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1

Blue eye color in humans may be caused by a perfectly associated founder mutation in a regulatory element located within the HERC2 gene inhibiting OCA2 expression.  

PubMed

The human eye color is a quantitative trait displaying multifactorial inheritance. Several studies have shown that the OCA2 locus is the major contributor to the human eye color variation. By linkage analysis of a large Danish family, we finemapped the blue eye color locus to a 166 Kbp region within the HERC2 gene. By association analyses, we identified two SNPs within this region that were perfectly associated with the blue and brown eye colors: rs12913832 and rs1129038. Of these, rs12913832 is located 21.152 bp upstream from the OCA2 promoter in a highly conserved sequence in intron 86 of HERC2. The brown eye color allele of rs12913832 is highly conserved throughout a number of species. As shown by a Luciferase assays in cell cultures, the element significantly reduces the activity of the OCA2 promoter and electrophoretic mobility shift assays demonstrate that the two alleles bind different subsets of nuclear extracts. One single haplotype, represented by six polymorphic SNPs covering half of the 3' end of the HERC2 gene, was found in 155 blue-eyed individuals from Denmark, and in 5 and 2 blue-eyed individuals from Turkey and Jordan, respectively. Hence, our data suggest a common founder mutation in an OCA2 inhibiting regulatory element as the cause of blue eye color in humans. In addition, an LOD score of Z = 4.21 between hair color and D14S72 was obtained in the large family, indicating that RABGGTA is a candidate gene for hair color. PMID:18172690

Eiberg, Hans; Troelsen, Jesper; Nielsen, Mette; Mikkelsen, Annemette; Mengel-From, Jonas; Kjaer, Klaus W; Hansen, Lars

2008-03-01

2

Blue eye color in humans may be caused by a perfectly associated founder mutation in a regulatory element located within the HERC2 gene inhibiting OCA2 expression  

Microsoft Academic Search

The human eye color is a quantitative trait displaying multifactorial inheritance. Several studies have shown that the OCA2 locus is the major contributor to the human eye color variation. By linkage analysis of a large Danish family, we finemapped\\u000a the blue eye color locus to a 166 Kbp region within the HERC2 gene. By association analyses, we identified two SNPs within

Hans Eiberg; Jesper Troelsen; Mette Nielsen; Annemette Mikkelsen; Klaus W. Kjaer; Lars Hansen

2008-01-01

3

Frequent intragenic deletion of the P gene in Tanzanian patients with Type II oculocutaneous albinism (OCA2)  

SciTech Connect

Type II oculocutaneous albinism (OCA2) is an autosomal recessive disorder in which the biosynthesis of melanin pigment is reduced in the skin, hair, and eyes. OCA2, which results from mutations of the P gene, is the most frequent type of albinism in African and African-American patients. OCA2 is especially frequent in Tanzania, where it occurs with an incidence of {approximately}1/1,400. We have identified abnormalities of the P gene in each of 13 unrelated patients with OCA2 from Tanzania. One of these, a deletion of exon 7, is strongly predominant, accounting for {approximately}77% of mutant alleles in this group of patients. 20 refs., 2 figs.

Spritz, R.; Fukai, K.; Holmes, S.A. [Univ. of Wisconsin, Madison, WI (United States)] [and others

1995-06-01

4

Functional characterization of two novel splicing mutations in the OCA2 gene associated with oculocutaneous albinism type II.  

PubMed

Oculocutaneous albinism (OCA) is characterized by hypopigmentation of the skin, hair and eye, and by ophthalmologic abnormalities caused by a deficiency in melanin biosynthesis. OCA type II (OCA2) is one of the four commonly-recognized forms of albinism, and is determined by mutation in the OCA2 gene. In the present study, we investigated the molecular basis of OCA2 in two siblings and one unrelated patient. The mutational screening of the OCA2 gene identified two hitherto-unknown putative splicing mutations. The first one (c.1503+5G>A), identified in an Italian proband and her affected sibling, lies in the consensus sequence of the donor splice site of OCA2 intron 14 (IVS14+5G>A), in compound heterozygosity with a frameshift mutation, c.1450_1451insCTGCCCTGACA, which is predicted to determine the premature termination of the polypeptide chain (p.I484Tfs*19). In-silico prediction of the effect of the IVS14+5G>A mutation on splicing showed a score reduction for the mutant splice site and indicated the possible activation of a newly-created deep-intronic acceptor splice site. The second mutation is a synonymous transition (c.2139G>A, p.K713K) involving the last nucleotide of exon 20. This mutation was found in a young African albino patient in compound heterozygosity with a previously-reported OCA2 missense mutation (p.T404M). In-silico analysis predicted that the mutant c.2139G>A allele would result in the abolition of the splice donor site. The effects on splicing of these two novel mutations were investigated using an in-vitro hybrid-minigene approach that led to the demonstration of the causal role of the two mutations and to the identification of aberrant transcript variants. PMID:24361966

Rimoldi, Valeria; Straniero, Letizia; Asselta, Rosanna; Mauri, Lucia; Manfredini, Emanuela; Penco, Silvana; Gesu, Giovanni P; Del Longo, Alessandra; Piozzi, Elena; Soldà, Giulia; Primignani, Paola

2014-03-01

5

Genotyping of five single nucleotide polymorphisms in the OCA2 and HERC2 genes associated with blue-brown eye color in the Japanese population.  

PubMed

Human eye color is a polymorphic phenotype influenced by multiple genes. It has recently been reported that three single nucleotide polymorphisms (SNPs) within intron 1 of the OCA2 gene (rs7495174, rs4778241, rs4778138) and two SNPs in intron 86 (rs12913832) and the 3' UTR region (rs1129038) of the HERC2 gene--located in the upstream of the OCA2 locus--have a high statistical association with human eye color. The present study is the first to examine in detail the genotype and haplotype frequencies for these five SNPs in an Asian (Japanese) population (n = 523) comprising solely brown-eyed individuals. Comparison of the genotype and haplotype distributions in Japanese with those in African and European subjects revealed significant differences between Japanese and other populations. Analysis of haplotypes consisting of four SNPs at the HERC2-OCA2 locus (rs12913832/rs7495174/rs4778241/rs4778138) showed that the most frequent haplotype in the Japanese population is A-GAG (0.568), while the frequency of this haplotype is rather low in the European population, even in the brown-eyed group (0.167). The haplotype distribution in the Japanese population was significantly different from that in the brown-eyed European group (F(ST) = 0.18915). PMID:19472299

Iida, Reiko; Ueki, Misuzu; Takeshita, Haruo; Fujihara, Junko; Nakajima, Tamiko; Kominato, Yoshihiko; Nagao, Masataka; Yasuda, Toshihiro

2009-07-01

6

A Potential Benefit of Albinism in Astyanax Cavefish: Downregulation of the oca2 Gene Increases Tyrosine and Catecholamine Levels as an Alternative to Melanin Synthesis  

PubMed Central

Albinism, the loss of melanin pigmentation, has evolved in a diverse variety of cave animals but the responsible evolutionary mechanisms are unknown. In Astyanax mexicanus, which has a pigmented surface dwelling form (surface fish) and several albino cave-dwelling forms (cavefish), albinism is caused by loss of function mutations in the oca2 gene, which operates during the first step of the melanin synthesis pathway. In addition to albinism, cavefish have evolved differences in behavior, including feeding and sleep, which are under the control of the catecholamine system. The catecholamine and melanin synthesis pathways diverge after beginning with the same substrate, L-tyrosine. Here we describe a novel relationship between the catecholamine and melanin synthesis pathways in Astyanax. Our results show significant increases in L-tyrosine, dopamine, and norepinephrine in pre-feeding larvae and adult brains of Pachón cavefish relative to surface fish. In addition, norepinephrine is elevated in cavefish adult kidneys, which contain the teleost homologs of catecholamine synthesizing adrenal cells. We further show that the oca2 gene is expressed during surface fish development but is downregulated in cavefish embryos. A key finding is that knockdown of oca2 expression in surface fish embryos delays the development of pigmented melanophores and simultaneously increases L-tyrosine and dopamine. We conclude that a potential evolutionary benefit of albinism in Astyanax cavefish may be to provide surplus L-tyrosine as a precursor for the elevated catecholamine synthesis pathway, which could be important for adaptation to the challenging cave environment. PMID:24282555

Parkhurst, Amy; Jeffery, William R.

2013-01-01

7

Genetic analysis of three important genes in pigmentation and melanoma susceptibility: CDKN2A, MC1R and HERC2/OCA2.  

PubMed

The CDKN2A gene is regarded as the major familial malignant melanoma (MM) susceptibility gene. Human pigmentation is one of the main modulators of individual risk of developing MM. Therefore, the genes involved in the determination of skin colour and tanning response are potentially implicated in MM predisposition and may be useful predictors of MM risk in the general population. The human melanocortin-1 receptor gene (MC1R) plays a crucial role in pigmentation and also appears to be important in MM. The OCA2 gene has emerged as a new and significant determinant of human iris colour variation. We present a case-control study in Spanish population including 390 consecutive patients with melanoma and 254 control subjects. Sequence analysis of the entire coding region and genotyping of 5 tag-SNPs in the genomic region of MC1R was performed. We identified 27 variants, two reaching statistical significance [R160W (OR: 4.18, 95% CI: 1.24-14.04, P = 0.02) and D294H (OR: 3.10, 95% CI: 1.37-7.01, P = 0.01)] and we detected two novel non-synonymous changes: V92L and T308M. Odds ratio for carrying two functional variants was 4.25 (95% CI: 2.30-7.84, P = 3.63 x 10(-6)). Haplotypes of the entire MC1R region have been established, and we observed an enrichment of a rare European haplotype similar to African values carrying variants V92M and I155T. In addition, three potentially functional SNPs were selected in p16/CDKN2A and in the promoter region of OCA2/HERC2. Our data for CDKN2A gene did not reach statistically significant results for any of the two studied alleles. We found that the variant allele A > G of OCA2/HERC2 (rs12913832) was associated with pigmentation features: eye, hair and skin colour; P-values = 1.8 x 10(-29), 9.2 x 10(-16), 1.1 x 10(-3), respectively, validating previous results. PMID:20629734

Ibarrola-Villava, Maider; Fernandez, Lara P; Pita, Guillermo; Bravo, Jerónimo; Floristan, Uxua; Sendagorta, Elena; Feito, Marta; Avilés, José A; Martin-Gonzalez, Manuel; Lázaro, Pablo; Benítez, Javier; Ribas, Gloria

2010-09-01

8

Genetic variation in regulatory DNA elements: the case of OCA2 transcriptional regulation.  

PubMed

Mutations within the OCA2 gene or the complete absence of the OCA2 protein leads to oculocutaneous albinism type 2. The OCA2 protein plays a central role in melanosome biogenesis, and it is a strong determinant of the eumelanin content in melanocytes. Transcript levels of the OCA2 gene are strongly correlated with pigmentation intensities. Recent studies demonstrated that the transcriptional level of OCA2 is to a large extent determined by the noncoding SNP rs12913832 located 21.5 kb upstream of the OCA2 gene promoter. In this review, we discuss current hypotheses and the available data on the mechanism of OCA2 transcriptional regulation and how this is influenced by genetic variation. Finally, we will explore how future epigenetic studies can be used to advance our insight into the functional biology that connects genetic variation to human pigmentation. PMID:24387780

Visser, Mijke; Kayser, Manfred; Grosveld, Frank; Palstra, Robert-Jan

2014-03-01

9

oca2 regulation of chromatophore differentiation and number is cell type specific in zebrafish.  

PubMed

We characterized a zebrafish mutant that displays defects in melanin synthesis and in the differentiation of melanophores and iridophores of the skin and retinal pigment epithelium. Positional cloning and candidate gene sequencing link this mutation to a 410-kb region on chromosome 6, containing the oculocutaneous albinism 2 (oca2) gene. Quantification of oca2 mutant melanophores shows a reduction in the number of differentiated melanophores compared with wildtype siblings. Consistent with the analysis of mouse Oca2-deficient melanocytes, zebrafish mutant melanophores have immature melanosomes which are partially rescued following treatment with vacuolar-type ATPase inhibitor/cytoplasmic pH modifier, bafilomycin A1. Melanophore-specific gene expression is detected at the correct time and in anticipated locations. While oca2 zebrafish display unpigmented gaps on the head region of mutants 3 days post-fertilization, melanoblast quantification indicates that oca2 mutants have the correct number of melanoblasts, suggesting a differentiation defect explains the reduced melanophore number. Unlike melanophores, which are reduced in number in oca2 mutants, differentiated iridophores are present at significantly higher numbers. These data suggest distinct mechanisms for oca2 in establishing differentiated chromatophore number in developing zebrafish. PMID:24330346

Beirl, Alisha J; Linbo, Tor H; Cobb, Marea J; Cooper, Cynthia D

2014-03-01

10

Human eye colour and HERC2, OCA2 and MATP.  

PubMed

Prediction of human eye colour by forensic genetic methods is of great value in certain crime investigations. Strong associations between blue/brown eye colour and the SNP loci rs1129038 and rs12913832 in the HERC2 gene were recently described. Weaker associations between eye colour and other genetic markers also exist. In 395 randomly selected Danes, we investigated the predictive values of various combinations of SNP alleles in the HERC2, OCA2 and MATP (SLC45A2) genes and compared the results to the eye colours as they were described by the individuals themselves. The highest predictive value of typing either the HERC2 SNPs rs1129038 and/or rs12913832 that are in strong linkage disequilibrium was observed when eye colour was divided into two groups, (1) blue, grey and green (light) and (2) brown and hazel (dark). Sequence variations in rs11636232 and rs7170852 in HERC2, rs1800407 in OCA2 and rs16891982 in MATP showed additional association with eye colours in addition to the effect of HERC2 rs1129038. Diplotype analysis of three sequence variations in HERC2 and one sequence variation in OCA2 showed the best discrimination between light and dark eye colours with a likelihood ratio of 29.3. PMID:20457063

Mengel-From, Jonas; Børsting, Claus; Sanchez, Juan J; Eiberg, Hans; Morling, Niels

2010-10-01

11

Interactions between HERC2, OCA2 and MC1R may influence human pigmentation phenotype.  

PubMed

Human pigmentation is a polygenic trait which may be shaped by different kinds of gene-gene interactions. Recent studies have revealed that interactive effects between HERC2 and OCA2 may be responsible for blue eye colour determination in humans. Here we performed a population association study, examining important polymorphisms within the HERC2 and OCA2 genes. Furthermore, pooling these results with genotyping data for MC1R, ASIP and SLC45A2 obtained for the same population sample we also analysed potential genetic interactions affecting variation in eye, hair and skin colour. Our results confirmed the association of HERC2 rs12913832 with eye colour and showed that this SNP is also significantly associated with skin and hair colouration. It is also concluded that OCA2 rs1800407 is independently associated with eye colour. Finally, using various approaches we were able to show that there is an interaction between MC1R and HERC2 in determination of skin and hair colour in the studied population sample. PMID:19208107

Branicki, Wojciech; Brudnik, Urszula; Wojas-Pelc, Anna

2009-03-01

12

Analysis of cultured human melanocytes based on polymorphisms within the SLC45A2/MATP, SLC24A5/NCKX5, and OCA2/P loci.  

PubMed

Single nucleotide polymorphisms (SNPs) within the SLC45A2/MATP, SLC24A5/NCKX5, and OCA2/P genes have been associated with natural variation of pigmentation traits in human populations. Here, we describe the characterization of human primary melanocytic cells genotyped for polymorphisms within the MATP, NCKX5, or OCA2 loci. On the basis of genotype, these cultured cells reflect the phenotypes observed by others in terms of both melanin content and tyrosinase (TYR) activity when comparing skin designated as either "White" or "Black". We found a statistically significant association of MATP-374L (darker skin) with higher TYR protein abundance that was not observed for any NCKX5-111 or OCA2 rs12913832 allele. MATP-374L/L homozygous strains displayed significantly lower MATP transcript levels compared to MATP-374F/F homozygous cells, but this did not reach statistical significance based on NCKX5 or OCA2 genotype. Similarly, we observed significantly increased levels of OCA2 mRNA in rs12913832-T (brown eye) homozygotes compared to rs12913832-C (blue eye) homozygous strains, which was not observed for MATP or NCKX5 gene transcripts. In genotype-phenotype associations performed on a collection of 226 southern European individuals using these same SNPs, we were able to show strong correlations in MATP-L374F, OCA2, and melanocortin-1 receptor with skin, eye, and hair color variation, respectively. PMID:18650849

Cook, Anthony L; Chen, Wei; Thurber, Amy E; Smit, Darren J; Smith, Aaron G; Bladen, Timothy G; Brown, Darren L; Duffy, David L; Pastorino, Lorenza; Bianchi-Scarra, Giovanna; Leonard, J Helen; Stow, Jennifer L; Sturm, Richard A

2009-02-01

13

HERC2 rs12913832 modulates human pigmentation by attenuating chromatin-loop formation between a long-range enhancer and the OCA2 promoter.  

PubMed

Pigmentation of skin, eye, and hair reflects some of the most evident common phenotypes in humans. Several candidate genes for human pigmentation are identified. The SNP rs12913832 has strong statistical association with human pigmentation. It is located within an intron of the nonpigment gene HERC2, 21 kb upstream of the pigment gene OCA2, and the region surrounding rs12913832 is highly conserved among animal species. However, the exact functional role of HERC2 rs12913832 in human pigmentation is unknown. Here we demonstrate that the HERC2 rs12913832 region functions as an enhancer regulating OCA2 transcription. In darkly pigmented human melanocytes carrying the rs12913832 T-allele, we detected binding of the transcription factors HLTF, LEF1, and MITF to the HERC2 rs12913832 enhancer, and a long-range chromatin loop between this enhancer and the OCA2 promoter that leads to elevated OCA2 expression. In contrast, in lightly pigmented melanocytes carrying the rs12913832 C-allele, chromatin-loop formation, transcription factor recruitment, and OCA2 expression are all reduced. Hence, we demonstrate that allelic variation of a common noncoding SNP located in a distal regulatory element not only disrupts the regulatory potential of this element but also affects its interaction with the relevant promoter. We provide the key mechanistic insight that allele-dependent differences in chromatin-loop formation (i.e., structural differences in the folding of gene loci) result in differences in allelic gene expression that affects common phenotypic traits. This concept is highly relevant for future studies aiming to unveil the functional basis of genetically determined phenotypes, including diseases. PMID:22234890

Visser, Mijke; Kayser, Manfred; Palstra, Robert-Jan

2012-03-01

14

HERC2 rs12913832 modulates human pigmentation by attenuating chromatin-loop formation between a long-range enhancer and the OCA2 promoter  

PubMed Central

Pigmentation of skin, eye, and hair reflects some of the most evident common phenotypes in humans. Several candidate genes for human pigmentation are identified. The SNP rs12913832 has strong statistical association with human pigmentation. It is located within an intron of the nonpigment gene HERC2, 21 kb upstream of the pigment gene OCA2, and the region surrounding rs12913832 is highly conserved among animal species. However, the exact functional role of HERC2 rs12913832 in human pigmentation is unknown. Here we demonstrate that the HERC2 rs12913832 region functions as an enhancer regulating OCA2 transcription. In darkly pigmented human melanocytes carrying the rs12913832 T-allele, we detected binding of the transcription factors HLTF, LEF1, and MITF to the HERC2 rs12913832 enhancer, and a long-range chromatin loop between this enhancer and the OCA2 promoter that leads to elevated OCA2 expression. In contrast, in lightly pigmented melanocytes carrying the rs12913832 C-allele, chromatin-loop formation, transcription factor recruitment, and OCA2 expression are all reduced. Hence, we demonstrate that allelic variation of a common noncoding SNP located in a distal regulatory element not only disrupts the regulatory potential of this element but also affects its interaction with the relevant promoter. We provide the key mechanistic insight that allele-dependent differences in chromatin-loop formation (i.e., structural differences in the folding of gene loci) result in differences in allelic gene expression that affects common phenotypic traits. This concept is highly relevant for future studies aiming to unveil the functional basis of genetically determined phenotypes, including diseases. PMID:22234890

Visser, Mijke; Kayser, Manfred; Palstra, Robert-Jan

2012-01-01

15

A Three-Single-Nucleotide Polymorphism Haplotype in Intron 1 of OCA2 Explains Most Human Eye-Color Variation  

PubMed Central

We have previously shown that a quantitative-trait locus linked to the OCA2 region of 15q accounts for 74% of variation in human eye color. We conducted additional genotyping to clarify the role of the OCA2 locus in the inheritance of eye color and other pigmentary traits associated with skin-cancer risk in white populations. Fifty-eight synonymous and nonsynonymous exonic single-nucleotide polymorphisms (SNPs) and tagging SNPs were typed in a collection of 3,839 adolescent twins, their siblings, and their parents. The highest association for blue/nonblue eye color was found with three OCA2 SNPs: rs7495174 T/C, rs6497268 G/T, and rs11855019 T/C (P values of 1.02×10-61, 1.57×10-96, and 4.45×10-54, respectively) in intron 1. These three SNPs are in one major haplotype block, with TGT representing 78.4% of alleles. The TGT/TGT diplotype found in 62.2% of samples was the major genotype seen to modify eye color, with a frequency of 0.905 in blue or green compared with only 0.095 in brown eye color. This genotype was also at highest frequency in subjects with light brown hair and was more frequent in fair and medium skin types, consistent with the TGT haplotype acting as a recessive modifier of lighter pigmentary phenotypes. Homozygotes for rs11855019 C/C were predominantly without freckles and had lower mole counts. The minor population impact of the nonsynonymous coding-region polymorphisms Arg305Trp and Arg419Gln associated with nonblue eyes and the tight linkage of the major TGT haplotype within the intron 1 of OCA2 with blue eye color and lighter hair and skin tones suggest that differences within the 5? proximal regulatory control region of the OCA2 gene alter expression or messenger RNA–transcript levels and may be responsible for these associations. PMID:17236130

Duffy, David L.; Montgomery, Grant W.; Chen, Wei; Zhao, Zhen Zhen; Le, Lien; James, Michael R.; Hayward, Nicholas K.; Martin, Nicholas G.; Sturm, Richard A.

2007-01-01

16

Association of the OCA2 Polymorphism His615Arg with Melanin Content in East Asian Populations: Further Evidence of Convergent Evolution of Skin Pigmentation  

PubMed Central

The last decade has witnessed important advances in our understanding of the genetics of pigmentation in European populations, but very little is known about the genes involved in skin pigmentation variation in East Asian populations. Here, we present the results of a study evaluating the association of 10 Single Nucleotide Polymorphisms (SNPs) located within 5 pigmentation candidate genes (OCA2, DCT, ADAM17, ADAMTS20, and TYRP1) with skin pigmentation measured quantitatively in a sample of individuals of East Asian ancestry living in Canada. We show that the non-synonymous polymorphism rs1800414 (His615Arg) located within the OCA2 gene is significantly associated with skin pigmentation in this sample. We replicated this result in an independent sample of Chinese individuals of Han ancestry. This polymorphism is characterized by a derived allele that is present at a high frequency in East Asian populations, but is absent in other population groups. In both samples, individuals with the derived G allele, which codes for the amino acid arginine, show lower melanin levels than those with the ancestral A allele, which codes for the amino acid histidine. An analysis of this non-synonymous polymorphism using several programs to predict potential functional effects provides additional support for the role of this SNP in skin pigmentation variation in East Asian populations. Our results are consistent with previous research indicating that evolution to lightly-pigmented skin occurred, at least in part, independently in Europe and East Asia. PMID:20221248

Edwards, Melissa; Bigham, Abigail; Tan, Jinze; Li, Shilin; Gozdzik, Agnes; Ross, Kendra; Jin, Li; Parra, Esteban J.

