading to biosynthesis of proanthocyanidins and accumulation of inorganic elements. We show that enhanced FR and anthocyanin precursors appear in developing brown fibres well before the brown colour is detectible, demonstrating for the first time that the polymerized proanthocyanidins that constitute the brown colour are not the source of enhanced FR. Identifying the particular colourless metabolite that provides Lc1 cotton with enhanced FR could help minimize the use of synthetic chemical flame retardant additives in textiles. PMID:27567364

The GhTT2_A07 gene is linked to the brown colour and natural flame retardancy phenotypes of Lc1 cotton (Gossypium hirsutum L.) fibres.

PubMed

Hinchliffe, Doug J; Condon, Brian D; Thyssen, Gregory; Naoumkina, Marina; Madison, Crista A; Reynolds, Michael; Delhom, Christopher D; Fang, David D; Li, Ping; McCarty, Jack

2016-10-01

Some naturally coloured brown cotton fibres from accessions of Gossypium hirsutum L. can be used to make textiles with enhanced flame retardancy (FR). Several independent brown fibre loci have been identified and mapped to chromosomes, but the underlying genes have not yet been identified, and the mechanism of lint fibre FR is not yet fully understood. In this study, we show that both the brown colour and enhanced FR of the Lc1 lint colour locus are linked to a 1.4Mb inversion on chromosome A07 that is immediately upstream of a gene with similarity to Arabidopsis TRANSPARENT TESTA 2 (TT2). As a result of the alternative upstream sequence, the transcription factor GhTT2_A07 is highly up-regulated in developing fibres. In turn, genes in the phenylpropanoid metabolic pathway are activated, leading to biosynthesis of proanthocyanidins and accumulation of inorganic elements. We show that enhanced FR and anthocyanin precursors appear in developing brown fibres well before the brown colour is detectible, demonstrating for the first time that the polymerized proanthocyanidins that constitute the brown colour are not the source of enhanced FR. Identifying the particular colourless metabolite that provides Lc1 cotton with enhanced FR could help minimize the use of synthetic chemical flame retardant additives in textiles. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Mapping and genomic targeting of the major leaf shape gene (L) in Upland cotton (Gossypium hirsutum L.).

PubMed

Andres, Ryan J; Bowman, Daryl T; Kaur, Baljinder; Kuraparthy, Vasu

2014-01-01

A major leaf shape locus (L) was mapped with molecular markers and genomically targeted to a small region in the D-genome of cotton. By using expression analysis and candidate gene mapping, two LMI1 -like genes are identified as possible candidates for leaf shape trait in cotton. Leaf shape in cotton is an important trait that influences yield, flowering rates, disease resistance, lint trash, and the efficacy of foliar chemical application. The leaves of okra leaf cotton display a significantly enhanced lobing pattern, as well as ectopic outgrowths along the lobe margins when compared with normal leaf cotton. These phenotypes are the hallmark characteristics of mutations in various known modifiers of leaf shape that culminate in the mis/over-expression of Class I KNOX genes. To better understand the molecular and genetic processes underlying leaf shape in cotton, a normal leaf accession (PI607650) was crossed to an okra leaf breeding line (NC05AZ21). An F2 population of 236 individuals confirmed the incompletely dominant single gene nature of the okra leaf shape trait in Gossypium hirsutum L. Molecular mapping with simple sequence repeat markers localized the leaf shape gene to 5.4 cM interval in the distal region of the short arm of chromosome 15. Orthologous mapping of the closely linked markers with the sequenced diploid D-genome (Gossypium raimondii) tentatively resolved the leaf shape locus to a small genomic region. RT-PCR-based expression analysis and candidate gene mapping indicated that the okra leaf shape gene (L (o) ) in cotton might be an upstream regulator of Class I KNOX genes. The linked molecular markers and delineated genomic region in the sequenced diploid D-genome will assist in the future high-resolution mapping and map-based cloning of the leaf shape gene in cotton.

Ameliorative effects of l-carnitine on rats raised on a diet supplemented with lead acetate.

PubMed

El-Sherbini, El-Said; El-Sayed, Gehad; El Shotory, Rehab; Gheith, Nervana; Abou-Alsoud, Mohamed; Harakeh, Steve Mustapha; Karrouf, Gamal I

2017-09-01

Lead intoxication has been a major health hazard in humans. It affects people at all ages. Its toxicity is associated with various organs of the body and affects different metabolic pathways. Based on histological data, l-carnitine reduced the severity of tissue damage produced as a result of exposure of rats to lead acetate. The main objective of this study was to evaluate the underlying mechanism of protection offered by l-carnitine against lead acetate intoxication using male Sprague-Dawley rats. Forty male Sprague-Dawley rats were randomly divided into four groups with ten rats in each. The first group (G1) served as the control group and animals received standard diet only. The second group (G2) received lead acetate in their diet. The third group (G3) was the l-carnitine treated group and received the normal standard diet supplemented with l-carnitine. While the fourth group (G4) had a diet supplemented with both lead acetate and l-carnitine. At the end of each experiment, blood (serum and whole blood) were collected from each animal and analyzed for the following parameters: serum testosterone levels, serum nitric oxide and serum malondialdehyde. This is in addition to looking at the enzymatic activities of two important enzymes (superoxide dismutase and catalase) and on (glutathione reductase) which are indicative of the antioxidant activities in the whole blood. The results indicated that l-carnitine will counteract the undesirable effects of lead intoxication. It exerted its antioxidant potential by reducing the production of ROS and scavenging free radicals by maintaining and protecting the level of the of antioxidant enzymes SOD, CAT and glutathione peroxidase. Conclusion: l-Carnitine may play an important role in reversing the undesirable effects of lead intoxication. Future studies should be conducted to see whether such an effect is applicable in humans exposed to lead poising.

The GMD1 and GMD2 genes of Arabidopsis encode isoforms of GDP-D-mannose 4,6-dehydratase with cell type-specific expression patterns.

PubMed

Bonin, Christopher P; Freshour, Glenn; Hahn, Michael G; Vanzin, Gary F; Reiter, Wolf-Dieter

2003-06-01

l-Fucose (l-Fuc) is a monosaccharide constituent of plant cell wall polysaccharides and glycoproteins. The committing step in the de novo synthesis of l-Fuc is catalyzed by GDP-d-mannose 4,6-dehydratase, which, in Arabidopsis, is encoded by the GMD1 and GMD2 (MUR1) genes. To determine the functional significance of this genetic redundancy, the expression patterns of both genes were investigated via promoter-beta-glucuronidase fusions and immunolocalization of a Fuc-containing epitope. GMD2 is expressed in most cell types of the root, with the notable exception of the root tip where strong expression of GMD1 is observed. Within shoot organs, GMD1::GUS expression is confined to stipules and pollen grains leading to fucosylation of the walls of these cell types in the mur1 mutant. These results suggest that GMD2 represents the major housekeeping gene for the de novo synthesis of GDP-l-Fuc, whereas GMD1 expression is limited to a number of specialized cell types. We conclude that the synthesis of GDP-l-Fuc is controlled in a cell-autonomous manner by differential expression of two isoforms of the same enzyme.

Curd development associated gene (CDAG1) in cauliflower (Brassica oleracea L. var. botrytis) could result in enlarged organ size and increased biomass.

PubMed

Li, Hui; Liu, Qian; Zhang, Qingli; Qin, Erjun; Jin, Chuan; Wang, Yu; Wu, Mei; Shen, Guangshuang; Chen, Chengbin; Song, Wenqin; Wang, Chunguo

2017-01-01

The curd is a specialized organ and the most important product organ of cauliflower (Brassica oleracea L. var. botrytis). However, the mechanism underlying the regulation of curd formation and development remains largely unknown. In the present study, a novel homologous gene containing the Organ Size Related (OSR) domain, namely, CDAG1 (Curd Development Associated Gene 1) was identified in cauliflower. Quantitative analysis indicated that CDAG1 showed significantly higher transcript levels in young tissues. Functional analysis demonstrated that the ectopic overexpression of CDAG1 in Arabidopsis and cauliflower could significantly promote organ growth and result in larger organ size and increased biomass. Organ enlargement was predominantly due to increased cell number. In addition, 228 genes involved in the CDAG1-mediated regulatory network were discovered by transcriptome analysis. Among these genes, CDAG1 was confirmed to inhibit the transcriptional expression of the endogenous OSR genes, ARGOS and ARL, while a series of ethylene-responsive transcription factors (ERFs) were found to increased expression in 35S:CDAG1 transgenic Arabidopsis plants. This implies that CDAG1 may function in the ethylene-mediated signal pathway. These findings provide new insight into the function of OSR genes, and suggest potential applications of CDAG1 in breeding high-yielding crops. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Unprecedented genomic diversity of AhR1 and AhR2 genes in Atlantic salmon (Salmo salar L.).

PubMed

Hansson, Maria C; Wittzell, Håkan; Persson, Kerstin; von Schantz, Torbjörn

2004-06-24

Aryl hydrocarbon receptor (AhR) genes encode proteins involved in mediating the toxic responses induced by several environmental pollutants. Here, we describe the identification of the first two AhR1 (alpha and beta) genes and two additional AhR2 (alpha and beta) genes in the tetraploid species Atlantic salmon (Salmo salar L.) from a cosmid library screening. Cosmid clones containing genomic salmon AhR sequences were isolated using a cDNA clone containing the coding region of the Atlantic salmon AhR2gamma as a probe. Screening revealed 14 positive clones, from which four were chosen for further analyses. One of the cosmids contained genomic AhR sequences that were highly similar to the rainbow trout (Oncorhynchus mykiss) AhR2alpha and beta genes. SMART RACE amplified two complete, highly similar but not identical AhR type 2 sequences from salmon cDNA, which from phylogenetic analyses were determined as the rainbow trout AhR2alpha and beta orthologs. The salmon AhR2alpha and beta encode proteins of 1071 and 1058 residues, respectively, and encompass characteristic AhR sequence elements like a basic-helix-loop-helix (bHLH) and two PER-ARNT-SIM (PAS) domains. Both genes are transcribed in liver, spleen and muscle tissues of adult salmon. A second cosmid contained partial sequences, which were identical to the previously characterized AhR2gamma gene. The last two cosmids contained partial genomic AhR sequences, which were more similar to other AhR type 1 fish genes than the four characterized salmon AhR2 genes. However, attempts to amplify the corresponding complete cDNA sequences of the inserts proved very difficult, suggesting that these genes are non-functional or very weakly transcribed in the examined tissues. Phylogenetic analyses of the conserved regions did, however, clearly indicate that these two AhRs belong to the AhR type 1 clade and have been assigned as the Atlantic salmon AhR1alpha and AhR1beta genes. Taken together, these findings demonstrate that

The left end of rat L1 (L1Rn, long interspersed repeated) DNA which is a CpG island can function as a promoter.

PubMed Central

Nur, I; Pascale, E; Furano, A V

1988-01-01

Here we report that the 600 bp promoter-like region at the left end of a newly isolated and characterized rat L1 DNA element can activate the prokaryotic chloramphenicol acyltransferase gene in a rat cell line. Activation only occurs when the promoter region is oriented to the transferase gene as it is to the L1 protein encoding sequences and is 75% inhibited by methylation of just 5 of the 22 CpGs present in the promoter. The G + C rich promoter contains enough CpGs to qualify it as a CpG island, but in contrast to other CpG islands, genomic L1 promoters are fully methylated in both somatic cell and sperm DNA as judged by restriction enzyme analysis. Partial demethylation of the genomic promoters by treatment with 5-azacytidine failed to produce discrete L1 transcripts. The relationship of methylation to the evolutionary history and fate of the rat L1 promoter is discussed. Images PMID:2459662

Molecular aspects of zygotic embryogenesis in sunflower (Helianthus annuus L.): correlation of positive histone marks with HaWUS expression and putative link HaWUS/HaL1L.

PubMed

Salvini, Mariangela; Fambrini, Marco; Giorgetti, Lucia; Pugliesi, Claudio

2016-01-01

The link HaWUS/ HaL1L , the opposite transcriptional behavior, and the decrease/increase in positive histone marks bond to both genes suggest an inhibitory effect of WUS on HaL1L in sunflower zygotic embryos. In Arabidopsis, a group of transcription factors implicated in the earliest events of embryogenesis is the WUSCHEL-RELATED HOMEOBOX (WOX) protein family including WUSCHEL (WUS) and other 14 WOX protein, some of which contain a conserved WUS-box domain in addition to the homeodomain. WUS transcripts appear very early in embryogenesis, at the 16-cell embryo stage, but gradually become restricted to the center of the developing shoot apical meristem (SAM) primordium and continues to be expressed in cells of the niche/organizing center of SAM and floral meristems to maintain stem cell population. Moreover, WUS has decisive roles in the embryonic program presumably promoting the vegetative-to-embryonic transition and/or maintaining the identity of the embryonic stem cells. However, data on the direct interaction between WUS and key genes for seed development (as LEC1 and L1L) are not collected. The novelty of this report consists in the characterization of Helianthus annuus WUS (HaWUS) gene and in its analysis regarding the pattern of the methylated lysine 4 (K4) of the Histone H3 and of the acetylated histone H3 during the zygotic embryo development. Also, a parallel investigation was performed for HaL1L gene since two copies of the WUS-binding site (WUSATA), previously identified on HaL1L nucleotide sequence, were able to be bound by the HaWUS recombinant protein suggesting a not described effect of HaWUS on HaL1L transcription.

A novel RNA binding protein affects rbcL gene expression and is specific to bundle sheath chloroplasts in C4 plants

PubMed Central

2013-01-01

Background Plants that utilize the highly efficient C4 pathway of photosynthesis typically possess kranz-type leaf anatomy that consists of two morphologically and functionally distinct photosynthetic cell types, the bundle sheath (BS) and mesophyll (M) cells. These two cell types differentially express many genes that are required for C4 capability and function. In mature C4 leaves, the plastidic rbcL gene, encoding the large subunit of the primary CO2 fixation enzyme Rubisco, is expressed specifically within BS cells. Numerous studies have demonstrated that BS-specific rbcL gene expression is regulated predominantly at post-transcriptional levels, through the control of translation and mRNA stability. The identification of regulatory factors associated with C4 patterns of rbcL gene expression has been an elusive goal for many years. Results RLSB, encoded by the nuclear RLSB gene, is an S1-domain RNA binding protein purified from C4 chloroplasts based on its specific binding to plastid-encoded rbcL mRNA in vitro. Co-localized with LSU to chloroplasts, RLSB is highly conserved across many plant species. Most significantly, RLSB localizes specifically to leaf bundle sheath (BS) cells in C4 plants. Comparative analysis using maize (C4) and Arabidopsis (C3) reveals its tight association with rbcL gene expression in both plants. Reduced RLSB expression (through insertion mutation or RNA silencing, respectively) led to reductions in rbcL mRNA accumulation and LSU production. Additional developmental effects, such as virescent/yellow leaves, were likely associated with decreased photosynthetic function and disruption of associated signaling networks. Conclusions Reductions in RLSB expression, due to insertion mutation or gene silencing, are strictly correlated with reductions in rbcL gene expression in both maize and Arabidopsis. In both plants, accumulation of rbcL mRNA as well as synthesis of LSU protein were affected. These findings suggest that specific accumulation

Analysis of the Arabidopsis IRX9/IRX9-L and IRX14/IRX14-L Pairs of Glycosyltransferase Genes Reveals Critical Contributions to Biosynthesis of the Hemicellulose Glucuronoxylan1[C][W

PubMed Central

Wu, Ai-Min; Hörnblad, Emma; Voxeur, Aline; Gerber, Lorenz; Rihouey, Christophe; Lerouge, Patrice; Marchant, Alan

2010-01-01

The hemicellulose glucuronoxylan (GX) is a major component of plant secondary cell walls. However, our understanding of GX synthesis remains limited. Here, we identify and analyze two new genes from Arabidopsis (Arabidopsis thaliana), IRREGULAR XYLEM9-LIKE (IRX9-L) and IRX14-LIKE (IRX14-L) that encode glycosyltransferase family 43 members proposed to function during xylan backbone elongation. We place IRX9-L and IRX14-L in a genetic framework with six previously described glycosyltransferase genes (IRX9, IRX10, IRX10-L, IRX14, FRAGILE FIBER8 [FRA8], and FRA8 HOMOLOG [F8H]) and investigate their function in GX synthesis. Double-mutant analysis identifies IRX9-L and IRX14-L as functional homologs of IRX9 and IRX14, respectively. Characterization of irx9 irx10 irx14 fra8 and irx9-L irx10-L irx14-L f8h quadruple mutants allows definition of a set of genes comprising IRX9, IRX10, IRX14, and FRA8 that perform the main role in GX synthesis during vegetative development. The IRX9-L, IRX10-L, IRX14-L, and F8H genes are able to partially substitute for their respective homologs and normally perform a minor function. The irx14 irx14-L double mutant virtually lacks xylan, whereas irx9 irx9-L and fra8 f8h double mutants form lowered amounts of GX displaying a greatly reduced degree of backbone polymerization. Our findings reveal two distinct sets of four genes each differentially contributing to GX biosynthesis. PMID:20424005

Minimal influence of G-protein null mutations on ozone-induced changes in gene expression, foliar injury, gas exchange and peroxidase activity in Arabidopsis thaliana L

PubMed Central

Booker, Fitzgerald; Burkey, Kent; Morgan, Patrick; Fiscus, Edwin; Jones, Alan

2016-01-01

Ozone (O3) uptake by plants leads to an increase in reactive oxygen species (ROS) in the intercellular space of leaves and induces signalling processes reported to involve the membrane-bound heterotrimeric G-protein complex. Therefore, potential G-protein-mediated response mechanisms to O3 were compared between Arabidopsis thaliana L. lines with null mutations in the α- and β-subunits (gpa1-4, agb1-2 and gpa1-4/agb1-2) and Col-0 wild-type plants. Plants were treated with a range of O3 concentrations (5, 125, 175 and 300 nL L−1) for 1 and 2 d in controlled environment chambers. Transcript levels of GPA1, AGB1 and RGS1 transiently increased in Col-0 exposed to 125 nL L−1 O3 compared with the 5 nL L−1 control treatment. However, silencing of α and β G-protein genes resulted in little alteration of many processes associated with O3 injury, including the induction of ROS-signalling genes, increased leaf tissue ion leakage, decreased net photosynthesis and stomatal conductance, and increased peroxidase activity, especially in the leaf apoplast. These results indicated that many responses to O3 stress at physiological levels were not detectably influenced by α and β G-proteins. PMID:21988569

Identification of the Pr1 Gene Product Completes the Anthocyanin Biosynthesis Pathway of Maize

PubMed Central

Sharma, Mandeep; Cortes-Cruz, Moises; Ahern, Kevin R.; McMullen, Michael; Brutnell, Thomas P.; Chopra, Surinder

2011-01-01

In maize, mutations in the pr1 locus lead to the accumulation of pelargonidin (red) rather than cyanidin (purple) pigments in aleurone cells where the anthocyanin biosynthetic pathway is active. We characterized pr1 mutation and isolated a putative F3′H encoding gene (Zmf3′h1) and showed by segregation analysis that the red kernel phenotype is linked to this gene. Genetic mapping using SNP markers confirms its position on chromosome 5L. Furthermore, genetic complementation experiments using a CaMV 35S::ZmF3′H1 promoter–gene construct established that the encoded protein product was sufficient to perform a 3′-hydroxylation reaction. The Zmf3′h1-specific transcripts were detected in floral and vegetative tissues of Pr1 plants and were absent in pr1. Four pr1 alleles were characterized: two carry a 24 TA dinucleotide repeat insertion in the 5′-upstream promoter region, a third has a 17-bp deletion near the TATA box, and a fourth contains a Ds insertion in exon1. Genetic and transcription assays demonstrated that the pr1 gene is under the regulatory control of anthocyanin transcription factors red1 and colorless1. The cloning and characterization of pr1 completes the molecular identification of all genes encoding structural enzymes of the anthocyanin pathway of maize. PMID:21385724

Phosphorylation of nuclear factor erythroid 2-like 2 (NFE2L2) in mammary tissue of Holstein cows during the periparturient period is associated with mRNA abundance of antioxidant gene networks.

PubMed

Han, L Q; Zhou, Z; Ma, Y; Batistel, F; Osorio, J S; Loor, J J

2018-04-18

Changes in the production of reactive oxygen species in the mammary gland of dairy cows during the periparturient period could lead to oxidative stress and potentially impair mammary function. Phosphorylation of the transcription factor nuclear factor erythroid 2-like 2 (NFE2L2), also known as nuclear factor-E2-related factor 2, controls mRNA abundance of genes encoding antioxidant proteins and enzymes. The hypothesis was that NFE2L2 phosphorylation status and target gene mRNA abundance in the mammary gland of dairy cows is altered around parturition. Total NFE2L2 protein, phosphorylated protein (p-NFE2L2), and ratio of p-NFE2L2 to NFE2L2 along with mRNA abundance of 24 genes related to the NFE2L2 signaling pathway, apoptosis, and cell proliferation were measured in mammary tissue samples from Holstein cows at -30, 1, 15, and 30 d relative to parturition. Although total NFE2L2 protein abundance did not differ, p-NFE2L2 and p-NFE2L2-to-NFE2L2 ratio were greater after parturition. The upregulation of DNA damage inducible transcript 3 (DDIT3) postpartum indicated a localized oxidative stress state. Among genes evaluated, thioredoxin (TXN), glutathione peroxidase 1 (GPX1), and glutathione S-transferase mu 1 (GSTM1) had the highest (37.1, 15.1, and 4.8% of total mRNA measured, respectively) abundance. The mRNA abundance of various target genes with detoxifying enzymatic functions and free radical scavenging activities [glutamate-cysteine ligase catalytic subunit (GCLC); glutathione reductase (GSR); ferrochelatase (FECH); TXN; thioredoxin reductase 1 (TXNRD1); and NAD(P)H quinone dehydrogenase 1 (NQO1)] were consistently upregulated (linear effect of time) as parturition approached and lactation began. Among the transcription regulators, NFE2L2 had the highest mRNA abundance (7.3% of total mRNA measured). Abundance of NFE2L2 and other transcription factors [nuclear factor kappa B subunit 1 (NFKB1), retinoid X receptor α (RXRA), and mitogen-activated protein kinase 14

Lead phytotoxicity on wheat (Triticum aestivum L.) seed germination and seedlings growth.