2010-01-01

17

Association Between a Germline OCA2 Polymorphism at Chromosome 15q13.1 and Estrogen Receptor-Negative Breast Cancer Survival  

PubMed Central

Background Traditional prognostic factors for survival and treatment response of patients with breast cancer do not fully account for observed survival variation. We used available genotype data from a previously conducted two-stage, breast cancer susceptibility genome-wide association study (ie, Studies of Epidemiology and Risk factors in Cancer Heredity [SEARCH]) to investigate associations between variation in germline DNA and overall survival. Methods We evaluated possible associations between overall survival after a breast cancer diagnosis and 10?621 germline single-nucleotide polymorphisms (SNPs) from up to 3761 patients with invasive breast cancer (including 647 deaths and 26?978 person-years at risk) that were genotyped previously in the SEARCH study with high-density oligonucleotide microarrays (ie, hypothesis-generating set). Associations with all-cause mortality were assessed for each SNP by use of Cox regression analysis, generating a per rare allele hazard ratio (HR). To validate putative associations, we used patient genotype information that had been obtained with 5? nuclease assay or mass spectrometry and overall survival information for up to 14?096 patients with invasive breast cancer (including 2303 deaths and 70?019 person-years at risk) from 15 international case–control studies (ie, validation set). Fixed-effects meta-analysis was used to generate an overall effect estimate in the validation dataset and in combined SEARCH and validation datasets. All statistical tests were two-sided. Results In the hypothesis-generating dataset, SNP rs4778137 (C>G) of the OCA2 gene at 15q13.1 was statistically significantly associated with overall survival among patients with estrogen receptor–negative tumors, with the rare G allele being associated with increased overall survival (HR of death per rare allele carried = 0.56, 95% confidence interval [CI] = 0.41 to 0.75, P = 9.2 × 10?5). This association was also observed in the validation dataset (HR of death per rare allele carried = 0.88, 95% CI = 0.78 to 0.99, P = .03) and in the combined dataset (HR of death per rare allele carried = 0.82, 95% CI = 0.73 to 0.92, P = 5 × 10?4). Conclusion The rare G allele of the OCA2 polymorphism, rs4778137, may be associated with improved overall survival among patients with estrogen receptor–negative breast cancer. PMID:20308648

Tyrer, Jonathan; Fasching, Peter A.; Beckmann, Matthias W.; Ekici, Arif B.; Schulz-Wendtland, Rudiger; Bojesen, Stig E.; Nordestgaard, B?rge G.; Flyger, Henrik; Milne, Roger L.; Arias, Jose Ignacio; Menendez, Primitiva; Benitez, Javier; Chang-Claude, Jenny; Hein, Rebecca; Wang-Gohrke, Shan; Nevanlinna, Heli; Heikkinen, Tuomas; Aittomaki, Kristiina; Blomqvist, Carl; Margolin, Sara; Mannermaa, Arto; Kosma, Veli-Matti; Kataja, Vesa; Beesley, Jonathan; Chen, Xiaoqing; Chenevix-Trench, Georgia; Couch, Fergus J.; Olson, Janet E.; Fredericksen, Zachary S.; Wang, Xianshu; Giles, Graham G.; Severi, Gianluca; Baglietto, Laura; Southey, Melissa C.; Devilee, Peter; Tollenaar, Rob A. E. M.; Seynaeve, Caroline; Garcia-Closas, Montserrat; Lissowska, Jolanta; Sherman, Mark E.; Bolton, Kelly L.; Hall, Per; Czene, Kamila; Cox, Angela; Brock, Ian W.; Elliott, Graeme C.; Reed, Malcolm W. R.; Greenberg, David; Anton-Culver, Hoda; Ziogas, Argyrios; Humphreys, Manjeet; Easton, Douglas F.; Caporaso, Neil E.; Pharoah, Paul D. P.

2010-01-01

18

Albinism in phylogenetically and geographically distinct populations of Astyanax cavefish arises through the same loss-of-function Oca2 allele  

PubMed Central

The Mexican tetra, Astyanax mexicanus, comprises 29 populations of cave-adapted fish distributed across a vast karst region in northeastern Mexico. These populations have a complex evolutionary history, having descended from ‘old' and ‘young' ancestral surface-dwelling stocks that invaded the region ?6.7 and ?2.8 MYa, respectively. This study investigates a set of captive, pigmented Astyanax cavefish collected from the Micos cave locality in 1970, in which albinism appeared over the past two decades. We combined novel coloration analyses, coding sequence comparisons and mRNA expression level studies to investigate the origin of albinism in captive-bred Micos cavefish. We discovered that albino Micos cavefish harbor two copies of a loss-of-function ocular and cutaneous albinism type II (Oca2) allele previously identified in the geographically distant Pachón cave population. This result suggests that phylogenetically young Micos cavefish and phylogenetically old Pachón cave fish inherited this Oca2 allele from the ancestral surface-dwelling taxon. This likely resulted from the presence of the loss-of-function Oca2 haplotype in the ‘young' ancestral surface-dwelling stock that colonized the Micos cave and also introgressed into the ancient Pachón cave population. The appearance of albinism in captive Micos cavefish, caused by the same loss-of-function allele present in Pachón cavefish, implies that geographically and phylogenetically distinct cave populations can evolve the same troglomorphic phenotype from standing genetic variation present in the ancestral taxon. PMID:23572122

Gross, J B; Wilkens, H

2013-01-01

19

Contrasting signals of positive selection in genes involved in human skin color variation from tests based on SNP scans and resequencing  

E-print Network

involved in pigmentation traits. However, it is unclear how well the signatures discovered by such haplotype-based test statistics can be reproduced in tests based on full resequencing data. Four genes (oculocutaneous albinism II (OCA2), tyrosinase...

de Gruijter, Johanna Maria; Lao, Oscar; Vermeulen, Mark; Xue, Yali; Woodwark, Cara; Gillson, Christopher J; Coffey, Alison J; Ayub, Qasim; Mehdi, S QASIM; Kayser, Manfred; Tyler-Smith, Chris

2011-12-01

20

Inhibition of polyadenylation reduces inflammatory gene induction.  

PubMed

Cordycepin (3' deoxyadenosine) has long been used in the study of in vitro assembled polyadenylation complexes, because it terminates the poly(A) tail and arrests the cleavage complex. It is derived from caterpillar fungi, which are highly prized in Chinese traditional medicine. Here we show that cordycepin specifically inhibits the induction of inflammatory mRNAs by cytokines in human airway smooth muscle cells without affecting the expression of control mRNAs. Cordycepin treatment results in shorter poly(A) tails, and a reduction in the efficiency of mRNA cleavage and transcription termination is observed, indicating that the effects of cordycepin on 3' processing in cells are similar to those described in in vitro reactions. For the CCL2 and CXCL1 mRNAs, the effects of cordycepin are post-transcriptional, with the mRNA disappearing during or immediately after nuclear export. In contrast, although the recruitment of RNA polymerase II to the IL8 promoter is also unaffected, the levels of nascent transcript are reduced, indicating a defect in transcription elongation. We show that a reporter construct with 3' sequences from a histone gene is unaffected by cordycepin, while CXCL1 sequences confer cordycepin sensitivity to the reporter, demonstrating that polyadenylation is indeed required for the effect of cordycepin on gene expression. In addition, treatment with another polyadenyation inhibitor and knockdown of poly(A) polymerase ? also specifically reduced the induction of inflammatory mRNAs. These data demonstrate that there are differences in the 3' processing of inflammatory and housekeeping genes and identify polyadenylation as a novel target for anti-inflammatory drugs. PMID:23118416

Kondrashov, Alexander; Meijer, Hedda A; Barthet-Barateig, Adeline; Parker, Hannah N; Khurshid, Asma; Tessier, Sarah; Sicard, Marie; Knox, Alan J; Pang, Linhua; De Moor, Cornelia H

2012-12-01

21

Inhibition of polyadenylation reduces inflammatory gene induction  

PubMed Central

Cordycepin (3? deoxyadenosine) has long been used in the study of in vitro assembled polyadenylation complexes, because it terminates the poly(A) tail and arrests the cleavage complex. It is derived from caterpillar fungi, which are highly prized in Chinese traditional medicine. Here we show that cordycepin specifically inhibits the induction of inflammatory mRNAs by cytokines in human airway smooth muscle cells without affecting the expression of control mRNAs. Cordycepin treatment results in shorter poly(A) tails, and a reduction in the efficiency of mRNA cleavage and transcription termination is observed, indicating that the effects of cordycepin on 3? processing in cells are similar to those described in in vitro reactions. For the CCL2 and CXCL1 mRNAs, the effects of cordycepin are post-transcriptional, with the mRNA disappearing during or immediately after nuclear export. In contrast, although the recruitment of RNA polymerase II to the IL8 promoter is also unaffected, the levels of nascent transcript are reduced, indicating a defect in transcription elongation. We show that a reporter construct with 3? sequences from a histone gene is unaffected by cordycepin, while CXCL1 sequences confer cordycepin sensitivity to the reporter, demonstrating that polyadenylation is indeed required for the effect of cordycepin on gene expression. In addition, treatment with another polyadenyation inhibitor and knockdown of poly(A) polymerase ? also specifically reduced the induction of inflammatory mRNAs. These data demonstrate that there are differences in the 3? processing of inflammatory and housekeeping genes and identify polyadenylation as a novel target for anti-inflammatory drugs. PMID:23118416

Kondrashov, Alexander; Meijer, Hedda A.; Barthet-Barateig, Adeline; Parker, Hannah N.; Khurshid, Asma; Tessier, Sarah; Sicard, Marie; Knox, Alan J.; Pang, Linhua; de Moor, Cornelia H.

2012-01-01

22

Expression of a truncated tomato polygalacturonase gene inhibits expression of the endogenous gene in transgenic plants  

Microsoft Academic Search

Tomato plants were transformed with a chimaeric polygalacturonase (PG) gene, designed to produce a truncated PG transcript constitutively. In these plants expression of the endogenous PG gene was inhibited during ripening, resulting in a substantial reduction in PG mRNA and enzyme accumulation. This inhibition was comparable to that achieved previously using antisense genes. The expression of the truncated gene in

C. J. S. Smith; C. F. Watson; C. R. Bird; J. Ray; W. Schuch; D. Grierson

1990-01-01

23

Antisense RNA inhibition of polygalacturonase gene expression in transgenic tomatoes  

Microsoft Academic Search

Regulation of expression of specific genes by antisense RNA is a naturally occurring mechanism in bacteria1,2, although gene regulation by this mechanism has not yet been observed in higher eukaryotes. However, antisense RNA has been shown to reduce expression of specific genes when injected into frog oocytes3 and Drosophila embryos4. Inhibition of expression of artificially introduced genes has been demonstrated

C. J. S. Smith; C. F. Watson; J. Ray; C. R. Bird; P. C. Morris; W. Schuch; D. Grierson

1988-01-01

24

Antisense Oligodeoxynucleotide Inhibition of HIV Gene Expression.  

National Technical Information Service (NTIS)

The goal of this work is to develop novel, efficacious, injectable, gene-specific therapeutics for treatment of human immunodeficiency virus (HIV) infection. These products will be nuclease resistant, stereospecific antisense inhibitors of human immunodef...

E. Wickstrom

1989-01-01

25

A Single SNP in an Evolutionary Conserved Region within Intron 86 of the HERC2 Gene Determines Human Blue-Brown Eye Color  

PubMed Central

We have previously demonstrated that haplotypes of three single nucleotide polymorphisms (SNPs) within the first intron of the OCA2 gene are extremely strongly associated with variation in human eye color. In the present work, we describe additional fine association mapping of eye color SNPs in the intergenic region upstream of OCA2 and within the neighboring HERC2 (hect domain and RLD2) gene. We screened an additional 92 SNPs in 300–3000 European individuals and found that a single SNP in intron 86 of HERC2, rs12913832, predicted eye color significantly better (ordinal logistic regression R2 = 0.68, association LOD = 444) than our previous best OCA2 haplotype. Comparison of sequence alignments of multiple species showed that this SNP lies in the center of a short highly conserved sequence and that the blue-eye-associated allele (frequency 78%) breaks up this conserved sequence, part of which forms a consensus binding site for the helicase-like transcription factor (HLTF). We were also able to demonstrate the OCA2 R419Q, rs1800407, coding SNP acts as a penetrance modifier of this new HERC2 SNP for eye color, and somewhat independently, of melanoma risk. We conclude that the conserved region around rs12913832 represents a regulatory region controlling constitutive expression of OCA2 and that the C allele at rs12913832 leads to decreased expression of OCA2, particularly within iris melanocytes, which we postulate to be the ultimate cause of blue eye color. PMID:18252222

Sturm, Richard A.; Duffy, David L.; Zhao, Zhen Zhen; Leite, Fabio P.N.; Stark, Mitchell S.; Hayward, Nicholas K.; Martin, Nicholas G.; Montgomery, Grant W.

2008-01-01

26

A single SNP in an evolutionary conserved region within intron 86 of the HERC2 gene determines human blue-brown eye color.  

PubMed

We have previously demonstrated that haplotypes of three single nucleotide polymorphisms (SNPs) within the first intron of the OCA2 gene are extremely strongly associated with variation in human eye color. In the present work, we describe additional fine association mapping of eye color SNPs in the intergenic region upstream of OCA2 and within the neighboring HERC2 (hect domain and RLD2) gene. We screened an additional 92 SNPs in 300-3000 European individuals and found that a single SNP in intron 86 of HERC2, rs12913832, predicted eye color significantly better (ordinal logistic regression R(2) = 0.68, association LOD = 444) than our previous best OCA2 haplotype. Comparison of sequence alignments of multiple species showed that this SNP lies in the center of a short highly conserved sequence and that the blue-eye-associated allele (frequency 78%) breaks up this conserved sequence, part of which forms a consensus binding site for the helicase-like transcription factor (HLTF). We were also able to demonstrate the OCA2 R419Q, rs1800407, coding SNP acts as a penetrance modifier of this new HERC2 SNP for eye color, and somewhat independently, of melanoma risk. We conclude that the conserved region around rs12913832 represents a regulatory region controlling constitutive expression of OCA2 and that the C allele at rs12913832 leads to decreased expression of OCA2, particularly within iris melanocytes, which we postulate to be the ultimate cause of blue eye color. PMID:18252222

Sturm, Richard A; Duffy, David L; Zhao, Zhen Zhen; Leite, Fabio P N; Stark, Mitchell S; Hayward, Nicholas K; Martin, Nicholas G; Montgomery, Grant W

2008-02-01

27

Curcumin inhibits ultraviolet light induced human immunodeficiency virus gene expression.  

PubMed

Recently, we reported that the herbal drug St. John's Wort is a potent inhibitor of UV-induced HIV-LTR activation in stably transfected HIVcat/HeLa cells. Our previous studies have demonstrated that the activation of p38 MAP kinase (stress-activated protein kinase-2) and NF-kappaB are both required for a full UV-induced HIV gene expression response. In this study we have investigated the mechanism by which curcumin inhibits UV-activated HIV-LTR gene expression. We found that treatment of HIVcat/HeLa cells with micromolar concentrations of curcumin completely abolished UV activation of HIV gene expression. Curcumin treatment at similar doses as those used to inhibit HIV gene expression also effectively blocked UV activation of NF-kappaB, as demonstrated by electrophoretic mobility shift assay. In contrast, curcumin did not inhibit UV-induced phosphorylation of p38 MAP kinase. This observation was also supported by findings that curcumin did not inhibit UV-induced phosphorylation of CREB/ATF-1 and ATF-2. Although curcumin was ineffective in preventing UV-induced p44/42 MAP kinase phosphorylation, the JNK (1 and 2) and AP-1 activation were efficiently blocked by curcumin in HeLa cells. We conclude that the mechanism by which curcumin modulates UV activation of HIV-LTR gene expression mainly involves the inhibition of NF-kappaB activation. PMID:14674708

Taher, Mohiuddin M; Lammering, Guido; Hershey, Chad; Valerie, Kristoffer

2003-12-01

28

Nanoparticle-mediated p53 gene therapy for tumor inhibition  

PubMed Central

The p53 tumor suppressor gene is mutated in 50% of human cancers, resulting in more aggressive disease with greater resistance to chemotherapy and radiation therapy. Advances in gene therapy technologies offer a promising approach to restoring p53 function. We have developed polymeric nanoparticles (NPs), based on poly (lactic-co-glycolic acid), that provide sustained intracellular delivery of plasmid DNA, resulting in sustained gene expression without vector-associated toxicity. Our previous studies with p53 gene-loaded NPs (p53NPs) demonstrated sustained antiproliferative effects in cancer cells in vitro. The objective of this study was to evaluate the efficacy of p53NPs in vivo. Tumor xenografts in mice were established with human p53-null prostate cancer cells. Animals were treated with p53NPs by either local (intratumoral injection) or systemic (intravenous) administration. Controls included saline, p53 DNA alone, and control NPs. Mice treated with local injections of p53NPs demonstrated significant tumor inhibition and improved animal survival compared with controls. Tumor inhibition corresponded to sustained and greater p53 gene and protein expression in tumors treated with p53NPs than with p53 DNA alone. A single intravenous dose of p53NPs was successful in reducing tumor growth and improving animal survival, although not to the same extent as with local injections. Imaging studies showed that NPs accumulate in tumor tissue after intravenous injection; however, further improvement in tumor targeting efficiency of p53NPs may be needed for better outcome. In conclusion, the NP-mediated p53 gene therapy is effective in tumor growth inhibition. NPs may be developed as nonviral vectors for cancer and other genetic diseases. PMID:22553503

Sharma, Blanka; Ma, Wenxue; Adjei, Isaac Morris; Panyam, Jayanth; Dimitrijevic, Sanja

2012-01-01

29

Inhibition of GLI1 gene activation by Patched1  

PubMed Central

Patched1 (PTCH1) is a human tumour suppressor that acts as an HH (Hedgehog) receptor protein and is important for embryonic patterning. PTCH1 mediates its effects through SMO (Smoothened) and represses the expression of HH target genes such as the transcription factor GLI1 (glioma 1) as well as PTCH1. Up-regulation of these genes has been observed in several cancer forms, including basal cell carcinoma, digestive track tumours and small cell lung cancer. The fact that PTCH1 down-regulates its own expression via ‘negative feedback’ is an important feature in HH signalling, as it keeps the balance between HH and PTCH1 activities that are essential for normal development. In the present study, we provide evidence that a novel mechanism allowing PTCH1 to maintain this balance may also exist. We show that gene activation by GLI1, the transcriptional effector of the pathway, can be down-regulated by PTCH1 without involvement of the canonical cascade of HH signalling events. Specifically, the SMO antagonist cyclopamine has no appreciable effects in blocking this PTCH1-mediated inhibition. Moreover, the negative GLI1 regulator SUFU (Suppressor of Fused) was also found to be dispensable. Additionally, deletion mapping of PTCH1 has revealed that the domains encompassed by amino acids 180–786 and 1058–1210 are of highest significance in inhibiting GLI1 gene activation. This contrasts with the importance of the PTCH1 C-terminal domain for HH signalling. PMID:16229683

Rahnama, Fahimeh; Shimokawa, Takashi; Lauth, Matthias; Finta, Csaba; Kogerman, Priit; Teglund, Stephan; Toftgard, Rune; Zaphiropoulos, Peter G.

2005-01-01

30

Three Genome-wide Association Studies and a Linkage Analysis Identify HERC2 as a Human Iris Color Gene  

PubMed Central

Human iris color was one of the first traits for which Mendelian segregation was established. To date, the genetics of iris color is still not fully understood and is of interest, particularly in view of forensic applications. In three independent genome-wide association (GWA) studies of a total of 1406 persons and a genome-wide linkage study of 1292 relatives, all from the Netherlands, we found that the 15q13.1 region is the predominant region involved in human iris color. There were no other regions showing consistent genome-wide evidence for association and linkage to iris color. Single nucleotide polymorphisms (SNPs) in the HERC2 gene and, to a lesser extent, in the neighboring OCA2 gene were independently associated to iris color variation. OCA2 has been implicated in iris color previously. A replication study within two populations confirmed that the HERC2 gene is a new and significant determinant of human iris color variation, in addition to OCA2. Furthermore, HERC2 rs916977 showed a clinal allele distribution across 23 European populations, which was significantly correlated to iris color variation. We suggest that genetic variants regulating expression of the OCA2 gene exist in the HERC2 gene or, alternatively, within the 11.7 kb of sequence between OCA2 and HERC2, and that most iris color variation in Europeans is explained by those two genes. Testing markers in the HERC2-OCA2 region may be useful in forensic applications to predict eye color phenotypes of unknown persons of European genetic origin. PMID:18252221

Kayser, Manfred; Liu, Fan; Janssens, A. Cecile J.W.; Rivadeneira, Fernando; Lao, Oscar; van Duijn, Kate; Vermeulen, Mark; Arp, Pascal; Jhamai, Mila M.; van IJcken, Wilfred F.J.; den Dunnen, Johan T.; Heath, Simon; Zelenika, Diana; Despriet, Dominiek D.G.; Klaver, Caroline C.W.; Vingerling, Johannes R.; de Jong, Paulus T.V.M.; Hofman, Albert; Aulchenko, Yurii S.; Uitterlinden, Andre G.; Oostra, Ben A.; van Duijn, Cornelia M.

2008-01-01

31

In Silico Analysis of miRNA-Mediated Gene Regulation in OCA and OA Genes.  

PubMed

Albinism is an autosomal recessive genetic disorder due to low secretion of melanin. The oculocutaneous albinism (OCA) and ocular albinism (OA) genes are responsible for melanin production and also act as a potential targets for miRNAs. The role of miRNA is to inhibit the protein synthesis partially or completely by binding with the 3'UTR of the mRNA thus regulating gene expression. In this analysis, we predicted the genetic variation that occurred in 3'UTR of the transcript which can be a reason for low melanin production thus causing albinism. The single nucleotide polymorphisms (SNPs) in 3'UTR cause more new binding sites for miRNA which binds with mRNA which leads to inhibit the translation process either partially or completely. The SNPs in the mRNA of OCA and OA genes can create new binding sites for miRNA which may control the gene expression and lead to hypopigmentation. We have developed a computational procedure to determine the SNPs in the 3'UTR region of mRNA of OCA (TYR, OCA2, TYRP1 and SLC45A2) and OA (GPR143) genes which will be a potential cause for albinism. We identified 37 SNPs in five genes that are predicted to create 87 new binding sites on mRNA, which may lead to abrogation of the translation process. Expression analysis confirms that these genes are highly expressed in skin and eye regions. It is well supported by enrichment analysis that these genes are mainly involved in eye pigmentation and melanin biosynthesis process. The network analysis also shows how the genes are interacting and expressing in a complex network. This insight provides clue to wet-lab researches to understand the expression pattern of OCA and OA genes and binding phenomenon of mRNA and miRNA upon mutation, which is responsible for inhibition of translation process at genomic levels. PMID:25060099

Kamaraj, Balu; Gopalakrishnan, Chandrasekhar; Purohit, Rituraj

2014-12-01

32

Inhibition of host cell encapsulation through inhibiting immune gene expression by the parasitic wasp venom calreticulin.  