PubMed

Lamhamdi, Mostafa; Bakrim, Ahmed; Aarab, Ahmed; Lafont, René; Sayah, Fouad

2011-02-01

Lead (Pb) is an environmental pollutant extremely toxic to plants and other living organisms including humans. To assess Pb phytotoxicity, experiments focusing on germination of wheat seeds were germinated in a solution containing Pb (NO(3))(2) (0.05; 0.1; 0.5; 1g/L) during 6 days. Lead accumulation in seedlings was positively correlated with the external concentrations, and negatively correlated with morphological parameters of plant growth. Lead increased lipid peroxidation, enhanced soluble protein concentrations and induced a significant accumulation of proline in roots. Esterase activity was enhanced in the presence of lead, whereas α-amylase activity was significantly inhibited. Antioxidant enzymes activities, such as, ascorbate peroxidase, peroxidase, superoxide dismutase, catalase and glutathione S-transferase were generally significantly increased in the presence of lead in a dose-dependent manner. The present results thus provide a model system to screen for natural compounds able to counteract the deleterious effects of lead. Copyright © 2010 Académie des sciences. Published by Elsevier SAS. All rights reserved.

Does angiotensin-converting enzyme-1 (ACE-1) gene polymorphism lead to chronic kidney disease among hypertensive patients?

PubMed

Sarkar, Taposh; Singh, Narinder Pal; Kar, Premashish; Husain, Syed Akhtar; Kapoor, Seema; Pollipalli, Sunil Kumar; Kumar, Anish; Garg, Neena

2016-06-01

Hypertension is one of the important contributing factors linked with both causation and development of kidney disease. It is a multifactorial, polygenic, and complex disorder due to interaction of several risk genes with environmental factors. The present study was aimed to explore genetic polymorphism in ACE-1 gene as a risk factor for CKD among hypertensive patients. Three hundred patients were enrolled in the study. Ninety were hypertensive patients with CKD taken as cases, whereas 210 hypertensive patients without CKD were taken as controls. Demographic data including age, sex, Body mass index (BMI), and other risk factors were also recorded. DNA was extracted from blood by salting out method. Genotyping of ACE gene was done by PCR technique. All the statistical analysis was done by using Epi Info and SPSS version 16 software (SPSS Inc., Chicago, IL). Mean age was higher in the control group (p < 0.05). Variables among two groups were compared out of which age, BMI, hemoglobin (Hb) was found to be statistically significant whereas other variables like systolic blood pressure, triglyceride and low-density lipoprotein were not. Blood urea and serum creatinine levels were statistically significant in the two genotypes (p < 0.05). Total and HDL cholesterol were statistically significant for DD genotype of ACE gene (OR = 1.42, 95% CI = 0.72-2.81). Similarly, the risk for CKD among hypertensive patients was also associated with D allele of ACE gene (OR = 1.25, 95% CI = 0.86-1.79). It is concluded that ACE-DD genotype may be a risk factor for the causation and development of chronic kidney failure among hypertensive patients.

The genes and enzymes of the carotenoid metabolic pathway in Vitis vinifera L.

PubMed Central

2012-01-01

Background Carotenoids are a heterogeneous group of plant isoprenoids primarily involved in photosynthesis. In plants the cleavage of carotenoids leads to the formation of the phytohormones abscisic acid and strigolactone, and C13-norisoprenoids involved in the characteristic flavour and aroma compounds in flowers and fruits and are of specific importance in the varietal character of grapes and wine. This work extends the previous reports of carotenoid gene expression and photosynthetic pigment analysis by providing an up-to-date pathway analysis and an important framework for the analysis of carotenoid metabolic pathways in grapevine. Results Comparative genomics was used to identify 42 genes putatively involved in carotenoid biosynthesis/catabolism in grapevine. The genes are distributed on 16 of the 19 chromosomes and have been localised to the physical map of the heterozygous ENTAV115 grapevine sequence. Nine of the genes occur as single copies whereas the rest of the carotenoid metabolic genes have more than one paralogue. The cDNA copies of eleven corresponding genes from Vitis vinifera L. cv. Pinotage were characterised, and four where shown to be functional. Microarrays provided expression profiles of 39 accessions in the metabolic pathway during three berry developmental stages in Sauvignon blanc, whereas an optimised HPLC analysis provided the concentrations of individual carotenoids. This provides evidence of the functioning of the lutein epoxide cycle and the respective genes in grapevine. Similarly, orthologues of genes leading to the formation of strigolactone involved in shoot branching inhibition were identified: CCD7, CCD8 and MAX1. Moreover, the isoforms typically have different expression patterns, confirming the complex regulation of the pathway. Of particular interest is the expression pattern of the three VvNCEDs: Our results support previous findings that VvNCED3 is likely the isoform linked to ABA content in berries. Conclusions The

L-glutamine Induces Expression of Listeria monocytogenes Virulence Genes

PubMed Central

Lobel, Lior; Burg-Golani, Tamar; Sigal, Nadejda; Rose, Jessica; Livnat-Levanon, Nurit; Lewinson, Oded; Herskovits, Anat A.

2017-01-01

The high environmental adaptability of bacteria is contingent upon their ability to sense changes in their surroundings. Bacterial pathogen entry into host poses an abrupt and dramatic environmental change, during which successful pathogens gauge multiple parameters that signal host localization. The facultative human pathogen Listeria monocytogenes flourishes in soil, water and food, and in ~50 different animals, and serves as a model for intracellular infection. L. monocytogenes identifies host entry by sensing both physical (e.g., temperature) and chemical (e.g., metabolite concentrations) factors. We report here that L-glutamine, an abundant nitrogen source in host serum and cells, serves as an environmental indicator and inducer of virulence gene expression. In contrast, ammonia, which is the most abundant nitrogen source in soil and water, fully supports growth, but fails to activate virulence gene transcription. We demonstrate that induction of virulence genes only occurs when the Listerial intracellular concentration of L-glutamine crosses a certain threshold, acting as an on/off switch: off when L-glutamine concentrations are below the threshold, and fully on when the threshold is crossed. To turn on the switch, L-glutamine must be present, and the L-glutamine high affinity ABC transporter, GlnPQ, must be active. Inactivation of GlnPQ led to complete arrest of L-glutamine uptake, reduced type I interferon response in infected macrophages, dramatic reduction in expression of virulence genes, and attenuated virulence in a mouse infection model. These results may explain observations made with other pathogens correlating nitrogen metabolism and virulence, and suggest that gauging of L-glutamine as a means of ascertaining host localization may be a general mechanism. PMID:28114430

Salicornia europaea L. Na⁺/H⁺ antiporter gene improves salt tolerance in transgenic alfalfa (Medicago sativa L.).

PubMed

Zhang, L Q; Niu, Y D; Huridu, H; Hao, J F; Qi, Z; Hasi, A

2014-07-24

In order to obtain a salt-tolerant perennial alfalfa (Medicago sativa L.), we transferred the halophyte Salicornia europaea L. Na(+)/H(+) antiporter gene, SeNHX1, to alfalfa by using the Agrobacterium-mediated transformation method. The transformants were confirmed by both PCR and RT-PCR analyses. Of 197 plants that were obtained after transformation, 36 were positive by PCR analysis using 2 primer pairs for the CaMV35S-SeNHX1 and SeNHX1-Nos fragments; 6 plants survived in a greenhouse. RT-PCR analysis revealed that SeNHX1 was expressed in 5 plants. The resultant transgenic alfalfa had better salt tolerance. After stress treatment for 21 days with 0.6% NaCl, the chlorophyll and MDA contents in transgenic plants were lower, but proline content and SOD, POD, and CAT activities were higher than those in wild-type plants. These results suggest that the salt tolerance of transgenic alfalfa was improved by the overexpression of the SeNHX1 gene.

IFN regulatory factor 1 restricts hepatitis E virus replication by activating STAT1 to induce antiviral IFN-stimulated genes.

PubMed

Xu, Lei; Zhou, Xinying; Wang, Wenshi; Wang, Yijin; Yin, Yuebang; Laan, Luc J W van der; Sprengers, Dave; Metselaar, Herold J; Peppelenbosch, Maikel P; Pan, Qiuwei

2016-10-01

IFN regulatory factor 1 (IRF1) is one of the most important IFN-stimulated genes (ISGs) in cellular antiviral immunity. Although hepatitis E virus (HEV) is a leading cause of acute hepatitis worldwide, how ISGs counteract HEV infection is largely unknown. This study was conducted to investigate the effect of IRF1 on HEV replication. Multiple cell lines were used in 2 models that harbor HEV. In different HEV cell culture systems, IRF1 effectively inhibited HEV replication. IRF1 did not trigger IFN production, and chromatin immunoprecipitation sequencing data analysis revealed that IRF1 bound to the promoter region of signal transducers and activators of transcription 1 (STAT1). Functional assay confirmed that IRF1 could drive the transcription of STAT1, resulting in elevation of total and phosphorylated STAT1 proteins and further activating the transcription of a panel of downstream antiviral ISGs. By pharmacological inhibitors and RNAi-mediated gene-silencing approaches, we revealed that antiviral function of IRF1 is dependent on the JAK-STAT cascade. Furthermore, induction of ISGs and the anti-HEV effect of IRF1 overlapped that of IFNα, but was potentiated by ribavirin. We demonstrated that IRF1 effectively inhibits HEV replication through the activation of the JAK-STAT pathway, and the subsequent transcription of antiviral ISGs, but independent of IFN production.-Xu, L., Zhou, X., Wang, W., Wang, Y., Yin, Y., van der Laan, L. J. W., Sprengers, D., Metselaar, H. J., Peppelenbosch, M. P., Pan, Q. IFN regulatory factor 1 restricts hepatitis E virus replication by activating STAT1 to induce antiviral IFN-stimulated genes. © FASEB.

Mechanism for the decrease in the FIP1L1-PDGFRalpha protein level in EoL-1 cells by histone deacetylase inhibitors.

PubMed

Ishihara, Kenji; Kaneko, Motoko; Kitamura, Hajime; Takahashi, Aki; Hong, Jang Ja; Seyama, Toshio; Iida, Koji; Wada, Hiroshi; Hirasawa, Noriyasu; Ohuchi, Kazuo

2008-01-01

Acetylation and deacetylation of proteins occur in cells in response to various stimuli, and are reversibly catalyzed by histone acetyltransferase and histone deacetylase (HDAC), respectively. EoL-1 cells have an FIP1L1-PDGFRA fusion gene that causes transformation of eosinophilic precursor cells into leukemia cells. The HDAC inhibitors apicidin and n-butyrate suppress the proliferation of EoL-1 cells and induce differentiation into eosinophils by a decrease in the protein level of FIP1L1-PDGFRalpha without affecting the mRNA level for FIP1L1-PDGFRA. In this study, we analyzed the mechanism by which the protein level of FIP1L1-PDGFRalpha is decreased by apicidin and n-butyrate. EoL-1 cells were incubated in the presence of the HDAC inhibitors apicidin, trichostatin A or n-butyrate. The protein levels of FIP1L1-PDGFRalpha and phosphorylated eIF-2alpha were determined by Western blotting. Actinomycin D and cycloheximide were used to block RNA synthesis and protein synthesis, respectively, in the chasing experiment of the amount of FIP1L1-PDGFRalpha protein. When apicidin- and n-butyrate-treated EoL-1 cells were incubated in the presence of actinomycin D, the decrease in the protein level of FIP1L1-PDGFRalpha was significantly enhanced when compared with controls. In contrast, the protein levels were not changed by cycloheximide among these groups. Apicidin and n-butyrate induced the continuous phosphorylation of eIF-2alpha for up to 8 days. The decrease in the level of FIP1L1-PDGFRalpha protein by continuous inhibition of HDAC may be due to the decrease in the translation rate of FIP1L1-PDGFRA. Copyright 2008 S. Karger AG, Basel.

How Alterations in the Cdt1 Expression Lead to Gene Amplification in Breast Cancer

DTIC Science & Technology

2011-07-01

absence of extrinsic DNA damage. We measured the TLS activity by measuring the mutation frequency in a supF gene (in a shuttle vector) subjected to UV...induced DNA damage before its introduction into the cells. Error-prone TLS activity will mutate the supF gene , which is scored by a blue-white colony...Figure 4A). Sequencing of the mutant supF genes , revealed a mutation spectrum consistent with error prone TLS (Supplemental Table 1). Significantly

L-Asparaginase from Streptomyces griseus NIOT-VKMA29: optimization of process variables using factorial designs and molecular characterization of L-asparaginase gene

NASA Astrophysics Data System (ADS)

Meena, Balakrishnan; Anburajan, Lawrance; Sathish, Thadikamala; Vijaya Raghavan, Rangamaran; Dharani, Gopal; Valsalan Vinithkumar, Nambali; Kirubagaran, Ramalingam

2015-07-01

Marine actinobacteria are known to be a rich source for novel metabolites with diverse biological activities. In this study, a potential extracellular L-asparaginase was characterised from the Streptomyces griseus NIOT-VKMA29. Box-Behnken based optimization was used to determine the culture medium components to enhance the L-asparaginase production. pH, starch, yeast extract and L-asparagine has a direct correlation for enzyme production with a maximum yield of 56.78 IU mL-1. A verification experiment was performed to validate the experiment and more than 99% validity was established. L-Asparaginase biosynthesis gene (ansA) from Streptomyces griseus NIOT-VKMA29 was heterologously expressed in Escherichia coli M15 and the enzyme production was increased threefold (123 IU mL-1) over the native strain. The ansA gene sequences reported in this study encloses several base substitutions with that of reported sequences in GenBank, resulting in altered amino acid sequences of the translated protein.

Interpreting and managing blood lead levels < 10 microg/dL in children and reducing childhood exposures to lead: recommendations of CDC's Advisory Committee on Childhood Lead Poisoning Prevention.

PubMed

2007-11-02

Lead is a common environmental contaminant, and exposure to lead is a preventable risk that exists in all areas of the United States. Lead is associated with negative outcomes in children, including impaired cognitive, motor, behavioral, and physical abilities. In 1991, CDC defined the blood lead level (BLL) that should prompt public health actions as 10 microg/dL. Concurrently, CDC also recognized that a BLL of 10 microg/dL did not define a threshold for the harmful effects of lead. Research conducted since 1991 has strengthened the evidence that children's physical and mental development can be affected at BLLs < or =10 microg/dL. This report summarizes the findings of a review of clinical interpretation and management of BLLs < or =10 microg/dL conducted by CDC's Advisory Committee on Childhood Lead Poisoning Prevention. This report provides information to help clinicians understand BLLs < or =10 microg/dL, identifies gaps in knowledge concerning lead levels in this range, and outlines strategies to reduce childhood exposures to lead. In addition, this report summarizes scientific data relevant to counseling, blood lead screening, and lead exposure risk assessment. To aid in the interpretation of BLLs, clinicians should understand the laboratory error range for blood lead values and, if possible, select a laboratory that achieves routine performance within +/-2 microg/dL. Clinicians should obtain an environmental history on all children they examine, provide families with lead prevention counseling, and follow blood lead screening recommendations established for their areas. As local and patient circumstances permit, clinicians should consider early referral to developmental programs for children at high risk for exposure to lead and consider more frequent rescreening of children with BLLs approaching 10 microg/dL, depending on the potential for exposure to lead, child age, and season of testing. In addition, clinicians should direct parents to agencies and

The organization of the fuc regulon specifying L-fucose dissimilation in Escherichia coli K12 as determined by gene cloning.

PubMed

Chen, Y M; Zhu, Y; Lin, E C

1987-12-01

In Escherichia coli the six known genes specifying the utilization of L-fucose as carbon and energy source cluster at 60.2 min and constitute a regulon. These genes include fucP (encoding L-fucose permease), fucI (encoding L-fucose isomerase), fucK (encoding L-fuculose kinase), fucA (encoding L-fuculose 1-phosphate aldolase), fucO (encoding L-1,2-propanediol oxidoreductase), and fucR (encoding the regulatory protein). In this study the fuc genes were cloned and their positions on the chromosome were established by restriction endonuclease and complementation analyses. Clockwise, the gene order is: fucO-fucA-fucP-fucI-fucK-fucR. The operons comprising the structural genes and the direction of transcription were determined by complementation analysis and Southern blot hybridization. The fucPIK and fucA operons are transcribed clockwise. The fucO operon is transcribed counterclockwise. The fucR gene product activates the three structural operons in trans.

Statistics and Epidemiology of Lead Poisoning (FY 72-L1).

ERIC Educational Resources Information Center

Morrison, John H., Jr.; And Others

This report is the first in a quarterly series which will contain statistics and epidemiologic notes on lead poisoning at both the national and local levels. This report contains (a) statistics on childhood lead poisoning; (b) a status report on the Community Lead Poisoning Data System, which was designed to assist local lead control programs and…

Lead (Pb) bioaccumulation; genera Bacillus isolate S1 and SS19 as a case study

NASA Astrophysics Data System (ADS)

Arifiyanto, Achmad; Apriyanti, Fitria Dwi; Purwaningsih, Puput; Kalqutny, Septian Hary; Agustina, Dyah; Surtiningsih, Tini; Shovitri, Maya; Zulaika, Enny

2017-06-01

Lead (Pb) includes a group of large heavy metal in nature was toxic either on animal or human and did not provide an advantage function biologically. Bacillus isolates S1 and SS19 known resistant to lead up to 50 mg / L PbCl2. In this research will be examined whether genera Bacillus isolates S1 and SS19 could accumulate metal lead (Pb), their capability in accumulating and profile protein differences when the bacteria genera Bacillus isolates S1 and SS19 get exposed metal lead (Pb). Inoculum at age ± 9 hours are used, with a Nutrient Broth (NB) containing 50, 75 and 100 mg / L PbCl2. Inductively Coupled Plasma Atomic Emission Spectrometry (ICP) used to assessed Pb2+ concentrations. Bioaccumulation levels of Pb2+ by Bacillus isolate S1 and SS19 related to the distinction of beginning concentration to the final concentration. Bacillus isolate S1 achieved 53% and 51% bioaccumulation efficiency rate in lead presence concentration (75 and 100 mg/L) and 51% (50 mg/L). Another way Bacillus isolate SS19 was able to accumulate 57% (50 mg/L PbCl2) and kept stable on 36% bioaccumulation efficiency rate (75 and 100 mg/L PbCl2). Regarding SDS-PAGE electrophoresis protein profile result, protein in ± 127 kDa, molecule mass detected in the presence of Lead for Bacillus isolate S1.

Expression and function of AtMBD4L, the single gene encoding the nuclear DNA glycosylase MBD4L in Arabidopsis.

PubMed

Nota, Florencia; Cambiagno, Damián A; Ribone, Pamela; Alvarez, María E

2015-06-01

DNA glycosylases recognize and excise damaged or incorrect bases from DNA initiating the base excision repair (BER) pathway. Methyl-binding domain protein 4 (MBD4) is a member of the HhH-GPD DNA glycosylase superfamily, which has been well studied in mammals but not in plants. Our knowledge on the plant enzyme is limited to the activity of the Arabidopsis recombinant protein MBD4L in vitro. To start evaluating MBD4L in its biological context, we here characterized the structure, expression and effects of its gene, AtMBD4L. Phylogenetic analysis indicated that AtMBD4L belongs to one of the seven families of HhH-GPD DNA glycosylase genes existing in plants, and is unique on its family. Two AtMBD4L transcripts coding for active enzymes were detected in leaves and flowers. Transgenic plants expressing the AtMBD4L:GUS gene confined GUS activity to perivascular leaf tissues (usually adjacent to hydathodes), flowers (anthers at particular stages of development), and the apex of immature siliques. MBD4L-GFP fusion proteins showed nuclear localization in planta. Interestingly, overexpression of the full length MBD4L, but not a truncated enzyme lacking the DNA glycosylase domain, induced the BER gene LIG1 and enhanced tolerance to oxidative stress. These results suggest that endogenous MBD4L acts on particular tissues, is capable of activating BER, and may contribute to repair DNA damage caused by oxidative stress. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Characterization of the duplicate L-SIGN and DC-SIGN genes in miiuy croaker and evolutionary analysis of L-SIGN in fishes.

PubMed

Shu, Chang; Wang, Shanchen; Xu, Tianjun

2015-05-01

Dendritic cell-specific ICAM-3-grabbing non-integrin (DC-SIGN/CD209) and liver/lymph node-specific ICAM-grabbing non-integrin (L-SIGN/CD299) which are homologues of DC-SIGN are important members in C-type lectin receptors family as key molecules to recognize and eliminate pathogens in the innate immune system. DC-SIGN and L-SIGN have become hot topics in recent studies which both served as cell adhesion and phagocytic pathogen recognition receptors in mammals. However, there have been almost no studies of DC-SIGN and L-SIGN structure and characters in fish, only DC-SIGN in the zebrafish had been studied. In our study, we identified and characterized the full-length miiuy croaker (Miichthys miiuy) DC-SIGN (mmDC-SIGN) and L-SIGN (mmL-SIGN) genes. The sequence analysis results showed that mmDC-SIGN and mmL-SIGN have the same domains with other vertebrates except primates, and share some conserved motifs in CRD among all the vertebrates which play a crucial role in interacting with Ca(2+) and for recognizing mannose-containing motifs. Gene synteny of DC-SIGN and L-SIGN were analyzed for the first time and gene synteny of L-SIGN was conserved among the five fishes. Interestingly, one gene next to L-SIGN from gene synteny had high similarity with L-SIGN gene that was described as L-SIGN-like in fish species. While only one L-SIGN gene existed in other vertebrates, two L-SIGN in fish may be in consequence of the fish-specific genome duplication to adapt the specific environment. The evolutionary analysis showed that the ancestral lineages of L-SIGN gene in fishes experienced purifying selection and the current lineages of L-SIGN gene in fishes underwent positive selection, indicating that the ancestral lineages and current lineages of L-SIGN gene in fishes underwent different evolutionary patterns. Both mmDC-SIGN and mmL-SIGN were expressed in all tested tissues and ubiquitously up-regulated in infected liver, spleen and kidney at different sampling time points

Comparison of gene co-networks reveals the molecular mechanisms of the rice (Oryza sativa L.) response to Rhizoctonia solani AG1 IA infection.

PubMed

Zhang, Jinfeng; Zhao, Wenjuan; Fu, Rong; Fu, Chenglin; Wang, Lingxia; Liu, Huainian; Li, Shuangcheng; Deng, Qiming; Wang, Shiquan; Zhu, Jun; Liang, Yueyang; Li, Ping; Zheng, Aiping

2018-05-05

Rhizoctonia solani causes rice sheath blight, an important disease affecting the growth of rice (Oryza sativa L.). Attempts to control the disease have met with little success. Based on transcriptional profiling, we previously identified more than 11,947 common differentially expressed genes (TPM > 10) between the rice genotypes TeQing and Lemont. In the current study, we extended these findings by focusing on an analysis of gene co-expression in response to R. solani AG1 IA and identified gene modules within the networks through weighted gene co-expression network analysis (WGCNA). We compared the different genes assigned to each module and the biological interpretations of gene co-expression networks at early and later modules in the two rice genotypes to reveal differential responses to AG1 IA. Our results show that different changes occurred in the two rice genotypes and that the modules in the two groups contain a number of candidate genes possibly involved in pathogenesis, such as the VQ protein. Furthermore, these gene co-expression networks provide comprehensive transcriptional information regarding gene expression in rice in response to AG1 IA. The co-expression networks derived from our data offer ideas for follow-up experimentation that will help advance our understanding of the translational regulation of rice gene expression changes in response to AG1 IA.

The waaL gene mutation compromised the inhabitation of Enterobacter sp. Ag1 in the mosquito gut environment.

PubMed

Pei, Dong; Jiang, Jinjin; Yu, Wanqin; Kukutla, Phanidhar; Uentillie, Alejandro; Xu, Jiannong

2015-08-27

The mosquito gut harbors a variety of bacteria that are dynamically associated with mosquitoes in various contexts. However, little is known about bacterial factors that affect bacterial inhabitation in the gut microbial community. Enterobacter sp. Ag1 is a predominant Gram negative bacterium in the mosquito midgut. In a mutant library that was generated using transposon Tn5-mediated mutagenesis, a mutant was identified, in which the gene waaL was disrupted by the Tn5 insertion. The waaL encodes O antigen ligase, which is required for the attachment of O antigen to the outer core oligosaccharide of the lipopolysaccharide (LPS). The waaL(-) mutation caused the O antigen repeat missing in the LPS. The normal LPS structure was restored when the mutant was complemented with a plasmid containing waaL gene. The waaL(-) mutation did not affect bacterial proliferation in LB culture, the mutant cells grew at a rate the same as the wildtype (wt) cells. However, when waaL(-) strain were co-cultured with the wt strain or complemented strain, the mutant cells proliferated with a slower rate, indicating that the mutants were less competitive than wt cells in a community setting. Similarly, in a co-feeding assay, when fluorescently tagged wt strain and waaL(-) strain were orally co-introduced into the gut of Anopheles stephensi mosquitoes, the mutant cells were less prevalent in both sugar-fed and blood-fed guts. The data suggest that the mutation compromised the bacterial inhabitation in the gut community. Besides, the mutant was more sensitive to oxidative stress, demonstrated by lower survival rate upon exposure to 20 mM H₂O₂. Lack of the O antigen structure in LPS of Enterobacter compromised the effective growth in co-culture and co-feeding assays. In addition, O-antigen was involved in protection against oxidative stress. The findings suggest that intact LPS is crucial for the bacteria to steadily stay in the gut microbial community.

Cloning and characterization of a novel NAC family gene CarNAC1 from chickpea (Cicer arietinum L.).

PubMed

Peng, Hui; Yu, Xingwang; Cheng, Huiying; Shi, Qinghua; Zhang, Hua; Li, Jiangui; Ma, Hao

2010-01-01

The plant-specific NAC (for NAM, ATAF1,2 and CUC2) proteins have been found to play important roles in plant development and stress responses. In this study, a NAC gene CarNAC1 (for Cicer arietinum L. NAC gene 1) was isolated from a cDNA library constructed with chickpea seedling leaves treated by polyethylene glycol. CarNAC1 encoded a putative protein with 239 amino acids and contained 3 exons and 2 introns within genomic DNA sequence. CarNAC1 had a conserved NAC domain in the N-terminus and the CarNAC1:GFP (green fluorescent protein) fusion protein was localized in the nucleus of onion epidermal cells. Additionally, CarNAC1 exhibited the trans-activation activity which was mapped to the C-terminus. The CarNAC1 transcript was detected in many chickpea organs including seedling leaves, stems, roots, flowers, and young pods, but less accumulated in young seeds. CarNAC1 was induced by leaf age and showed changes in expression during seed development and germination. Furthermore, the expression of CarNAC1 was strongly induced by drought, salt, cold, wounding, H(2)O(2), ethephon, salicylic acid, indole-3-acetic acid, and gibberellin. Our results suggest that CarNAC1 encodes a novel NAC-domain protein and may be a transcriptional activator involved in plant development and various stress responses.

Development and validation of breeder-friendly KASPar markers for er1, a powdery mildew resistance gene in pea (Pisum sativum L.)

USDA-ARS?s Scientific Manuscript database

Powdery mildew of pea is caused by Erysiphe pisi DC and is a serious threat to pea (Pisum sativum L.) production throughout much of the world. Development and utilization of genetic resistance to powdery mildew is considered an effective and sustainable strategy to manage this disease. One gene, er1...

The p.L302P mutation in the lysosomal enzyme gene SMPD1 is a risk factor for Parkinson disease

PubMed Central

Gan-Or, Ziv; Ozelius, Laurie J.; Bar-Shira, Anat; Saunders-Pullman, Rachel; Mirelman, Anat; Kornreich, Ruth; Gana-Weisz, Mali; Raymond, Deborah; Rozenkrantz, Liron; Deik, Andres; Gurevich, Tanya; Gross, Susan J.; Schreiber-Agus, Nicole; Giladi, Nir; Bressman, Susan B.

2013-01-01

Objective: To study the possible association of founder mutations in the lysosomal storage disorder genes HEXA, SMPD1, and MCOLN1 (causing Tay-Sachs, Niemann-Pick A, and mucolipidosis type IV diseases, respectively) with Parkinson disease (PD). Methods: Two PD patient cohorts of Ashkenazi Jewish (AJ) ancestry, that included a total of 938 patients, were studied: a cohort of 654 patients from Tel Aviv, and a replication cohort of 284 patients from New York. Eight AJ founder mutations in the HEXA, SMPD1, and MCOLN1 genes were analyzed. The frequencies of these mutations were compared to AJ control groups that included large published groups undergoing prenatal screening and 282 individuals matched for age and sex. Results: Mutation frequencies were similar in the 2 groups of patients with PD. The SMPD1 p.L302P was strongly associated with a highly increased risk for PD (odds ratio 9.4, 95% confidence interval 3.9–22.8, p < 0.0001), as 9/938 patients with PD were carriers of this mutation compared to only 11/10,709 controls. Conclusions: The SMPD1 p.L302P mutation is a novel risk factor for PD. Although it is rare on a population level, the identification of this mutation as a strong risk factor for PD may further elucidate PD pathogenesis and the role of lysosomal pathways in disease development. PMID:23535491

Lead suppresses chimeric human transferrin gene expression in transgenic mouse liver.

PubMed

Adrian, G S; Rivera, E V; Adrian, E K; Lu, Y; Buchanan, J; Herbert, D C; Weaker, F J; Walter, C A; Bowman, B H

1993-01-01

The major iron-transport protein in serum is transferrin (TF) which also has the capacity to transport other metals. This report presents evidence that synthesis of human TF can be regulated by the metal lead. Transgenic mice carrying chimeric human TF-chloramphenicol acetyl transferase (CAT) genes received lead or sodium salts by intraperitoneal injections or in drinking water. Transgene expression in liver was suppressed 31 to 50% by the lead treatment. Lead regulates human TF transgenes at the mRNA level since liver CAT enzyme activity, CAT protein, and TF-CAT mRNA levels were all suppressed. The dosages of lead did not alter synthesis of the other liver proteins, mouse TF and albumin, as measured by Northern blot analysis of total liver RNA and rocket immunoelectrophoresis of mouse sera. Moderate levels of lead exposure were sufficient to evoke the human TF transgene response; blood lead levels in mice that received lead acetate in drinking water ranged from 30 micrograms/dl to 56 micrograms/dl. In addition to suppressing expression of TF-CAT genes in transgenic mice, lead also suppressed synthesis of TF protein in cultured human hepatoma HepG2 cells. The regulation of human TF apparently differs from the regulation of mouse TF which is unresponsive to lead exposure.

Transcriptome profiling of sleeping, waking, and sleep deprived adult heterozygous Aldh1L1 - eGFP-L10a mice.

PubMed

Bellesi, Michele; de Vivo, Luisa; Tononi, Giulio; Cirelli, Chiara

2015-12-01

Transcriptomic studies revealed that hundreds of mRNAs show differential expression in the brains of sleeping relative to awake rats, mice, flies, and sparrows. Although these results have offered clues regarding the molecular consequences of sleep and sleep loss, their functional significance thus far has been limited. This is probably because the previous studies pooled transcripts from all brain cells, including neurons and glia. In Bellesi et al. 2015 [1], we used the translating ribosome affinity purification technology (TRAP) and microarray analysis to obtain a genome-wide mRNA profiling of astrocytes as a function of sleep and wake. We used bacterial artificial chromosome (BAC) transgenic mice expressing eGFP tagged ribosomal protein L10a under the promoter of the Aldh1L1 gene, a highly expressed astrocytic gene. Using this approach, we could extract only the astrocytic mRNAs, and only those already committed to be translated into proteins (L10a is part of the translational machinery). Here, we report a detailed description of the protocol used in the study [1]. Array data have been submitted to NCBI GEO under accession number (GSE69079).

Endothelin-1 gene regulation

PubMed Central

Stow, Lisa R.; Jacobs, Mollie E.; Wingo, Charles S.; Cain, Brian D.

2011-01-01

Over two decades of research have demonstrated that the peptide hormone endothelin-1 (ET-1) plays multiple, complex roles in cardiovascular, neural, pulmonary, reproductive, and renal physiology. Differential and tissue-specific production of ET-1 must be tightly regulated in order to preserve these biologically diverse actions. The primary mechanism thought to control ET-1 bioavailability is the rate of transcription from the ET-1 gene (edn1). Studies conducted on a variety of cell types have identified key transcription factors that govern edn1 expression. With few exceptions, the cis-acting elements bound by these factors have been mapped in the edn1 regulatory region. Recent evidence has revealed new roles for some factors originally believed to regulate edn1 in a tissue or hormone-specific manner. In addition, other mechanisms involved in epigenetic regulation and mRNA stability have emerged as important processes for regulated edn1 expression. The goal of this review is to provide a comprehensive overview of the specific factors and signaling systems that govern edn1 activity at the molecular level.—Stow, L. R., Jacobs, M. E., Wingo, C. S., Cain, B. D. Endothelin-1 gene regulation. PMID:20837776

Transcriptomic and physiological analyses of Medicago sativa L. roots in response to lead stress

PubMed Central

Zhang, Shichao; Guo, Qiang; Jin, Yan; Chen, Jingjing; Ma, Hongxia

2017-01-01

Lead (Pb) is one of the nonessential and toxic metals that threaten the environment and human health. Medicago sativa L. is a legume with high salt tolerance and high biomass production. It is not only a globally important forage crop but is also an ideal plant for phytoremediation. However, the biological and molecular mechanisms that respond to heavy metals are still not well defined in M. sativa. In this study, de novo and strand-specific RNA-sequencing was performed to identify genes involved in the Pb stress response in M. sativa roots. A total of 415,350 unigenes were obtained from the assembled cDNA libraries, among which 5,416 were identified as significantly differentially expressed genes (DEGs) (false discovery rate < 0.005) between cDNA libraries from control and Pb-treated plants. Gene ontology and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses of the DEGs showed they mainly clustered with terms associated with binding, transport, membranes, and the pathways related to signal and energy metabolism. Moreover, a number of candidate genes included antioxidant enzymes, metal transporters, and transcription factors involved in heavy metal response were upregulated under Pb stress. Quantitative real-time PCR(qRT-PCR) validation of the expression patterns of 10 randomly selected candidate DEGs were consistent with the transcriptome analysis results. Thus, this study offers new information towards the investigation of biological changes and molecular mechanisms related to Pb stress response in plants. PMID:28388670

The Aux/IAA gene rum1 involved in seminal and lateral root formation controls vascular patterning in maize (Zea mays L.) primary roots.

PubMed

Zhang, Yanxiang; Paschold, Anja; Marcon, Caroline; Liu, Sanzhen; Tai, Huanhuan; Nestler, Josefine; Yeh, Cheng-Ting; Opitz, Nina; Lanz, Christa; Schnable, Patrick S; Hochholdinger, Frank

2014-09-01

The maize (Zea mays L.) Aux/IAA protein RUM1 (ROOTLESS WITH UNDETECTABLE MERISTEMS 1) controls seminal and lateral root initiation. To identify RUM1-dependent gene expression patterns, RNA-Seq of the differentiation zone of primary roots of rum1 mutants and the wild type was performed in four biological replicates. In total, 2 801 high-confidence maize genes displayed differential gene expression with Fc ≥2 and FDR ≤1%. The auxin signalling-related genes rum1, like-auxin1 (lax1), lax2, (nam ataf cuc 1 nac1), the plethora genes plt1 (plethora 1), bbm1 (baby boom 1), and hscf1 (heat shock complementing factor 1) and the auxin response factors arf8 and arf37 were down-regulated in the mutant rum1. All of these genes except nac1 were auxin-inducible. The maize arf8 and arf37 genes are orthologues of Arabidopsis MP/ARF5 (MONOPTEROS/ARF5), which controls the differentiation of vascular cells. Histological analyses of mutant rum1 roots revealed defects in xylem organization and the differentiation of pith cells around the xylem. Moreover, histochemical staining of enlarged pith cells surrounding late metaxylem elements demonstrated that their thickened cell walls displayed excessive lignin deposition. In line with this phenotype, rum1-dependent mis-expression of several lignin biosynthesis genes was observed. In summary, RNA-Seq of RUM1-dependent gene expression in maize primary roots, in combination with histological and histochemical analyses, revealed the specific regulation of auxin signal transduction components by RUM1 and novel functions of RUM1 in vascular development. © The Author 2014. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Gene-breaking: A new paradigm for human retrotransposon-mediated gene evolution

PubMed Central

Wheelan, Sarah J.; Aizawa, Yasunori; Han, Jeffrey S.; Boeke, Jef D.

2005-01-01

The L1 retrotransposon is the most highly successful autonomous retrotransposon in mammals. This prolific genome parasite may on occasion benefit its host through genome rearrangements or adjustments of host gene expression. In examining possible effects of L1 elements on host gene expression, we investigated whether a full-length L1 element inserted in the antisense orientation into an intron of a cellular gene may actually split the gene's transcript into two smaller transcripts: (1) a transcript containing the upstream exons and terminating in the major antisense polyadenylation site (MAPS) of the L1, and (2) a transcript derived from the L1 antisense promoter (ASP) that includes the downstream exons of the gene. Bioinformatic analysis and experimental follow-up provide evidence for this L1 “gene-breaking” hypothesis. We identified three human genes apparently “broken” by L1 elements, as well as 12 more candidate genes. Most of the inserted L1 elements in our 15 candidate genes predate the human/chimp divergence. If indeed split, the transcripts of these genes may in at least one case encode potentially interacting proteins, and in another case may encode novel proteins. Gene-breaking represents a new mechanism through which L1 elements remodel mammalian genomes. PMID:16024818

LMI1-like genes involved in leaf margin development of Brassica napus.

PubMed

Ni, Xiyuan; Liu, Han; Huang, Jixiang; Zhao, Jianyi

2017-06-01

In rapeseed (Brassica napus L.), leaf margins are variable and can be entire, serrate, or lobed. In our previous study, the lobed-leaf gene (LOBED-LEAF 1, BnLL1) was mapped to a 32.1 kb section of B. napus A10. Two LMI1-like genes, BnaA10g26320D and BnaA10g26330D, were considered the potential genes that controlled the lobed-leaf trait in rapeseed. In the present study, these two genes and another homologous gene (BnaC04g00850D) were transformed into Arabidopsis thaliana (L.) Heynh. plants to identify their functions. All three LMI1-like genes of B. napus produced serrate leaf margins. The expression analysis indicated that the expression level of BnaA10g26320D determined the difference between lobed- and entire-leaved lines in rapeseed. Therefore, it is likely that BnaA10g26320D corresponds to BnLL1.

E3 ubiquitin ligase gene CMPG1-V from Haynaldia villosa L. contributes to powdery mildew resistance in common wheat (Triticum aestivum L.).

PubMed

Zhu, Yanfei; Li, Yingbo; Fei, Fei; Wang, Zongkuan; Wang, Wei; Cao, Aizhong; Liu, Yuan; Han, Shuang; Xing, Liping; Wang, Haiyan; Chen, Wei; Tang, Sanyuan; Huang, Xiahe; Shen, Qianhua; Xie, Qi; Wang, Xiue

2015-10-01

Powdery mildew is one of the most devastating wheat fungal diseases. A diploid wheat relative, Haynaldia villosa L., is highly resistant to powdery mildew, and its genetic resource of resistances, such as the Pm21 locus, is now widely used in wheat breeding. Here we report the cloning of a resistance gene from H. villosa, designated CMPG1-V, that encodes a U-box E3 ubiquitin ligase. Expression of the CMPG1-V gene was induced in the leaf and stem of H. villosa upon inoculation with Blumeria graminis f. sp. tritici (Bgt) fungus, and the presence of Pm21 is essential for its rapid induction of expression. CMPG1-V has conserved key residues for E3 ligase, and possesses E3 ligase activity in vitro and in vivo. CMPG1-V is localized in the nucleus, endoplasmic reticulum, plasma membrane and partially in trans-Golgi network/early endosome vesicles. Transgenic wheat over-expressing CMPG1-V showed improved broad-spectrum powdery mildew resistance at seedling and adult stages, associated with an increase in expression of salicylic acid-responsive genes, H2 O2 accumulation, and cell-wall protein cross-linking at the Bgt infection sites, and the expression of CMPG1-V in H. villosa was increased when treated with salicylic acid, abscisic acid and H2 O2 . These results indicate the involvement of E3 ligase in defense responses to Bgt fungus in wheat, particularly in broad-spectrum disease resistance, and suggest association of reactive oxidative species and the phytohormone pathway with CMPG1-V-mediated powdery mildew resistance. © 2015 The Authors The Plant Journal © 2015 John Wiley & Sons Ltd.

Selection of reference genes for quantitative gene expression normalization in flax (Linum usitatissimum L.).

PubMed

Huis, Rudy; Hawkins, Simon; Neutelings, Godfrey

2010-04-19

Quantitative real-time PCR (qRT-PCR) is currently the most accurate method for detecting differential gene expression. Such an approach depends on the identification of uniformly expressed 'housekeeping genes' (HKGs). Extensive transcriptomic data mining and experimental validation in different model plants have shown that the reliability of these endogenous controls can be influenced by the plant species, growth conditions and organs/tissues examined. It is therefore important to identify the best reference genes to use in each biological system before using qRT-PCR to investigate differential gene expression. In this paper we evaluate different candidate HKGs for developmental transcriptomic studies in the economically-important flax fiber- and oil-crop (Linum usitatissimum L). Specific primers were designed in order to quantify the expression levels of 20 different potential housekeeping genes in flax roots, internal- and external-stem tissues, leaves and flowers at different developmental stages. After calculations of PCR efficiencies, 13 HKGs were retained and their expression stabilities evaluated by the computer algorithms geNorm and NormFinder. According to geNorm, 2 Transcriptional Elongation Factors (TEFs) and 1 Ubiquitin gene are necessary for normalizing gene expression when all studied samples are considered. However, only 2 TEFs are required for normalizing expression in stem tissues. In contrast, NormFinder identified glyceraldehyde-3-phosphate dehydrogenase (GADPH) as the most stably expressed gene when all samples were grouped together, as well as when samples were classed into different sub-groups.qRT-PCR was then used to investigate the relative expression levels of two splice variants of the flax LuMYB1 gene (homologue of AtMYB59). LuMYB1-1 and LuMYB1-2 were highly expressed in the internal stem tissues as compared to outer stem tissues and other samples. This result was confirmed with both geNorm-designated- and Norm

Genetic Variants in the Apoptosis Gene BCL2L1 Improve Response to Interferon-Based Treatment of Hepatitis C Virus Genotype 3 Infection

PubMed Central

Clausen, Louise Nygaard; Weis, Nina; Ladelund, Steen; Madsen, Lone; Lunding, Suzanne; Tarp, Britta; Christensen, Peer Brehm; Krarup, Henrik Bygum; Møller, Axel; Gerstoft, Jan; Clausen, Mette Rye; Benfield, Thomas

2015-01-01

Genetic variation upstream of the apoptosis pathway has been associated with outcome of hepatitis C virus (HCV) infection. We investigated genetic polymorphisms in the intrinsic apoptosis pathway to assess their influence on sustained virological response (SVR) to pegylated interferon-α and ribavirin (pegIFN/RBV) treatment of HCV genotypes 1 and 3 infections. We conducted a candidate gene association study in a prospective cohort of 201 chronic HCV-infected individuals undergoing treatment with pegIFN/RBV. Differences between groups were compared in logistic regression adjusted for age, HCV viral load and interleukin 28B genotypes. Four single nucleotide polymorphisms (SNPs) located in the B-cell lymphoma 2-like 1 (BCL2L1) gene were significantly associated with SVR. SVR rates were significantly higher for carriers of the beneficial rs1484994 CC genotypes. In multivariate logistic regression, the rs1484994 SNP combined CC + TC genotypes were associated with a 3.4 higher odds ratio (OR) in SVR for the HCV genotype 3 (p = 0.02). The effect estimate was similar for genotype 1, but the association did not reach statistical significance. In conclusion, anti-apoptotic SNPs in the BCL2L1 gene were predictive of SVR to pegIFN/RBV treatment in HCV genotypes 1 and 3 infected individuals. These SNPs may be used in prediction of SVR, but further studies are needed. PMID:25648321

Increased Plp1 gene expression leads to massive microglial cell activation and inflammation throughout the brain

PubMed Central

Tatar, Carrie L; Appikatla, Sunita; Bessert, Denise A; Paintlia, Ajaib S; Singh, Inderjit; Skoff, Robert P

2010-01-01

PMD (Pelizaeus–Merzbacher disease) is a rare neurodegenerative disorder that impairs motor and cognitive functions and is associated with a shortened lifespan. The cause of PMD is mutations of the PLP1 [proteolipid protein 1 gene (human)] gene. Transgenic mice with increased Plp1 [proteolipid protein 1 gene (non-human)] copy number model most aspects of PMD patients with duplications. Hypomyelination and demyelination are believed to cause the neurological abnormalities in mammals with PLP1 duplications. We show, for the first time, intense microglial reactivity throughout the grey and white matter of a transgenic mouse line with increased copy number of the native Plp1 gene. Activated microglia in the white and grey matter of transgenic mice are found as early as postnatal day 7, before myelin commences in normal cerebra. This finding indicates that degeneration of myelin does not cause the microglial response. Microglial numbers are doubled due to in situ proliferation. Compared with the jp (jimpy) mouse, which has much more oligodendrocyte death and hardly any myelin, microglia in the overexpressors show a more dramatic microglial reactivity than jp, especially in the grey matter. Predictably, many classical markers of an inflammatory response, including TNF-α (tumour necrosis factor-α) and IL-6, are significantly up-regulated manyfold. Because inflammation is believed to contribute to axonal degeneration in multiple sclerosis and other neurodegenerative diseases, inflammation in mammals with increased Plp1 gene dosage may also contribute to axonal degeneration described in patients and rodents with PLP1 increased gene dosage. PMID:20885931

Improved transcription and translation with L-leucine stimulation of mTORC1 in Roberts syndrome.