PubMed

Parasitoid wasps inject venom into the host to protect their offspring against host immune responses. In our previous study, we identified a calreticulin (CRT) in Pteromalus puparum venom. In this study, we expressed the wild-type and the coiled-coil domain deletion mutant P. puparum calreticulins (PpCRTs) in Escherichia coli and prepared polyclonal antibody in rabbit against PpCRT. Western blot analysis showed that PpCRT protein was not only present in the venom but also in all the tissues tested. Real time PCR results indicated that PpCRT mRNA was highly expressed in the venom gland. The transcript level of PpCRT in the venom gland was peaked at 2 days post-eclosion, while the PpCRT protein in the venom was maintained at a constant level. Both recombinant wild-type and mutant PpCRT proteins could bind to the surface of P. puparum eggs. Recombinant PpCRT inhibited hemocyte spreading and cellular encapsulation of the host Pieris rapae in vitro, and the coiled-coil domain is important for the inhibitory function of PpCRT. Immunocytochemistry results showed that PpCRT entered P. rapae hemocytes, and the coiled-coil domain played a role in this process. After injection of recombinant PpCRT into P. rapae pupae, real time PCR results showed that PpCRT inhibited transcript levels of host encapsulation-related genes, including calreticulin and scavenger receptor genes. In conclusion, our results suggest that P. puparum venom protects its offspring against host cellular immune responses via its functional component PpCRT to inhibit the expression of host cellular response-related genes. PMID:23933213

Wang, Lei; Fang, Qi; Qian, Cen; Wang, Fei; Yu, Xiao-Qiang; Ye, Gongyin

2013-10-01

33

Gene Therapy Inhibiting Neointimal Vascular Lesion: In vivo Transfer of Endothelial Cell Nitric Oxide Synthase Gene  

NASA Astrophysics Data System (ADS)

It is postulated that vascular disease involves a disturbance in the homeostatic balance of factors regulating vascular tone and structure. Recent developments in gene transfer techniques have emerged as an exciting therapeutic option to treat vascular disease. Several studies have established the feasibility of direct in vivo gene transfer into the vasculature by using reporter genes such as ?-galactosidase or luciferase. To date no study has documented therapeutic effects with in vivo gene transfer of a cDNA encoding a functional enzyme. This study tests the hypothesis that endothelium-derived nitric oxide is an endogenous inhibitor of vascular lesion formation. After denudation by balloon injury of the endothelium of rat carotid arteries, we restored endothelial cell nitric oxide synthase (ec-NOS) expression in the vessel wall by using the highly efficient Sendai virus/liposome in vivo gene transfer technique. ec-NOS gene transfection not only restored NO production to levels seen in normal untreated vessels but also increased vascular reactivity of the injured vessel. Neointima formation at day 14 after balloon injury was inhibited by 70%. These findings provide direct evidence that NO is an endogenous inhibitor of vascular lesion formation in vivo (by inhibiting smooth muscle cell proliferation and migration) and suggest the possibility of ec-NOS transfection as a potential therapeutic approach to treat neointimal hyperplasia.

von der Leyen, Heiko E.; Gibbons, Gary H.; Morishita, Ryuichi; Lewis, Neil P.; Zhang, Lunan; Nakajima, Masatoshi; Kaneda, Yasufumi; Cooke, John P.; Dzau, Victor J.

1995-02-01

34

Cycloheximide inhibition of delayed early gene expression in baculovirus-infected cells  

E-print Network

The baculovirus protein IE I is required for the transactivation of many early viral genes in transient expression assays. However, cycloheximide inhibition studies have failed to reveal a dependence of early gene transcription on expression of IE I...

Ross, Larry Dale

2012-06-07

35

Efficient shRNA-Mediated Inhibition of Gene Expression in Zebrafish  

E-print Network

Despite the broad repertoire of loss of function (LOF) tools available for use in the zebrafish, there remains a need for a simple and rapid method that can inhibit expression of genes at later stages. RNAi would fulfill ...

Sive, Hazel L.

36

BMP7 Gene Transfer via Gold Nanoparticles into Stroma Inhibits Corneal Fibrosis In Vivo  

E-print Network

This study examined the effects of BMP7 gene transfer on corneal wound healing and fibrosis inhibition in vivo using a rabbit model. Corneal haze in rabbits was produced with the excimer laser performing -9 diopters ...

Tandon, Ashish

37

Brain contains inhibiting factors specific to the large T-antigen gene  

Microsoft Academic Search

SV40 large T-antigen (LTa) gene-induced immortalized rat dopamine-producing nerve cells (1RB3AN27), which produce LTa protein and divide in vitro, do not divide and do not produce LTa protein when transplanted into striatum of adult rats. This suggests the presence of LTa gene-inhibiting factors in brain. Here we report that rat brain soluble fraction (SF) contains factors which specifically inhibit LTa

Edward D Clarkson; Francisco G La Rosa; Judith Edwards-Prasad; Sanjay Kumar; Arun Kumar; William Cole; Curt R Freed; Kedar N Prasad

1998-01-01

38

Adherens Junction Formation Inhibits Lentivirus Entry and Gene Transfer  

PubMed Central

Although cellular signaling pathways that affect lentivirus infection have been investigated, the role of cell-cell interactions in lentiviral gene delivery remains elusive. In the course of our studies we observed that lentiviral gene transfer was a strong function of the position of epithelial cells within colonies. While peripheral cells were transduced efficiently, cells in the center of colonies were resistant to gene transfer. In addition, gene delivery was enhanced significantly under culture conditions that disrupted adherens junctions (AJ) but decreased upon AJ formation. In agreement, gene knockdown and gain-of-function approaches showed that ?-catenin, a key component of the AJ complex prevented lentivirus gene transfer. Using a doxycycline regulatable system we showed that expression of dominant negative E-cadherin enhanced gene transfer in a dose-dependent manner. In addition, dissolution of AJ by doxycycline increased entry of lentiviral particles into the cell cytoplasm in a dose-dependent manner. Taken together our results demonstrate that AJ formation renders cells non-permissive to lentiviral gene transfer and may facilitate development of simple means to enhance gene delivery or combat virus infection. PMID:24236116

Padmashali, Roshan; You, Hui; Karnik, Nikhila; Lei, Pedro; Andreadis, Stelios T.

2013-01-01

39

Inhibition of Stat1-Mediated Gene Activation by PIAS1  

Microsoft Academic Search

STAT (signal transducer and activator of transcription) proteins are latent cytoplasmic transcription factors that become activated by tyrosine phosphorylation in response to cytokine stimulation. Tyrosine phosphorylated STATs dimerize and translocate into the nucleus to activate specific genes. Different members of the STAT protein family have distinct functions in cytokine signaling. Biochemical and genetic analysis has demonstrated that Stat1 is essential

Bin Liu; Jiayu Liao; Xiaoping Rao; Steven A. Kushner; Chan D. Chung; David D. Chang; Ke Shuai

1998-01-01

40

Tumor suppressor gene PDCD4 negatively regulates autophagy by inhibiting the expression of autophagy-related gene ATG5  

PubMed Central

PDCD4 (programmed cell death protein 4), a suppressor of gene transcription and translation, plays a crucial inhibitory role in several types of human tumors. However, its underlying mechanisms remain unclear. Autophagy, an evolutionarily conserved catabolic process, maintains cellular homeostasis under stress conditions such as starvation and plays a crucial role in tumor initiation and progression. We report here that PDCD4 inhibits autophagy in multiple cell types both in vitro and in vivo, which in turn contributes to its tumor suppressor activity. Importantly, PDCD4 inhibits the expression of an essential autophagy related gene, ATG5 and the formation of an ATG12–ATG5 complex, and its ma3 domains are required for PDCD4-mediated inhibition of autophagy. Unlike most tumor suppressors that act as positive or dual regulators of autophagy, our findings indicate that PDCD4 negatively regulates autophagy by targeting ATG5, which provides a novel mechanism of tumor suppression. PMID:23486359

Song, Xingguo; Zhang, Xia; Wang, Xiaoyan; Zhu, Faliang; Guo, Chun; Wang, Qun; Shi, Yongyu; Wang, Jianing; Chen, Youhai; Zhang, Lining

2013-01-01

41

Hypoxic inhibition of nonsense-mediated RNA decay regulates gene expression and the integrated stress response.  

PubMed

Nonsense-mediated RNA decay (NMD) rapidly degrades both mutated mRNAs and nonmutated cellular mRNAs in what is thought to be a constitutive fashion. Here we demonstrate that NMD is inhibited in hypoxic cells and that this inhibition is dependent on phosphorylation of the alpha subunit of eukaryotic initiation factor 2 (eIF2alpha). eIF2alpha phosphorylation is known to promote translational and transcriptional up-regulation of genes important for the cellular response to stress. We show that the mRNAs of several of these stress-induced genes are NMD targets and that the repression of NMD stabilizes these mRNAs, thus demonstrating that the inhibition of NMD augments the cellular stress response. Furthermore, hypoxia-induced formation of cytoplasmic stress granules is also dependent on eIF2alpha phosphorylation, and components of the NMD pathway are relocalized to these granules in hypoxic cells, providing a potential mechanism for the hypoxic inhibition of NMD. Our demonstration that NMD is inhibited in hypoxic cells reveals that the regulation of NMD can dynamically alter gene expression and also establishes a novel mechanism for hypoxic gene regulation. PMID:18362164

Gardner, Lawrence B

2008-06-01

42

Genomic targets, and histone acetylation and gene expression profiling of neural HDAC inhibition  

PubMed Central

Histone deacetylase inhibitors (HDACis) have been shown to potentiate hippocampal-dependent memory and synaptic plasticity and to ameliorate cognitive deficits and degeneration in animal models for different neuropsychiatric conditions. However, the impact of these drugs on hippocampal histone acetylation and gene expression profiles at the genomic level, and the molecular mechanisms that underlie their specificity and beneficial effects in neural tissue, remains obscure. Here, we mapped four relevant histone marks (H3K4me3, AcH3K9,14, AcH4K12 and pan-AcH2B) in hippocampal chromatin and investigated at the whole-genome level the impact of HDAC inhibition on acetylation profiles and basal and activity-driven gene expression. HDAC inhibition caused a dramatic histone hyperacetylation that was largely restricted to active loci pre-marked with H3K4me3 and AcH3K9,14. In addition, the comparison of Chromatin immunoprecipitation sequencing and gene expression profiles indicated that Trichostatin A-induced histone hyperacetylation, like histone hypoacetylation induced by histone acetyltransferase deficiency, had a modest impact on hippocampal gene expression and did not affect the transient transcriptional response to novelty exposure. However, HDAC inhibition caused the rapid induction of a homeostatic gene program related to chromatin deacetylation. These results illuminate both the relationship between hippocampal gene expression and histone acetylation and the mechanism of action of these important neuropsychiatric drugs. PMID:23821663

Lopez-Atalaya, Jose P.; Ito, Satomi; Valor, Luis M.; Benito, Eva; Barco, Angel

2013-01-01

43

Urokinase plasminogen activator gene deficiency inhibits fracture cartilage remodeling.  

PubMed

Urokinase plasminogen activator (uPA) regulates a proteolytic cascade of extracellular matrix degradation that functions in tissue development and tissue repair. The development and remodeling of the skeletal extracellular matrix during wound healing suggests that uPA might regulate bone development and repair. To determine whether uPA functions regulate bone development and repair, we examined the basal skeletal phenotype and endochondral bone fracture repair in uPA-deficient mice. The skeletal phenotype of uPA knockout mice was compared with that of control mice under basal conditions by dual-energy X-ray absorptiometry and micro-CT analysis, and during femur fracture repair by micro-CT and histological examination of the fracture callus. No effects of uPA gene deficiency were observed in the basal skeletal phenotype of the whole body or the femur. However, uPA gene deficiency resulted in increased fracture callus cartilage abundance during femur fracture repair at 14 days healing. The increase in cartilage corresponded to reduced tartrate-resistant acid phosphatase (TRAP) staining for osteoclasts in the uPA knockout fracture callus at this time, consistent with impaired osteoclast-mediated remodeling of the fracture cartilage. CD31 staining was reduced in the knockout fracture tissues at this time, suggesting that angiogenesis was also reduced. Osteoclasts also colocalized with CD31 expression in the endothelial cells of the fracture tissues during callus remodeling. These results indicate that uPA promotes remodeling of the fracture cartilage by osteoclasts that are associated with angiogenesis and suggest that uPA promotes angiogenesis and remodeling of the fracture cartilage at this time of bone fracture repair. PMID:23700285

Popa, Nicoleta L; Wergedal, Jon E; Lau, K-H William; Mohan, Subburaman; Rundle, Charles H

2014-03-01

44

Sequence of psi , a gene on the symbiotic plasmid of Rhizobium phaseoli which inhibits exopolysaccharide synthesis and nodulation and demonstration that its transcription is inhibited by psr , another gene on the symbiotic plasmid  

Microsoft Academic Search

A gene termed psi (polysaccharide inhibition), located close to the nodulation genes of the Rhizobium phaseoli symbiotic plasmid pRP2JI inhibited exopolysaccharide synthesis (EPS) and nodulation ability (Nod) in Rhizobium when it was cloned in a multicopy plasmid. The sequence of psi showed that it specified a polypeptide of mol. wt. 10000 that may be associated with the membrane of Rhizobium.

D. Borthakur; A. W. B. Johnston

1987-01-01

45

Delivery and Inhibition of Reporter Genes by Small Interfering RNAs in a Mouse Skin Model  

Microsoft Academic Search

RNA interference offers the potential of a novel therapeutic approach for treating skin disorders. To this end, we investigated delivery of nucleic acids, including a plasmid expressing the reporter gene luciferase, to mouse skin by intradermal injection into footpads using in vivo bioluminescence imaging over multiple time points. In order to evaluate the ability of RNA interference to inhibit skin

Qian Wang; Heini Ilves; Pauline Chu; Christopher H. Contag; Devin Leake; Brian H. Johnston; Roger L. Kaspar

2007-01-01

46

Curcumin, the active constituent of turmeric, inhibits amyloid peptide-induced cytochemokine gene expression  

E-print Network

Curcumin, the active constituent of turmeric, inhibits amyloid peptide-induced cytochemokine gene-a and IL-1b) and chemokines (MIP-1b, MCP-1 and IL-8) in monocytes. We determined whether curcumin expression of cytochemokines. We show that curcumin (12.5­25 lM) sup- presses the activation of Egr-1 DNA

Giri, Ranjit K.

47

RNAi Mediated Tiam1 Gene Knockdown Inhibits Invasion of Retinoblastoma  

PubMed Central

T lymphoma invasion and metastasis protein (Tiam1) is up-regulated in variety of cancers and its expression level is related to metastatic potential of the type of cancer. Earlier, Tiam1 was shown to be overexpressed in retinoblastoma (RB) and we hypothesized that it was involved in invasiveness of RB. This was tested by silencing Tiam1 in RB cell lines (Y79 and Weri-Rb1) using siRNA pool, targeting different regions of Tiam1 mRNA. The cDNA microarray of Tiam1 silenced cells showed gene regulations altered by Tiam1 were predominantly on the actin cytoskeleton interacting proteins, apoptotic initiators and tumorogenic potential targets. The silenced phenotype resulted in decreased growth and increased apoptosis with non-invasive characteristics. Transfection of full length and N-terminal truncated construct (C1199) clearly revealed membrane localization of Tiam1 and not in the case of C580 construct. F-actin staining showed the interaction of Tiam1 with actin in the membrane edges that leads to ruffling, and also imparts varying invasive potential to the cell. The results obtained from our study show for the first time that Tiam1 modulates the cell invasion, mediated by actin cytoskeleton remodeling in RB. PMID:23950931

Biswas, Jyotirmay; Kanwar, Rupinder K.; Kanwar, Jagat R.; Krishnakumar, Subramanian

2013-01-01

48

Organization and sequence of the human P gene and identification of a new family of transport proteins  

SciTech Connect

We have determined the structure, nucleotide sequence, and polymorphisms of the human P gene. Mutations of the P gene result in type H oculocutaneous albinism (OCA2) in humans and pink-eyed dilution (p) in mice. We find that the human P gene is quite large, consisting of 25 exons spanning 250 to 600 kb in chromosome segment 15q11-q13. The P polypeptide appears to define a novel family of small molecule transporters and may be involved in transport of tyrosine, the precursor to melanin synthesis, within the melanocyte. These results provide the basis for analyses of patients with OCA2 and may point toward eventual pharmacologic treatment of this and related disorders of pigmentation. 40 refs., 5 figs., 3 tabs.

Lee, S.T.; Fukai, K.; Spritz, R.A. [Univ. of Wisconsin School of Medicine, Madison, WI (United States)] [and others] [Univ. of Wisconsin School of Medicine, Madison, WI (United States); and others

1995-03-20

49

Ultrasound-mediated interferon {beta} gene transfection inhibits growth of malignant melanoma  

SciTech Connect

Highlights: {yields} Successful ultrasound-mediated transfection of melanoma (C32) cells with IFN-{beta} genes both in vitro and in vivo. {yields} Ultrasound-mediated IFN-{beta} transfection inhibited proliferation of melanoma cells in vitro. {yields} Ultrasound-mediated IFN-{beta} transfection inhibited melanoma tumor growth in vivo. -- Abstract: We investigated the effects of ultrasound-mediated transfection (sonotransfection) of interferon {beta} (IFN-{beta}) gene on melanoma (C32) both in vitro and in vivo. C32 cells were sonotransfected with IFN-{beta} in vitro. Subcutaneous C32 tumors in mice were sonicated weekly immediately after intra-tumor injection with IFN-{beta} genes mixed with microbubbles. Successful sonotransfection with IFN-{beta} gene in vitro was confirmed by ELISA, which resulted in C32 growth inhibition. In vivo, the growth ratio of tumors transfected with IFN-{beta} gene was significantly lower than the other experimental groups. These results may lead to a new method of treatment against melanoma and other hard-to-treat cancers.

Yamaguchi, Kazuki [Department of Dermatology, Fukuoka University School of Medicine, 7-45-1 Nanakuma, Jonan-ku, Fukuoka City 814-0180 (Japan) [Department of Dermatology, Fukuoka University School of Medicine, 7-45-1 Nanakuma, Jonan-ku, Fukuoka City 814-0180 (Japan); Department of Anatomy, Fukuoka University School of Medicine, 7-45-1 Nanakuma, Jonan-ku, Fukuoka City 814-0180 (Japan); Feril, Loreto B., E-mail: ferilism@yahoo.com [Department of Anatomy, Fukuoka University School of Medicine, 7-45-1 Nanakuma, Jonan-ku, Fukuoka City 814-0180 (Japan); Tachibana, Katsuro [Department of Anatomy, Fukuoka University School of Medicine, 7-45-1 Nanakuma, Jonan-ku, Fukuoka City 814-0180 (Japan)] [Department of Anatomy, Fukuoka University School of Medicine, 7-45-1 Nanakuma, Jonan-ku, Fukuoka City 814-0180 (Japan); Takahashi, Akira; Matsuo, Miki; Endo, Hitomi [Department of Dermatology, Fukuoka University School of Medicine, 7-45-1 Nanakuma, Jonan-ku, Fukuoka City 814-0180 (Japan)] [Department of Dermatology, Fukuoka University School of Medicine, 7-45-1 Nanakuma, Jonan-ku, Fukuoka City 814-0180 (Japan); Harada, Yoshimi [Department of Anatomy, Fukuoka University School of Medicine, 7-45-1 Nanakuma, Jonan-ku, Fukuoka City 814-0180 (Japan)] [Department of Anatomy, Fukuoka University School of Medicine, 7-45-1 Nanakuma, Jonan-ku, Fukuoka City 814-0180 (Japan); Nakayama, Juichiro [Department of Dermatology, Fukuoka University School of Medicine, 7-45-1 Nanakuma, Jonan-ku, Fukuoka City 814-0180 (Japan)] [Department of Dermatology, Fukuoka University School of Medicine, 7-45-1 Nanakuma, Jonan-ku, Fukuoka City 814-0180 (Japan)

2011-07-22

50

A macrophage factor inhibits adipocyte gene expression: an in vitro model of cachexia.  

PubMed

Certain infections and malignancies in mammals cause the development of a condition known as cachexia in which the animal continues to lose weight, often while consuming an adequate diet. When macrophages are stimulated with an endotoxin, they produce a factor or factors, termed cachectin, that inhibits the activity of fat-producing (lipogenic) enzymes in cultured adipocytes. This effect may reflect one of the physiological bases for cachexia. In the present study, clones of complementary DNA from genes whose expression is increased during the differentiation of adipocytes were used to study the molecular basis of cachectin's actions. In the presence of cachectin, the expression of the corresponding genes was reversibly and specifically inhibited. Furthermore, when mature adipocytes were exposed to cachectin, the messenger RNA's of those genes diminished and rapidly approached the levels present before differentiation. PMID:3839597

Torti, F M; Dieckmann, B; Beutler, B; Cerami, A; Ringold, G M

1985-08-30

51

Neuropeptide Y gene transfection inhibits post-epileptic hippocampal synaptic reconstruction?  

PubMed Central

Exogenous neuropeptide Y has antiepileptic effects; however, the underlying mechanism and optimal administration method for neuropeptide Y are still unresolved. Previous studies have used intracerebroventricular injection of neuropeptide Y into animal models of epilepsy. In this study, a recombinant adeno-associated virus expression vector carrying the neuropeptide Y gene was injected into the lateral ventricle of rats, while the ipsilateral hippocampus was injected with kainic acid to establish the epileptic model. After transfection of neuropeptide Y gene, mossy fiber sprouting in the hippocampal CA3 region of epileptic rats was significantly suppressed, hippocampal synaptophysin (p38) mRNA and protein expression were inhibited, and epileptic seizures were reduced. These experimental findings indicate that a recombinant adeno-associated virus expression vector carrying the neuropeptide Y gene reduces mossy fiber sprouting and inhibits abnormal synaptophysin expression, thereby suppressing post-epileptic synaptic reconstruction.

Zhang, Fan; Zhao, Wenqing; Li, Wenling; Dong, Changzheng; Zhang, Xinying; Wu, Jiang; Li, Na; Liang, Chuandong

2013-01-01

52

Variants in melanogenesis-related genes associate with skin cancer risk among Japanese populations.  