PubMed

Xu, Baoshan; Gogol, Madelaine; Gaudenz, Karin; Gerton, Jennifer L

2016-01-05

Roberts syndrome (RBS) is a human developmental disorder caused by mutations in the cohesin acetyltransferase ESCO2. We previously reported that mTORC1 signaling was depressed and overall translation was reduced in RBS cells and zebrafish models for RBS. Treatment of RBS cells and zebrafish RBS models with L-leucine partially rescued mTOR function and protein synthesis, correlating with increased cell division and improved development. In this study, we use RBS cells to model mTORC1 repression and analyze transcription and translation with ribosome profiling to determine gene-level effects of L-leucine. L-leucine treatment partially rescued translational efficiency of ribosomal subunits, translation initiation factors, snoRNA production, and mitochondrial function in RBS cells, consistent with these processes being mTORC1 controlled. In contrast, other genes are differentially expressed independent of L-leucine treatment, including imprinted genes such as H19 and GTL2, miRNAs regulated by GTL2, HOX genes, and genes in nucleolar associated domains. Our study distinguishes between gene expression changes in RBS cells that are TOR dependent and those that are independent. Some of the TOR independent gene expression changes likely reflect the architectural role of cohesin in chromatin looping and gene expression. This study reveals the dramatic rescue effects of L-leucine stimulation of mTORC1 in RBS cells and supports that normal gene expression and translation requires ESCO2 function.

E1B-55K mediated regulation of RNF4 STUbL promotes HAdV gene expression.

PubMed

Müncheberg, Sarah; Hay, Ron T; Ip, Wing H; Meyer, Tina; Weiß, Christina; Brenke, Jara; Masser, Sawinee; Hadian, Kamyar; Dobner, Thomas; Schreiner, Sabrina

2018-04-25

HAdV E1B-55K is a multifunctional regulator of productive viral replication and oncogenic transformation in non-permissive mammalian cells. These functions depend on E1B-55K's posttranslational modification with the SUMO protein and its binding to HAdV E4orf6. Both early viral proteins recruit specific host factors to form an E3 Ubiquitin ligase complex that targets antiviral host substrates for proteasomal degradation. Recently, we reported that the PML-NB-associated factor Daxx represses efficient HAdV productive infection and is proteasomally degraded via a SUMO-E1B-55K-dependent, E4orf6-independent pathway, the details of which remained to be established.RNF4, a cellular SUMO-targeted Ubiquitin ligase (STUbL), induces ubiquitinylation of specific SUMOylated proteins and plays an essential role during DNA repair. Here, we show that E1B-55K recruits RNF4 to the insoluble nuclear matrix fraction of the infected cell to support RNF4/Daxx association, promoting Daxx PTM, and thus inhibiting this antiviral factor. Removing RNF4 from infected cells using RNAi resulted in blocking the proper establishment of viral replication centers and significantly diminished viral gene expression. These results provide a model for how HAdV antagonize the antiviral host responses by exploiting the functional capacity of cellular STUbLs. Thus, RNF4 and its STUbL function represent a positive factor during lytic infection and a novel candidate for future therapeutic antiviral intervention strategies. IMPORTANCE Daxx is a PML-NB-associated transcription factor, which was recently shown to repress efficient HAdV productive infection. To counteract this antiviral measurement during infection, Daxx is degraded via a novel pathway including viral E1B-55K and host proteasomes. This virus-mediated degradation is independent of the classical HAdV E3 Ubiquitin ligase complex, which is essential during viral infection to target other host antiviral substrates. To maintain productive viral life

Enrichment of the Antibiotic Resistance Gene tet(L) in an Alkaline Soil Fertilized With Plant Derived Organic Manure.

PubMed

Peng, Shuang; Dolfing, Jan; Feng, Youzhi; Wang, Yiming; Lin, Xiangui

2018-01-01

Fifteen antibiotic resistance genes (ARGs) and intI1 , a gene involved in horizontal gene transfer (HGT) of ARGs, were quantified in three different soil samples from a 22 year old field experiment that had received inorganic fertilizer (NPK), organic manure (OM; a mixture of wheat straw, soybean oil cake and cotton cake), and control fields that had received no fertilizer and manure (CK). Tet (L) was the most abundant ARG in OM, which also contained considerable levels of intI1 . Molecular analysis of yearly collected archived soils over the past 22 years showed that tet (L) and intI1 were higher in OM soils than in NPK soils. The relative abundance of tet (L) was essentially constant during these years, while the level of intI1 in OM soils decreased over time. The main genotype of tet (L) was the same in archived and in fresh soil, OM, and irrigation water. Phylogenetic analysis of the 16S rRNA genes of tetracycline-resistant bacteria (TRB) isolates indicated that the Firmucutes carrying tet (L) in OM were similar to those in the OM soil, suggesting that OM transferred TRB into the OM soils where they survived. Almost all of the TRB isolated from OM carried tet (L) and belonged to the Firmicutes . Survival of bacteria from the organic manure that carried tet (L) may be the cause of the increased level of tet (L) in OM soil.

Characterization of the equine 2'-5' oligoadenylate synthetase 1 (OAS1) and ribonuclease L (RNASEL) innate immunity genes

PubMed Central

Rios, Jonathan J; Perelygin, Andrey A; Long, Maureen T; Lear, Teri L; Zharkikh, Andrey A; Brinton, Margo A; Adelson, David L

2007-01-01

Background The mammalian OAS/RNASEL pathway plays an important role in antiviral host defense. A premature stop-codon within the murine Oas1b gene results in the increased susceptibility of mice to a number of flaviviruses, including West Nile virus (WNV). Mutations in either the OAS1 or RNASEL genes may also modulate the outcome of WNV-induced disease or other viral infections in horses. Polymorphisms in the human OAS gene cluster have been previously utilized for case-control analysis of virus-induced disease in humans. No polymorphisms have yet been identified in either the equine OAS1 or RNASEL genes for use in similar case-control studies. Results Genomic sequence for equine OAS1 was obtained from a contig assembly generated from a shotgun subclone library of CHORI-241 BAC 100I10. Specific amplification of regions of the OAS1 gene from 13 horses of various breeds identified 33 single nucleotide polymorphisms (SNP) and two microsatellites. RNASEL cDNA sequences were determined for 8 mammals and utilized in a phylogenetic analysis. The chromosomal location of the RNASEL gene was assigned by FISH to ECA5p17-p16 using two selected CHORI-241 BAC clones. The horse genomic RNASEL sequence was assembled. Specific amplification of regions of the RNASEL gene from 13 horses identified 31 SNPs. Conclusion In this report, two dinucleotide microsatellites and 64 single nucleotide polymorphisms within the equine OAS1 and RNASEL genes were identified. These polymorphisms are the first to be reported for these genes and will facilitate future case-control studies of horse susceptibility to infectious diseases. PMID:17822564

Over-Expression of Arabidopsis EDT1 Gene Confers Drought Tolerance in Alfalfa (Medicago sativa L.)

PubMed Central

Zheng, Guangshun; Fan, Cunying; Di, Shaokang; Wang, Xuemin; Xiang, Chengbin; Pang, Yongzhen

2017-01-01

Alfalfa (Medicago sativa L.) is an important legume forage crop with great economic value. However, as the growth of alfalfa is seriously affected by an inadequate supply of water, drought is probably the major abiotic environmental factor that most severely affects alfalfa production worldwide. In an effort to enhance alfalfa drought tolerance, we transformed the Arabidopsis Enhanced Drought Tolerance 1 (AtEDT1) gene into alfalfa via Agrobacterium-mediated transformation. Compared with wild type plants, drought stress treatment resulted in higher survival rates and biomass, but reduced water loss rates in the transgenic plants. Furthermore, transgenic alfalfa plants had increased stomatal size, but reduced stomatal density, and these stomatal changes contributed greatly to reduced water loss from leaves. Importantly, transgenic alfalfa plants exhibited larger root systems with larger root lengths, root weight, and root diameters than wild type plants. The transgenic alfalfa plants had reduced membrane permeability and malondialdehyde content, but higher soluble sugar and proline content, higher superoxide dismutase activity, higher chlorophyll content, enhanced expression of drought-responsive genes, as compared with wild type plants. Notably, transgenic alfalfa plants grew better in a 2-year field trial and showed enhanced growth performance with increased biomass yield. All of our morphological, physiological, and molecular analyses demonstrated that the ectopic expression of AtEDT1 improved growth and enhanced drought tolerance in alfalfa. Our study provides alfalfa germplasm for use in forage improvement programs, and may help to increase alfalfa production in arid lands. PMID:29326737

Over-Expression of Arabidopsis EDT1 Gene Confers Drought Tolerance in Alfalfa (Medicago sativa L.).

PubMed

Zheng, Guangshun; Fan, Cunying; Di, Shaokang; Wang, Xuemin; Xiang, Chengbin; Pang, Yongzhen

2017-01-01

Alfalfa ( Medicago sativa L.) is an important legume forage crop with great economic value. However, as the growth of alfalfa is seriously affected by an inadequate supply of water, drought is probably the major abiotic environmental factor that most severely affects alfalfa production worldwide. In an effort to enhance alfalfa drought tolerance, we transformed the Arabidopsis Enhanced Drought Tolerance 1 ( AtEDT1 ) gene into alfalfa via Agrobacterium -mediated transformation. Compared with wild type plants, drought stress treatment resulted in higher survival rates and biomass, but reduced water loss rates in the transgenic plants. Furthermore, transgenic alfalfa plants had increased stomatal size, but reduced stomatal density, and these stomatal changes contributed greatly to reduced water loss from leaves. Importantly, transgenic alfalfa plants exhibited larger root systems with larger root lengths, root weight, and root diameters than wild type plants. The transgenic alfalfa plants had reduced membrane permeability and malondialdehyde content, but higher soluble sugar and proline content, higher superoxide dismutase activity, higher chlorophyll content, enhanced expression of drought-responsive genes, as compared with wild type plants. Notably, transgenic alfalfa plants grew better in a 2-year field trial and showed enhanced growth performance with increased biomass yield. All of our morphological, physiological, and molecular analyses demonstrated that the ectopic expression of AtEDT1 improved growth and enhanced drought tolerance in alfalfa. Our study provides alfalfa germplasm for use in forage improvement programs, and may help to increase alfalfa production in arid lands.

Vitamin D receptor Fok1 polymorphism and blood lead concentration in children.

PubMed Central

Haynes, Erin N; Kalkwarf, Heidi J; Hornung, Richard; Wenstrup, Richard; Dietrich, Kim; Lanphear, Bruce P

2003-01-01

Variation in blood lead concentration is caused by a complex interaction of environmental, social, nutritional, and genetic factors. We evaluated the association between blood lead concentration and a vitamin D receptor (VDR) gene polymorphism. Environmental samples and blood were analyzed for lead, nutritional and behavioral factors were assessed, and VDR -Fok1 genotype was determined in 245 children. We found a significant interaction between floor dust lead and genotype on blood lead concentration. For every 1 microg/ft(2) increase in floor dust, children with VDR -FF genotype had a 1.1% increase in blood lead [95% confidence interval (CI), 0.69-1.5], VDR -Ff, 0.53% increase (95% CI, 0.1-0.92), and VDR -ff, 3.8% increase (95% CI, 1.2-6.3); however, at floor dust levels < 10 microg/ft(2), children with VDR -ff had the lowest blood lead concentrations. These data suggest that VDR -Fok1 is an effect modifier of the relationship of floor dust lead exposure and blood lead concentration. PMID:14527848

Tumor-recruited M2 macrophages promote gastric and breast cancer metastasis via M2 macrophage-secreted CHI3L1 protein.

PubMed

Chen, Yulei; Zhang, Siyuan; Wang, Qizhi; Zhang, Xiaobo

2017-02-01

The macrophage, one of the several key immune cell types, is believed to be involved in tumorigenesis. However, the mechanism of macrophages promoting tumor progression is largely unknown. The differentially secreted proteins of M1 and M2 macrophages were analyzed by mass spectrometry. We performed GST pull-down assay for the identification of cell-membrane receptors that interact with chitinase 3-like protein 1 (CHI3L1) protein. The mouse model was used to validate the function of CHI3L1 in cancer metastasis in vivo. Protein phosphorylation and gene expression were performed to study the signaling pathway activation of cancer cells after CHI3L1 treatment. M2 macrophage-secreted CHI3L1 promoted the metastasis of gastric and breast cancer cells in vitro and in vivo. The CHI3L1 protein functioned by interacting with interleukin-13 receptor α2 chain (IL-13Rα2) molecules on the plasma membranes of cancer cells. Activation of IL-13Rα2 by CHI3L1 triggered the activation of the mitogen-activated protein kinase signaling pathway, leading to the upregulated expression of matrix metalloproteinase genes, which promoted tumor metastasis. The results of this study indicated that the level of CHI3L1 protein in the sera of patients with gastric or breast cancer was significantly elevated compared with those of healthy donors. Our study revealed a novel aspect of macrophages with respect to cancer metastasis and showed that CHI3L1 could be a marker of metastatic gastric and breast cancer in patients.

L-Theanine Content and Related Gene Expression: Novel Insights into Theanine Biosynthesis and Hydrolysis among Different Tea Plant (Camellia sinensis L.) Tissues and Cultivars

PubMed Central

Liu, Zhi-Wei; Wu, Zhi-Jun; Li, Hui; Wang, Yong-Xin; Zhuang, Jing

2017-01-01

L-Theanine content has tissues and cultivars specificity in tea plant (Camellia sinensis L.), the correlations of theanine metabolic related genes expression profiles with theanine contents were explored in this study. L-theanine contents in the bud and 1st leaf, 2nd leaf, 3rd leaf, old leaf, stem, and lateral root were determined by HPLC from three C. sinensis cultivars, namely ‘Huangjinya’, ‘Anjibaicha’, and ‘Yingshuang’, respectively. The theanine contents in leaves and root of ‘Huangjinya’ were the highest, followed by ‘Anjibaicha’, and ‘Yingshuang’. The theanine contents in the leaves reduced as the leaf mature gradually, and in stem were the least. Seventeen genes encoding enzymes involved in theanine metabolism were identified from GenBank and our tea transcriptome database, including CsTS1, CsTS2, CsGS1, CsGS2, CsGOGAT-Fe, CsGOGAT-NAD(P)H, CsGDH1, CsGDH2, CsALT, CsSAMDC, CsADC, CsCuAO, CsPAO, CsNiR, CsNR, CsGGT1, and CsGGT3. The transcript profiles of those seventeen genes in the different tissues of three tea plant cultivars were analyzed comparatively. Among the different cultivars, the transcript levels of most selected genes in ‘Huangjinya’ were significantly higher than that in the ‘Anjibaicha’ and ‘Yingshuang’. Among the different tissues, the transcript levels of CsTS2, CsGS1, and CsGDH2 almost showed positive correlation with the theanine contents, while the other genes showed negative correlation with the theanine contents in most cases. The theanine contents showed correlations with related genes expression levels among cultivars and tissues of tea plant, and were determined by the integrated effect of the metabolic related genes. PMID:28439281

LRH-1 and PTF1-L coregulate an exocrine pancreas-specific transcriptional network for digestive function.

PubMed

Holmstrom, Sam R; Deering, Tye; Swift, Galvin H; Poelwijk, Frank J; Mangelsdorf, David J; Kliewer, Steven A; MacDonald, Raymond J

2011-08-15

We have determined the cistrome and transcriptome for the nuclear receptor liver receptor homolog-1 (LRH-1) in exocrine pancreas. Chromatin immunoprecipitation (ChIP)-seq and RNA-seq analyses reveal that LRH-1 directly induces expression of genes encoding digestive enzymes and secretory and mitochondrial proteins. LRH-1 cooperates with the pancreas transcription factor 1-L complex (PTF1-L) in regulating exocrine pancreas-specific gene expression. Elimination of LRH-1 in adult mice reduced the concentration of several lipases and proteases in pancreatic fluid and impaired pancreatic fluid secretion in response to cholecystokinin. Thus, LRH-1 is a key regulator of the exocrine pancreas-specific transcriptional network required for the production and secretion of pancreatic fluid.

Correlation of toxicity with lead content in root tip cells (Allium cepa L.).

PubMed

Carruyo, Ingrid; Fernández, Yusmary; Marcano, Letty; Montiel, Xiomara; Torrealba, Zaida

2008-12-01

The present study determines lead content in onion root tip cells (Allium cepa L.), correlating it with its toxicity. The treatment was carried at 25 +/- 0.5 degrees C using aqueous solutions of lead chloride at 0.1, 0.25, 0.50, 0.75, and 1 ppm for 12, 24, 48, and 72 h. For each treatment, a control where the lead solution was substituted by distilled water was included. After treatment, the meristems were fixed with a mixture of alcohol-acetic acid (3:1) and colored according to the technique of Feulgen. Lead content was quantified by graphite furnace absorption atomic spectrometry. The lead content in the roots ranged from 3.25 to 244.72 microg/g dry weight, with a direct relation with the concentration and time of exposure. A significant negative correlation was presented (r = -0.3629; p < 0.01) among lead content and root growth increment, and a positive correlation (r = 0.7750; p < 0.01) with the induction of chromosomic aberrations. In conclusion, lead is able to induce a toxic effect in the exposed roots, correlated with its content.

Functional conservation and diversification of the soybean maturity gene E1 and its homologs in legumes.

PubMed

Zhang, Xingzheng; Zhai, Hong; Wang, Yaying; Tian, Xiaojie; Zhang, Yupeng; Wu, Hongyan; Lü, Shixiang; Yang, Guang; Li, Yuqiu; Wang, Lu; Hu, Bo; Bu, Qingyun; Xia, Zhengjun

2016-07-13

Gene regulatory networks involved in flowering time and photoperiodic responses in legumes remain unknown. Although the major maturity gene E1 has been successfully deciphered in soybean, knowledge on the functional conservation of this gene is limited to a certain extent to E1 homologs in legumes. The ectopic expression of Phvul.009G204600 (PvE1L), an E1 homolog from common bean, delayed the onset of flowering in soybean. By contrast, the ectopic expression of Medtr2g058520 (MtE1L) from Medicago truncatula did not affect the flowering of soybean. Characterization of the late-flowering mte1l mutant indicated that MtE1L promoted flowering in Medicago truncatula. Moreover, all transgenic E1, PvE1L and MtE1L soybean lines exhibited phenotypic changes in terms of plant height. Transgenic E1 or PvE1L plants were taller than the wild-type, whereas transgenic MtE1L plants produced dwarf phenotype with few nodes and short internode. Thus, functional conservation and diversification of E1 family genes from legumes in the regulation of flowering and plant growth may be associated with lineage specification and genomic duplication.

The yeast Hot1 transcription factor is critical for activating a single target gene, STL1

PubMed Central

Bai, Chen; Tesker, Masha; Engelberg, David

2015-01-01

Transcription factors are commonly activated by signal transduction cascades and induce expression of many genes. They therefore play critical roles in determining the cell's fate. The yeast Hog1 MAP kinase pathway is believed to control the transcription of hundreds of genes via several transcription factors. To identify the bona fide target genes of Hog1, we inducibly expressed the spontaneously active variant Hog1D170A+F318L in cells lacking the Hog1 activator Pbs2. This system allowed monitoring the effects of Hog1 by itself. Expression of Hog1D170A+F318L in pbs2∆ cells imposed induction of just 105 and suppression of only 26 transcripts by at least twofold. We looked for the Hog1-responsive element within the promoter of the most highly induced gene, STL1 (88-fold). A novel Hog1 responsive element (HoRE) was identified and shown to be the direct target of the transcription factor Hot1. Unexpectedly, we could not find this HoRE in any other yeast promoter. In addition, the only gene whose expression was abolished in hot1∆ cells was STL1. Thus Hot1 is essential for transcription of just one gene, STL1. Hot1 may represent a class of transcription factors that are essential for transcription of a very few genes or even just one. PMID:25904326

L1C signal design options

USGS Publications Warehouse

Betz, J.W.; Cahn, C.R.; Dafesh, P.A.; Hegarty, C.J.; Hudnut, K.W.; Jones, A.J.; Keegan, R.; Kovach, K.; Lenahan, L.S.; Ma, H.H.; Rushanan, J.J.; Stansell, T.A.; Wang, C.C.; Yi, S.K.

2006-01-01

Design activities for a new civil signal centered at 1575.42 MHz, called L1C, began in 2003, and the Phase 1 effort was completed in 2004. The L1C signal design has evolved and matured during a Phase 2 design activity that began in 2005. Phase 2 has built on the initial design activity, guided by responses to international user surveys conducted during Phase 1. A common core of signal characteristics has been developed to provide advances in robustness and performance. The Phase 2 activity produced five design options, all drawing upon the core signal characteristics, while representing different blends of characteristics and capabilities. A second round of international user surveys was completed to solicit advice concerning these design options. This paper provides an update of the L1C design process, and describes the current L1C design options. Initial performance estimates are presented for each design option, displaying trades between signal tracking robustness, the speed and robustness of clock and ephemeris data, and the rate and robustness of other data message contents. Planned remaining activities are summarized, leading to optimization of the L1C design.

Prkar1a gene knockout in the pancreas leads to neuroendocrine tumorigenesis.