PubMed

Human skin color is known to be associated with the risk of cutaneous cancer. Some reports indicated that pigmentation-related gene variants were associated with cutaneous cancer risk in Caucasian populations, but there are no similar reports in East Asian populations. This study aimed to evaluate the association between pigmentation-related genes and the risk of skin cancer in Japanese populations. We studied the associations between 12 variants of four pigmentation-related genes and melanin index variations in 198 Japanese patients with skin cancer and compared these findings to those of 500 Japanese controls by using multiple logistic regression analysis. Furthermore, we analyzed an independent sample of 107 Japanese patients with skin cancer. A non-synonymous variant, H615R in the oculocutaneous albinism 2 gene (OCA2), was associated with the risk of malignant melanoma in the Yamagata group (odds ratio [OR], 0.38; 95% confidence interval [CI], 0.17-0.86; P = 0.020). Another non-synonymous variant, A481T in OCA2, was associated with the risk of squamous cell carcinoma and actinic keratosis in the Osaka group (OR, 3.16; 95% CI, 1.41-7.04; P = 0.005). In malignant melanoma cases, the minor allele in OCA2 H615R might have induced the development of lesions in sun-exposed skin (OR, 26.32; 95% CI, 1.96-333; P = 0.014). Our results suggest that some OCA2 variants are definite risk factors for the onset of cutaneous cancer in Japanese populations. PMID:24617981

Yoshizawa, Junko; Abe, Yuko; Oiso, Naoki; Fukai, Kazuyoshi; Hozumi, Yutaka; Nakamura, Tomohiro; Narita, Tomohiko; Motokawa, Tomonori; Wakamatsu, Kazumasa; Ito, Shosuke; Kawada, Akira; Tamiya, Gen; Suzuki, Tamio

2014-04-01

53

Structural features of GmIRCHS, candidate of the I gene inhibiting seed coat pigmentation in soybean: implications for inducing endogenous RNA silencing of chalcone synthase genes  

Microsoft Academic Search

Most commercial soybean varieties have yellow seeds due to loss of pigmentation in the seed coat. The I gene inhibits pigmentation over the entire seed coat, resulting in a uniform yellow color of mature harvested seeds. We previously\\u000a demonstrated that the inhibition of seed coat pigmentation by the I gene results from post-transcriptional gene silencing (PTGS) of chalcone synthase (CHS)

Atsushi Kasai; Kosuke Kasai; Setsuzo Yumoto; Mineo Senda

2007-01-01

54

Antisense oligodeoxynucleotide to the cystic fibrosis gene inhibits anion transport in normal cultured sweat duct cells  

SciTech Connect

The authors have tested the hypothesis that the cystic fibrosis (CF) gene product, called the CF transmembrane conductance regulator (CFTR), mediates anion transport in normal human sweat duct cells. Sweat duct cells in primary culture were treated with oligodeoxynucleotides that were antisense to the CFTR gene transcript in order to block the expression of the wild-type CFTR. Anion transport in CFTR transcript antisense-treated cells was then assessed with a halide-specific dye, 6-methoxy-N-(3-sulfopropryl)quinolinium, and fluorescent digital imaging microscopy to monitor halide influx and efflux from single sweat duct cells. Antisense oligodeoxynucleotide treatment for 24 hr virtually abolished Cl{sup {minus}} transport in sweat duct cells compared with untreated cells or control cells treated with sense oligodeoxynucleotides. Br{sup {minus}} uptake into sweat duct cells was also blocked after a 24-hr CFTR transcript antisense treatments, but not after treatments for only 4 hr. Lower concentrations of antisense oligodeoxynucleotides were less effective at inhibiting Cl{sup {minus}} transport. These results indicate that oligodeoxynucleotides that are antisense to CFTR transcript inhibit sweat duct Cl{sup {minus}} permeability in both a time-dependent and dose-dependent manner. This approach provides evidence that inhibition of the expression of the wild-type CFTR gene in a normal, untransfected epithelial cell results in an inhibition of Cl{sup {minus}} permeability.

Sorscher, E.J.; Kirk, K.L.; Weaver, M.L.; Jilling, T.; Blalock, J.E.; LeBoeuf, R.D. (Univ of Alabama, Birmingham (United States))

1991-09-01

55

Differential Gene Expression for Investigation of Escherichia coli Biofilm Inhibition by Plant Extract Ursolic Acid  

PubMed Central

After 13,000 samples of compounds purified from plants were screened, a new biofilm inhibitor, ursolic acid, has been discovered and identified. Using both 96-well microtiter plates and a continuous flow chamber with COMSTAT analysis, 10 ?g of ursolic acid/ml inhibited Escherichia coli biofilm formation 6- to 20-fold when added upon inoculation and when added to a 24-h biofilm; however, ursolic acid was not toxic to E. coli, Pseudomonas aeruginosa, Vibrio harveyi, and hepatocytes. Similarly, 10 ?g of ursolic acid/ml inhibited biofilm formation by >87% for P. aeruginosa in both complex and minimal medium and by 57% for V. harveyi in minimal medium. To investigate the mechanism of this nontoxic inhibition on a global genetic basis, DNA microarrays were used to study the gene expression profiles of E. coli K-12 grown with or without ursolic acid. Ursolic acid at 10 and 30 ?g/ml induced significantly (P < 0.05) 32 and 61 genes, respectively, and 19 genes were consistently induced. The consistently induced genes have functions for chemotaxis and mobility (cheA, tap, tar, and motAB), heat shock response (hslSTV and mopAB), and unknown functions (such as b1566 and yrfHI). There were 31 and 17 genes repressed by 10 and 30 ?g of ursolic acid/ml, respectively, and 12 genes were consistently repressed that have functions in cysteine synthesis (cysK) and sulfur metabolism (cysD), as well as unknown functions (such as hdeAB and yhaDFG). Ursolic acid inhibited biofilms without interfering with quorum sensing, as shown with the V. harveyi AI-1 and AI-2 reporter systems. As predicted by the differential gene expression, deleting motAB counteracts ursolic acid inhibition (the paralyzed cells no longer become too motile). Based on the differential gene expression, it was also discovered that sulfur metabolism (through cysB) affects biofilm formation (in the absence of ursolic acid). PMID:16000817

Ren, Dacheng; Zuo, Rongjun; Gonzalez Barrios, Andres F.; Bedzyk, Laura A.; Eldridge, Gary R.; Pasmore, Mark E.; Wood, Thomas K.

2005-01-01

56

Gene expression profiling in equine polysaccharide storage myopathy revealed inflammation, glycogenesis inhibition, hypoxia and mitochondrial dysfunctions  

PubMed Central

Background Several cases of myopathies have been observed in the horse Norman Cob breed. Muscle histology examinations revealed that some families suffer from a polysaccharide storage myopathy (PSSM). It is assumed that a gene expression signature related to PSSM should be observed at the transcriptional level because the glycogen storage disease could also be linked to other dysfunctions in gene regulation. Thus, the functional genomic approach could be conducted in order to provide new knowledge about the metabolic disorders related to PSSM. We propose exploring the PSSM muscle fiber metabolic disorders by measuring gene expression in relationship with the histological phenotype. Results Genotypying analysis of GYS1 mutation revealed 2 homozygous (AA) and 5 heterozygous (GA) PSSM horses. In the PSSM muscles, histological data revealed PAS positive amylase resistant abnormal polysaccharides, inflammation, necrosis, and lipomatosis and active regeneration of fibers. Ultrastructural evaluation revealed a decrease of mitochondrial number and structural disorders. Extensive accumulation of an abnormal polysaccharide displaced and partially replaced mitochondria and myofibrils. The severity of the disease was higher in the two homozygous PSSM horses. Gene expression analysis revealed 129 genes significantly modulated (p < 0.05). The following genes were up-regulated over 2 fold: IL18, CTSS, LUM, CD44, FN1, GST01. The most down-regulated genes were the following: mitochondrial tRNA, SLC2A2, PRKC?, VEGF?. Data mining analysis showed that protein synthesis, apoptosis, cellular movement, growth and proliferation were the main cellular functions significantly associated with the modulated genes (p < 0.05). Several up-regulated genes, especially IL18, revealed a severe muscular inflammation in PSSM muscles. The up-regulation of glycogen synthase kinase-3 (GSK3?) under its active form could be responsible for glycogen synthase (GYS1) inhibition and hypoxia-inducible factor (HIF1?) destabilization. Conclusion The main disorders observed in PSSM muscles could be related to mitochondrial dysfunctions, glycogenesis inhibition and the chronic hypoxia of the PSSM muscles. PMID:19664222

Barrey, Eric; Mucher, Elodie; Jeansoule, Nicolas; Larcher, Thibaut; Guigand, Lydie; Herszberg, Bérénice; Chaffaux, Stéphane; Guérin, Gérard; Mata, Xavier; Benech, Philippe; Canale, Marielle; Alibert, Olivier; Maltere, Péguy; Gidrol, Xavier

2009-01-01

57

The Latency-Related Gene of Bovine Herpesvirus 1 Inhibits Programmed Cell Death  

PubMed Central

Although viral gene expression occurs in the peripheral nervous system during acute infection, bovine herpesvirus 1 (BHV-1) gene expression is extinguished, many neurons survive, and latency ensues. The only abundant viral transcript expressed during latency is the latency-related (LR) RNA, which is alternatively spliced in trigeminal ganglia during acute infection (L. Devireddy and C. Jones, J. Virol. 72:7294–7301, 1998). A subset of neurons express a protein encoded by the LR gene and the LR protein (LRP) is associated with cyclin-dependent kinase 2 (Cdk2)/cyclin complexes during productive infection (Y. Jiang, A. Hossain, M. T. Winkler, T. Holt, A. Doster, and C. Jones, J. Virol. 72:8133–8142, 1998). LR gene products inhibit cell cycle progression, perhaps as a result of LRP interacting with Cdk2/cyclin complexes. During acute infection, expression of cyclin A occurs in trigeminal ganglionic neurons (L. M. Schang, A. Hossain, and C. Jones, J. Virol. 70:3807–3814, 1996). Inappropriate expression of G1- and S-phase cyclins can initiate programmed cell death (PCD), apoptosis, in neurons, suggesting that LR gene products inhibit PCD. To test this hypothesis, we modified an assay to measure PCD frequency in transiently transfected cells. C6-ceramide, fumonisin B1 (FB1), or etoposide was used to initiate PCD following transfection of cells with plasmids expressing LR gene products and the ?-galactosidase gene. Transfected cells that survived were quantified by counting ?-galactosidase-positive cells. Plasmids that expressed LR gene products promoted survival of monkey kidney (CV-1), human lung (IMR-90), or mouse neuroblastoma (neuro-2A) cells after induction of PCD. Plasmids with termination codons at the beginning of LR open reading frames or deletion of sequences that mediate splicing of LR RNA did not promote cell survival following PCD induction. We hypothesize that LR gene products play a role in promoting survival of postmitotic neurons during acute infection or reactivation. PMID:10559283

Ciacci-Zanella, Janice; Stone, Melissa; Henderson, Gail; Jones, Clinton

1999-01-01

58

Inhibition of HCV 3a core gene through Silymarin and its fractions  

PubMed Central

Hepatitis C is a major health problem affecting 270 million individuals in world including Pakistan. Current treatment regimen, interferon alpha and ribavirin only cure half of patients due to side effects and high cost. Results In the present study Silybum marianum (Milk thistle) seeds were collected, extracted and analyzed against HCV 3a core gene by transiently transfecting the liver cells with HCV core plasmid. Our results demonstrated that Silymarin (SM) dose dependently inhibit the expression or function of HCV core gene at a non toxic concentration while the GAPDH remained constant. To identify the active ingredient, SM was fractioned by thin layer chromatography (TLC), column chromatography and HPLC. Purified fractions were tested for HCV core gene and western blotting results showed that two factions of SM (S1 and S2) inhibit HCV 3a core expression or function in liver cells Conclusion Our results suggest SM and its fractions (S1 and S2) inhibit HCV core gene of 3a genotype and combination of SM and its fractions with interferon will be a better option to treat HCV infection PMID:21453551

2011-01-01

59

Subinhibitory Clindamycin Differentially Inhibits Transcription of Exoprotein Genes in Staphylococcus aureus  

Microsoft Academic Search

It has long been known that certain antibiotics, at subinhibitory concentrations, differentially inhibit the synthesis of a-hemolysin and other staphylococcal virulence factors. In this report, we show that subinhibitory clindamycin (SBCL) eliminates production of nearly all exoproteins by Staphylococcus aureus but has virtually no effect on cytoplasmic proteins. The effect was abolished by a gene conferring resistance to macrolides-linco- samides-streptogramin

SILVIA HERBERT; PETER BARRY; RICHARD P. NOVICK

2001-01-01

60

Ingestion of bacterially expressed double-stranded RNA inhibits gene expression in planarians  

Microsoft Academic Search

population that is present in the adult planarian. The study of these organisms, classic experimental models for investigating metazoan regeneration, has been revitalized by the application of modern molecular biological approaches. The identification of thousands of unique planarian ESTs, coupled with large-scale whole-mount in situ hybridization screens, and the ability to inhibit planarian gene expression through double-stranded RNA-mediated genetic inter-

Phillip A. Newmark; Peter W. Reddien; Francesc Cebria; Alejandro Sanchez Alvarado

2003-01-01

61

HEPATOCYTE GROWTH FACTOR INHIBITS VEGF-FORKHEAD-DEPENDENT GENE EXPRESSION IN ENDOTHELIAL CELLS  

PubMed Central

Objective Recently, we reported that the forkhead transcription factor, FKHR/FOXO1, is required for vascular endothelial growth factor (VEGF)-mediated upregulation of a number of genes in endothelial cells. Here, we tested the hypothesis that hepatocyte growth factor (HGF), a potent activator of PI3K-Akt in endothelial cells, is capable of depleting the nucleus of FKHR/FOXO1 and thus inhibiting VEGF induction of this class of genes. Methods and Results Incubation of human coronary artery endothelial cells with HGF induced prolonged PI3K/Akt-dependent phosphorylation and nuclear exclusion of FKHR/FOXO1. HGF-mediated inhibition of FKHR/FOXO1 activity resulted in secondary attenuation of VEGF-induced expression of FKHR/FOXO1-dependent genes including vascular cell adhesion molecule-1, manganese superoxide dismutase, endothelial specific molecule-1, CBP/p300 interacting transactivator with ED-rich tail-2, bone morphogenetic protein-2, matrix metalloproteinase-10 and MGC5618. At a functional level, pre-incubation of HGF resulted in inhibition of VEGF-induced VCAM-1-mediated monocyte adhesion to endothelial cells. HGF-mediated inhibition of VEGF-inducible VCAM-1 expression and monocyte adhesion was reversed by overexpression of constitutively active phosphorylation-resistant triple mutant (TM)-FKHR. Conclusion These findings suggest that physiological agonists of PI3K-Akt signaling pathway may modulate VEGF-FKHR/FOXO1-dependent gene expression in endothelial cells. The data underscore the importance of the “set point” of the endothelial cell when considering mechanisms of signal transduction. PMID:18787186

Abid, Md. Ruhul; Nadeau, Robert J.; Spokes, Katherine C.; Minami, Takashi; Li, Dan; Shih, Shou-Ching; Aird, William C.

2009-01-01

62

Ibuprofen-induced inhibition of cyclooxygenase isoform gene expression and regression of rat mammary carcinomas  

Microsoft Academic Search

A single dose of 75 mg\\/kg 7,12 dimethylbenz[a]anthracene was administered to 50-day-old virgin female Sprague–Dawley rats and 100 days later, animals were randomized and provided with Teklad rodent chow mixed with a dose of 25 mg\\/rat\\/day ibuprofen for 35 days. Ibuprofen treatment reduced tumor volume (P<0.05) and significantly inhibited gene expression of both cyclooxygenase-1 and cyclooxygenase-2 (P<0.02). These results indicate

Fredika M Robertson; Michelle L Parrett; Farahnaz S Joarder; Mary Ross; Hussein M Abou-Issa; Galal Alshafie; Randall E Harris

1998-01-01

63

Relief of amplification inhibition in PCR with bovine serum albumin or T4 gene 32 protein  

Microsoft Academic Search

The benefits of adding bovine serum albumin (BSA) or T4 gene 32 proteins (gp32) to PCR were evaluated with reaction mixtures containing substances that inhibit amplification. Whereas 10- to 1,000-fold more FeClâ, hemin, fulvic acids, humic acids, tannic acids, or extracts from feces, freshwater, or marine water were accommodated in PCR when either 400 ng of BSA per μl was

CAROL A. KREADER

1996-01-01

64

Inhibition of virus replication and induction of human tetherin gene expression by equine IFN-?1.  

PubMed

Type I interferons (IFNs) play important roles in the defense of host cells against viral infection by inducing the expression of a diverse range of antiviral factors. IFNs from different animals likely share similar features with human IFNs, and some of them have cross-species activities. Equine IFN-? was proved effective in both equine and human cells. However, the previous studies mostly focused on the inhibition of virus induced cytopathic effects. In this study, we used virus-specific assays to demonstrate the antiviral activities of equine IFN-?1 in both equine and human cells. Equine IFN-?1 inhibited the expression of viral structural proteins and the production of virions of equine infectious anemia virus (EIAV) and equine arteritis virus (EAV) in equine cells. In addition, equine IFN-?1 inhibited the production of EIAV virus-like particles (VLP) from human 293T cells. An IFN-inducible human gene, tetherin, was induced in 293T cells by equine IFN-?1. Its induction correlated with the inhibition of VLP release from the cell membrane. This result indicates that equine IFN-?1 shares a similar mechanism of action with human IFN-? in regulating antiviral genes expression in human cells. PMID:24144682

Hu, Zhe; Wu, Xingliang; Ge, Jinying; Wang, Xiaojun

2013-11-15

65

Multiple pigmentation gene polymorphisms account for a substantial proportion of risk of cutaneous malignant melanoma  

PubMed Central

We have previously described the role of red hair (Melanocortin 1 Receptor, MC1R) and blue eye (Oculocutaneous Albinism Type 2, OCA2) gene polymorphisms in modulating risk of cutaneous malignant melanoma (CMM) in a highly sun-exposed population of European descent. A number of recent studies, including genome-wide association studies (GWAS), have identified numerous polymorphisms controlling human hair, eye and skin colour. In this paper, we test a selected set of polymorphisms in pigmentation loci (ASIP, TYR, TYRP1, MC1R, OCA2, IRF4, SLC24A4, SLC45A2) for association with CMM risk in a large Australian population-based case control study. Variants in IRF4 and SLC24A4, despite being strongly associated with pigmentation in our sample, did not modify CMM risk, but the other six did. Three SNPs (rs28777, rs35391, rs16891982) in the MATP gene (SLC45A2) exhibited the strongest crude association with risk, but this was attenuated to approximately the same effect size as that of a MC1R red hair color allele by controlling for ancestry of cases and controls. We also detected significant epistatic interactions between SLC45A2 and OCA2 alleles, and MC1R and ASIP alleles. Overall, these measured variants account for 12% of the familial risk of CMM in our population. PMID:19710684

Duffy, David L.; Zhao, Z. Z.; Sturm, Richard A.; Hayward, Nicholas K.; Martin, Nicholas G.; Montgomery, Grant W.

2013-01-01

66

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

PubMed

We have previously developed an in vitro model of rat brainstem explants. The latter release sizable amounts of calcitonin gene-related peptide (CGRP); basal release can be stimulated by such secretagogues as high KCl concentrations, veratridine or capsaicine. In this paradigm we investigated the activity of the analgesic agent tapentadol; the effects of tapentadol were compared to those of a classical opioid receptor agonist, morphine, and the selective noradrenaline reuptake inhibitor reboxetine. Morphine inhibited basal CGRP release, with statistical significance from 1 nM onward and maximal (-44%) inhibition at 100 ?M. Morphine also inhibited K(+)-stimulated peptide release, with a significant effect from 1 ?M and maximal (-39%) decrease at 100 ?M, but failed to inhibit release stimulated by 10 ?M capsaicin. At variance, reboxetine had no effect on baseline CGRP outflow, but was able to inhibit both K(+)-stimulated [significant inhibition from 1 ?M onward and maximal (-37%) decrease at 100 ?M], and capsaicin-stimulated release [significant effect from 1 ?M and maximal (-31%) decrease at 100 ?M]. Likewise, tapentadol had no effect on baseline CGRP release up to 100 ?M, but decreased secretion stimulated by 56 mM KCl or capsaicin, with significant effects from 0.1 and 1 ?M respectively; maximal inhibition over 56 mM KCl and capsaicin stimuli was -29% and -31%, respectively. Naloxone antagonized the effect of morphine, but not those of reboxetine and tapentadol, on K(+)-stimulated CGRP secretion. In conclusion the present study provides consistent pharmacological evidence that tapentadol acts as a noradrenaline reuptake inhibitor agent in this experimental model. PMID:24662320

Greco, Maria Cristina; Lisi, Lucia; Currò, Diego; Navarra, Pierluigi; Tringali, Giuseppe

2014-06-01

67

Gene expression profile of Xenopus A6 cells cultured under random positioning machine shows downregulation of ion transporter genes and inhibition of dome formation  

NASA Astrophysics Data System (ADS)

Random positioning machine (RPM) devices that generate a simulated microgravity environment of approximately 0 g prevent the formation of dome structures in Xenopus kidney-derived A6 cells. In the present study, the gene expression profile of A6 cells cultured under RPM was determined using the Xenopus 22K scale microarray, and those genes up- or downregulated twofold or more were investigated. We identified 29 genes (up, 25 genes; down, 4 genes) on day 5, 68 genes (up, 25 genes; down, 43 genes) on day 8, 111 genes (up, 69 genes; down, 42 genes) on day 10, and 283 genes (up, 153 genes; down, 130 genes) on day 15 of culture under RPM. These genes were classified according to categories described in the KOG database, such as "extracellular structure", "cytoskeleton", and "transcription". Almost all the genes involved in "inorganic ion transport and metabolism" were downregulated under RPM. Our study further investigated some of these including the epithelial Na + channel (ENaC) and Na +/K +-ATPase transporter genes. A specific inhibitor of Na +/K +-ATPases, ouabain, inhibited dome formation in the A6 cells, even under control culturing conditions of 1 g (the static condition). Together these data suggested that downregulation of sodium ion transporter gene expression plays a significant role in the RPM-dependent prevention of the dome formation in kidney epithelial cells.

Ikuzawa, Masayuki; Akiduki, Saori; Asashima, Makoto

68

BMP7 Gene Transfer via Gold Nanoparticles into Stroma Inhibits Corneal Fibrosis In Vivo  

PubMed Central

This study examined the effects of BMP7 gene transfer on corneal wound healing and fibrosis inhibition in vivo using a rabbit model. Corneal haze in rabbits was produced with the excimer laser performing -9 diopters photorefractive keratectomy. BMP7 gene was introduced into rabbit keratocytes by polyethylimine-conjugated gold nanoparticles (PEI2-GNPs) transfection solution single 5-minute topical application on the eye. Corneal haze and ocular health in live animals was gauged with stereo- and slit-lamp biomicroscopy. The levels of fibrosis [?-smooth muscle actin (?SMA), F-actin and fibronectin], immune reaction (CD11b and F4/80), keratocyte apoptosis (TUNEL), calcification (alizarin red, vonKossa and osteocalcin), and delivered-BMP7 gene expression in corneal tissues were quantified with immunofluorescence, western blotting and/or real-time PCR. Human corneal fibroblasts (HCF) and in vitro experiments were used to characterize the molecular mechanism mediating BMP7’s anti-fibrosis effects. PEI2-GNPs showed substantial BMP7 gene delivery into rabbit keratocytes in vivo (2×104 gene copies/ug DNA). Localized BMP7 gene therapy showed a significant corneal haze decrease (1.68±0.31 compared to 3.2±0.43 in control corneas; p<0.05) in Fantes grading scale. Immunostaining and immunoblot analyses detected significantly reduced levels of ?SMA (46±5% p<0.001) and fibronectin proteins (48±5% p<0.01). TUNEL, CD11b, and F4/80 assays revealed that BMP7 gene therapy is nonimmunogenic and nontoxic for the cornea. Furthermore, alizarin red, vonKossa and osteocalcin analyses revealed that localized PEI2-GNP-mediated BMP7 gene transfer in rabbit cornea does not cause calcification or osteoblast recruitment. Immunofluorescence of BMP7-transefected HCFs showed significantly increased pSmad-1/5/8 nuclear localization (>88%; p<0.0001), and immunoblotting of BMP7-transefected HCFs grown in the presence of TGF? demonstrated significantly enhanced pSmad-1/5/8 (95%; p<0.001) and Smad6 (53%, p<0.001), and decreased ?SMA (78%; p<0.001) protein levels. These results suggest that localized BMP7 gene delivery in rabbit cornea modulates wound healing and inhibits fibrosis in vivo by counter balancing TGF?1-mediated profibrotic Smad signaling. PMID:23799103

Tandon, Ashish; Sharma, Ajay; Rodier, Jason T.; Klibanov, Alexander M.; Rieger, Frank G.; Mohan, Rajiv R.

2013-01-01

69

Antisense Oligodeoxynucleotide Inhibition as an Alternative and Convenient Method for Gene Function Analysis in Pollen Tubes  

PubMed Central

Antisense oligodeoxynucleotide (A-ODN) inhibition works well in animal cells. However, there have been few successful examples to date of its application in plants, and more specifically whether the technique can be used in pollen tubes as a model of plant cell growth. NtGNL1 plays an important role in pollen tube development and was thus selected as an indicator to assess the biological effects of A-ODN. An A-ODN inhibition technique was used to down-regulate NtGNL1 expression in tobacco pollen tubes and showed that A-ODNs could quickly enter pollen tubes through the thick wall and cell membrane and effectively block NtGNL1 expression. Phenotype analysis revealed that the down-regulation of NtGNL1 by A-ODNs resulted in abnormalities in endocytosis and subsequent vesicle trafficking, similar to the phenotypes of pollen tubes treated with NtGNL1 RNAi. This investigation confirmed that A-ODNs could specifically inhibit target gene expression, and furthermore demonstrated that A-ODN functioned in a concentration- and duration-dependent manner, because A-ODNs could be degraded when incubated with pollen tubes. Thus, the A-ODN technique was successfully used for gene function analysis in pollen tubes and appears to be an alternative and convenient technique when the in vitro pollen tube is used as the study model. This technique will greatly facilitate investigations on the molecular mechanism(s) underlying pollen tube growth. PMID:23527102

Liao, Fanglei; Wang, Lu; Yang, Li-Bo; Zhang, Liyao; Peng, Xiongbo; Sun, Meng-xiang

2013-01-01

70

eNOS Gene Transfer Inhibits Smooth Muscle Cell Migration and MMP-2 and MMP-9 Activity  

Microsoft Academic Search

Vascular smooth muscle cell (SMC) migration is a critical step in the development of neointima after angioplasty. Matrix metalloproteinases (MMPs) degrade the basement membrane and the extracellular matrix, facilitating SMC migration. Transfer of the endothelial nitric oxide synthase (eNOS) gene to the injury site inhibits neointima formation. Neither the signaling pathways leading to NO-mediated inhibition of SMC migration and proliferation

Milind V. Gurjar; Ram V. Sharma; Ramesh C. Bhalla

2010-01-01

71

Ebola virus inhibits induction of genes by double-stranded RNA in endothelial cells.  

PubMed

Fatal cases of filoviral infection are accompanied by a marked immunosuppression. Endothelial cells play a vital role in the host immune response through the expression of several immunomodulatory genes in addition to the expression of the antiviral genes, 2',5'-oligoadenylate synthetase [2'-5'(A)N], and the double-stranded RNA (dsRNA)-activated protein kinase (PKR). dsRNA, an intermediate generated during viral replication and gene transcription of many viruses, leads to the induction of immunomodulatory genes in endothelial cells. In this report, we show that induction of the major histocompatibility complex class I family of genes, 2'-5'(A)N, interleukin-6 (IL-6), PKR, interferon (IFN)-regulatory factor-1, and intercellular adhesion molecule-1 (ICAM-1) by dsRNA in human umbilical vein endothelial cells is suppressed by infection with the filovirus Ebola-Zaire (EZ). In contrast, induction of IL-6 and ICAM-1 by IL-1 is intact in EZ-infected cells. Gel shift analysis demonstrates that dsRNA-induced protein binding to IFN-responsive elements is strongly suppressed by EZ-IFN, whereas NF-kappa B activation by dsRNA remains intact. We previously reported that IFN signaling is suppressed by EZ infection, and these data strongly suggest that elements shared between IFN and dsRNA signaling are being inhibited by EZ. Inhibition of IFN and dsRNA responsiveness could play a role in the immunosuppression seen in EZ infections and would play a role in the pathogenesis of disease caused by EZ. PMID:9875327

Harcourt, B H; Sanchez, A; Offermann, M K

1998-12-01

72

Human Cytomegalovirus Inhibition by Cardiac Glycosides: Evidence for Involvement of the hERG Gene  

PubMed Central

Infection with human cytomegalovirus (HCMV) continues to be a major threat for pregnant women and the immunocompromised population. Although several anti-HCMV therapies are available, the development of new anti-HCMV agents is highly desired. There is growing interest in identifying compounds that might inhibit HCMV by modulating the cellular milieu. Interest in cardiac glycosides (CG), used in patients with congestive heart failure, has increased because of their established anticancer and their suggested antiviral activities. We report that the several CG—digoxin, digitoxin, and ouabain—are potent inhibitors of HCMV at nM concentrations. HCMV inhibition occurred prior to DNA replication, but following binding to its cellular receptors. The levels of immediate early, early, and late viral proteins and cellular NF-?B were significantly reduced in CG-treated cells. The activity of CG in infected cells correlated with the expression of the potassium channel gene, hERG. CMV infection upregulated hERG, whereas CG significantly downregulated its expression. Infection with mouse CMV upregulated mouse ERG (mERG), but treatment with CG did not inhibit virus replication or mERG transcription. These findings suggest that CG may inhibit HCMV by modulating human cellular targets associated with hERG and that these compounds should be studied for their antiviral activities. PMID:22777050

Kapoor, Arun; Cai, Hongyi; Forman, Michael; He, Ran; Shamay, Meir

2012-01-01

73

Relief of amplification inhibition in PCR with bovine serum albumin or T4 gene 32 protein  

SciTech Connect

The benefits of adding bovine serum albumin (BSA) or T4 gene 32 proteins (gp32) to PCR were evaluated with reaction mixtures containing substances that inhibit amplification. Whereas 10- to 1,000-fold more FeCl{sub 3}, hemin, fulvic acids, humic acids, tannic acids, or extracts from feces, freshwater, or marine water were accommodated in PCR when either 400 ng of BSA per {mu}l was included in the reactions, neither BSA nor gp32 relieved interference significantly when minimum inhibitory levels of bile salts, bilirubin, EDTA, NaCl, sodium dodecyl sulfate, or Triton X-100 were present. Use of BSA and gp32 together offered no more relief of inhibition than either alone at its optimal level, and neither protein had any noticeable effect on amplification in the absence of inhibitors. 21 refs., 3 figs.

Kreader, C.A. [Environmental Protection Agency, Cincinnati, OH (United States)

1996-03-01

74

Autophagy inhibits viral genome replication and gene expression stages in West Nile virus infection.  

PubMed

Autophagy is a lysosomal degradation pathway that is implicated in many viral infections. However, its role in West Nile virus (WNV) infection remains controversial. In the present study, we examined the relationship between WNV infection and autophagy in infected cells. We demonstrated that LC3-II expression, a molecular marker for autophagosomal membranes, was enhanced in WNV-infected cells 6h post-infection. LC3-II expression was further enhanced in WNV-inoculated cells when treated with a lysosomal protease inhibitor. Meanwhile, WNV replication in cells lacking Atg5, an essential factor for autophagy, was increased compared with replication in wild-type cells. In addition, WNV replication was inhibited in cells lacking Atg5 when they were transfected with an ATG5 expression plasmid. These results suggest an antiviral role for autophagy in WNV-infected cells. We also examined which viral replication stages were affected by autophagy by using a Tat-beclin 1 peptide to induce autophagy and pseudo-infectious WNV reporter virus particles (WNV-RVPs) that monitor viral genome replication and gene expression stages via GFP expression. We found that autophagy induction in HeLa cells by Tat-beclin 1 peptide 3h after WNV inoculation inhibited viral replication, and GFP expression was significantly inhibited in wild-type cells when compared with cells lacking Atg5. Taken together, these results suggest that autophagy is induced by WNV infection, and that this induction inhibits WNV replication at the viral genome replication and gene expression stages. PMID:25091564

Kobayashi, Shintaro; Orba, Yasuko; Yamaguchi, Hiroki; Takahashi, Kenta; Sasaki, Michihito; Hasebe, Rie; Kimura, Takashi; Sawa, Hirofumi

2014-10-13

75

In vitro expression of Escherichia coli ribosomal protein genes: autogenous inhibition of translation.  

PubMed Central

Escherichia coli ribosomal protein L1 (0.5 micro M) was found to inhibit the synthesis of both proteins of the L11 operon, L11 and L1, but not the synthesis of other proteins directed by lambda rifd 18 DNA. Similarly, S4 (1 micro M) selectively inhibited the synthesis of three proteins of the alpha operon, S13, S11, and S4, directed by lambda spcI DNA or a restriction enzyme fragment obtained from this DNA. S8 (3.6 micro M) also showed preferential inhibitory effects on the synthesis of some proteins encoded in the spc operon, L24 and L5 (and probably S14 and S8), directed by lambda spcl DNA or a restriction enzyme fragment carrying the genes for these proteins. The inhibitory effect of L1 was observed only with L1 and not with other proteins examined, including S4 and S8. Similarly, the effect of S4 was not observed with L1 or S8, and that of S8 was not seen with L1 or S4. Inhibition was shown to take place at the level of translation rather than transcription. Thus, at least some ribosomal proteins (L1 S4, and S8) have the ability to cause selective translational inhibition of the synthesis of certain ribosomal proteins whose genes are in the same operon as their own. These results support the hypothesis that certain free ribosomal proteins not assembled into ribosomes act as "autogenous" feedback inhibitors to regulate the synthesis of ribosomal proteins. Images PMID:6445562

Yates, J L; Arfsten, A E; Nomura, M

1980-01-01

76

Inhibition of flower pigmentation by antisense CHS genes: promoter and minimal sequence requirements for the antisense effect  

Microsoft Academic Search

Introduction of a constitutive antisense full-length chalcone synthase (CHS) cDNA gene in petunia can result in an inhibition of flower pigmentation. We have evaluated some of the factors which may be important for the effectiveness of an antisense CHS gene.

Alexander R. van der Krol; Leon A. Mur; Pieter de Lange; Joseph N. M. Mol; Antoine R. Stuitje

1990-01-01

77

The inhibiting effect of 1·4 recombinant P chromosome of wheat- Agropyron cristatum addition line on the Ph gene  

Microsoft Academic Search

P chromosomes may carry a genetic system that inhibits the Ph gene in wheat. Abnormal chromosome synapsis in wheat-Agropyron cristatum addition line II-21-2 (additional 1·4 recombinant P chromosome) was observed in this study. The results of cytogenetics and\\u000a Ph1 gene amplification showed that the Ph1 gene was normal and the average number of quadrivalents or hexavalents was determined to be

GuoHui Yang; XinMing Yang; RuiHui Wang; AiNong Gao; LiHui Li; WeiHua Liu

2010-01-01

78

The roles of the bacteriophage T4 r genes in lysis inhibition and fine-structure genetics: a new perspective.  

PubMed Central

Seldom has the study of a set of genes contributed more to our understanding of molecular genetics than has the characterization of the rapid-lysis genes of bacteriophage T4. For example, T4 rII mutants were used to define gene structure and mutagen effects at the molecular level and to help unravel the genetic code. The large-plaque morphology of these mutants reflects a block in expressing lysis inhibition (LIN), the ability to delay lysis for several hours in response to sensing external related phages attacking the cell, which is a unique and highly adaptive attribute of the T4 family of phages. However, surprisingly little is known about the mechanism of LIN, or how the various r genes affect its expression. Here, we review the extensive old literature about the r genes and the lysis process and try to sort out the major players affecting lysis inhibition. We confirm that superinfection can induce lysis inhibition even while infected cells are lysing, suggesting that the signal response is virtually instantaneous and thus probably the result of post-translational regulation. We identify the rI gene as ORF tk.-2, based on sequence analysis of canonical rI mutants. The rI gene encodes a peptide of 97 amino acids (Mr = 11.1 kD; pI = 4.8) that probably is secreted into the periplasmic space. This gene is widely conserved among T-even phage. We then present a model for LIN, postulating that rI is largely responsible for regulating the gpt holin protein in response to superinfection. The evidence suggests that the rIIA and B genes are not directly involved in lysis inhibition; rather, when they are absent, an alternate pathway for lysis develops which depends on the presence of genes from any of several possible prophages and is not sensitive to lysis inhibition. PMID:9560373

Paddison, P; Abedon, S T; Dressman, H K; Gailbreath, K; Tracy, J; Mosser, E; Neitzel, J; Guttman, B; Kutter, E

1998-01-01

79

Genome-Wide RNAi Screening Identifies Genes Inhibiting the Migration of Glioblastoma Cells  

PubMed Central

Glioblastoma Multiforme (GBM) cells are highly invasive, infiltrating into the surrounding normal brain tissue, making it impossible to completely eradicate GBM tumors by surgery or radiation. Increasing evidence also shows that these migratory cells are highly resistant to cytotoxic reagents, but decreasing their migratory capability can re-sensitize them to chemotherapy. These evidences suggest that the migratory cell population may serve as a better therapeutic target for more effective treatment of GBM. In order to understand the regulatory mechanism underlying the motile phenotype, we carried out a genome-wide RNAi screen for genes inhibiting the migration of GBM cells. The screening identified a total of twenty-five primary hits; seven of them were confirmed by secondary screening. Further study showed that three of the genes, FLNA, KHSRP and HCFC1, also functioned in vivo, and knocking them down caused multifocal tumor in a mouse model. Interestingly, two genes, KHSRP and HCFC1, were also found to be correlated with the clinical outcome of GBM patients. These two genes have not been previously associated with cell migration. PMID:23593504

Yang, Jian; Fan, Jing; Li, Ying; Li, Fuhai; Chen, Peikai; Fan, Yubo; Xia, Xiaofeng; Wong, Stephen T.

2013-01-01

80

The tuberous sclerosis-1 (TSC1) gene product hamartin suppresses cell growth and augments the expression of the TSC2 product tuberin by inhibiting its ubiquitination  

Microsoft Academic Search

We report here that overexpression of the tuberous sclerosis-1 (TSC1) gene product hamartin results in the inhibition of growth, as well as changes in cell morphology. Growth inhibition was associated with an increase in the endogenous level of the product of the tuberous sclerosis-2 (TSC2) gene, tuberin. As overexpression of tuberin inhibits cell growth, and hamartin is known to bind

Giovanna Benvenuto; Shaowei Li; Samantha J Brown; Richard Braverman; William C Vass; Jeremy P Cheadle; Dicky JJ Halley; Julian R Sampson; Ralf Wienecke; Jeffrey E DeClue

2000-01-01

81

AAV-Mediated Gene Targeting Is Significantly Enhanced by Transient Inhibition of Nonhomologous End Joining or the Proteasome In Vivo  

PubMed Central

Abstract Recombinant adeno-associated virus (rAAV) vectors have clear potential for use in gene targeting but low correction efficiencies remain the primary drawback. One approach to enhancing efficiency is a block of undesired repair pathways like nonhomologous end joining (NHEJ) to promote the use of homologous recombination. The natural product vanillin acts as a potent inhibitor of NHEJ by inhibiting DNA-dependent protein kinase (DNA-PK). Using a homology containing rAAV vector, we previously demonstrated in vivo gene repair frequencies of up to 0.1% in a model of liver disease hereditary tyrosinemia type I. To increase targeting frequencies, we administered vanillin in combination with rAAV. Gene targeting frequencies increased up to 10-fold over AAV alone, approaching 1%. Fah?/?Ku70?/? double knockout mice also had increased gene repair frequencies, genetically confirming the beneficial effects of blocking NHEJ. A second strategy, transient proteasomal inhibition, also increased gene-targeting frequencies but was not additive to NHEJ inhibition. This study establishes the benefit of transient NHEJ inhibition with vanillin, or proteasome blockage with bortezomib, for increasing hepatic gene targeting with rAAV. Functional metabolic correction of a clinically relevant disease model was demonstrated and provided evidence for the feasibility of gene targeting as a therapeutic strategy. PMID:22486314

Paulk, Nicole K.; Loza, Laura Marquez; Finegold, Milton J.

2012-01-01

82

Fibroblast growth factor 7 inhibits cholesterol 7?-hydroxylase gene expression in hepatocytes.  

PubMed

Cholesterol 7?-hydroxylase (CYP7A1) is the initial and rate-limiting enzyme for bile acid synthesis. Transcription of the CYP7A1 gene is regulated by bile acids, nuclear receptors and cytokines. Fibroblast growth factor 7 (FGF7) secreted from activated hepatic stellate cells (HSC) during chronic liver fibrosis regulates hepatocyte survival and liver regeneration. In the carbon tetrachloride (CCl(4))-induced fibrotic mouse liver, we demonstrated that the expression of CYP7A1 was largely decreased while the expression of FGF7 was significantly increased. We further demonstrated that FGF7 inhibited CYP7A1 gene expression in hepatocytes. Knockdown study by short interfering RNA, kinase inhibition and phosphorylation assays revealed that the suppression of CYP7A1 expression by FGF7 was mediated by FGFR2 and its downstream JNK signaling cascade. The FGF7 neutralizing antibody restored CYP7A1 expression in Hep3B cells treated with conditioned medium from HSC. In summary, the data suggest that FGF7 is a novel regulator of CYP7A1 expression in hepatocytes and may prevent hepatocytes from accumulating toxic bile acids during liver injury and fibrosis. PMID:22713451

Sun, Zhichao; Yu, Xuemei; Wu, Weibin; Jia, Dongwei; Chen, Yinle; Ji, Lingling; Liu, Xijun; Peng, Xiaomin; Li, Yintao; Yang, Lili; Ruan, Yuanyuan; Gu, Jianxin; Ren, Shifang; Zhang, Songwen

2012-07-13

83

Azidothymidine inhibits NF-?B and induces Epstein-Barr virus gene expression in Burkitt lymphoma  

PubMed Central

The antiviral compound azidothymidine (AZT), alone or in combination with other agents, induces apoptosis in early-passage, Epstein-Barr virus–positive Burkitt lymphoma (EBV+ BL) lines and has clinical activity in EBV+ BL. We report here a mechanism of AZT's antitumor activity. The nuclei of these cells contain activated nuclear factor-?B (NF-?B) subunits p50, c-Rel, RelB, and p52, but not p65. Treatment of primary EBV+ BL lines with AZT inhibited NF-?B within 1 to 2 hours. This was followed by up-regulation of EBV gene expression including viral thymidine kinase (vTK) and apoptosis. Subclones of EBV+ BL cells that demonstrated activated p65 were resistant to AZT. In EBV+ BLs, AZT but not ganciclovir (GCV) was highly phosphorylated to its monophosphate form (AZT-MP). Phosphorylation, as well as apoptosis, was markedly enhanced in the presence of hydroxyurea. AZT inhibits NF-?B and up-regulates EBV gene expression in primary EBV+ BLs. AZT with hydroxyurea may represent an inexpensive, targeted regimen for endemic BL. PMID:15790788

Kurokawa, Motoki; Ghosh, Subrata K.; Ramos, Juan Carlos; Mian, Abdul M.; Toomey, Ngoc L.; Cabral, Lisa; Whitby, Denise; Barber, Glen N.; Dittmer, Dirk P.; Harrington, William J.

2005-01-01

84

Borna disease virus P protein inhibits nitric oxide synthase gene expression in astrocytes  

SciTech Connect

Borna disease virus (BDV) is one of the potential infectious agents involved in the development of central nervous system (CNS) diseases. Neurons and astrocytes are the main targets of BDV infection, but little is known about the roles of BDV infection in the biological effects of astrocytes. Here we reported that BDV inhibits the activation of inducible nitric oxide synthase (iNOS) in murine astrocytes induced by bacterial LPS and PMA. To determine which protein of BDV is responsible for the regulation of iNOS expression, we co-transfected murine astrocytes with reporter plasmid iNOS-luciferase and plasmid expressing individual BDV proteins. Results from analyses of reporter activities revealed that only the phosphoprotein (P) of BDV had an inhibitory effect on the activation of iNOS. In addition, P protein inhibits nitric oxide production through regulating iNOS expression. We also reported that the nuclear factor kappa B (NF-{kappa}B) binding element, AP-1 recognition site, and interferon-stimulated response element (ISRE) on the iNOS promoter were involved in the repression of iNOS gene expression regulated by the P protein. Functional analysis indicated that sequences from amino acids 134 to 174 of the P protein are necessary for the regulation of iNOS. These data suggested that BDV may suppress signal transduction pathways, which resulted in the inhibition of iNOS activation in astrocytes.

Peng Guiqing [State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430072 (China); Zhang Fengmin [College of Basic Medical Science, Harbin Medical University, Harbin 150081 (China); Zhang Qi; Wu Kailang; Zhu Fan [State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430072 (China); Wu Jianguo [State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan 430072 (China)], E-mail: jwu@edu.edu.cn

2007-09-30

85

PAAn-1b and PAAn-E: two phosphorothioate antisense oligodeoxynucleotides inhibit human aromatase gene expression.  

PubMed

Estrogen-dependent diseases, especially breast cancers, are frequently treated with aromatase inhibitors. Another more recent strategy is the antisense technology. In this study, after predicting aromatase mRNA secondary structure, we describe the design, the efficiency, and the toxicity of two antisense phosphorothioate oligodeoxynucleotides (PAAn-1b and PAAn-E) directed toward aromatase mRNA. Indeed, 2 microM PAAn-1b and PAAn-E encapsulated with 54 microM polyethylenimine inhibit aromatase activity by 71 and 79%, respectively, in transfected 293 cells, with IC50 values of 0.2 and 0.6 microM. The mechanism of inhibition appears to be specific after using sense and scramble oligodeoxynucleotides as controls and largely decreases aromatase mRNA and protein amounts. Moreover, PAAn-1b and PAAn-E are not cytotoxic for 293 cells. This study finally provides a new strategy for aromatase inhibition. It offers new tools for studying aromatase gene expression and its role in cancer for instance, and this could be of help for the therapy of estrogen-dependent diseases. PMID:9875210

Auvray, P; Sourdaine, P; Séralini, G E

1998-12-01

86

Gene 5. 5 protein of bacteriophaze T7 inhibits the nucleoid protein H-NS of Escherichia coli  

SciTech Connect

Gene 5.5 of coliphage T7 is one of the most highly expressed genes during T7 infection. Gene 5.5 protein, purified from cells overexpressing the cloned gene, purifies with the nucleoid protein H-NS of Escherichia coli during three chromatographic steps. A fusion protein of gene 5.5 protein and maltose binding protein also purifies with H-NS. The fusion protein binds to the DNA-H-NS complex and abolishes H-NS-mediated inhibition of transcription by Escherichia coli and T7 RNA polymerases in vitro. Expression of gene 5.5 also relieves the repression of the Escherichia coli proU promoter by H-NS in vivo. The change of leucine to proline at residue 30 of gene 5.5 protein abolishes the interaction between gene 5.5 protein and H-NS. 30 refs., 4 figs., 1 tab.