PubMed

Saloustros, Emmanouil; Salpea, Paraskevi; Starost, Matthew; Liu, Sissi; Faucz, Fabio R; London, Edra; Szarek, Eva; Song, Woo-Jin; Hussain, Mehboob; Stratakis, Constantine A

2017-01-01

Carney complex (CNC) is a rare disease associated with multiple neoplasias, including a predisposition to pancreatic tumors; it is caused most frequently by the inactivation of the PRKAR1A gene, a regulator of the cyclic AMP (cAMP)-dependent kinase (PKA). The method used was to create null alleles of prkar1a in mouse cells expressing pdx1 (Δ-Prkar1a). We found that these mice developed endocrine or mixed endocrine/acinar cell carcinomas with 100% penetrance by the age of 4-5 months. Malignant behavior of the tumors was seen as evidenced by stromal invasion and metastasis to locoregional lymph nodes. Histologically, most tumors exhibited an organoid pattern as seen in the islet-cell tumors. Biochemically, the lesions exhibited high PKA activity, as one would expect from deleting prkar1a The primary neuroendocrine nature of these tumor cells was confirmed by immunohistochemical staining and electron microscopy, the latter revealing the characteristic granules. Although the Δ-Prkar1a mice developed hypoglycemia after overnight fasting, insulin and glucagon levels in the plasma were normal. Negative immunohistochemical staining for the most commonly produced peptides (insulin, c-peptide, glucagon, gastrin and somatostatin) suggested that these tumors were non-functioning. We hypothesize that the recently identified multipotent pdx1+/insulin- cell in adult pancreas, gives rise to endocrine or mixed endocrine/acinar pancreatic malignancies with complete prkar1a deficiency. In conclusion, this mouse model supports the role of prkar1a as a tumor suppressor gene in the pancreas and points to the PKA pathway as a possible therapeutic target for these lesions. © 2017 Society for Endocrinology.

L1 Retrotransposon Heterogeneity in Ovarian Tumor Cell Evolution.

PubMed

Nguyen, Thu H M; Carreira, Patricia E; Sanchez-Luque, Francisco J; Schauer, Stephanie N; Fagg, Allister C; Richardson, Sandra R; Davies, Claire M; Jesuadian, J Samuel; Kempen, Marie-Jeanne H C; Troskie, Robin-Lee; James, Cini; Beaven, Elizabeth A; Wallis, Tristan P; Coward, Jermaine I G; Chetty, Naven P; Crandon, Alexander J; Venter, Deon J; Armes, Jane E; Perrin, Lewis C; Hooper, John D; Ewing, Adam D; Upton, Kyle R; Faulkner, Geoffrey J

2018-06-26

LINE-1 (L1) retrotransposons are a source of insertional mutagenesis in tumor cells. However, the clinical significance of L1 mobilization during tumorigenesis remains unclear. Here, we applied retrotransposon capture sequencing (RC-seq) to multiple single-cell clones isolated from five ovarian cancer cell lines and HeLa cells and detected endogenous L1 retrotransposition in vitro. We then applied RC-seq to ovarian tumor and matched blood samples from 19 patients and identified 88 tumor-specific L1 insertions. In one tumor, an intronic de novo L1 insertion supplied a novel cis-enhancer to the putative chemoresistance gene STC1. Notably, the tumor subclone carrying the STC1 L1 mutation increased in prevalence after chemotherapy, further increasing STC1 expression. We also identified hypomethylated donor L1s responsible for new L1 insertions in tumors and cultivated cancer cells. These congruent in vitro and in vivo results highlight L1 insertional mutagenesis as a common component of ovarian tumorigenesis and cancer genome heterogeneity. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

nm23-H1 gene driven by hTERT promoter induces inhibition of invasive phenotype and metastasis of lung cancer xenograft in mice.

PubMed

Fan, Yu; Yao, Yibing; Li, Lu; Wu, Zhihao; Xu, Feng; Hou, Mei; Wu, Heng; Shen, Yali; Wan, Haisu; Zhou, Qinghua

2013-02-01

Lung cancer is the leading cause of cancer death in both men and women worldwide. Tumor metastasis is an essential aspect of lung cancer progression and patient death. The nm23-H1 gene has been extensively investigated as a metastasis suppressor gene. Our previous studies have revealed: that a significant relationship exists between the low-level expression nm23-H1 in primary non-small cell lung cancer (NSCLC) with increased metastasis and a poor prognosis; that L9981-nm23-H1 cells (a nm23-H1 transfactant cell) exhibited lower cell proliferation rates, more G0/G1 phase growth, and an increase in apoptosis with a dramatic decrease in the tumor cells' ability to invade than L9981 cells did; and that L9981- nm23-H1 cells also demonstrated a significantly reduced lymph node and distant metastatic capacity in vivo than L9981 cells did in nude mice. In this study, we construct a plasmid containing the nm23-H1 gene, which was driven by the human telomerase reverse transcriptase (hTERT) promoter. We evaluated the anti-invasion and anti-metastatic effects of pGL3-hTP-nm23 on L9981, a human large cell lung cancer cell line with nm23-H1 negative expression, by transwell assay in vitro and bioluminescence in nude mice models. The toxicity of pGL3-hTP-nm23 and its effects on tumor growth were evaluated in nude mice models after gene therapy. The cell cycles, apoptosis, and proliferation of the nm23-H1 transfactant were also detected by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT assay) and flow cytometry (FCM). The results showed that the hTERT-promoter dramatically drives nm23-H1 gene expression, and induces inhibition of cell growth and migration in L9981-luc cells and MRC-5 cells in vitro. nm23-H1 also significantly inhibited the tumorigenesis and distant metastasis of L9981-luc cell in vivo. Moreover, no obvious side effect was detected in normal mouse tissues after intratumoral injection of the vector. The treatment of the nm23-H1 gene driven by h

Enhanced bioremediation of lead-contaminated soil by Solanum nigrum L. with Mucor circinelloides.

PubMed

Sun, Liqun; Cao, Xiufeng; Li, Min; Zhang, Xu; Li, Xinxin; Cui, Zhaojie

2017-04-01

Strain selected from mine tailings in Anshan for Pb bioremediation was characterized at the genetic level by internal transcribed spacer (ITS) sequencing. Results revealed that the strain belongs to Mucor circinelloides. Bioremediation of lead-contaminated soil was conducted using Solanum nigrum L. combined with M. circinelloides. The removal efficacy was in the order microbial/phytoremediation > phytoremediation > microbial remediation > control. The bioremediation rates were 58.6, 47.2, and 40.2% in microbial/phytoremediation, microbial remediation, and phytoremediation groups, respectively. Inoculating soil with M. circinelloides enhanced Pb removal and S. nigrum L. growth. The bioaccumulation factor (BF, 1.43), enrichment factor (EF, 1.56), and translocation factor (TF, 1.35) were higher than unit, suggesting an efficient ability of S. nigrum L. in Pb bioremediation. Soil fertility was increased after bioremediation according to change in enzyme activities. The results indicated that inoculating S. nigrum L. with M. circinelloides enhanced its efficiency for phytoremediation of soil contaminated with Pb.

OsNucleolin1-L Expression in Arabidopsis Enhances Photosynthesis via Transcriptome Modification under Salt Stress Conditions.

PubMed

Udomchalothorn, Thanikarn; Plaimas, Kitiporn; Sripinyowanich, Siriporn; Boonchai, Chutamas; Kojonna, Thammaporn; Chutimanukul, Panita; Comai, Luca; Buaboocha, Teerapong; Chadchawan, Supachitra

2017-04-01

OsNUC1 encodes rice nucleolin, which has been shown to be involved in salt stress responses. Expression of the full-length OsNUC1 gene in Arabidopsis resulted in hypersensitivity to ABA during germination. Transcriptome analysis of the transgenic lines, in comparison with the wild type, revealed that the RNA abundance of >1,900 genes was significantly changed under normal growth conditions, while under salt stress conditions the RNAs of 999 genes were found to be significantly regulated. Gene enrichment analysis showed that under normal conditions OsNUC1 resulted in repression of genes involved in photosynthesis, while in salt stress conditions OsNUC1 increased expression of the genes involved in the light-harvesting complex. Correspondingly, the net rate of photosynthesis of the transgenic lines was increased under salt stress. Transgenic rice lines with overexpression of the OsNUC1-L gene were generated and tested for photosynthetic performance under salt stress conditions. The transgenic rice lines treated with salt stress at the booting stage had a higher photosynthetic rate and stomatal conductance in flag leaves and second leaves than the wild type. Moreover, higher contents of Chl a and carotenoids were found in flag leaves of the transgenic rice. These results suggest a role for OsNUC1 in the modification of the transcriptome, especially the gene transcripts responsible for photosynthesis, leading to stabilization of photosynthesis under salt stress conditions. © The Author 2017. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Structural organization of the porcine and human genes coding for a leydig cell-specific insulin-like peptide (LEY I-L) and chromosomal localization of the human gene (INSL3)

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

Burkhardt E.; Adham, I.M.; Brosig, B.

1994-03-01

Leydig insulin-like protein (LEY I-L) is a member of the insulin-like hormone superfamily. The LEY I-L gene (designated INSL3) is expressed exclusively in prenatal and postnatal Leydig cells. The authors report here the cloning and nucleotide sequence of porcine and human LEY I-L genes including the 5[prime] regions. Both genes consist of two exons and one intron. The organization of the LEY I-L gene is similar to that of insulin and relaxin. The transcription start site in the porcine and human LEY I-L gene is localized 13 and 14 bp upstream of the translation start site, respectively. Alignment of themore » 5[prime] flanking regions of both genes reveals that the first 107 nucleotides upstream of the transcription start site exhibit an overall sequence similarity of 80%. This conserved region contains a consensus TATAA box, a CAAT-like element (GAAT), and a consensus SP1 sequence (GGGCGG) at equivalent positions in both genes and therefore may play a role in regulation of expression of the LEY I-L gene. The porcine and human genome contains a single copy of the LEY I-L gene. By in situ hybridization, the human gene was assigned to bands p13.2-p12 of the short arm of chromosome 19. 25 refs., 6 figs.« less

The BlueGene/L supercomputer

NASA Astrophysics Data System (ADS)

Bhanota, Gyan; Chen, Dong; Gara, Alan; Vranas, Pavlos

2003-05-01

The architecture of the BlueGene/L massively parallel supercomputer is described. Each computing node consists of a single compute ASIC plus 256 MB of external memory. The compute ASIC integrates two 700 MHz PowerPC 440 integer CPU cores, two 2.8 Gflops floating point units, 4 MB of embedded DRAM as cache, a memory controller for external memory, six 1.4 Gbit/s bi-directional ports for a 3-dimensional torus network connection, three 2.8 Gbit/s bi-directional ports for connecting to a global tree network and a Gigabit Ethernet for I/O. 65,536 of such nodes are connected into a 3-d torus with a geometry of 32×32×64. The total peak performance of the system is 360 Teraflops and the total amount of memory is 16 TeraBytes.

Copolymers of poly-L-lysine with serine and tryptophan form stable DNA vectors: implications for receptor-mediated gene transfer.

PubMed

Gómez-Valadés, A G; Molas, M; Vidal-Alabró, A; Bermúdez, J; Bartrons, R; Perales, J C

2005-01-20

Inefficient gene transfer and poor stability in physiological medium are important shortcomings for receptor-mediated gene transfer vectors. Here, we evaluate vectors formulated with random copolymers of L-lysine/L-serine (3:1) and L-lysine/L-tryptophan (4:1), focusing on both their biophysical and functional characterization. By means of dynamic light scattering (DLS) and transmission electron microscopy (TEM), we demonstrate that poly-L-lysine (pK), poly-L-lysine-L-tryptophan (pKW) and poly-L-lysine-L-serine (pKS) are able to form compacted, small particles when mixed with plasmid DNA in the absence of salt. Upon dilution in physiological medium, copolymers of both lys/ser and lys/trp do not aggregate, in contrast with poly-L-lysine DNA complexes as determined by scattering, DLS and TEM measurements. Tight packing, as demonstrated by resistance to heparin, SDS and trypsin treatments, is also featured in tryptophan-containing complexes. Successful receptor-mediated endocytosis gene transfer using galactosylated copolymers into cells expressing the asiagloglycoprotein receptor correlated with lack of aggregation. Particles obtained using galactosylated poly-L-lysine-L-tryptophan (Gal-pKW) copolymer demonstrated specific receptor-mediated gene transfer since reporter gene activity dropped in the presence of an excess ligand in the culture medium during transfection. Although copolymers of galactosylated poly-L-lysine-L-serine (Gal-pKS) do not aggregate in the presence of salt, they are not able to internalize in a specific receptor-mediated endocytosis fashion. The introduction of bulky aromatic/hydrophobic (tryptophan) or hydrophillic (serine) moieties into the positively charged vectors allows the compacted particles to disperse into salt-containing medium avoiding salt-induced aggregation. Moreover, tryptophan-containing particles are able to mediate specific gene transfer via receptor-mediated endocytosis.

The divergently transcribed genes encoding yeast ribosomal proteins L46 and S24 are activated by shared RPG-boxes.

PubMed Central

Kraakman, L S; Mager, W H; Maurer, K T; Nieuwint, R T; Planta, R J

1989-01-01

Transcription of the majority of the ribosomal protein (rp) genes in yeast is activated through common cis-acting elements, designated RPG-boxes. These elements have been shown to act as specific binding sites for the protein factor TUF/RAP1/GRF1 in vitro. Two such elements occur in the intergenic region separating the divergently transcribed genes encoding L46 and S24. To investigate whether the two RPG-boxes mediate transcription activation of both the L46 and S24 gene, two experimental strategies were followed: cloning of the respective genes on multicopy vectors and construction of fusion genes. Cloning of the L46 + S24 gene including the intergenic region in a multicopy yeast vector indicated that both genes are transcriptionally active. Using constructs in which only the S24 or the L46 gene is present, with or without the intergenic region, we obtained evidence that the intergenic region is indispensable for transcription activation of either gene. To demarcate the element(s) responsible for this activation, fusions of the intergenic region in either orientation to the galK reporter gene were made. Northern analysis of the levels of hybrid mRNA demonstrated that the intergenic region can serve as an heterologous promoter when it is in the 'S24-orientation'. Surprisingly, however, when fused in the reverse orientation the intergenic region did hardly confer transcription activity on the fusion gene. Furthermore, a 274 bp FnuDII-FnuDII fragment from the intergenic region that contains the RPG-boxes, could replace the naturally occurring upstream activation site (UASrpg) of the L25 rp-gene only when inserted in the 'S24-orientation'. Removal of 15 bp from the FnuDII fragment appeared to be sufficient to obtain transcription activation in the 'L46 orientation' as well. Analysis of a construct in which the RPG-boxes were selectively deleted from the promoter region of the L46 gene indicated that the RPG-boxes are needed for efficient transcriptional activation of

The divergently transcribed genes encoding yeast ribosomal proteins L46 and S24 are activated by shared RPG-boxes.

PubMed

Kraakman, L S; Mager, W H; Maurer, K T; Nieuwint, R T; Planta, R J

1989-12-11

Transcription of the majority of the ribosomal protein (rp) genes in yeast is activated through common cis-acting elements, designated RPG-boxes. These elements have been shown to act as specific binding sites for the protein factor TUF/RAP1/GRF1 in vitro. Two such elements occur in the intergenic region separating the divergently transcribed genes encoding L46 and S24. To investigate whether the two RPG-boxes mediate transcription activation of both the L46 and S24 gene, two experimental strategies were followed: cloning of the respective genes on multicopy vectors and construction of fusion genes. Cloning of the L46 + S24 gene including the intergenic region in a multicopy yeast vector indicated that both genes are transcriptionally active. Using constructs in which only the S24 or the L46 gene is present, with or without the intergenic region, we obtained evidence that the intergenic region is indispensable for transcription activation of either gene. To demarcate the element(s) responsible for this activation, fusions of the intergenic region in either orientation to the galK reporter gene were made. Northern analysis of the levels of hybrid mRNA demonstrated that the intergenic region can serve as an heterologous promoter when it is in the 'S24-orientation'. Surprisingly, however, when fused in the reverse orientation the intergenic region did hardly confer transcription activity on the fusion gene. Furthermore, a 274 bp FnuDII-FnuDII fragment from the intergenic region that contains the RPG-boxes, could replace the naturally occurring upstream activation site (UASrpg) of the L25 rp-gene only when inserted in the 'S24-orientation'. Removal of 15 bp from the FnuDII fragment appeared to be sufficient to obtain transcription activation in the 'L46 orientation' as well. Analysis of a construct in which the RPG-boxes were selectively deleted from the promoter region of the L46 gene indicated that the RPG-boxes are needed for efficient transcriptional activation of

Contrastive Analysis of English and Japanese Demonstratives from the Perspective of L1 and L2 Acquisition.

ERIC Educational Resources Information Center

Niimura, Tomomi; Hayashi, Brenda

1996-01-01

Presents a contrastive analysis of English and Japanese demonstratives based on the first- (L1) and second-language (L2) data of an earlier study. First, the traditional explanations and their alternative models for English and Japanese are presented, then, all models are tested with the L1 and L2 data, which leads to a discussion of the different…

Molecular analysis of the L1CAM gene in patients with X-linked hydrocephalus demonstrates eight novel mutations and suggests non-allelic heterogeneity of the trait.

PubMed

Gu, S M; Orth, U; Zankl, M; Schröder, J; Gal, A

1997-08-22

Eight novel mutations were identified in the gene encoding L1CAM, a neural cell adhesion protein, in patients/families with X-linked hydrocephalus (XHC) providing additional evidence for extreme allelic heterogeneity of the trait. The two nonsense mutations (Gln440Ter and Gln1042Ter) result most likely in functional null-alleles and complete absence of L1CAM at the cell surface. The four missense mutations (Leu482Pro, Ser542Pro, Met741Thr, and Val752Met) as well as delSer526 may considerably alter the structure of L1CAM. Interestingly, a missense mutation in an XHC family predicting the Val768Ile change in the second fibronectin type III domain of L1CAM was found not only in the two affected cousins and their obligate carrier mothers but also in two unaffected male relatives of the patients. Several possible explanations of this finding are discussed; the most likely being that Val768Ile is a rare non-pathogenic variant. If this were indeed the case, our data suggest that the XHC in this family is not due to a mutation of the L1CAM gene, i.e., that, in addition to the extreme allelic heterogeneity of XHC, a non-allelic form of genetic heterogeneity may also exist in this trait.

[Physical mapping of the genes px and cld coding peroxidase and cold-regulated protein in maize (Zea mays L.)].

PubMed

Ning, S B; Wang, L; Song, Y C

2000-01-01

Peroxidase plays a key role in plant disease resistance, cold stress and some developmental processes, and cold-regulated protein functions necessarily in reaction of plants on cold or heat stress. Recent studies showed that these processes in plant cells were involved in programmed cell death (PCD). Using a biotin-labelled in situ hybridization (ISH) technique, we physically mapped the genes px and cld coding peroxidase and cold-regulated protein respectively onto maize chromosomes. Both DAB and fluorescence detection systems gave the identical results, the probe uaz235 corresponding to gene px was localized onto the long arm of chromosome 2 (2L) and 7L, and csu19 corresponding to gene cld was hybridized onto 4L and 5L. The percentage distances (from the hybridization sites to centromeres) of uaz235 in 2L and 7L were 45.4 +/- 1.3 and 67.4 +/- 3.7 respectively, and those of csu19 in 4L and 5L were 68.6 +/- 2.6 and 58.2 +/- 1.6 respectively. The physical positions of px in 2L and cld in 4L coincide with those in their genetic map pattern. The results also show that both of these genes have duplicated sites in maize genome.

Lead Exposure during Early Human Development and DNA Methylation of Imprinted Gene Regulatory Elements in Adulthood

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

Li, Yue; Xie, Changchun; Murphy, Susan K.

Here, lead exposure during early development causes neurodevelopmental disorders by unknown mechanisms. Epidemiologic studies have focused recently on determining associations between lead exposure and global DNA methylation; however, such approaches preclude the identification of loci that may alter human disease risk. The objective of this study was to determine whether maternal, postnatal, and early childhood lead exposure can alter the differentially methylated regions (DMRs) that control the monoallelic expression of imprinted genes involved in metabolism, growth, and development. Questionnaire data and serial blood lead levels were obtained from 105 participants (64 females, 41 males) of the Cincinnati Lead Study frommore » birth to 78 months. When participants were adults, we used Sequenom EpiTYPER assays to test peripheral blood DNA to quantify CpG methylation in peripheral blood leukocytes at DMRs of 22 human imprinted genes. Statistical analyses were conducted using linear regression. Mean blood lead concentration from birth to 78 months was associated with a significant decrease in PEG3 DMR methylation (β = –0.0014; 95% CI: –0.0023, –0.0005, p = 0.002), stronger in males (β = –0.0024; 95% CI: –0.0038, –0.0009, p = 0.003) than in females (β = –0.0009; 95% CI: –0.0020, 0.0003, p = 0.1). Elevated mean childhood blood lead concentration was also associated with a significant decrease in IGF2/H19 (β = –0.0013; 95% CI: –0.0023, –0.0003, p = 0.01) DMR methylation, but primarily in females, (β = –0.0017; 95% CI: –0.0029, –0.0006, p = 0.005) rather than in males, (β = –0.0004; 95% CI: –0.0023, 0.0015, p = 0.7). Elevated blood lead concentration during the neonatal period was associated with higher PLAGL1/HYMAI DMR methylation regardless of sex (β = 0.0075; 95% CI: 0.0018, 0.0132, p = 0.01). The magnitude of associations between cumulative lead exposure and CpG methylation remained unaltered from 30 to 78 months. Our

Lead Exposure during Early Human Development and DNA Methylation of Imprinted Gene Regulatory Elements in Adulthood

DOE PAGES

Li, Yue; Xie, Changchun; Murphy, Susan K.; ...

2015-06-26

Here, lead exposure during early development causes neurodevelopmental disorders by unknown mechanisms. Epidemiologic studies have focused recently on determining associations between lead exposure and global DNA methylation; however, such approaches preclude the identification of loci that may alter human disease risk. The objective of this study was to determine whether maternal, postnatal, and early childhood lead exposure can alter the differentially methylated regions (DMRs) that control the monoallelic expression of imprinted genes involved in metabolism, growth, and development. Questionnaire data and serial blood lead levels were obtained from 105 participants (64 females, 41 males) of the Cincinnati Lead Study frommore » birth to 78 months. When participants were adults, we used Sequenom EpiTYPER assays to test peripheral blood DNA to quantify CpG methylation in peripheral blood leukocytes at DMRs of 22 human imprinted genes. Statistical analyses were conducted using linear regression. Mean blood lead concentration from birth to 78 months was associated with a significant decrease in PEG3 DMR methylation (β = –0.0014; 95% CI: –0.0023, –0.0005, p = 0.002), stronger in males (β = –0.0024; 95% CI: –0.0038, –0.0009, p = 0.003) than in females (β = –0.0009; 95% CI: –0.0020, 0.0003, p = 0.1). Elevated mean childhood blood lead concentration was also associated with a significant decrease in IGF2/H19 (β = –0.0013; 95% CI: –0.0023, –0.0003, p = 0.01) DMR methylation, but primarily in females, (β = –0.0017; 95% CI: –0.0029, –0.0006, p = 0.005) rather than in males, (β = –0.0004; 95% CI: –0.0023, 0.0015, p = 0.7). Elevated blood lead concentration during the neonatal period was associated with higher PLAGL1/HYMAI DMR methylation regardless of sex (β = 0.0075; 95% CI: 0.0018, 0.0132, p = 0.01). The magnitude of associations between cumulative lead exposure and CpG methylation remained unaltered from 30 to 78 months. Our

Candidate gene association mapping of Sclerotinia stalk rot resistance in sunflower (Helianthus annuus L.) uncovers the importance of COI1 homologs.

PubMed

Talukder, Zahirul I; Hulke, Brent S; Qi, Lili; Scheffler, Brian E; Pegadaraju, Venkatramana; McPhee, Kevin; Gulya, Thomas J

2014-01-01

Functional markers for Sclerotinia basal stalk rot resistance in sunflower were obtained using gene-level information from the model species Arabidopsis thaliana. Sclerotinia stalk rot, caused by Sclerotinia sclerotiorum, is one of the most destructive diseases of sunflower (Helianthus annuus L.) worldwide. Markers for genes controlling resistance to S. sclerotiorum will enable efficient marker-assisted selection (MAS). We sequenced eight candidate genes homologous to Arabidopsis thaliana defense genes known to be associated with Sclerotinia disease resistance in a sunflower association mapping population evaluated for Sclerotinia stalk rot resistance. The total candidate gene sequence regions covered a concatenated length of 3,791 bp per individual. A total of 187 polymorphic sites were detected for all candidate gene sequences, 149 of which were single nucleotide polymorphisms (SNPs) and 38 were insertions/deletions. Eight SNPs in the coding regions led to changes in amino acid codons. Linkage disequilibrium decay throughout the candidate gene regions declined on average to an r (2) = 0.2 for genetic intervals of 120 bp, but extended up to 350 bp with r (2) = 0.1. A general linear model with modification to account for population structure was found the best fitting model for this population and was used for association mapping. Both HaCOI1-1 and HaCOI1-2 were found to be strongly associated with Sclerotinia stalk rot resistance and explained 7.4 % of phenotypic variation in this population. These SNP markers associated with Sclerotinia stalk rot resistance can potentially be applied to the selection of favorable genotypes, which will significantly improve the efficiency of MAS during the development of stalk rot resistant cultivars.

PD-1/PD-L1 pathway inhibitors in advanced prostate cancer.

PubMed

Isaacsson Velho, Pedro; Antonarakis, Emmanuel S

2018-05-01

Pharmacological inhibition of immune checkpoint receptors or their ligands represents a transformative breakthrough in the management of multiple cancers. However, immune checkpoint inhibitors have yet to be FDA-approved for the management of metastatic prostate cancer (PCa), the commonest non-cutaneous malignancy in men. Areas covered: We review our current understanding of the PD-1/PD-L1 pathway in cancer, the use of anti-PD-1/PD-L1 therapeutics in PCa, and potential subgroups of PCa patients who may derive the greatest benefit from these agents (such as men with tumors that have expression of PD-L1 and/or high mutational load). We also review the prior and current clinical trials evaluating the blockade of PD-1/PD-L1 in PCa, highlighting some of the key ongoing studies of greatest relevance to the field. Expert commentary: Clinical trials investigating PD-1/PD-L1 inhibitors should be encouraged in patients with PCa. While it is unlikely that immune checkpoint monotherapies will produce long-lasting responses in a substantial proportion of patients, there is early evidence of activity in some patient subsets. These subgroups may include those with high PD-L1 expression, those with hypermutated or microsatellite-unstable tumors, and those enriched for germline and/or somatic DNA-repair gene mutations (e.g. intraductal/ductal histology, primary Gleason pattern 5, and perhaps AR-V7-positive tumors).

Identification of host genes leading to West Nile virus encephalitis in mice brain using RNA-seq analysis

PubMed Central

Kumar, Mukesh; Belcaid, Mahdi; Nerurkar, Vivek R.

2016-01-01

Differential host responses may be critical determinants of distinct pathologies of West Nile virus (WNV) NY99 (pathogenic) and WNV Eg101 (non-pathogenic) strains. We employed RNA-seq technology to analyze global differential gene expression in WNV-infected mice brain and to identify the host cellular factors leading to lethal encephalitis. We identified 1,400 and 278 transcripts, which were differentially expressed after WNV NY99 and WNV Eg101 infections, respectively, and 147 genes were common to infection with both the viruses. Genes that were up-regulated in infection with both the viruses were mainly associated with interferon signaling. Genes associated with inflammation and cell death/apoptosis were only expressed after WNV NY99 infection. We demonstrate that differences in the activation of key pattern recognition receptors resulted in the induction of unique innate immune profiles, which corresponded with the induction of interferon and inflammatory responses. Pathway analysis of differentially expressed genes indicated that after WNV NY99 infection, TREM-1 mediated activation of toll-like receptors leads to the high inflammatory response. In conclusion, we have identified both common and specific responses to WNV NY99 and WNV Eg101 infections as well as genes linked to potential resistance to infection that may be targets for therapeutics. PMID:27211830

Cucumber (Cucumis sativus L.) Nitric Oxide Synthase Associated Gene1 (CsNOA1) Plays a Role in Chilling Stress

PubMed Central

Liu, Xingwang; Liu, Bin; Xue, Shudan; Cai, Yanlinq; Qi, Wenzhu; Jian, Chen; Xu, Shuo; Wang, Ting; Ren, Huazhong

2016-01-01

Nitric oxide (NO) is a gaseous signaling molecule in plants, transducing information as a result of exposure to low temperatures. However, the underlying molecular mechanism linking NO with chilling stress is not well understood. Here, we functionally characterized the cucumber (Cucumis sativus L.) nitric oxide synthase-associated gene, NITRIC OXIDE ASSOCIATED 1 (CsNOA1). Expression analysis of CsNOA1, using quantitative real-time PCR, in situ hybridization, and a promoter::β-glucuronidase (GUS) reporter assay, revealed that it is expressed mainly in the root and shoot apical meristem (SAM), and that expression is up-regulated by low temperatures. A CsNOA1-GFP fusion protein was found to be localized in the mitochondria, and ectopic expression of CsNOA1 in the A. thaliana noa1 mutant partially rescued the normal phenotype. When overexpressing CsNOA1 in the Atnoa1 mutant under normal condition, no obvious phenotypic differences was observed between its wild type and transgenic plants. However, the leaves from mutant plant grown under chilling conditions showed hydrophanous spots and wilting. Physiology tolerance markers, chlorophyll fluorescence parameter (Fv/Fm), and electrolyte leakage, were observed to dramatically change, compared mutant to overexpressing lines. Transgenic cucumber plants revealed that the gene is required by seedlings to tolerate chilling stress: constitutive over-expression of CsNOA1 led to a greater accumulation of soluble sugars, starch, and an up-regulation of Cold-regulatory C-repeat binding factor3 (CBF3) expression as well as a lower chilling damage index (CI). Conversely, suppression of CsNOA1 expression resulted in the opposite phenotype and a reduced NO content compared to wild type plants. Those results suggest that CsNOA1 regulates cucumber seedlings chilling tolerance. Additionally, under normal condition, we took several classic inhibitors to perform, and detect endogenous NO levels in wild type cucumber seedling. The results

Identification of Regulatory Genes Implicated in Continuous Flowering of Longan (Dimocarpus longan L.)

PubMed Central

Jia, Tianqi; Wei, Danfeng; Meng, Shan; Allan, Andrew C.; Zeng, Lihui

2014-01-01

Longan (Dimocarpus longan L.) is a tropical/subtropical fruit tree of significant economic importance in Southeast Asia. However, a lack of transcriptomic and genomic information hinders research on longan traits, such as the control of flowering. In this study, high-throughput RNA sequencing (RNA-Seq) was used to investigate differentially expressed genes between a unique longan cultivar ‘Sijimi’(S) which flowers throughout the year and a more typical cultivar ‘Lidongben’(L) which flowers only once in the season, with the aim of identifying candidate genes associated with continuous flowering. 36,527 and 40,982 unigenes were obtained by de novo assembly of the clean reads from cDNA libraries of L and S cultivars. Additionally 40,513 unigenes were assembled from combined reads of these libraries. A total of 32,475 unigenes were annotated by BLAST search to NCBI non-redundant protein (NR), Swiss-Prot, Clusters of Orthologous Groups (COGs) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases. Of these, almost fifteen thousand unigenes were identified as significantly differentially expressed genes (DEGs) by using Reads Per kb per Million reads (RPKM) method. A total of 6,415 DEGs were mapped to 128 KEGG pathways, and 8,743 DEGs were assigned to 54 Gene Ontology categories. After blasting the DEGs to public sequence databases, 539 potential flowering-related DEGs were identified. In addition, 107 flowering-time genes were identified in longan, their expression levels between two longan samples were compared by RPKM method, of which the expression levels of 15 were confirmed by real-time quantitative PCR. Our results suggest longan homologues of SHORT VEGETATIVE PHASE (SVP), GIGANTEA (GI), F-BOX 1 (FKF1) and EARLY FLOWERING 4 (ELF4) may be involved this flowering trait and ELF4 may be a key gene. The identification of candidate genes related to continuous flowering will provide new insight into the molecular process of regulating flowering time in woody

Proteomic analysis of rodent ribosomes revealed heterogeneity including ribosomal proteins L10-like, L22-like 1, and L39-like.

PubMed

Sugihara, Yoshihiko; Honda, Hiroki; Iida, Tomoharu; Morinaga, Takuma; Hino, Shingo; Okajima, Tetsuya; Matsuda, Tsukasa; Nadano, Daita

2010-03-05

Heterogeneity of ribosome structure, due to variations in ribosomal protein composition, has been shown to be of physiological significance in plants and yeast. Mammalian genomics have demonstrated numerous genes that are paralogous to genes encoding ribosomal proteins. Although the vast majority are considered to be pseudogenes, mRNA expression of a few paralogues, such as human ribosomal protein L39-like/L39-2, has been reported. In the present study, ribosomes from the liver, mammary gland, and testis of rodents were analyzed using a combination of two-dimensional gel electrophoresis under radical-free and highly reducing conditions, and mass spectrometry. This system allowed identification of 78 ribosomal proteins and Rack1 from a single gel. The degree of heterogeneity was far less than that reported for plant and yeast ribosomes, and was in accord with published biochemical and genetic data for mammalian ribosomes. Nevertheless, an uncharacterized paralogue of ribosomal protein L22, ribosomal protein L22-like 1, was identified as a minor ribosomal component. Ribosomal proteins L10-like and L39-like, paralogues of ribosomal proteins L10 and L39, respectively, were found in ribosomes only from the testis. Reverse transcription-polymerase chain reaction yielded supportive evidence for specific expression of L10-like and L39-like in the testis. Newly synthesized L39-like is likely to be transported to the nucleolus, where ribosome biosynthesis occurs, and then incorporated into translating ribosomes in the cytoplasm. Heterogeneity of mammalian testicular ribosomes is structurally non-negligible, and may offer valuable insights into the function of the customized ribosome.

[Inheritance of bc1 gene in intersubspecific hybrids of rice (Oryza sativa L.)].

PubMed

Lü, Chuan-Gen; Zong, Shou-Yu; Zhao, Ling; Qi, Qing-Ming; Zou, Jiang-Shi; Ikehashi, Hiroshi

2004-10-01

Distorted segregation of the brittle culm-1 gene (bc1) on rice chromosome 3 was found with greatly increased or decreased frequency of bc1 bc1 genotype in inter-subspecific hybrids, although the gene normally transmitted to its offspring following the Mendelian Law in intra-subspecific hybrids. In a combination of Kamairazu//Ketan Nangka/Kamairazu,an increased frequency of bc1 bc1 in F1, normal segregation in F2, and increased and decreased frequency in a few F3 and F4 lines were observed. In a cross of IR36/Kamairazu, decreased frequency in F2, both normal and decreased segregations in F3 and F4, and a few lines of increased ratio in F4 were found. In F2 of Ketan Nangka/IR36//Kamairazu, increased and decreased and normal segregations were all observed. There was no significant correlation between the frequency of bc1 bc1 and pollen fertility. It implied that distorted segregation of bc1 was caused by selective fertilization of male gametes, which were governed by gametophyte genes of ga2, ga3 and ga14 on chromosome 3.

Epigenetic analysis leads to identification of HNF1B as a subtype-specific susceptibility gene for ovarian cancer

PubMed Central

Shen, Hui; Fridley, Brooke L.; Song, Honglin; Lawrenson, Kate; Cunningham, Julie M.; Ramus, Susan J.; Cicek, Mine S.; Tyrer, Jonathan; Stram, Douglas; Larson, Melissa C.; Köbel, Martin; Ziogas, Argyrios; Zheng, Wei; Yang, Hannah P.; Wu, Anna H.; Wozniak, Eva L.; Woo, Yin Ling; Winterhoff, Boris; Wik, Elisabeth; Whittemore, Alice S.; Wentzensen, Nicolas; Weber, Rachel Palmieri; Vitonis, Allison F.; Vincent, Daniel; Vierkant, Robert A.; Vergote, Ignace; Van Den Berg, David; Van Altena, Anne M.; Tworoger, Shelley S.; Thompson, Pamela J.; Tessier, Daniel C.; Terry, Kathryn L.; Teo, Soo-Hwang; Templeman, Claire; Stram, Daniel O.; Southey, Melissa C.; Sieh, Weiva; Siddiqui, Nadeem; Shvetsov, Yurii B.; Shu, Xiao-Ou; Shridhar, Viji; Wang-Gohrke, Shan; Severi, Gianluca; Schwaab, Ira; Salvesen, Helga B.; Rzepecka, Iwona K.; Runnebaum, Ingo B.; Rossing, Mary Anne; Rodriguez-Rodriguez, Lorna; Risch, Harvey A.; Renner, Stefan P.; Poole, Elizabeth M.; Pike, Malcolm C.; Phelan, Catherine M.; Pelttari, Liisa M.; Pejovic, Tanja; Paul, James; Orlow, Irene; Omar, Siti Zawiah; Olson, Sara H.; Odunsi, Kunle; Nickels, Stefan; Nevanlinna, Heli; Ness, Roberta B.; Narod, Steven A.; Nakanishi, Toru; Moysich, Kirsten B.; Monteiro, Alvaro N.A.; Moes-Sosnowska, Joanna; Modugno, Francesmary; Menon, Usha; McLaughlin, John R.; McGuire, Valerie; Matsuo, Keitaro; Adenan, Noor Azmi Mat; Massuger, Leon F.A. G.; Lurie, Galina; Lundvall, Lene; Lubiński, Jan; Lissowska, Jolanta; Levine, Douglas A.; Leminen, Arto; Lee, Alice W.; Le, Nhu D.; Lambrechts, Sandrina; Lambrechts, Diether; Kupryjanczyk, Jolanta; Krakstad, Camilla; Konecny, Gottfried E.; Kjaer, Susanne Krüger; Kiemeney, Lambertus A.; Kelemen, Linda E.; Keeney, Gary L.; Karlan, Beth Y.; Karevan, Rod; Kalli, Kimberly R.; Kajiyama, Hiroaki; Ji, Bu-Tian; Jensen, Allan; Jakubowska, Anna; Iversen, Edwin; Hosono, Satoyo; Høgdall, Claus K.; Høgdall, Estrid; Hoatlin, Maureen; Hillemanns, Peter; Heitz, Florian; Hein, Rebecca; Harter, Philipp; Halle, Mari K.; Hall, Per; Gronwald, Jacek; Gore, Martin; Goodman, Marc T.; Giles, Graham G.; Gentry-Maharaj, Aleksandra; Garcia-Closas, Montserrat; Flanagan, James M.; Fasching, Peter A.; Ekici, Arif B.; Edwards, Robert; Eccles, Diana; Easton, Douglas F.; Dürst, Matthias; du Bois, Andreas; Dörk, Thilo; Doherty, Jennifer A.; Despierre, Evelyn; Dansonka-Mieszkowska, Agnieszka; Cybulski, Cezary; Cramer, Daniel W.; Cook, Linda S.; Chen, Xiaoqing; Charbonneau, Bridget; Chang-Claude, Jenny; Campbell, Ian; Butzow, Ralf; Bunker, Clareann H.; Brueggmann, Doerthe; Brown, Robert; Brooks-Wilson, Angela; Brinton, Louise A.; Bogdanova, Natalia; Block, Matthew S.; Benjamin, Elizabeth; Beesley, Jonathan; Beckmann, Matthias W.; Bandera, Elisa V.; Baglietto, Laura; Bacot, François; Armasu, Sebastian M.; Antonenkova, Natalia; Anton-Culver, Hoda; Aben, Katja K.; Liang, Dong; Wu, Xifeng; Lu, Karen; Hildebrandt, Michelle A.T.; Schildkraut, Joellen M.; Sellers, Thomas A.; Huntsman, David; Berchuck, Andrew; Chenevix-Trench, Georgia; Gayther, Simon A.; Pharoah, Paul D.P.; Laird, Peter W.; Goode, Ellen L.; Pearce, Celeste Leigh

2013-01-01

HNF1B is overexpressed in clear cell epithelial ovarian cancer, and we observed epigenetic silencing in serous epithelial ovarian cancer, leading us to hypothesize that variation in this gene differentially associates with epithelial ovarian cancer risk according to histological subtype. Here we comprehensively map variation in HNF1B with respect to epithelial ovarian cancer risk and analyse DNA methylation and expression profiles across histological subtypes. Different single-nucleotide polymorphisms associate with invasive serous (rs7405776 odds ratio (OR) = 1.13, P = 3.1 × 10−10) and clear cell (rs11651755 OR = 0.77, P = 1.6 × 10−8) epithelial ovarian cancer. Risk alleles for the serous subtype associate with higher HNF1B-promoter methylation in these tumours. Unmethylated, expressed HNF1B, primarily present in clear cell tumours, coincides with a CpG island methylator phenotype affecting numerous other promoters throughout the genome. Different variants in HNF1B associate with risk of serous and clear cell epithelial ovarian cancer; DNA methylation and expression patterns are also notably distinct between these subtypes. These findings underscore distinct mechanisms driving different epithelial ovarian cancer histological subtypes. PMID:23535649

L-rhamnose induces browning in 3T3-L1 white adipocytes and activates HIB1B brown adipocytes.

PubMed

Choi, Minji; Mukherjee, Sulagna; Kang, Nam Hyeon; Barkat, Jameel Lone; Parray, Hilal Ahmad; Yun, Jong Won

2018-06-01

Induction of the brown adipocyte-like phenotype in white adipocytes (browning) is considered as a novel strategy to fight obesity due to the ability of brown adipocytes to increase energy expenditure. Here, we report that L-rhamnose induced browning by elevating expression levels of beige-specific marker genes, including Cd137, Cited1, Tbx1, Prdm16, Tmem26, and Ucp1, in 3T3-L1 adipocytes. Moreover, L-rhamnose markedly elevated expression levels of proteins involved in thermogenesis both in 3T3-L1 white and HIB1B brown adipocytes. L-rhamnose treatment in 3T3-L1 adipocytes also significantly elevated protein levels of p-HSL, p-AMPK, ACOX, and CPT1 as well as reduced levels of ACC, FAS, C/EBPα, and PPARγ, suggesting its possible role in enhancement of lipolysis and lipid catabolism as well as reduced adipogenesis and lipogenesis, respectively. The quick technique of efficient molecular docking provided insight into the strong binding of L-rhamnose to the fat-digesting glycine residue of β 3 -adrenergic receptor (AR), indicating strong involvement of L-rhamnose in fat metabolism. Further examination of the molecular mechanism of L-rhamnose revealed that it induced browning of 3T3-L1 adipocytes via coordination of multiple signaling pathways through β 3 -AR, SIRT1, PKA, and p-38. To the best of our knowledge, this is the first study to demonstrate that L-rhamnose plays multiple modulatory roles in the induction of white fat browning, activation of brown adipocytes, as well as promotion of lipid metabolism, thereby demonstrating its therapeutic potential for treatment of obesity. © 2018 IUBMB Life, 70(6):563-573, 2018. © 2018 International Union of Biochemistry and Molecular Biology.

Inactivation of the survival motor neuron gene, a candidate gene for human spinal muscular atrophy, leads to massive cell death in early mouse embryos

PubMed Central

Schrank, Bertold; Götz, Rudolf; Gunnersen, Jennifer M.; Ure, Janice M.; Toyka, Klaus V.; Smith, Austin G.; Sendtner, Michael

1997-01-01

Proximal spinal muscular atrophy is an autosomal recessive human disease of spinal motor neurons leading to muscular weakness with onset predominantly in infancy and childhood. With an estimated heterozygote frequency of 1/40 it is the most common monogenic disorder lethal to infants; milder forms represent the second most common pediatric neuromuscular disorder. Two candidate genes—survival motor neuron (SMN) and neuronal apoptosis inhibitory protein have been identified on chromosome 5q13 by positional cloning. However, the functional impact of these genes and the mechanism leading to a degeneration of motor neurons remain to be defined. To analyze the role of the SMN gene product in vivo we generated SMN-deficient mice. In contrast to the human genome, which contains two copies, the mouse genome contains only one SMN gene. Mice with homozygous SMN disruption display massive cell death during early embryonic development, indicating that the SMN gene product is necessary for cellular survival and function. PMID:9275227

[Effect of DOT1L gene silence on proliferation of acute monocytic leukemia cell line THP-1].