Liu, Q.; Richardson, C.C. (Harvard Medical School, Boston, MA (United States))

1993-03-01

87

Black Raspberry Components Inhibit Proliferation, Induce Apoptosis and Modulate Gene Expression in Rat Esophageal Epithelial Cells  

PubMed Central

We have shown that a diet containing freeze-dried black raspberries (BRB) inhibits the development of chemically-induced cancer in the rat esophagus. To provide insights into possible mechanisms by which BRB inhibit esophageal carcinogenesis, we evaluated an ethanol (EtOH) extract of BRB, and two component anthocyanins (cyanidin-3-O-glucoside and cyanidin-3-O-rutinoside) in BRB, for their effects on growth, apoptosis and gene expression in rat esophageal epithelial cell lines. The EtOH extract and both anthocyanins selectively caused significant growth inhibition and induction of apoptosis in a highly tumorigenic cell line (RE-149 DHD) but not in a weakly tumorigenic line (RE-149). The uptake of anthocyanins from the EtOH extract into RE-149 DHD cells far exceeded their uptake into RE-149 cells, which may have accounted for the selective effects of the extract on growth and apoptosis of RE-149 DHD cells. The growth inhibitory and pro-apoptotic effects were enhanced by the daily addition of the EtOH extract and the anthocyanins to the medium. Interestingly, the EtOH extract did not alter cyclooxygenase-2 (COX-2) and nitric oxide synthase (i-NOS) expression in RE-149 DHD cells whereas, both anthocyanins down-regulated the expressions of these genes. This differential effect may have been related to the relative amounts of anthocyanins in the extract versus when they were added individually to the medium. We conclude that the selective effects of the EtOH extract on growth and apoptosis of highly tumorigenic rat esophageal epithelial cells in vitro may be due to preferential uptake and retention of its component anthocyanins, and this may also be responsible for the greater inhibitory effects of freeze-dried whole berries on tumor cells in vivo. PMID:20155622

Zikri, Nancy N.; Riedl, Kenneth M.; Wang, Li-Shu; Lechner, John F.; Schwartz, Steven J.; Stoner, Gary D.

2010-01-01

88

Fibroblast growth factor 7 inhibits cholesterol 7{alpha}-hydroxylase gene expression in hepatocytes  

SciTech Connect

Highlights: Black-Right-Pointing-Pointer FGF7 strongly and rapidly down-regulates the expression of CYP7A1 in hepatocytes. Black-Right-Pointing-Pointer FGF7 suppresses the expression of CYP7A1 via FGFR2 and downstream JNK activation. Black-Right-Pointing-Pointer Blocking FGF7 abrogates HSC-induced inhibition of CYP7A1 expression in hepatocytes. -- Abstract: Cholesterol 7{alpha}-hydroxylase (CYP7A1) is the initial and rate-limiting enzyme for bile acid synthesis. Transcription of the CYP7A1 gene is regulated by bile acids, nuclear receptors and cytokines. Fibroblast growth factor 7 (FGF7) secreted from activated hepatic stellate cells (HSC) during chronic liver fibrosis regulates hepatocyte survival and liver regeneration. In the carbon tetrachloride (CCl{sub 4})-induced fibrotic mouse liver, we demonstrated that the expression of CYP7A1 was largely decreased while the expression of FGF7 was significantly increased. We further demonstrated that FGF7 inhibited CYP7A1 gene expression in hepatocytes. Knockdown study by short interfering RNA, kinase inhibition and phosphorylation assays revealed that the suppression of CYP7A1 expression by FGF7 was mediated by FGFR2 and its downstream JNK signaling cascade. The FGF7 neutralizing antibody restored CYP7A1 expression in Hep3B cells treated with conditioned medium from HSC. In summary, the data suggest that FGF7 is a novel regulator of CYP7A1 expression in hepatocytes and may prevent hepatocytes from accumulating toxic bile acids during liver injury and fibrosis.

Sun, Zhichao [Department of Biochemistry and Molecular Biology, Shanghai Medical College, Fudan University, Shanghai (China)] [Department of Biochemistry and Molecular Biology, Shanghai Medical College, Fudan University, Shanghai (China); Yu, Xuemei [Department of Endocrinology, Fengxian Central Hospital, Shanghai (China)] [Department of Endocrinology, Fengxian Central Hospital, Shanghai (China); Wu, Weibin; Jia, Dongwei; Chen, Yinle; Ji, Lingling; Liu, Xijun; Peng, Xiaomin [Department of Biochemistry and Molecular Biology, Shanghai Medical College, Fudan University, Shanghai (China)] [Department of Biochemistry and Molecular Biology, Shanghai Medical College, Fudan University, Shanghai (China); Li, Yintao [Institute of Endocrinology and Diabetology, Huashan Hospital, Shanghai Medical College, Fudan University, Shanghai (China)] [Institute of Endocrinology and Diabetology, Huashan Hospital, Shanghai Medical College, Fudan University, Shanghai (China); Yang, Lili [Department of Endocrinology, Fengxian Central Hospital, Shanghai (China)] [Department of Endocrinology, Fengxian Central Hospital, Shanghai (China); Ruan, Yuanyuan; Gu, Jianxin [Department of Biochemistry and Molecular Biology, Shanghai Medical College, Fudan University, Shanghai (China)] [Department of Biochemistry and Molecular Biology, Shanghai Medical College, Fudan University, Shanghai (China); Ren, Shifang, E-mail: renshifang@fudan.edu.cn [Department of Biochemistry and Molecular Biology, Shanghai Medical College, Fudan University, Shanghai (China)] [Department of Biochemistry and Molecular Biology, Shanghai Medical College, Fudan University, Shanghai (China); Zhang, Songwen, E-mail: songwenzhang@fudan.edu.cn [Department of Biochemistry and Molecular Biology, Shanghai Medical College, Fudan University, Shanghai (China)] [Department of Biochemistry and Molecular Biology, Shanghai Medical College, Fudan University, Shanghai (China)

2012-07-13

89

Gene expression analysis of the mechanism of inhibition of Desulfovibrio vulgaris Hildenborough by nitrate-reducing, sulfide-oxidizing bacteria.  

PubMed

Sulfate-reducing bacteria (SRB) are inhibited by nitrate-reducing, sulfide-oxidizing bacteria (NR-SOB) in the presence of nitrate. This inhibition has been attributed either to an increase in redox potential or to production of nitrite by the NR-SOB. Nitrite specifically inhibits the final step in the sulfate reduction pathway. When the NR-SOB Thiomicrospira sp. strain CVO was added to mid-log phase cultures of the SRB Desulfovibrio vulgaris Hildenborough in the presence of nitrate, sulfate reduction was inhibited. Strain CVO reduced nitrate and oxidized sulfide, with transient production of nitrite. Sulfate reduction by D. vulgaris resumed once nitrite was depleted. A DNA macroarray with open reading frames encoding enzymes involved in energy metabolism of D. vulgaris was used to study the effects of NR-SOB on gene expression. Shortly following addition of strain CVO, D. vulgaris genes for cytochrome c nitrite reductase and hybrid cluster proteins Hcp1 and Hcp2 were upregulated. Genes for sulfate reduction enzymes, except those for dissimilatory sulfite reductase, were downregulated. Genes for the membrane-bound electron transferring complexes QmoABC and DsrMKJOP were downregulated and unaffected, respectively, whereas direct addition of nitrite downregulated both operons. Overall the gene expression response of D. vulgaris upon exposure to strain CVO and nitrate resembled that observed upon direct addition of nitrite, indicating that inhibition of SRB is primarily due to nitrite production by NR-SOB. PMID:16104868

Haveman, Shelley A; Greene, E Anne; Voordouw, Gerrit

2005-09-01

90

Neuropeptide S receptor (NPSR1) gene variation modulates response inhibition and error monitoring.  

PubMed

The neuropeptide S (NPS) system has been suggested to contribute to the pathogenesis of anxiety. In order to further characterize the cognitive-neurophysiological relevance of neuropeptide S in the etiology of anxiety, the influence of a functional neuropeptide S receptor gene (NPSR1) variant on response inhibition and error monitoring was investigated under consideration of the dimensional phenotype of anxiety sensitivity (AS). In a sample of N=97 healthy probands, event-related potential (ERP) measurement using a modified Flanker task was applied allowing for a distinct neurophysiological examination of processes related to response inhibition (Nogo-N2, Nogo-P3) and error monitoring (Ne/ERN). All subjects were genotyped for the functional NPSR1 A/T (Asn(107)Ile) variant (rs324981) and characterized for anxiety sensitivity using the Anxiety Sensitivity Index (ASI). Carriers of the NPSR1 T allele displayed intensified response inhibition (Nogo-P3) and error monitoring (Ne/ERN), which was in both cases paralleled by the behavioral data. Furthermore, anxiety sensitivity was found to be higher in NPSR1 T allele carriers and to correlate with Nogo-P3 and Ne/ERN. A mediation analysis revealed the ERN to mediate the effect between NPSR1 genotype and anxiety sensitivity. In summary, the more active NPSR1 T allele may confer enhanced response inhibition and increased error monitoring and might drive particularly error monitoring as a neurophysiological endophenotype of anxiety as reflected by increased anxiety sensitivity. These findings further corroborate a major role of the neuropeptide S system in the pathogenesis of anxiety and suggest a potentially beneficial use of therapeutic agents targeting the NPS system in anxiety disorders. PMID:23319044

Beste, Christian; Konrad, Carsten; Uhlmann, Christina; Arolt, Volker; Zwanzger, Peter; Domschke, Katharina

2013-05-01

91

Interferon selectively inhibits the expression of mitochondrial genes: a novel pathway for interferon-mediated responses.  

PubMed Central

As an approach to identifying genes involved in physiological actions of interferons we used differential probes to screen a cDNA library from mouse L-929 cells treated with interferon alpha/beta. We identified two negatively regulated mRNA species which have been examined by analysis of the corresponding mRNAs and by DNA sequencing. Comparison with the GenBank database showed that these cDNA clones corresponded to mitochondrially encoded genes for cytochrome b and subunit I of cytochrome c oxidase. A further cDNA encompassing three mitochondrial genes was used as a probe to show that a third mRNA, NADH dehydrogenase subunit 5, was also down-regulated by interferon while a fourth, NADH dehydrogenase subunit 6, was unaffected. Expression of cytochrome b was also inhibited in mouse NIH 3T3 cells treated with interferon alpha/beta and in human Daudi lymphoblastoid cells treated with interferon alpha. The ability of interferon to reduce mitochondrial mRNA levels could be blocked by cycloheximide suggesting that these effects are mediated by an interferon-responsive nuclear gene which encodes a product capable of regulating mitochondrial gene expression. Analysis of proteins synthesized in the presence of emetine, a specific inhibitor of cytoplasmic translation, showed that the synthesis of several mitochondrial translation products, including cytochrome b, was reduced after treatment with interferon. Our results reveal a novel effect of interferon on cellular physiology which could have important consequences for understanding the effects of interferons as well as suggesting new mechanisms for the regulation of mitochondrial biogenesis and function. Images Fig. 1. Fig. 2. Fig. 3. Fig. 5. Fig. 6. Fig. 7. PMID:2176148

Shan, B; Vazquez, E; Lewis, J A

1990-01-01

92

Inhibition of Interleukin-2 Gene Expression by Human Herpesvirus 6B U54 Tegument Protein.  

PubMed

Human herpesvirus 6B (HHV-6B) is a ubiquitous pathogen causing lifelong infections in approximately 95% of humans worldwide. To persist within its host, HHV-6B has developed several immune evasion mechanisms, such as latency, during which minimal proteins are expressed, and the ability to disturb innate and adaptive immune responses. The primary cellular targets of HHV-6B are CD4(+) T cells. Previous studies by Flamand et al. (L. Flamand, J. Gosselin, I. Stefanescu, D. Ablashi, and J. Menezes, Blood 85:1263-1271, 1995) reported on the capacity of HHV-6A as well as UV-irradiated HHV-6A to inhibit interleukin-2 (IL-2) synthesis in CD4(+) lymphocytes, suggesting that viral structural components could be responsible for this effect. In the present study, we identified the HHV-6B U54 tegument protein (U54) as being capable of inhibiting IL-2 expression. U54 binds the calcineurin (CaN) phosphatase enzyme, causing improper dephosphorylation and nuclear translocation of NFAT (nuclear factor of activated T cells) proteins, resulting in suboptimal IL-2 gene transcription. The U54 GISIT motif (amino acids 293 to 297), analogous to the NFAT PXIXIT motif, contributed to the inhibition of NFAT activation. IMPORTANCE Human herpesvirus 6A (HHV-6A) and HHV-6B are associated with an increasing number of pathologies. These viruses have developed strategies to avoid the immune response allowing them to persist in the host. Several studies have illustrated mechanisms by which HHV-6A and HHV-6B are able to disrupt host defenses (reviewed in L. Dagna, J. C. Pritchett, and P. Lusso, Future Virol. 8:273-287, 2013, doi:10.2217/fvl.13.7). Previous work informed us that HHV-6A is able to suppress synthesis of interleukin-2 (IL-2), a key immune growth factor essential for adequate T lymphocyte proliferation and expansion. We obtained evidence that HHV-6B also inhibits IL-2 gene expression and identified the mechanisms by which it does so. Our work led us to the identification of U54, a virion-associated tegument protein, as being responsible for suppression of IL-2. Consequently, we have identified HHV-6B U54 protein as playing a role in immune evasion. These results further contribute to our understanding of HHV-6 interactions with its human host and the efforts deployed to ensure its long-term persistence. PMID:25122797

Iampietro, Mathieu; Morissette, Guillaume; Gravel, Annie; Flamand, Louis

2014-11-01

93

psi , a plasmid-linked Rhizobium phaseoli gene that inhibits exopolysaccharide production and which is required for symbiotic nitrogen fixation  

Microsoft Academic Search

A strain of R. phaseoli cured of its symbiotic plasmid, pRP2JI, retained the ability to make exopolysaccharide (EPS). However, a region of pRP2JI, when cloned at an increased copy number in wide host-range vectors and transferred to this and other strains of Rhizobium, inhibited EPS synthesis. The gene responsible was termed psi (polysaccharide inhibition) and was located in a region

D. Borthakur; J. A. Downie; A. W. B. Johnston; J. W. Lamb

1985-01-01

94

Human interleukin-10 gene inhibits acute rejection by triggering apoptosis in allograft vascular transplantation  

PubMed Central

The aim this study is to explore effect of IL-10 on apoptosis of VSMCs in allograft arterial transplantation rats, and to investigate mechanism. SD rats were divied into three groups, including control group (CN, with physiological saline), blank vector group (BV, with blank adenovirus) and combined gene group (CG, with adenovirus carried IL-10 gene). The isolated donor vascular was transfected with the adenovirus carried hIL-10 gene for 30 minutes by immersing method. Forty-five days post transplantation, the grafts were harvested. The allografts pathologioc changes were observed and the size of vascular intima and middle layer of allografts were measured. The expression of hIL-10 was detected by RT-PCR, ELISA and immunohistochemistry, respectively. The repression of Fas/Fasl in artery allografts was also examined by immunohistochemistry method. The results indicated that 45 days after transplantation, the intimal and middle hyperplasia ratio in CG group was significantly lower than that in CN and BV group (P < 0.05). The transgene expression of human interleukin-10 was significantly enhanced in CG group compared to CN and BV group by ELISA, RT-PCR and immunohistochemistry (P < 0.05). The expression of Fas/FasL was higher in CG group compared with the other groups (P < 0.05). The level of apoptotic smooth muscle cells were significantly increased in CG group compared to CN and BV group (P < 0.05). In conclusion, adenovirus mediated IL-10 expression could up-regulate Fas/FasL expression, induce smooth muscle cell apoptosis and alleviate angiosclerosis process. The IL-10 gene transfer to allograft artery could inhibit acute rejection reaction of allograft vascular transplantation. PMID:25337228

Liu, Haibo; Yang, Shunzhang; Sun, Xuejun; Chen, Tianbao

2014-01-01

95

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

PubMed Central

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

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

2011-01-01

96

Dynamic Telomerase Gene Suppression via Network Effects of GSK3 Inhibition  

PubMed Central

Background Telomerase controls telomere homeostasis and cell immortality and is a promising anti-cancer target, but few small molecule telomerase inhibitors have been developed. Reactivated transcription of the catalytic subunit hTERT in cancer cells controls telomerase expression. Better understanding of upstream pathways is critical for effective anti-telomerase therapeutics and may reveal new targets to inhibit hTERT expression. Methodology/Principal Findings In a focused promoter screen, several GSK3 inhibitors suppressed hTERT reporter activity. GSK3 inhibition using 6-bromoindirubin-3?-oxime suppressed hTERT expression, telomerase activity and telomere length in several cancer cell lines and growth and hTERT expression in ovarian cancer xenografts. Microarray analysis, network modelling and oligonucleotide binding assays suggested that multiple transcription factors were affected. Extensive remodelling involving Sp1, STAT3, c-Myc, NF?B, and p53 occurred at the endogenous hTERT promoter. RNAi screening of the hTERT promoter revealed multiple kinase genes which affect the hTERT promoter, potentially acting through these factors. Prolonged inhibitor treatments caused dynamic expression both of hTERT and of c-Jun, p53, STAT3, AR and c-Myc. Conclusions/Significance Our results indicate that GSK3 activates hTERT expression in cancer cells and contributes to telomere length homeostasis. GSK3 inhibition is a clinical strategy for several chronic diseases. These results imply that it may also be useful in cancer therapy. However, the complex network effects we show here have implications for either setting. PMID:19649288

Bilsland, Alan E.; Hoare, Stacey; Stevenson, Katrina; Plumb, Jane; Gomez-Roman, Natividad; Cairney, Claire; Burns, Sharon; Lafferty-Whyte, Kyle; Roffey, Jon; Hammonds, Tim; Keith, W. Nicol

2009-01-01

97

Inhibition of Corticosteroid-Binding Globulin Gene Expression by Glucocorticoids Involves C/EBP?  

PubMed Central

Corticosteroid-binding globulin (CBG), a negative acute phase protein produced primarily in the liver, is responsible for the transport of glucocorticoids (GCs). It also modulates the bioavailability of GCs, as only free or unbound steroids are biologically active. Fluctuations in CBG levels therefore can directly affect GC bioavailability. This study investigates the molecular mechanism whereby GCs inhibit the expression of CBG. GCs regulate gene expression via the glucocorticoid receptor (GR), which either directly binds to DNA or acts indirectly via tethering to other DNA-bound transcription factors. Although no GC-response elements (GRE) are present in the Cbg promoter, putative binding sites for C/EBP?, able to tether to the GR, as well as HNF3? involved in GR signaling, are present. C/EBP?, but not HNF3?, was identified as an important mediator of DEX-mediated inhibition of Cbg promoter activity by using specific deletion and mutant promoter reporter constructs of Cbg. Furthermore, knockdown of C/EBP? protein expression reduced DEX-induced repression of CBG mRNA, confirming C/EBP?’s involvement in GC-mediated CBG repression. Chromatin immunoprecipitation (ChIP) after DEX treatment indicated increased co-recruitment of C/EBP? and GR to the Cbg promoter, while C/EBP? knockdown prevented GR recruitment. Together, the results suggest that DEX repression of CBG involves tethering of the GR to C/EBP?. PMID:25335188

Verhoog, Nicolette; Allie-Reid, Fatima; Vanden Berghe, Wim; Smith, Carine; Haegeman, Guy; Hapgood, Janet; Louw, Ann

2014-01-01

98

Inhibition of HIV-1 gene expression by Ciclopirox and Deferiprone, drugs that prevent hypusination of eukaryotic initiation factor 5A  

PubMed Central

Background Eukaryotic translation initiation factor eIF5A has been implicated in HIV-1 replication. This protein contains the apparently unique amino acid hypusine that is formed by the post-translational modification of a lysine residue catalyzed by deoxyhypusine synthase and deoxyhypusine hydroxylase (DOHH). DOHH activity is inhibited by two clinically used drugs, the topical fungicide ciclopirox and the systemic medicinal iron chelator deferiprone. Deferiprone has been reported to inhibit HIV-1 replication in tissue culture. Results Ciclopirox and deferiprone blocked HIV-1 replication in PBMCs. To examine the underlying mechanisms, we investigated the action of the drugs on eIF5A modification and HIV-1 gene expression in model systems. At early times after drug exposure, both drugs inhibited substrate binding to DOHH and prevented the formation of mature eIF5A. Viral gene expression from HIV-1 molecular clones was suppressed at the RNA level independently of all viral genes. The inhibition was specific for the viral promoter and occurred at the level of HIV-1 transcription initiation. Partial knockdown of eIF5A-1 by siRNA led to inhibition of HIV-1 gene expression that was non-additive with drug action. These data support the importance of eIF5A and hypusine formation in HIV-1 gene expression. Conclusion At clinically relevant concentrations, two widely used drugs blocked HIV-1 replication ex vivo. They specifically inhibited expression from the HIV-1 promoter at the level of transcription initiation. Both drugs interfered with the hydroxylation step in the hypusine modification of eIF5A. These results have profound implications for the potential therapeutic use of these drugs as antiretrovirals and for the development of optimized analogs. PMID:19825182

Hoque, Mainul; Hanauske-Abel, Hartmut M; Palumbo, Paul; Saxena, Deepti; D'Alliessi Gandolfi, Darlene; Park, Myung Hee; Pe'ery, Tsafi; Mathews, Michael B

2009-01-01

99

Changes in rice allelopathy and rhizosphere microflora by inhibiting rice phenylalanine ammonia-lyase gene expression.  

PubMed

Gene expression of phenylalanine ammonia-lyase (PAL) in allelopathic rice PI312777 was inhibited by RNA interference (RNAi). Transgenic rice showed lower levels of PAL gene expression and PAL activity than wild type rice (WT). The concentrations of phenolic compounds were lower in the root tissues and root exudates of transgenic rice than in those of wild type plants. When barndyardgrass (BYG) was used as the receiver plant, the allelopathic potential of transgenic rice was reduced. The sizes of the bacterial and fungal populations in rice rhizospheric soil at the 3-, 5-, and 7-leaf stages were estimated by using quantitative PCR (qPCR), which showed a decrease in both populations at all stages of leaf development analyzed. However, PI312777 had a larger microbial population than transgenic rice. In addition, in T-RFLP studies, 14 different groups of bacteria were detected in WT and only 6 were detected in transgenic rice. This indicates that there was less rhizospheric bacterial diversity associated with transgenic rice than with WT. These findings collectively suggest that PAL functions as a positive regulator of rice allelopathic potential. PMID:23385369

Fang, Changxun; Zhuang, Yuee; Xu, Tiecheng; Li, Yingzhe; Li, Yue; Lin, Wenxiong

2013-02-01

100

Inhibition of Virulence Gene Expression in Staphylococcus aureus by Novel Depsipeptides from a Marine Photobacterium  

PubMed Central

During a global research expedition, more than five hundred marine bacterial strains capable of inhibiting the growth of pathogenic bacteria were collected. The purpose of the present study was to determine if these marine bacteria are also a source of compounds that interfere with the agr quorum sensing system that controls virulence gene expression in Staphylococcus aureus. Using a gene reporter fusion bioassay, we recorded agr interference as enhanced expression of spa, encoding Protein A, concomitantly with reduced expression of hla, encoding ?-hemolysin, and rnaIII encoding RNAIII, the effector molecule of agr. A marine Photobacterium produced compounds interfering with agr in S. aureus strain 8325-4, and bioassay-guided fractionation of crude extracts led to the isolation of two novel cyclodepsipeptides, designated solonamide A and B. Northern blot analysis confirmed the agr interfering activity of pure solonamides in both S. aureus strain 8325-4 and the highly virulent, community-acquired strain USA300 (CA-MRSA). To our knowledge, this is the first report of inhibitors of the agr system by a marine bacterium. PMID:22363239

Mansson, Maria; Nielsen, Anita; Kjærulff, Louise; Gotfredsen, Charlotte H.; Wietz, Matthias; Ingmer, Hanne; Gram, Lone; Larsen, Thomas O.

2011-01-01

101

Inhibition of virulence gene expression in Staphylococcus aureus by novel depsipeptides from a marine photobacterium.  

PubMed

During a global research expedition, more than five hundred marine bacterial strains capable of inhibiting the growth of pathogenic bacteria were collected. The purpose of the present study was to determine if these marine bacteria are also a source of compounds that interfere with the agr quorum sensing system that controls virulence gene expression in Staphylococcus aureus. Using a gene reporter fusion bioassay, we recorded agr interference as enhanced expression of spa, encoding Protein A, concomitantly with reduced expression of hla, encoding ?-hemolysin, and rnaIII encoding RNAIII, the effector molecule of agr. A marine Photobacterium produced compounds interfering with agr in S. aureus strain 8325-4, and bioassay-guided fractionation of crude extracts led to the isolation of two novel cyclodepsipeptides, designated solonamide A and B. Northern blot analysis confirmed the agr interfering activity of pure solonamides in both S. aureus strain 8325-4 and the highly virulent, community-acquired strain USA300 (CA-MRSA). To our knowledge, this is the first report of inhibitors of the agr system by a marine bacterium. PMID:22363239

Mansson, Maria; Nielsen, Anita; Kjærulff, Louise; Gotfredsen, Charlotte H; Wietz, Matthias; Ingmer, Hanne; Gram, Lone; Larsen, Thomas O

2011-12-01

102

Neferine inhibits angiotensin II-induced rat aortic smooth muscle cell proliferation predominantly by downregulating fractalkine gene expression  

PubMed Central

Neferine inhibits the angiotensin II (AngII)-induced proliferation of vascular smooth muscle cells (SMCs), but the underlying mechanism is unclear. The aim of this study was to explore the mechanism underlying the effect of neferine on the proliferation of vascular SMCs. Rat aortic SMCs (RASMCs) were used and fractalkine (Fkn) gene expression was measured by quantitative polymerase chain reaction and western blot analysis. The proliferation of RASMCs was analyzed by MTT assay and flow cytometry. It was revealed that AngII induced Fkn expression in a dose- and time-dependent manner. Fkn-knockdown with small interfering RNA attenuated the AngII-induced RASMC proliferation. Furthermore, neferine inhibited Fkn expression and attenuated the AngII-induced RASMC proliferation. These findings suggest that the Fkn gene may play an important role in AngII-induced RASMC proliferation and that neferine acts to attenuate AngII-induced RASMC proliferation by inhibiting Fkn expression.

ZHENG, LULU; CAO, YONGWEN; LIU, SHAO; PENG, ZHENYU; ZHANG, SAIDAN

2014-01-01

103

Overexpression of ERBB2 in human mammary epithelial cells signals inhibition of transcription of the E-cadherin gene.  

PubMed Central

Overexpression of the ERBB2 receptor in transfectants of a human mammary epithelial cell line (MTSV1-7) is associated with a reduced ability to undergo morphogenesis in vitro and with a decreased level of expression of the E-cadherin and alpha 2 integrin genes. The inhibition of expression of the adhesion molecules has been shown to be at the level of transcription by using nuclear run-on assays and by following transcription of a reporter gene fused to 5' sequences of the E-cadherin gene. To relate the effects on gene transcription to a functional ERBB2 protein, signaling from the receptor was inhibited by the antibody 4D5, which blocks phosphorylation of ERBB2 on tyrosine residues and association of the protein with the GRB2/Sem5 protein. After treatment with the antibody 4D5, the ERBB2 transfectants regain the ability to form three-dimensional structures in collagen gels and the rates of transcription of the genes encoding the E-cadherin and the alpha 2 integrin subunit are restored to the levels seen in MTSV1-7neo cells. These results demonstrate that the inhibition of morphogenesis and transcription of specific adhesion molecules in human mammary epithelial cells can be affected by signals generated by the ERBB2 receptor and suggest a role for ERBB2 overexpression in tumor progression and metastasis. Images PMID:7913748

D'souza, B; Taylor-Papadimitriou, J

1994-01-01

104

Inhibition of estrogen-responsive gene activation by the retinoid X receptor beta: evidence for multiple inhibitory pathways.  

PubMed Central

The retinoid X receptor beta (RXR beta; H-2RIIBP) forms heterodimers with various nuclear hormone receptors and binds multiple hormone response elements, including the estrogen response element (ERE). In this report, we show that endogenous RXR beta contributes to ERE binding activity in nuclear extracts of the human breast cancer cell line MCF-7. To define a possible regulatory role of RXR beta regarding estrogen-responsive transcription in breast cancer cells, RXR beta and a reporter gene driven by the vitellogenin A2 ERE were transfected into estrogen-treated MCF-7 cells. RXR beta inhibited ERE-driven reporter activity in a dose-dependent and element-specific fashion. This inhibition occurred in the absence of the RXR ligand 9-cis retinoic acid. The RXR beta-induced inhibition was specific for estrogen receptor (ER)-mediated ERE activation because inhibition was observed in ER-negative MDA-MB-231 cells only following transfection of the estrogen-activated ER. No inhibition of the basal reporter activity was observed. The inhibition was not caused by simple competition of RXR beta with the ER for ERE binding, since deletion mutants retaining DNA binding activity but lacking the N-terminal or C-terminal domain failed to inhibit reporter activity. In addition, cross-linking studies indicated the presence of an auxiliary nuclear factor present in MCF-7 cells that contributed to RXR beta binding of the ERE. Studies using known heterodimerization partners of RXR beta confirmed that RXR beta/triiodothyronine receptor alpha heterodimers avidly bind the ERE but revealed the existence of another triiodothyronine-independent pathway of ERE inhibition. These results indicate that estrogen-responsive genes may be negatively regulated by RXR beta through two distinct pathways. Images PMID:8384307

Segars, J H; Marks, M S; Hirschfeld, S; Driggers, P H; Martinez, E; Grippo, J F; Brown, M; Wahli, W; Ozato, K

1993-01-01

105

Gene expression profiles in engineered cardiac tissues respond to mechanical loading and inhibition of tyrosine kinases  

PubMed Central

Engineered cardiac tissues (ECTs) are platforms to investigate cardiomyocyte maturation and functional integration, the feasibility of generating tissues for cardiac repair, and as models for pharmacology and toxicology bioassays. ECTs rapidly mature in vitro to acquire the features of functional cardiac muscle and respond to mechanical load with increased proliferation and maturation. ECTs are now being investigated as platforms for in vitro models for human diseases and for pharmacologic screening for drug toxicities. We tested the hypothesis that global ECT gene expression patterns are complex and sensitive to mechanical loading and tyrosine kinase inhibitors similar to the maturing myocardium. We generated ECTs from day 14.5 rat embryo ventricular cells, as previously published, and then conditioned constructs after 5 days in culture for 48 h with mechanical stretch (5%, 0.5 Hz) and/or the p38 MAPK (p38 mitogen-activated protein kinase) inhibitor BIRB796. RNA was isolated from individual ECTs and assayed using a standard Agilent rat 4 × 44k V3 microarray and Pathway Analysis software for transcript expression fold changes and changes in regulatory molecules and networks. Changes in expression were confirmed by quantitative-polymerase chain reaction (q-PCR) for selected regulatory molecules. At the threshold of a 1.5-fold change in expression, stretch altered 1559 transcripts, versus 1411 for BIRB796, and 1846 for stretch plus BIRB796. As anticipated, top pathways altered in response to these stimuli include cellular development, cellular growth and proliferation; tissue development; cell death, cell signaling, and small molecule biochemistry as well as numerous other pathways. Thus, ECTs display a broad spectrum of altered gene expression in response to mechanical load and/or tyrosine kinase inhibition, reflecting a complex regulation of proliferation, differentiation, and architectural alignment of cardiomyocytes and noncardiomyocytes within ECT. PMID:24303162

Ye, Fei; Yuan, Fangping; Li, Xiaohong; Cooper, Nigel; Tinney, Joseph P; Keller, Bradley B

2013-01-01

106

Intraperitoneal administration of AAV9-shRNA inhibits target gene expression in the dorsal root ganglia of neonatal mice  

PubMed Central

Background There is considerable interest in inducing RNA interference (RNAi) in neurons to study gene function and identify new targets for disease intervention. Although short interfering RNAs (siRNAs) have been used to silence genes in neurons, in vivo delivery of RNAi remains a major challenge, especially by systemic administration. We have developed a highly efficient method for in vivo gene silencing in dorsal root ganglia (DRG) by using short hairpin RNA–expressing single-stranded adeno-associated virus 9 (ssAAV9-shRNA). Results Intraperitoneal administration of ssAAV9-shRNA to neonatal mice resulted in highly effective and specific silencing of a target gene in DRG. We observed an approximately 80% reduction in target mRNA in the DRG, and 74.7% suppression of the protein was confirmed by Western blot analysis. There were no major side effects, and the suppression effect lasted for more than three months after the injection of ssAAV9-shRNA. Conclusions Although we previously showed substantial inhibition of target gene expression in DRG via intrathecal ssAAV9-shRNA administration, here we succeeded in inhibiting target gene expression in DRG neurons via intraperitoneal injection of ssAAV9-shRNA. AAV9-mediated delivery of shRNA will pave the way for creating animal models for investigating the molecular biology of the mechanisms of pain and sensory ganglionopathies. PMID:23866078

2013-01-01

107

Effect of myeloperoxidase inhibition on gene expression profiles in HL-60 cells exposed to 1,2,4,-benzenetriol.  

PubMed

While it is known that benzene induces myeloid leukemia in humans, the mechanism has yet to be clarified. Previously, we suggested that myeloperoxidase (MPO) was the key enzyme because it promotes generation of powerful oxidant hypochlorous acid (HOCl) which, reacting with DNA, causes leukemogenesis. In this study, using a whole-human-genome oligonucleotide microarray to clarify the relationships between myelotoxicity of benzene and MPO, we analyzed the genome-wide expression profiles of HL-60 human promyelocytic cell lines exposed to 1,2,4-benzenetriol (BT) with or without MPO inhibition. The microarray analysis revealed that short (1 h) and longer (4 h) exposure to BT changed the expression in HL-60 cells of 1,213 or 1,214 genes associated with transcription, RNA metabolic processes, immune response, apoptosis, cell death, and biosynthetic processes (|Z-score|> 2.0), and that these changes were dramatically lessened by MPO-specific inhibition. The presence of functionally important genes and, specifically, genes related to apoptosis, carcinogenesis, regulation of transcription, immune responses, oxidative stress, and cell-cycle regulation were further validated by real-time RT-PCR. Gene expression profiles along with Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway annotation analysis suggest that BT-induced DNA halogenation by MPO is a primary reaction in the leukemogenesis associated with benzene. PMID:24530881

Miyahara, Emiko; Nishikawa, Takuro; Takeuchi, Toru; Yasuda, Kaori; Okamoto, Yasuhiro; Kawano, Yoshifumi; Horiuchi, Masahisa

2014-03-20

108

Ajoene, a Sulfur-Rich Molecule from Garlic, Inhibits Genes Controlled by Quorum Sensing  

PubMed Central

In relation to emerging multiresistant bacteria, development of antimicrobials and new treatment strategies of infections should be expected to become a high-priority research area. Quorum sensing (QS), a communication system used by pathogenic bacteria like Pseudomonas aeruginosa to synchronize the expression of specific genes involved in pathogenicity, is a possible drug target. Previous in vitro and in vivo studies revealed a significant inhibition of P. aeruginosa QS by crude garlic extract. By bioassay-guided fractionation of garlic extracts, we determined the primary QS inhibitor present in garlic to be ajoene, a sulfur-containing compound with potential as an antipathogenic drug. By comprehensive in vitro and in vivo studies, the effect of synthetic ajoene toward P. aeruginosa was elucidated. DNA microarray studies of ajoene-treated P. aeruginosa cultures revealed a concentration-dependent attenuation of a few but central QS-controlled virulence factors, including rhamnolipid. Furthermore, ajoene treatment of in vitro biofilms demonstrated a clear synergistic, antimicrobial effect with tobramycin on biofilm killing and a cease in lytic necrosis of polymorphonuclear leukocytes. Furthermore, in a mouse model of pulmonary infection, a significant clearing of infecting P. aeruginosa was detected in ajoene-treated mice compared to a nontreated control group. This study adds to the list of examples demonstrating the potential of QS-interfering compounds in the treatment of bacterial infections. PMID:22314537

Jakobsen, Tim Holm; van Gennip, Maria; Phipps, Richard Kerry; Shanmugham, Meenakshi Sundaram; Christensen, Louise Dahl; Alhede, Morten; Skindersoe, Mette Eline; Rasmussen, Thomas Bovbjerg; Friedrich, Karlheinz; Uthe, Friedrich; Jensen, Peter ?strup; Moser, Claus; Nielsen, Kristian Fog; Eberl, Leo; Larsen, Thomas Ostenfeld; Tanner, David; H?iby, Niels; Bjarnsholt, Thomas

2012-01-01

109

Gene selective mRNA cleavage inhibits the development of Plasmodium falciparum.  

PubMed

Unique peptide-morpholino oligomer (PMO) conjugates have been designed to bind and promote the cleavage of specific mRNA as a tool to inhibit gene function and parasite growth. The new conjugates were validated using the P. falciparum gyrase mRNA as a target (PfGyrA). Assays in vitro demonstrated a selective degradation of the PfGyrA mRNA directed by the external guide sequences, which are morpholino oligomers in the conjugates. Fluorescence microscopy revealed that labeled conjugates are delivered into Plasmodium-infected erythrocytes during all intraerythrocytic stages of parasite development. Consistent with the expression of PfGyrA in all stages of parasite development, proliferation assays showed that these conjugates have potent antimalarial activity, blocking early development, maturation, and replication of the parasite. The conjugates were equally effective against drug sensitive and resistant P. falciparum strains. The potency, selectivity, and predicted safety of PMO conjugates make this approach attractive for the development of a unique class of target-specific antimalarials and for large-scale functional analysis of the malarial genome. PMID:22474358

Augagneur, Yoann; Wesolowski, Donna; Tae, Hyun Seop; Altman, Sidney; Ben Mamoun, Choukri

2012-04-17

110

Indirubin derivatives alter DNA binding activity of the transcription factor NF-Y and inhibit MDR1 gene promoter.  

PubMed

Indirubin derivatives exert antitumor activity. However, their effects on the expression of multidrug resistance gene 1 (MDR1) have not been investigated. Here we found three derivatives that inhibit the MDR1 gene promoter. To investigate the effects of indirubins on the DNA binding of NF-Y, a major MDR1 gene transcription factor that recognizes an inverted CCAAT element in the promoter, gel mobility shift assay was performed using the element as a probe with nuclear extracts from NG108-15, MCF7, HepG2, C2C12, and SK-N-SH cells. Among 17 compounds, 5-methoxyindirubin inhibited the DNA binding of NF-Y significantly, whereas indirubin-3'-oxime and 7-methoxyindirubin 3'-oxime increased the binding considerably. After evaluating a suitable concentration of each compound for transcription analysis using living tumor cells, we performed a reporter gene assay using a reporter DNA plasmid containing EGFP cDNA fused to the MDR1 gene promoter region. Indirubin-3'-oxime exerted a significant inhibitory effect on the MDR1 promoter activity in MCF7 and HepG2 cells, and 5-methoxyindirubin inhibited the activity only in MCF7 cells; 7-methoxyindirubin 3'-oxime suppressed the activity in all of the cell lines. We further confirmed that the compounds reduced endogenous MDR1 transcription without any inhibitory effect on NF-Y expression. Moreover, each compound increased the doxorubicin sensitivity of MCF7 cells. These results indicate that each indirubin derivative acts on the DNA binding of NF-Y and represses the MDR1 gene promoter with tumor cell-type specificity. PMID:25066113

Tanaka, Toru; Ohashi, Sachiyo; Saito, Hiroaki; Higuchi, Takashi; Tabata, Keiichi; Kosuge, Yasuhiro; Suzuki, Takashi; Miyairi, Shinichi; Kobayashi, Shunsuke

2014-10-15

111

Mood-enhancing antidepressant St. John's wort inhibits the activation of human immunodeficiency virus gene expression by ultraviolet light.  

PubMed

Ultraviolet (UV) radiation is a potent activator of the human immunodeficiency virus (HIV) gene expression in a HeLa cell clone with stably integrated copies of the HIVcat reporter construct. Recently, we have shown that activation of p38 MAP kinase and NF-kappaB is necessary but not sufficient for triggering efficient HIV gene expression in response to UV. Here we demonstrate that St. John's wort is a potent inhibitor of the UV-induced activation of HIV gene expression in HeLa cells. Stably transfected HIVcat/HeLa cells were preincubated with different amounts (25-100 microl) of St. John's wort or gingko biloba extracts for 30 min, then irradiated with UV (30 J/m2). In contrast to ginkgo biloba, St. John's wort inhibited the UV-induced HIV gene expression in a dose-dependent manner. Furthermore, preincubation with St. John's wort (10, 20, and 30 microl) for 30 min before UV (30 J/m2) irradiation, PMA- and UV-induced NF-kappaB activation was completely blocked, whereas ginkgo biloba did not affect the PMA- and UV-induced NF-kappaB activation in HeLa cells. UV activation of p38 MAP kinase was not inhibited by St. John's wort or by ginkgo biloba. However, we found that p38 MAP kinase and JNK1 and -2 were activated by St. John's wort, but p44/42 MAP kinase was not activated by St. John's wort in HeLa cells. Hypericin an active ingredient in St. John's wort also inhibited the UV activation of HIV gene expression in HeLa cells. These results firmly confirm that St. John's wort is a potent inhibitor of the UV-induced activation of HIV gene expression in HeLa cells. PMID:12665247

Taher, Mohiuddin M; Lammering, Guido M; Hershey, Chad M; Valerie, Kristoffer C

2002-12-01

112

Inhibition of interferon-inducible gene expression by adenovirus E1A proteins: Block in transcriptional complex formation  

SciTech Connect

Infection with wild-type adenovirus 5, but not with a mutant lacking the E1A gene, prevented the induction by interferon (IFN) {alpha} of chloramphenicol acetyltransferase (CAT) activity in HeLaM cell lines that had been permanently transfected with chimeric CAT reporter genes driven by the transcriptional regulatory regions of the IFN-inducible CAT activity was observed in cells that were cotransfected with the same reporter genes and plasmids expressing either the E1A 289- or 243-amino acid protein. These proteins also prevented the induction of CAT activity by IFN-{gamma} from a cotransfected HLA-DR{alpha}-CAT gene. Experiments with E1A mutants mapped the inhibitory activity to amino acid residues 38-65 of these proteins. In a HeLa cell line permanently expressing the E1A 289-amino acid protein, the replication of vesicular stomatitis virus and encephalomyocarditis virus was not inhibited by IFN-{alpha}, suggesting a global blockade of IFN responses. The observed transcriptional inhibition could be attributed to the lack of formation of the crucial IFN-stimulated gene factor 3 (ISGF3) transcriptional complex. As shown by mobility shift assays, this complex was not formed in the nuclear extracts of IFN-treated adenovirus-infected cells or IFN-treated E1A-producing cells. These nuclear extracts were deficient in both ISGF3{alpha} and ISGF3{gamma} subunits. However, they did not block the formation of ISGF3 complex from exogenously added components.

Kalvakolanu, D.V.R.; Bandyopadhyay, S.K.; Harter, M.L.; Sen, G.C. (Cleveland Clinic Foundation, OH (United States))

1991-09-01

113

Mutation of hepatocyte nuclear factor-1? inhibits Pkhd1 gene expression and produces renal cysts in mice  

PubMed Central

Hepatocyte nuclear factor–1? (HNF-1?) is a Pit-1, Oct-1/2, UNC-86 (POU)/homeodomain-containing transcription factor that regulates tissue-specific gene expression in the liver, kidney, and other organs. Humans with autosomal dominant mutations of HNF-1? develop maturity-onset diabetes of the young type 5 (MODY5) and congenital cystic abnormalities of the kidney. Autosomal recessive polycystic kidney disease (ARPKD) is an inherited cystic disorder that produces renal failure in infants and children and is caused by mutations of PKHD1. The proximal promoter of the mouse Pkhd1 gene contains an evolutionarily conserved HNF-1–binding site that is located near a region of deoxyribonuclease hypersensitivity. HNF-1? and the structurally related HNF-1? bind specifically to the Pkhd1 promoter and stimulate gene transcription. Mutations of the HNF-1 site or expression of a dominant-negative HNF-1? mutant inhibit Pkhd1 promoter activity in transfected cells. Transgenic mice expressing a dominant-negative HNF-1? mutant under the control of a kidney-specific promoter develop renal cysts, similarly to humans with MODY5. Pkhd1 transcripts are absent in the cells lining the cysts but are present in morphologically normal surrounding tubules. These studies identify a link between two cystic disease genes, HNF1? (MODY5) and PKHD1 (ARPKD). HNF-1? directly regulates the transcription of Pkhd1, and inhibition of PKHD1 gene expression may contribute to the formation of renal cysts in humans with MODY5. PMID:15067314

Hiesberger, Thomas; Bai, Yun; Shao, Xinli; McNally, Brian T.; Sinclair, Angus M.; Tian, Xin; Somlo, Stefan; Igarashi, Peter

2004-01-01

114

Inhibition of human cytomegalovirus immediate-early gene expression by an antisense oligonucleotide complementary to immediate-early RNA.  