PubMed

Zhang, Yu-Juan; Li, Hua-Wen; Chang, Guo-Qiang; Zhang, Hong-Ju; Wang, Jian; Lin, Ya-Ni; Zhou, Jia-Xi; Li, Qing-Hua; Pang, Tian-Xiang

2013-08-01

This study was aimed to investigate the influence of short hairpin RNA (shRNA) on proliferation of human leukemia cell line THP-1. The shRNA targeting the site 732-752 of DOT1L mRNA was designed and chemically synthesized, then a single-vector lentiviral, tet-inducible shRNA-DOT1L system (Plko-Tet-On) was generated. Thereafter, the THP-1 cells with lentivirus were infected to create stable cell line with regulatable shRNA expression. The expression of DOT1L in the THP-1 cell line was assayed by RT-PCR. Effect of shRNA-DOT1L on the proliferation of THP-1 cells was detected with MTT method,and the change of colony forming potential of THP-1 cells was analyzed by colony forming unit test. Cell cycle distribution was tested by flow cytometry. The results indicated that the expression of DOT1L was statistically lower than that in the control groups. The proliferation and colony forming capacity of THP-1 cells were significantly inhibited. The percentage of cells at G0/G1 phase increased in THP-1/shRNA cells treated with Dox while the percentage of cells at S phase significantly decreased as compared with that in the control group. It is concluded that the shRNA targeting DOT1L can effectively inhibit the proliferation of acute monocytic leukemia cell line THP-1.

DNA adenine methylation of sams1 gene in symbiont-bearing Amoeba proteus.

PubMed

Jeon, Taeck J

2008-10-01

The expression of amoeba sams genes is switched from sams1 to sams2 when amoebae are infected with Legionella jeonii. To elucidate the mechanism for the inactivation of host sams1 gene by endosymbiotic bacteria, methylation states of the sams1 gene of D and xD amoebae was compared in this study. The sams1 gene of amoebae was methylated at an internal adenine residue of GATC site in symbiont-bearing xD amoebae but not in symbiont-free D amoebae, suggesting that the modification might have caused the inactivation of sams1 in xD amoebae. The sams1 gene of xD amoebae was inactivated at the transcriptional level. Analysis of DNA showed that adenine residues in L. jeonii sams were also methylated, implying that L. jeonii bacteria belong to a Dam methylase-positive strain. In addition, both SAM and Met appeared to act as negative regulators for the expression of sams1 whereas the expression of sams2 was not affected in amoebae.

Lead(II) complex formation with l-cysteine in aqueous solution

DOE PAGES

Jalilehvand, Farideh; Sisombath, Natalie S.; Schell, Adam C.; ...

2015-02-19

The lead(II) complexes formed with the multidentate chelator l-cysteine (H 2Cys) in an alkaline aqueous solution were studied using 207Pb, 13C, and 1H NMR, Pb L III-edge X-ray absorption, and UV–vis spectroscopic techniques, complemented by electrospray ion mass spectrometry (ESI-MS). The H 2Cys/Pb II mole ratios were varied from 2.1 to 10.0 for two sets of solutions with C PbII = 0.01 and 0.1 M, respectively, prepared at pH values (9.1–10.4) for which precipitates of lead(II) cysteine dissolved. At low H 2Cys/Pb II mole ratios (2.1–3.0), a mixture of the dithiolate [Pb(S,N-Cys) 2] 2– and [Pb(S,N,O-Cys)(S-HCys)] – complexes with averagemore » Pb–(N/O) and Pb–S distances of 2.42 ± 0.04 and 2.64 ± 0.04 Å, respectively, was found to dominate. At high concentration of free cysteinate (>0.7 M), a significant amount converts to the trithiolate [Pb(S,N-Cys)(S-HCys) 2] 2–, including a minor amount of a PbS 3-coordinated [Pb(S-HCys) 3] – complex. The coordination mode was evaluated by fitting linear combinations of EXAFS oscillations to the experimental spectra and by examining the 207Pb NMR signals in the chemical shift range δ Pb = 2006–2507 ppm, which became increasingly deshielded with increasing free cysteinate concentration. One-pulse magic-angle-spinning (MAS) 207Pb NMR spectra of crystalline Pb(aet) 2 (Haet = 2-aminoethanethiol or cysteamine) with PbS 2N 2 coordination were measured for comparison (δ iso = 2105 ppm). The UV–vis spectra displayed absorption maxima at 298–300 nm (S – → Pb II charge transfer) for the dithiolate PbS 2N(N/O) species; with increasing ligand excess, a shoulder appeared at ~330 nm for the trithiolate PbS 3N and PbS 3 (minor) complexes. Finally, the results provide spectroscopic fingerprints for structural models for lead(II) coordination modes to proteins and enzymes.« less

Bacillus subtilis 168 Contains Two Differentially Regulated Genes Encoding l-Asparaginase

PubMed Central

Fisher, Susan H.; Wray, Lewis V.

2002-01-01

Expression of the two Bacillus subtilis genes encoding l-asparaginase is controlled by independent regulatory factors. The ansZ gene (formerly yccC) was shown by mutational analysis to encode a functional l-asparaginase, the expression of which is activated during nitrogen-limited growth by the TnrA transcription factor. Gel mobility shift and DNase I footprinting experiments indicate that TnrA regulates ansZ expression by binding to a DNA site located upstream of the ansZ promoter. The expression of the ansA gene, which encodes the second l-asparaginase, was found to be induced by asparagine. The ansA repressor, AnsR, was shown to negatively regulate its own expression. PMID:11914346

Bacillus subtilis 168 contains two differentially regulated genes encoding L-asparaginase.

PubMed

Fisher, Susan H; Wray, Lewis V

2002-04-01

Expression of the two Bacillus subtilis genes encoding L-asparaginase is controlled by independent regulatory factors. The ansZ gene (formerly yccC) was shown by mutational analysis to encode a functional L-asparaginase, the expression of which is activated during nitrogen-limited growth by the TnrA transcription factor. Gel mobility shift and DNase I footprinting experiments indicate that TnrA regulates ansZ expression by binding to a DNA site located upstream of the ansZ promoter. The expression of the ansA gene, which encodes the second L-asparaginase, was found to be induced by asparagine. The ansA repressor, AnsR, was shown to negatively regulate its own expression.

Improvement of L-valine production at high temperature in Brevibacterium flavum by overexpressing ilvEBNrC genes.

PubMed

Hou, Xiaohu; Ge, Xiangyang; Wu, Di; Qian, He; Zhang, Weiguo

2012-01-01

Brevibacterium flavum ATCC14067 was engineered for L: -valine production by overexpression of different ilv genes; the ilvEBN(r)C genes from B. flavum NV128 provided the best candidate for L: -valine production. In traditional fermentation, L: -valine production reached 30.08 ± 0.92 g/L at 31°C in 72 h with a low conversion efficiency of 0.129 g/g. To further improve the L: -valine production and conversion efficiency based on the optimum temperatures of L: -valine biosynthesis enzymes (above 35°C) and the thermotolerance of B. flavum, the fermentation temperature was increased to 34, 37, and 40°C. As a result, higher metabolic rate and L: -valine biosynthesis enzymes activity were obtained at high temperature, and the maximum L: -valine production, conversion efficiency, and specific L: -valine production rate reached 38.08 ± 1.32 g/L, 0.241 g/g, and 0.133 g g(-1) h(-1), respectively, at 37°C in 48 h fermentation. The strategy for enhancing L: -valine production by overexpression of key enzymes in thermotolerant strains may provide an alternative approach to enhance branched-chain amino acids production with other strains.

Evaluation of the use of salivary lead levels as a surrogate of blood lead or plasma lead levels in lead exposed subjects.

PubMed

Barbosa, Fernando; Corrêa Rodrigues, Maria Heloísa; Buzalaf, Maria R; Krug, Francisco J; Gerlach, Raquel F; Tanus-Santos, José Eduardo

2006-10-01

We conducted a study to evaluate the use of parotid salivary lead (Pb-saliva) levels as a surrogate of the blood lead (Pb-B) or plasma lead levels (Pb-P) to diagnose lead exposure. The relationship between these biomarkers was assessed in a lead exposed population. Pb-saliva and Pb-P were determined by inductively coupled plasma mass spectrometry, while in whole blood lead was determined by graphite furnace atomic absorption spectrometry. We studied 88 adults (31 men and 57 women) from 18 to 60 years old. Pb-saliva levels varied from 0.05 to 4.4 microg/l, with a mean of 0.85 microg/l. Blood lead levels varied from 32.0 to 428.0 microg/l in men (mean 112.3 microg/l) and from 25.0 to 263.0 microg/l (mean 63.5 microg/l) in women. Corresponding Pb-Ps were 0.02-2.50 microg/l (mean 0.77 microg/l) and 0.03-1.6 microg/l (mean 0.42 microg/l) in men and women, respectively. A weak correlation was found between Log Pb-saliva and Log Pb-B (r=0.277, P<0.008), and between Log Pb-saliva and Log Pb-P (r=0.280, P=0.006). The Pb-saliva/Pb-P ratio ranged from 0.20 to 18.0. Age or gender does not affect Pb-saliva levels or Pb-saliva/Pb-P ratio. Taken together, these results suggest that salivary lead may not be used as a biomarker to diagnose lead exposure nor as a surrogate of plasma lead levels at least for low to moderately lead exposed population.

Sucrose non-ferment 1 related protein kinase 2 (SnRK2) genes could mediate the stress responses in potato (Solanum tuberosum L.).

PubMed

Bai, Jiangping; Mao, Juan; Yang, Hongyu; Khan, Awais; Fan, Aqi; Liu, Siyan; Zhang, Junlian; Wang, Di; Gao, Huijuan; Zhang, Jinlin

2017-05-15

The SnRKs (sucrose non-fermenting 1 related protein kinase) are a gene family coding for Ser/Thr protein kinases and play important roles in linking the tolerance and metabolic responses of plants to abiotic stresses. To date, no genome-wide characterization of the sucrose non-ferment 1 related protein kinase 2 (SnRK2) subfamily has been conducted in potato (Solanum tuberosum L.). In this study, eight StSnRK2 genes (StSnRK2.1- StSnRK2.8) were identified in the genome of the potato (Solanum tuberosum L.) cultivar 'Longshu 3', with similar characteristics to SnRK2 from other plant species in gene structure, motif distribution and secondary structures. The C-terminal regions were highly divergent among StSnRK2s, while they all carried the similar Ser/Thr protein kinase domain. The fluorescence of GFP fused with StSnRK2.1, StSnRK2.2, StSnRK2.6, StSnRK2.7 and StSnRK2.8 was detected in the nucleus and cytoplasm of onion epidermal cells with StSnRK2.3 and StSnRK2.4 mainly associated to the nucleus while StSnRK2.5 to subcellular organelles. Expression level analysis by qRT-PCR showed that StSnRK2.1, 2.2, 2.5 and 2.6 were more than 1 fold higher in the root than in the leaf, tuber and stem tissues. The expressions of StSnRK2.3, 2.7, and 2.8 were at least 1.5 folds higher in the leaf and stem than in the root, but lower in the tuber. The expression of StSnRK2.4 was also significantly (P < 0.05) higher in leaf, stem, and tuber than in the root. From the perspective of the relative expressions of StSnRK2 genes in potato, ABA treatment had a different effect from NaCl and PEG treatments. In the present study, we identified and characterized eight SnRK2s in the potato genome. The eight StSnRK2s exhibit similar gene structure and secondary structures in potato to the SnRK2s found in other plant species. The relative expression of eight genes varied among various tissues (roots, leaves, tubers, and stems) and abiotic stresses (ABA, NaCl and PEG-6000) with the prolongation of

Transcription of a vaccinia virus late promoter template: requirement for the product of the A2L intermediate-stage gene.

PubMed Central

Passarelli, A L; Kovacs, G R; Moss, B

1996-01-01

Evidence is presented that a 26-kDa protein encoded by the vaccinia virus A2L open reading frame, originally shown to be one of three intermediate-stage genes that together can transactivate late-stage gene expression in transfection assays (J. G. Keck, C. J. Baldick, and B. Moss, Cell 61:801-809, 1990), is required for in vitro transcription of a template with a late promoter. The critical step in this analysis was the preparation of an extract containing all the required factors except for the A2L protein. This extract was prepared from cells infected with a recombinant vaccinia virus expressing the bacteriophage T7 RNA polymerase in the presence of the DNA synthesis inhibitor cytosine arabinoside and transfected with plasmids containing the two other known transactivator genes, A1L and G8R, under T7 promoter control. Reaction mixtures made with extracts of these cells had background levels of late transcription activity, unless they were supplemented with extracts of cells transfected with the A2L gene. Active transcription mixtures were also made by mixing extracts from three sets of cells, each transfected with a gene (A1L, A2L, or G8R) encoding a separate factor, indicating the absence of any requirement for their coexpression. To minimize the possibility that the A2L protein functions indirectly by activating another viral or cellular protein, this gene was expressed in insect cells by using a baculovirus vector. The partially purified recombinant protein complemented the activity of A2L-deficient cell extracts. Recombinant A1L, A2L, and G8R proteins, all produced in insect cells, together complemented extracts from mammalian cells containing only viral early proteins, concordant with previous in vivo transfection data. PMID:8676468

Polymorphisms in DC-SIGN and L-SIGN genes are associated with HIV-1 vertical transmission in a Northeastern Brazilian population.

PubMed

da Silva, Ronaldo Celerino; Segat, Ludovica; Zanin, Valentina; Arraes, Luiz Claudio; Crovella, Sergio

2012-11-01

DC-SIGN and L-SIGN are receptors expressed on specialized macrophages in decidua, (Hofbauer and placental capillary endothelial cells), known to interact with several pathogens, including HIV-1. To disclose the possible involvement of these molecules in the susceptibility to HIV vertical transmission, we analyzed DC-SIGN and L-SIGN gene single nucleotide polymorphisms (SNPs) in 192 HIV-1 positive children and 58 HIV-1 negative children all born to HIV-1 positive mothers, as well as 96 healthy uninfected children not exposed to HIV-1, all from Northeast Brazil. The frequency of three SNPs in the DC-SIGN promoter (-139G>A, -201G>T and -336A>G) were significantly different when comparing HIV positive children with HIV-1 exposed uninfected children, indicating an association with susceptibility to HIV-1 vertical transmission. This genetic association suggests that DC-SIGN molecule may play a role in susceptibility to HIV-1 infection through vertical transmission. Copyright © 2012 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.

Paraoxonase 1 (PON1)-L55M among common variants in the coding region of the paraoxonase gene family may contribute to the glycemic control in type 2 diabetes.

PubMed

Mahrooz, Abdolkarim; Hashemi-Soteh, Mohammad Bagher; Heydari, Masoud; Boorank, Ruzbeh; Ramazani, Fatemeh; Mahmoudi, Ali; Kianmehr, Anvarsadat; Alizadeh, Ahad

2018-05-19

Genome studies have shown that the genes encoding paraoxonase 1 (PON1) and PON2 are associated with glucose metabolism. The goal of this study was to simultaneously evaluate the association between functional variants in PON1 and PON2 genes and susceptibility for type 2 diabetes (T2D) and determine whether they can affect glycemic control. We performed a case-control study with 145 newly diagnosed patients with T2D and 148 controls. The common variants including PON1-Q192R, PON1-L55M and PON2-S311C were genotyped by PCR-based RFLP. A mismatch-PCR/RFLP was applied for genotyping the PON2-A148G variant. The variant PON1-Q192R in males (OR = 2.55, 95%CI 1.16-5.69, p = 0.023) and PON2-A148G in females (OR = 1.56, 95%CI 1.00-2.44, p = 0.059) were associated with T2D. Compared with the LL genotypes of PON1-L55M, HbA1c levels were significantly lower in the LM genotypes (p = 0.01) and MM genotypes (p = 0.032) in patients. Multiple linear regression analyses showed that among the study variants only the PON1-L55M variant as an independent variable significantly associated with glycemic control. This variant significantly influenced glycemic control in patients with poor glycemic control so that it was better with the following order: LL < LM < MM. Based on gamma correlation, there was a significant inverse association between the number of M alleles of the PON1-L55M and HbA1c levels (r = -0.261, p = 0.001). Sex should be considered a confounding variable in association studies on the variants PON1-Q192R and PON2-A148G in T2D. Patients sharing the 55 M allele were prone to having good glycemic control. Our findings provide genetic evidence that the PON1-L55M variant may be a factor contributing to glycemic control. Copyright © 2018 Elsevier B.V. All rights reserved.

The role of the F-box gene TaFBA1 from wheat (Triticum aestivum L.) in drought tolerance.

PubMed

Zhou, Shumei; Sun, Xiudong; Yin, Suhong; Kong, Xiangzhu; Zhou, Shan; Xu, Ying; Luo, Yin; Wang, Wei

2014-11-01

Drought is one of the most important factors limiting plant growth and development. We identified a gene in wheat (Triticum aestivum L.) under drought stress named TaFBA1. TaFBA1 encodes a putative 325-amino-acid F-box protein with a conserved N-terminal F-box domain and a C-terminal AMN1 domain. Real-time RT-PCR analysis revealed that TaFBA1 transcript accumulation was upregulated by high-salinity, water stress, and abscisic acid (ABA) treatment. To evaluate the functions of TaFBA1 in the regulation of drought stress responses, we produced transgenic tobacco lines overexpressing TaFBA1. Under water stress conditions, the transgenic tobacco plants had a higher germination rate, higher relative water content, net photosynthesis rate (Pn), less chlorophyll loss, and less growth inhibition than WT. These results demonstrate the high tolerance of the transgenic plants to drought stress compared to the WT. The enhanced oxidative stress tolerance of these plants, which may be involved in their drought tolerance, was indicated by their lower levels of reactive oxygen species (ROS) accumulation, MDA content, and cell membrane damage under drought stress compared to WT. The antioxidant enzyme activities were higher in the transgenic plants than in WT, which may be related to the upregulated expression of some antioxidant genes via overexpression of TaFBA1. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

Mutations in 3 genes (MKS3, CC2D2A and RPGRIP1L) cause COACH syndrome (Joubert syndrome with congenital hepatic fibrosis)

PubMed Central

Doherty, D; Parisi, M A; Finn, L S; Gunay-Aygun, M; Al-Mateen, M; Bates, D; Clericuzio, C; Demir, H; Dorschner, M; van Essen, A J; Gahl, W A; Gentile, M; Gorden, N T; Hikida, A; Knutzen, D; Özyurek, H; Phelps, I; Rosenthal, P; Verloes, A; Weigand, H; Chance, P F; Dobyns, W B; Glass, I A

2011-01-01

Objective To identify genetic causes of COACH syndrome Background COACH syndrome is a rare autosomal recessive disorder characterised by Cerebellar vermis hypoplasia, Oligophrenia (developmental delay/mental retardation), Ataxia, Coloboma, and Hepatic fibrosis. The vermis hypoplasia falls in a spectrum of mid-hindbrain malformation called the molar tooth sign (MTS), making COACH a Joubert syndrome related disorder (JSRD). Methods In a cohort of 251 families with JSRD, 26 subjects in 23 families met criteria for COACH syndrome, defined as JSRD plus clinically apparent liver disease. Diagnostic criteria for JSRD were clinical findings (intellectual impairment, hypotonia, ataxia) plus supportive brain imaging findings (MTS or cerebellar vermis hypoplasia). MKS3/TMEM67 was sequenced in all subjects for whom DNA was available. In COACH subjects without MKS3 mutations, CC2D2A, RPGRIP1L and CEP290 were also sequenced. Results 19/23 families (83%) with COACH syndrome carried MKS3 mutations, compared to 2/209 (1%) with JSRD but no liver disease. Two other families with COACH carried CC2D2A mutations, one family carried RPGRIP1L mutations, and one lacked mutations in MKS3, CC2D2A, RPGRIP1L and CEP290. Liver biopsies from three subjects, each with mutations in one of the three genes, revealed changes within the congenital hepatic fibrosis/ductal plate malformation spectrum. In JSRD with and without liver disease, MKS3 mutations account for 21/232 families (9%). Conclusions Mutations in MKS3 are responsible for the majority of COACH syndrome, with minor contributions from CC2D2A and RPGRIP1L; therefore, MKS3 should be the first gene tested in patients with JSRD plus liver disease and/or coloboma, followed by CC2D2A and RPGRIP1L. PMID:19574260

Gene expression network regulated by DNA methylation and microRNA during microcystin-leucine arginine induced malignant transformation in human hepatocyte L02 cells.

PubMed

Chen, Hong-Qiang; Zhao, Ji; Li, Yan; He, Li-Xiong; Huang, Yu-Jing; Shu, Wei-Qun; Cao, Jia; Liu, Wen-Bin; Liu, Jin-Yi

2018-06-01

Microcystin (MC) is a cyclic heptapeptide compound which could lead to the development of hepatocellular carcinoma. However, the underlying epigenetic regulation mechanism is largely unknown. In this study, microcystin-LR (L: lysine, R: arginine, MC-LR) was used to induce the malignant transformation of human hepatocyte L02 cell line. The profile of gene expression, microRNA (miRNA) and DNA methylation were detected through high-throughput sequencing. Compared with control group, the expression of 826 genes and 187 miRNAs changed significantly in MC-LR treated group. DNA methylation sequencing analysis showed that 2592 CpG sites differentially methylated in promoter or the coding DNA sequence (CDS) of genes, while DNA methyltransferase 3 alpha (DNMT3a) and DNA methyltransferase 3 beta (DNMT3b) were dramatically up-regulated. Functional analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed that significantly changed mRNAs and microRNAs were mainly involved in the formation of cancer, proliferation, invasion, migration and metabolism. MiRNA-mRNA network and mRNA-mRNA network analysis showed that hsa-miR-320a, hsa-miR-331-3p, hsa-miR-26a-5p, hsa-miR-196a-5p, hsa-miR-221-3p, coiled-coil domain containing 180 (CCDC180), melanoma antigen gene family member D1 (MAGED1), membrane spanning 4-domains A7 (MS4A7), hephaestin like 1 (HEPHL1), BH3 (Bcl-2 homology 3)-like motif containing, cell death inducer (BLID), matrix metallopeptidase 13 (MMP13), guanylate binding protein 5 (GBP5), adipogenesis regulatory factor (ADIRF), formin homology 2 domain containing 1 (FHDC1), protein kinase CAMP-dependent type II regulatory subunit beta (PRKAR2B), nodium leak channel, non-selective (NALCN), myosin light chain kinase 3 (MYLK3), epidermal growth factor receptor (EGFR) and zinc finger protein 704 (ZNF704) were key miRNAs and genes in the malignant transformation induced by MC-LR in L02 cells. Moreover, we found that expression of MYLK3, EGFR and ZNF704 were

Gestational diabetes mellitus is associated with TCF7L2 gene polymorphisms independent of HLA-DQB1*0602 genotypes and islet cell autoantibodies

PubMed Central

Papadopoulou, A.; Lynch, K. F.; Shaat, N.; Håkansson, R.; Ivarsson, S. A.; Berntorp, K.; Agardh, C. D.; Lernmark, Å

2011-01-01

Aims To test whether the TCF7L2 gene was associated with gestational diabetes, whether the association between TCF7L2 and gestational diabetes was independent of HLA-DQB1*0602 and islet cell autoantibodies, as well as maternal age, number of pregnancies, family history of diabetes and the HLA-DQB1 genotypes, and to test whether the distribution of HLA-DQB1 alleles was affected by country of birth. Methods We genotyped the rs7903146, rs12255372 and rs7901695 single nucleotide polymorphisms of the TCF7L2 gene in 826 mothers with gestational diabetes and in 1185 healthy control subjects in the Diabetes Prediction in Skåne Study. The mothers were also typed for HLA-DQB1 genotypes and tested for islet cell autoantibodies against GAD65, insulinoma-associated antigen-2 and insulin. Results The heterozygous genotypes CT, GT and TC of the rs7903146 (T is risk for Type 2 diabetes), rs12255372 (T is risk for Type 2 diabetes) and rs7901695 (C is risk for Type 2 diabetes), respectively, as well as the homozygous genotypes TT, TT and CC of the rs7903146, rs12255372 and rs7901695, respectively, were strongly associated with gestational diabetes (P < 0.0001). These associations remained statistically significant after adjusting for maternal age, number of pregnancies, family history of diabetes and HLA-DQ genotypes and were independent of the presence of islet cell autoantibodies. No interaction was observed between TCF7L2 and HLA-DQB1*0602, which was shown to be negatively associated with gestational diabetes in mothers born in Sweden (P = 0.010). Conclusions The TCF7L2 was associated with susceptibility for gestational diabetes independently of the presence of HLA-DQB1*0602 and islet cell autoantibodies and other factors such as maternal age, number of pregnancies, family history of diabetes and other HLA-DQ genotypes. The HLA-DQB1*0602 was negatively associated with gestational diabetes in mothers born in Sweden. PMID:21672010

Characterization and expression analysis of a banana gene encoding 1-aminocyclopropane-1-carboxylate oxidase.

PubMed

Huang, P L; Do, Y Y; Huang, F C; Thay, T S; Chang, T W

1997-04-01

A cDNA encoding the banana 1-aminocyclopropane-1-carboxylate (ACC) oxidase has previously been isolated from a cDNA library that was constructed by extracting poly(A)+ RNA from peels of ripening banana. This cDNA, designated as pMAO2, has 1,199 bp and contains an open reading frame of 318 amino acids. In order to identify ripening-related promoters of the banana ACC oxidase gene, pMAO2 was used as a probe to screen a banana genomic library constructed in the lambda EMBL3 vector. The banana ACC oxidase MAO2 gene has four exons and three introns, with all of the boundaries between these introns and exons sharing a consensus dinucleotide sequence of GT-AG. The expression of MAO2 gene in banana begins after the onset of ripening (stage 2) and continuous into later stages of the ripening process. The accumulation of MAO2 mRNA can be induced by 1 microliter/l exogenous ethylene, and it reached steady state level when 100 microliters/l exogenous ethylene was present.

Expression of Genes Encoding the Enzymes for Glycogen and Trehalose Metabolism in L3 and L4 Larvae of Anisakis simplex.

PubMed

Łopieńska-Biernat, E; Zaobidna, E A; Dmitryjuk, M

2015-01-01

Trehalose and glycogen metabolism plays an important role in supporting life processes in many nematodes, including Anisakis simplex. Nematodes, cosmopolitan helminths parasitizing sea mammals and humans, cause a disease known as anisakiasis. The aim of this study was to investigate the expression of genes encoding the enzymes involved in the metabolism of trehalose and glycogen-trehalose-6-phosphate synthase (TPS), trehalose-6-phosphate phosphatase (TPP), glycogen synthase (GS), and glycogen phosphorylase (GP)-in stage L3 and stage L4 larvae of A. simplex. The expression of mRNA all four genes, tps, tpp, gs, and gp, was examined by real-time polymerase chain reaction. The A. simplex ribosomal gene (18S) was used as a reference gene. Enzymatic activity was determined. The expression of trehalose enzyme genes was higher in L3 than in L4 larvae, but an inverse relationship was noted for the expression of gs and gp genes.

Expression of Genes Encoding the Enzymes for Glycogen and Trehalose Metabolism in L3 and L4 Larvae of Anisakis simplex

PubMed Central

Łopieńska-Biernat, E.; Zaobidna, E. A.; Dmitryjuk, M.

2015-01-01

Trehalose and glycogen metabolism plays an important role in supporting life processes in many nematodes, including Anisakis simplex. Nematodes, cosmopolitan helminths parasitizing sea mammals and humans, cause a disease known as anisakiasis. The aim of this study was to investigate the expression of genes encoding the enzymes involved in the metabolism of trehalose and glycogen—trehalose-6-phosphate synthase (TPS), trehalose-6-phosphate phosphatase (TPP), glycogen synthase (GS), and glycogen phosphorylase (GP)—in stage L3 and stage L4 larvae of A. simplex. The expression of mRNA all four genes, tps, tpp, gs, and gp, was examined by real-time polymerase chain reaction. The A. simplex ribosomal gene (18S) was used as a reference gene. Enzymatic activity was determined. The expression of trehalose enzyme genes was higher in L3 than in L4 larvae, but an inverse relationship was noted for the expression of gs and gp genes. PMID:26783451

Two pathways for pyrrole formation in coumermycin A(1) biosynthesis: the central pyrrole moiety is formed from L-threonine.

PubMed

Siebenberg, Stefanie; Burkard, Nadja; Knuplesch, Anna; Gust, Bertolt; Grond, Stephanie; Heide, Lutz

2011-11-25

Coumermycin A(1) is an aminocoumarin antibiotic produced by Streptomyces rishiriensis. It contains three pyrrole rings, that is, two terminal 5-methyl-pyrrole-2-carboxyl moieties and a central 3-methylpyrrole-2,4-dicarboxylic acid moiety. The biosynthesis of the terminal pyrrole moieties has been elucidated previously. However, the biosynthetic precursors of the central pyrrole moiety have remained unknown, and none of the genes or enzymes involved in its formation has been identified. We now show that five genes, contained in a contiguous 4.7 kb region within the coumermycin biosynthetic gene cluster, are required for the biosynthesis of this central pyrrole moiety. Each of these genes was deleted individually, resulting in a strong reduction or an abolishment of coumermycin production. External feeding of the central pyrrole moiety restored coumermycin production. One of these genes shows similarity to L-threonine kinase genes. Feeding of [U-(13)C,(15) N]L-threonine and (13)C NMR analysis of the resulting compound unequivocally proved that threonine was incorporated intact into the central pyrrole (19 % enrichment) to provide the heterocyclic nitrogen as well as four of the seven carbons of this moiety. Therefore, this pyrrole is formed via a new, hitherto unknown biosynthetic pathway. A hypothesis for the reaction sequence leading to the central pyrrole moiety of coumermycin A(1) is presented. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Frequencies and expression levels of programmed death ligand 1 (PD-L1) in circulating tumor RNA (ctRNA) in various cancer types.

PubMed

Ishiba, Toshiyuki; Hoffmann, Andreas-Claudius; Usher, Joshua; Elshimali, Yahya; Sturdevant, Todd; Dang, Mai; Jaimes, Yolanda; Tyagi, Rama; Gonzales, Ronald; Grino, Mary; Pinski, Jacek K; Barzi, Afsaneh; Raez, Luis E; Eberhardt, Wilfried E; Theegarten, Dirk; Lenz, Heinz-Josef; Uetake, Hiroyuki; Danenberg, Peter V; Danenberg, Kathleen

2018-06-07

Precision medicine and prediction of therapeutic response requires monitoring potential biomarkers before and after treatment. Liquid biopsies provide noninvasive prognostic markers such as circulating tumor DNA and RNA. Circulating tumor RNA (ctRNA) in blood is also used to identify mutations in genes of interest, but additionally, provides information about relative expression levels of important genes. In this study, we analyzed PD-L1 expression in ctRNA isolated from various cancer types. Tumors inhibit antitumor response by modulating the immune checkpoint proteins programmed death ligand 1 (PD-L1) and its cognate receptor PD1. The expression of these genes has been implicated in evasion of immune response and resistance to targeted therapies. Blood samples were collected from gastric (GC), colorectal (CRC), lung (NSCLC), breast (BC), prostate cancer (PC) patients, and a healthy control group. ctRNA was purified from fractionated plasma, and following reverse transcription, levels of PD-L1 expression were analyzed using qPCR. PD-L1 expression was detected in the plasma ctRNA of all cancer types at varying frequencies but no PD-L1 mRNA was detected in cancer-free individuals. The frequencies of PD-L1 expression were significantly different among the various cancer types but the median relative PD-L1 expression values were not significantly different. In 12 cases where plasma and tumor tissue were available from the same patients, there was a high degree of concordance between expression of PD-L1 protein in tumor tissues and PD-L1 gene expression in plasma, and both methods were equally predictive of response to nivolumab. PD-L1 mRNA can be detected and quantitated in ctRNA of cancer patients. These results pave the way for further studies aimed at determining whether monitoring the levels of PD-L1 mRNA in blood can identify patients who are most likely to benefit from the conventional treatment. Copyright © 2018 Elsevier Inc. All rights reserved.

Genetic analysis and fine mapping of LH1 and LH2, a set of complementary genes controlling late heading in rice (Oryza sativa L.)

PubMed Central

Liu, Shuang; Wang, Feng; Gao, Li Jun; Li, Jin Hua; Li, Rong Bai; Gao, Han Liang; Deng, Guo Fu; Yang, Jin Shui; Luo, Xiao Jin

2012-01-01

Heading date in rice (Oryza sativa L.) is a critical agronomic trait with a complex inheritance. To investigate the genetic basis and mechanism of gene interaction in heading date, we conducted genetic analysis on segregation populations derived from crosses among the indica cultivars Bo B, Yuefeng B and Baoxuan 2. A set of dominant complementary genes controlling late heading, designated LH1 and LH2, were detected by molecular marker mapping. Genetic analysis revealed that Baoxuan 2 contains both dominant genes, while Bo B and Yuefeng B each possess either LH1 or LH2. Using larger populations with segregant ratios of 3 : 1, we fine-mapped LH1 to a 63-kb region near the centromere of chromosome 7 flanked by markers RM5436 and RM8034, and LH2 to a 177-kb region on the short arm of chromosome 8 between flanking markers Indel22468-3 and RM25. Some candidate genes were identified through sequencing of Bo B and Yuefeng B in these target regions. Our work provides a solid foundation for further study on gene interaction in heading date and has application in marker-assisted breeding of photosensitive hybrid rice in China. PMID:23341744

Genetic analysis and fine mapping of LH1 and LH2, a set of complementary genes controlling late heading in rice (Oryza sativa L.).

PubMed

Liu, Shuang; Wang, Feng; Gao, Li Jun; Li, Jin Hua; Li, Rong Bai; Gao, Han Liang; Deng, Guo Fu; Yang, Jin Shui; Luo, Xiao Jin

2012-12-01

Heading date in rice (Oryza sativa L.) is a critical agronomic trait with a complex inheritance. To investigate the genetic basis and mechanism of gene interaction in heading date, we conducted genetic analysis on segregation populations derived from crosses among the indica cultivars Bo B, Yuefeng B and Baoxuan 2. A set of dominant complementary genes controlling late heading, designated LH1 and LH2, were detected by molecular marker mapping. Genetic analysis revealed that Baoxuan 2 contains both dominant genes, while Bo B and Yuefeng B each possess either LH1 or LH2. Using larger populations with segregant ratios of 3 : 1, we fine-mapped LH1 to a 63-kb region near the centromere of chromosome 7 flanked by markers RM5436 and RM8034, and LH2 to a 177-kb region on the short arm of chromosome 8 between flanking markers Indel22468-3 and RM25. Some candidate genes were identified through sequencing of Bo B and Yuefeng B in these target regions. Our work provides a solid foundation for further study on gene interaction in heading date and has application in marker-assisted breeding of photosensitive hybrid rice in China.

Selection and validation of reference genes for quantitative gene expression analyses in various tissues and seeds at different developmental stages in Bixa orellana L.

PubMed

Moreira, Viviane S; Soares, Virgínia L F; Silva, Raner J S; Sousa, Aurizangela O; Otoni, Wagner C; Costa, Marcio G C

2018-05-01

Bixa orellana L., popularly known as annatto, produces several secondary metabolites of pharmaceutical and industrial interest, including bixin, whose molecular basis of biosynthesis remain to be determined. Gene expression analysis by quantitative real-time PCR (qPCR) is an important tool to advance such knowledge. However, correct interpretation of qPCR data requires the use of suitable reference genes in order to reduce experimental variations. In the present study, we have selected four different candidates for reference genes in B. orellana , coding for 40S ribosomal protein S9 (RPS9), histone H4 (H4), 60S ribosomal protein L38 (RPL38) and 18S ribosomal RNA (18SrRNA). Their expression stabilities in different tissues (e.g. flower buds, flowers, leaves and seeds at different developmental stages) were analyzed using five statistical tools (NormFinder, geNorm, BestKeeper, ΔCt method and RefFinder). The results indicated that RPL38 is the most stable gene in different tissues and stages of seed development and 18SrRNA is the most unstable among the analyzed genes. In order to validate the candidate reference genes, we have analyzed the relative expression of a target gene coding for carotenoid cleavage dioxygenase 1 (CCD1) using the stable RPL38 and the least stable gene, 18SrRNA , for normalization of the qPCR data. The results demonstrated significant differences in the interpretation of the CCD1 gene expression data, depending on the reference gene used, reinforcing the importance of the correct selection of reference genes for normalization.

The NK1 Receptor Antagonist L822429 Reduces Heroin Reinforcement

PubMed Central

Barbier, Estelle; Vendruscolo, Leandro F; Schlosburg, Joel E; Edwards, Scott; Juergens, Nathan; Park, Paula E; Misra, Kaushik K; Cheng, Kejun; Rice, Kenner C; Schank, Jesse; Schulteis, Gery; Koob, George F; Heilig, Markus

2013-01-01

Genetic deletion of the neurokinin 1 receptor (NK1R) has been shown to decrease the reinforcing properties of opioids, but it is unknown whether pharmacological NK1R blockade has the same effect. Here, we examined the effect of L822429, a rat-specific NK1R antagonist, on the reinforcing properties of heroin in rats on short (1 h: ShA) or long (12 h: LgA) access to intravenous heroin self-administration. ShA produces heroin self-administration rates that are stable over time, whereas LgA leads to an escalation of heroin intake thought to model important dependence-related aspects of addiction. L822429 reduced heroin self-administration and the motivation to consume heroin, measured using a progressive-ratio schedule, in both ShA and LgA rats. L822429 also decreased anxiety-like behavior in both groups, measured on the elevated plus maze, but did not affect mechanical hypersensitivity observed in LgA rats. Expression of TacR1 (the gene encoding NK1R) was decreased in reward- and stress-related brain areas both in ShA and LgA rats compared with heroin-naïve rats, but did not differ between the two heroin-experienced groups. In contrast, passive exposure to heroin produced increases in TacR1 expression in the prefrontal cortex and nucleus accumbens. Taken together, these results show that pharmacological NK1R blockade attenuates heroin reinforcement. The observation that animals with ShA and LgA to heroin were similarly affected by L822429 indicates that the SP/NK1R system is not specifically involved in neuroadaptations that underlie escalation resulting from LgA self-administration. Instead, the NK1R antagonist appears to attenuate acute, positively reinforcing properties of heroin and may be useful as an adjunct to relapse prevention in detoxified opioid-dependent subjects. PMID:23303056

Disruption of Transcriptional Coactivator Sub1 Leads to Genome-Wide Re-distribution of Clustered Mutations Induced by APOBEC in Active Yeast Genes

PubMed Central

Dhar, Alok; Polev, Dmitrii E.; Masharsky, Alexey E.; Rogozin, Igor B.; Pavlov, Youri I.

2015-01-01

Mutations in genomes of species are frequently distributed non-randomly, resulting in mutation clusters, including recently discovered kataegis in tumors. DNA editing deaminases play the prominent role in the etiology of these mutations. To gain insight into the enigmatic mechanisms of localized hypermutagenesis that lead to cluster formation, we analyzed the mutational single nucleotide variations (SNV) data obtained by whole-genome sequencing of drug-resistant mutants induced in yeast diploids by AID/APOBEC deaminase and base analog 6-HAP. Deaminase from sea lamprey, PmCDA1, induced robust clusters, while 6-HAP induced a few weak ones. We found that PmCDA1, AID, and APOBEC1 deaminases preferentially mutate the beginning of the actively transcribed genes. Inactivation of transcription initiation factor Sub1 strongly reduced deaminase-induced can1 mutation frequency, but, surprisingly, did not decrease the total SNV load in genomes. However, the SNVs in the genomes of the sub1 clones were re-distributed, and the effect of mutation clustering in the regions of transcription initiation was even more pronounced. At the same time, the mutation density in the protein-coding regions was reduced, resulting in the decrease of phenotypically detected mutants. We propose that the induction of clustered mutations by deaminases involves: a) the exposure of ssDNA strands during transcription and loss of protection of ssDNA due to the depletion of ssDNA-binding proteins, such as Sub1, and b) attainment of conditions favorable for APOBEC action in subpopulation of cells, leading to enzymatic deamination within the currently expressed genes. This model is applicable to both the initial and the later stages of oncogenic transformation and explains variations in the distribution of mutations and kataegis events in different tumor cells. PMID:25941824

Molecular cloning, characterization, and expression of an alfalfa (Medicago sativa L.) heme oxygenase-1 gene, MsHO1, which is pro-oxidants-regulated.

PubMed

Fu, Guang-Qing; Xu, Sheng; Xie, Yan-Jie; Han, Bin; Nie, Li; Shen, Wen-Biao; Wang, Ren

2011-07-01

It has been documented that plant heme oxygenase-1 (HO-1; EC 1.14.99.3) is both development- and stress-regulated, thus it plays a vital role in light signalling and stress responses. In this study, an alfalfa (Medica sativa L.) HO-1 gene MsHO1 was isolated and sequenced. It contains four exons and three introns within genomic DNA sequence and encodes a polypeptide with 283 amino acids. MsHO1 had a conserved HO signature sequence and showed high similarity to other HOs in plants, especially HO-1 isoform. The MsHO1:GFP fusion protein was localized in the chloroplast. Further biochemical activity analysis of mature MsHO1, which was expressed in Escherichia coli, showed that the Vmax was 48.78 nmol biliverdin-IXα (BV) h⁻¹ nmol⁻¹ protein with an apparent Km value for hemin of 2.33 μM, and the optimum Tm and pH were 37 °C and 7.2, respectively. Results of semi-quantitative RT-PCR and western blot showed that the expressions of MsHO1 were higher in alfalfa stems and leaves than those in germinating seeds and roots. Importantly, MsHO1 gene expression and protein level were induced significantly by some pro-oxidant compounds, including hemin and nitric oxide (NO) donor sodium nitroprusside (SNP). In conclusion, MsHO1 may play an important role in oxidative responses. Copyright © 2011 Elsevier Masson SAS. All rights reserved.

Mapping-by-sequencing of Ligon-lintless-1 (Li 1 ) reveals a cluster of neighboring genes with correlated expression in developing fibers of Upland cotton (Gossypium hirsutum L.).

PubMed

Thyssen, Gregory N; Fang, David D; Turley, Rickie B; Florane, Christopher; Li, Ping; Naoumkina, Marina

2015-09-01

Mapping-by-sequencing and SNP marker analysis were used to fine map the Ligon-lintless-1 ( Li 1 ) short fiber mutation in tetraploid cotton to a 255-kb region that contains 16 annotated proteins. The Ligon-lintless-1 (Li 1 ) mutant of cotton (Gossypium hirsutum L.) has been studied as a model for cotton fiber development since its identification in 1929; however, the causative mutation has not been identified yet. Here we report the fine genetic mapping of the mutation to a 255-kb region that contains only 16 annotated genes in the reference Gossypium raimondii genome. We took advantage of the incompletely dominant dwarf vegetative phenotype to identify 100 mutants (Li 1 /Li 1 ) and 100 wild-type (li 1 /li 1 ) homozygotes from a mapping population of 2567 F2 plants, which we bulked and deep sequenced. Since only homozygotes were sequenced, we were able to use a high stringency in SNP calling to rapidly narrow down the region harboring the Li 1 locus, and designed subgenome-specific SNP markers to test the population. We characterized the expression of all sixteen genes in the region by RNA sequencing of elongating fibers and by RT-qPCR at seven time points spanning fiber development. One of the most highly expressed genes found in this interval in wild-type fiber cells is 40-fold under-expressed at the day of anthesis (DOA) in the mutant fiber cells.  This gene is a major facilitator superfamily protein, part of the large family of proteins that includes auxin and sugar transporters. Interestingly, nearly all genes in this region were most highly expressed at DOA and showed a high degree of co-expression. Further characterization is required to determine if transport of hormones or carbohydrates is involved in both the dwarf and lintless phenotypes of Li 1 plants.

Polymorphisms and linkage analysis for ICAM-1 and the selectin gene cluster

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

Vora, D.K.; Rosenbloom, C.L.; Cottingham, R.W.

1994-06-01

Genetic polymorphisms in leukocyte and endothelial cell adhesion molecules may be important variables with regard to susceptibility to multifactorial disease processes that include an inflammatory component. For this reason, polymorphisms were sought for intercellular adhesion molecule-1 (ICAM-1; gene symbol ICAM1) and for the three genes in the selectin cluster, P-selectin, L-selectin, and E-selectin (gene symbols SELP, SELL, and SELE, respectively). Two amino acid polymorphisms were identified for ICAM-1; Gly or Arg at codon 241 and Lys or Glu at codon 469. Dinucleotide repeat polymorphisms were identified in the 3{prime}-untranslated region for ICAM-1 and in intron 9 for P-selectin. Restriction fragmentmore » length polymorphisms were found using cDNAs for each of the three selectin genes as probes; E-selectin with BglII, P-selectin with ScaI, and L-selectin with HincII. Linkage analysis was performed for the selectin gene cluster and for ICAM-1 using the CEPH families; ICAM-1 is very tightly linked to the LDL receptor on chromosome 19, and the selectin cluster is linked to markers at chromosome 1q23. 41 refs., 2 tabs.« less