PubMed Central

ISIS 2922 is a phosphorothioate oligonucleotide that is complementary to human cytomegalovirus (CMV) immediate-early (IE) RNA and that exhibits potent and specific antiviral activity against CMV in cell culture assays. Specific assay systems were developed to separately characterize the antisense and nonantisense components of the antiviral activity mediated by ISIS 2922. In U373 cells transformed with cDNA encoding the CMV IE 55-kDa (IE55) protein, expression was inhibited at nanomolar concentrations comparable to effective concentrations in antiviral assays. The specificity of inhibition was demonstrated by using control oligonucleotides incorporating progressive base changes to destabilize oligonucleotide-RNA base pairing and by showing a lack of inhibition of the CMV IE72 product expressed from the same promoter. Inhibition of IE55 protein expression correlated with a reduction in mRNA levels consistent with an RNase H-mediated termination event. Studies with virus-infected cells demonstrated that antisense and nonantisense mechanisms contribute to the antiviral activity of ISIS 2922. Base complementarity to target RNA was important for optimal activity in antiviral assays, but base changes affecting parameters other than hybridization affinity also influenced antiviral activity. Sequence-independent inhibition of virus adsorption to host cells by phosphorothioate oligonucleotides was also observed at high concentrations. Therefore, at least three different mechanisms may contribute to the antiviral activity of ISIS 2922 in cell culture: antisense-mediated inhibition of target gene expression; nonantisense, sequence-dependent inhibition of virus replication; and sequence-independent inhibition of virus adsorption to host cells. PMID:8878571

Anderson, K P; Fox, M C; Brown-Driver, V; Martin, M J; Azad, R F

1996-01-01

115

The OsFOR1 gene encodes a polygalacturonase-inhibiting protein (PGIP) that regulates floral organ number in rice  

Microsoft Academic Search

We have isolated a cDNA clone, OsFOR1, from the immature panicles of rice. The OsFOR1 (Oryza sativa floral organ regulator 1) gene encodes a protein that contains a leucine-rich repeat (LRR) domain. This domain comprises 10 tandem repeats of a canonical 24-amino acid LRR sequence. The structure and the number of LRRs for OsFOR1 are similar to those of polygalacturonase-inhibiting

Seonghoe Jang; Byongho Lee; Chanhong Kim; Soo-Jin Kim; Jieun Yim; Jong-Jin Han; Shinyoung Lee; Seong-Ryong Kim; Gynheung An

2003-01-01

116

Enhanced External Counterpulsation Inhibits Intimal Hyperplasia by Modifying Shear Stress-Responsive Gene Expression in Hypercholesterolemic Pigs  

Microsoft Academic Search

Background—Enhanced external counterpulsation (EECP) is a circulation assist device that may improve endothelial dysfunction by increasing shear stress. Chronic exposure of vascular endothelial cells and vascular smooth muscle cells to relatively high physiological shear stress has antiproliferative and vasoprotective effects. The present study hypothesizes that EECP inhibits intimal hyperplasia and atherogenesis by modifying shear stress-responsive gene expression. Methods and Results—Thirty-five

Yan Zhang; Xiaohong He; Xiaolin Chen; Hong Ma; Donghong Liu; Jinyun Luo; Zhimin Du; Yafei Jin; Yan Xiong; Jiangui He; Kuijian Wang; William E. Lawson; John C. K. Hui; Zhensheng Zheng; Guifu Wu

2010-01-01

117

Soluble pig lymphocyte activation gene-3 (LAG3; CD223) inhibits human-to-pig xenogeneic mixed lymphocyte reaction  

Microsoft Academic Search

Lymphocyte activation gene-3 (LAG-3; CD223) is a transmembrane protein that is structurally similar to CD4. Since LAG-3 has\\u000a a much higher binding affinity to MHC class II than that of CD4, several approaches using soluble LAG-3 were used to modulate\\u000a immune responses by activation or inhibition of MHC class II expressing antigen presenting cells. In this study, we constructed\\u000a soluble

Seon-Soo Kim; Hyun-Jung Byun; Sang-Hoon Kim; Han-Hyoung Lee; Suk Jun Lee; Sang Joon Kim; Chung-Gyu Park; Taehoon Chun

2010-01-01

118

Transcriptional Inhibition of Hypertrophic Scars by a Gene Silencer, Pyrrole–Imidazole Polyamide, Targeting the TGF-?1 Promoter  

Microsoft Academic Search

Synthetic pyrrole–imidazole (PI) polyamides bind to the minor groove of double-helical DNA with high affinity and specificity, and inhibit the transcription of corresponding genes. We examined the effects of a transforming growth factor (TGF)-?1-targeted PI polyamide (Polyamide) on hypertrophic skin scars in rats. Hypertrophic scars were created dorsally in rats by incisions. FITC-labeled Polyamide was injected to investigate its distribution

Hisayo Washio; Noboru Fukuda; Hiroyuki Matsuda; Hiroki Nagase; Takayoshi Watanabe; Yoshiaki Matsumoto; Tadashi Terui

2011-01-01

119

Nodulation inhibition by Rhizobium leguminosarum multicopy nodABC genes and analysis of early stages of plant infection.  

PubMed

During analysis of early events in the infection and nodulation of Vicia hirsuta roots inoculated with normal and mutant strains of Rhizobium leguminosarum and strains containing cloned nodulation (nod) genes, a number of novel observations were made. (i) Alternating zones of curled and straight root hairs were seen on roots of V. hirsuta inoculated with the wild-type strain of R. leguminosarum. This phasing of root hair curling was not seen if plants were grown under continuous light or continuous dark conditions. (ii) Reduced nodulation and delayed nodule initiation was observed with a strain carrying a Tn5 mutation in the nodE gene. In addition the phased root hair curling was absent, and root hair curling was observed along the length of the root. (iii) The nodABC genes cloned on a multicopy plasmid in a wild-type strain inhibited nodulation but induced a continuous root hair curling response. Those few nodules that eventually formed were found to contain bacteria which had lost the plasmid carrying the nodABC genes. (iv) With a strain of Rhizobium cured of its indigenous symbiotic plasmid, but containing the cloned nodABCDEF genes, continuous root hair curling on V. hirsuta was observed. However, no infection threads were observed, and surprisingly, it did appear that initial stages of nodule development occurred. Observations of thin sections of these early developing nodules indicated that early nodule meristematic divisions may have occurred but that no bacteria were found within the nodules and no infection threads were observed either within the nodule bumps or within any of the root hairs. It was concluded that for normal infections to occur, precise regulation of the nod genes is required and that overexpression of the root hair curling genes inhibits the normal infection process. PMID:3009408

Knight, C D; Rossen, L; Robertson, J G; Wells, B; Downie, J A

1986-05-01

120

Prostaglandin E2 and other cyclic AMP elevating agents inhibit interleukin 2 gene transcription by counteracting calcineurin-dependent pathways  

PubMed Central

We have previously shown that prostaglandin E2 and other cAMP elevating agents inhibit the nuclear transcription of the human IL-2 gene by interfering with a Ca(2+)-sensitive T cell signal transduction pathway. Calcineurin, a Ca2+/calmodulin-dependent 2B protein phosphatase, is an essential component of the T cell receptor signal transduction pathway leading to IL-2 gene expression. We have therefore tested the hypothesis that this phosphatase may be a target for the inhibitory effects of cAMP on IL-2 gene transcription. We report here that PGE2 markedly reduces the IL-2 promoter activity that is induced by a constitutively active form of calcineurin. In contrast to the complete inhibition of promoter activity produced by the immunosuppressants cyclosporin A and FK-506, this partial block suggests that PGE2 modulates downstream events needed for lymphokine gene activation. Overexpression of calcineurin in Jurkat cells decreases their apparent sensitivity to the inhibitory effects of PGE2 consistent with the fact that this enzyme plays a physiological role in dephosphorylating substrates of cAMP-dependent kinases in several tissues. These results provide evidence that cAMP-dependent pathways may antagonize calcineurin-regulated cascades for T cell activation in vivo, and suggest crosstalk between the Ca2+ and the cAMP signaling pathways during T cell activation. PMID:7693857

1993-01-01

121

Inhibition of the Ubiquitin-Proteasome System Prevents Vaccinia Virus DNA Replication and Expression of Intermediate and Late Genes?  

PubMed Central

The ubiquitin-proteasome system has a central role in the degradation of intracellular proteins and regulates a variety of functions. Viruses belonging to several different families utilize or modulate the system for their advantage. Here we showed that the proteasome inhibitors MG132 and epoxomicin blocked a postentry step in vaccinia virus (VACV) replication. When proteasome inhibitors were added after virus attachment, early gene expression was prolonged and the expression of intermediate and late genes was almost undetectable. By varying the time of the removal and addition of MG132, the adverse effect of the proteasome inhibitors was narrowly focused on events occurring 2 to 4 h after infection, the time of the onset of viral DNA synthesis. Further analyses confirmed that genome replication was inhibited by both MG132 and epoxomicin, which would account for the effect on intermediate and late gene expression. The virus-induced replication of a transfected plasmid was also inhibited, indicating that the block was not at the step of viral DNA uncoating. UBEI-41, an inhibitor of the ubiquitin-activating enzyme E1, also prevented late gene expression, supporting the role of the ubiquitin-proteasome system in VACV replication. Neither the overexpression of ubiquitin nor the addition of an autophagy inhibitor was able to counter the inhibitory effects of MG132. Further studies of the role of the ubiquitin-proteasome system for VACV replication may provide new insights into virus-host interactions and suggest potential antipoxviral drugs. PMID:19129442

Satheshkumar, P. S.; Anton, Luis C.; Sanz, Patrick; Moss, Bernard

2009-01-01

122

Reactivation of HIC-1 gene by saRNA inhibits clonogenicity and invasiveness in breast cancer cells  

PubMed Central

Hypermethylated in cancer 1 (HIC-1) is a tumor suppressor gene, which is epigenetically silenced in breast cancer. It is known that the loss of HIC-1, caused by promoter hypermethylation, is associated with tumor aggression and poor survival in breast carcinoma. It has been shown that small activating RNA (saRNA) targeting promoter sequences may induce gene re-expression. In the current study, saRNA was used to restore HIC-1 expression, and the effects on colony formation, invasiveness and the cell cycle in breast cancer cells were explored. dsHIC1-2998, an saRNA, exhibited activating efficacy on MCF-7 and MDA-MB-231 cancer cell lines. A clonogenicity assay showed that evident colony inhibition was induced via saRNA-mediated re-expression of HIC-1 in the two cancer cell lines. Reactivation of HIC-1 significantly inhibited cell migration and invasion, resulting in G0/G1 cell cycle arrest in these cell lines. These findings suggest that HIC-1 may be a potential target in gene therapy for the treatment of breast cancer. saRNA may function as a therapeutic option for upregulating tumor suppressor genes in breast cancer.

ZHAO, FENG; PAN, SHENGLI; GU, YAN; GUO, SHANYU; DAI, QIANCHENG; YU, YINGYAN; ZHANG, WEI

2015-01-01

123

Silkworm Apolipophorin Protein Inhibits Hemolysin Gene Expression of Staphylococcus aureus via Binding to Cell Surface Lipoteichoic Acids*  

PubMed Central

We previously reported that a silkworm hemolymph protein, apolipophorin (ApoLp), binds to the cell surface of Staphylococcus aureus and inhibits expression of the saePQRS operon encoding a two-component system, SaeRS, and hemolysin genes. In this study, we investigated the inhibitory mechanism of ApoLp on S. aureus hemolysin gene expression. ApoLp bound to lipoteichoic acids (LTA), an S. aureus cell surface component. The addition of purified LTA to liquid medium abolished the inhibitory effect of ApoLp against S. aureus hemolysin production. In an S. aureus knockdown mutant of ltaS encoding LTA synthetase, the inhibitory effects of ApoLp on saeQ expression and hemolysin production were attenuated. Furthermore, the addition of anti-LTA monoclonal antibody to liquid medium decreased the expression of S. aureus saeQ and hemolysin genes. In S. aureus strains expressing SaeS mutant proteins with a shortened extracellular domain, ApoLp did not decrease saeQ expression. These findings suggest that ApoLp binds to LTA on the S. aureus cell surface and inhibits S. aureus hemolysin gene expression via a two-component regulatory system, SaeRS. PMID:23873929

Omae, Yosuke; Hanada, Yuichi; Sekimizu, Kazuhisa; Kaito, Chikara

2013-01-01

124

Recent Transcription-induced Histone H3 Lysine 4 (H3K4) Methylation Inhibits Gene Reactivation*  

PubMed Central

Recent transcription of GAL genes transiently leaves an H3K4 methylation mark at their promoters, providing an epigenetic memory for the recent transcriptional activity. However, the physiological significance of this mark is enigmatic. In our study, we show that the transient H3K4 di- and trimethylation at recently transcribed GAL1 inhibited the reinduction of GAL1. The H3K4 methylation functioned by recruiting the Isw1 ATPase onto GAL1 and thereby limiting the action of RNA polymerase II during GAL1 reactivation. Strikingly, the H3K4 methylation was also observed at the promoters of inositol- and fatty acid-responsive genes after recent transcription and played a negative role in their reinduction. Taken together, our data present a new mechanism by which H3K4 methylation regulates gene transcription. PMID:21849496

Zhou, Bo O.; Zhou, Jin-Qiu

2011-01-01

125

Inhibition of PrPSc formation by lentiviral gene transfer of PrP containing dominant negative mutations  

PubMed Central

Summary Currently there is no treatment to cure Transmissible Spongiform Encephalopathies. By taking advantage of the prion “resistant” polymorphisms Q171R and E219K that naturally exist in sheep and humans, respectively, we have evaluated a lentiviral gene transfer therapeutic approach. Here we show that VSV-G (Vesicular Stomatitis Virus) pseudotyped FIV (Feline Immunodeficiency Virus) derived vectors carrying the mouse Prnp gene in which these mutations have been inserted, are able to inhibit prion replication in chronically prion infected cells. Since lentiviral tools are able to transduce post-mitotic cells such as neurons or cells of the lymphoreticular system, this result presents an insight into the development of gene or cell therapy approaches to prion disease. PMID:15494372

Crozet, Carole; Lin, Yea-Lih; Mettling, Clement; Mourton-Gilles, Chantal; Corbeau, Pierre; Lehmann, Sylvain; Perrier, Veronique

2004-01-01

126

Inhibition of protein translation by the DISC1-Boymaw fusion gene from a Scottish family with major psychiatric disorders.  

PubMed

The t(1; 11) translocation appears to be the causal genetic lesion with 70% penetrance for schizophrenia, major depression and other psychiatric disorders in a Scottish family. Molecular studies identified the disruption of the disrupted-in-schizophrenia 1 (DISC1) gene by chromosome translocation at chromosome 1q42. Our previous studies, however, revealed that the translocation also disrupted another gene, Boymaw (also termed DISC1FP1), on chromosome 11. After translocation, two fusion genes [the DISC1-Boymaw (DB7) and the Boymaw-DISC1 (BD13)] are generated between the DISC1 and Boymaw genes. In the present study, we report that expression of the DB7 fusion gene inhibits both intracellular NADH oxidoreductase activities and protein translation. We generated humanized DISC1-Boymaw mice with gene targeting to examine the in vivo functions of the fusion genes. Consistent with the in vitro studies on the DB7 fusion gene, protein translation activity is decreased in the hippocampus and in cultured primary neurons from the brains of the humanized mice. Expression of Gad67, Nmdar1 and Psd95 proteins are also reduced. The humanized mice display prolonged and increased responses to the NMDA receptor antagonist, ketamine, on various mouse genetic backgrounds. Abnormal information processing of acoustic startle and depressive-like behaviors are also observed. In addition, the humanized mice display abnormal erythropoiesis, which was reported to associate with depression in humans. Expression of the DB7 fusion gene may reduce protein translation to impair brain functions and thereby contribute to the pathogenesis of major psychiatric disorders. PMID:24908665

Ji, Baohu; Higa, Kerin K; Kim, Minjung; Zhou, Lynn; Young, Jared W; Geyer, Mark A; Zhou, Xianjin

2014-11-01

127

Anacardic acid inhibits estrogen receptor alpha-DNA binding and reduces target gene transcription and breast cancer cell proliferation.  

PubMed

Anacardic acid (AnAc; 2-hydroxy-6-alkylbenzoic acid) is a dietary and medicinal phytochemical with established anticancer activity in cell and animal models. The mechanisms by which AnAc inhibits cancer cell proliferation remain undefined. AnAc 24:1(omega5) was purified from geranium (Pelargonium x hortorum) and shown to inhibit the proliferation of estrogen receptor alpha (ERalpha)-positive MCF-7 and endocrine-resistant LCC9 and LY2 breast cancer cells with greater efficacy than ERalpha-negative primary human breast epithelial cells, MCF-10A normal breast epithelial cells, and MDA-MB-231 basal-like breast cancer cells. AnAc 24:1(omega5) inhibited cell cycle progression and induced apoptosis in a cell-specific manner. AnAc 24:1(omega5) inhibited estradiol (E(2))-induced estrogen response element (ERE) reporter activity and transcription of the endogenous E(2) target genes pS2, cyclin D1, and cathepsin D in MCF-7 cells. AnAc 24:1(omega5) did not compete with E(2) for ERalpha or ERbeta binding, nor did AnAc 24:1(omega5) reduce ERalpha or ERbeta steady-state protein levels in MCF-7 cells; rather, AnAc 24:1(omega5) inhibited ER-ERE binding in vitro. Virtual screening with the molecular docking software Surflex evaluated AnAc 24:1(omega5) interaction with ERalpha ligand binding (LBD) and DNA binding (DBD) domains in conjunction with experimental validation. Molecular modeling revealed AnAc 24:1(omega5) interaction with the ERalpha DBD but not the LBD. Chromatin immunoprecipitation experiments revealed that AnAc 24:1(omega5) inhibited E(2)-ERalpha interaction with the endogenous pS2 gene promoter region containing an ERE. These data indicate that AnAc 24:1(omega5) inhibits cell proliferation, cell cycle progression, and apoptosis in an ER-dependent manner by reducing ER-DNA interaction and inhibiting ER-mediated transcriptional responses. PMID:20197399

Schultz, David J; Wickramasinghe, Nalinie S; Ivanova, Margarita M; Isaacs, Susan M; Dougherty, Susan M; Imbert-Fernandez, Yoannis; Cunningham, Albert R; Chen, Chunyuan; Klinge, Carolyn M

2010-03-01

128

Anacardic acid inhibits estrogen receptor alpha-DNA binding and reduces target gene transcription and breast cancer cell proliferation  

PubMed Central

Anacardic acid (2-hydroxy-6-alkylbenzoic acid) is a dietary and medicinal phytochemical with established anticancer activity in cell and animal models. The mechanisms by which anacardic acid inhibits cancer cell proliferation remain undefined. Anacardic acid 24:1?5 (AnAc 24:1?5) was purified from geranium (Pelargonium × hortorum) and shown to inhibit the proliferation of estrogen receptor ? (ER?)-positive MCF-7 and endocrine-resistant LCC9 and LY2 breast cancer cells with greater efficacy than ER?-negative primary human breast epithelial cells, MCF-10A normal breast epithelial cells, and MDA-MB-231 basal-like breast cancer cells. AnAc 24:1?5 inhibited cell cycle progression and induced apoptosis in a cell-specific manner. AnAc 24:1?5 inhibited estradiol (E2)-induced estrogen response element (ERE) reporter activity and transcription of the endogenous E2-target genes: pS2, cyclin D1, and cathepsin D in MCF-7 cells. AnAc 24:1?5 did not compete with E2 for ER? or ER? binding, nor did AnAc 24:1?5 reduce ER? or ER? steady state protein levels in MCF-7 cells; rather, AnAc 24:1?5 inhibited ER-ERE binding in vitro. Virtual Screening with the molecular docking software Surflex evaluated AnAc 24:1?5 interaction with ER? ligand binding and DNA binding domains (LBD and DBD) in conjunction with experimental validation. Molecular modeling revealed AnAc 24:1?5 interaction with the ER? DBD but not the LBD. Chromatin immunoprecipitation (ChIP) experiments revealed that AnAc 24:1?5 inhibited E2-ER? interaction with the endogenous pS2 gene promoter region containing an ERE. These data indicate that AnAc 24:1?5 inhibits cell proliferation, cell cycle progression and apoptosis in an ER-dependent manner by reducing ER-DNA interaction and inhibiting ER-mediated transcriptional responses. PMID:20197399

Schultz, David J.; Wickramasinghe, Nalinie S.; Ivanova, Margarita M.; Isaacs, Susan M.; Dougherty, Susan M.; Imbert-Fernandez, Yoannis; Cunningham, Albert R.; Chen, Chunyuan; Klinge, Carolyn M.

2010-01-01

129

Retinoid X receptor and peroxisome proliferator-activated receptor-gamma agonists cooperate to inhibit matrix metalloproteinase gene expression  

PubMed Central

Introduction We recently described the ability of retinoid X receptor (RXR) ligand LG100268 (LG268) to inhibit interleukin-1-beta (IL-1-?)-driven matrix metalloproteinase-1 (MMP-1) and MMP-13 gene expression in SW-1353 chondrosarcoma cells. Other investigators have demonstrated similar effects in chondrocytes treated with rosiglitazone, a ligand for peroxisome proliferator-activated receptor-gamma (PPAR?), for which RXR is an obligate dimerization partner. The goals of this study were to evaluate the inhibition of IL-1-?-induced expression of MMP-1 and MMP-13 by combinatorial treatment with RXR and PPAR? ligands and to investigate the molecular mechanisms of this inhibition. Methods We used real-time reverse transcription-polymerase chain reaction to measure LG268- and rosiglitazone-mediated inhibition of MMP gene transcription in IL-1-?-treated SW-1353 chondrosarcoma cells. An in vitro collagen destruction assay was a functional readout of MMP collagenolytic activity. Luciferase reporter assays tested the function of a putative regulatory element in the promoters of MMP-1 and MMP-13, and chromatin immunoprecipitation (ChIP) assays detected PPAR? and changes in histone acetylation at this site. Post-translational modification of RXR and PPAR? by small ubiquitin-like modifier (SUMO) was assayed with immunoprecipitation and Western blot. Results Rosiglitazone inhibited MMP-1 and MMP-13 expression in IL-1-?-treated SW-1353 cells at the mRNA and heterogeneous nuclear RNA levels and blunted IL-1-?-induced collagen destruction in vitro. Combining LG268 and rosiglitazone had an additive inhibitory effect on MMP-1 and MMP-13 transcription and collagenolysis. IL-1-? inhibited luciferase expression in the MMP reporter assay, but rosiglitazone and LG268 had no effect. ChIP indicated that treatment with IL-1-?, but not LG268 and rosiglitazone, increased PPAR? at the proximal promoters of both MMPs. Finally, rosiglitazone or LG268 induced 'cross-SUMOylation' of both the target receptor and its binding partner, and IL-1-?-alone had no effect on SUMOylation of RXR and PPAR? but antagonized the ligand-induced SUMOylation of both receptors. Conclusions The PPAR? and RXR ligands rosiglitazone and LG268 may act through similar mechanisms, inhibiting MMP-1 and MMP-13 transcription. Combinatorial treatment activates each partner of the RXR:PPAR? heterodimer and inhibits IL-1-?-induced expression of MMP-1 and MMP-13 more effectively than either compound alone. We conclude that the efficacy of combined treatment with lower doses of each drug may minimize potential side effects of treatment with these compounds. PMID:19046432

Burrage, Peter S; Schmucker, Adam C; Ren, Yanqing; Sporn, Michael B; Brinckerhoff, Constance E

2008-01-01

130

Mutations in PRPF31 inhibit pre-mRNA splicing of rhodopsin gene and cause apoptosis of retinal cells.  

PubMed

Mutations in human PRPF31 gene have been identified in patients with autosomal dominant retinitis pigmentosa (adRP). To begin to understand mechanisms by which defects in this general splicing factor cause retinal degeneration, we examined the relationship between PRPF31 and pre-mRNA splicing of photoreceptor-specific genes. We used a specific anti-PRPF31 antibody to immunoprecipitate splicing complexes from retinal cells and identified the transcript of rhodopsin gene (RHO) among RNA species associated with PRPF31-containing complexes. Mutant PRPF31 proteins significantly inhibited pre-mRNA splicing of intron 3 in RHO gene. In primary retinal cell cultures, expression of the mutant PRPF31 proteins reduced rhodopsin expression and caused apoptosis of rhodopsin-positive retinal cells. This primary retinal culture assay provides an in vitro model to study photoreceptor cell death caused by PRPF31 mutations. Our results demonstrate that mutations in PRPF31 gene affect RHO pre-mRNA splicing and reveal a link between PRPF31 and RHO, two major adRP genes. PMID:15659613

Yuan, Liya; Kawada, Mariko; Havlioglu, Necat; Tang, Hao; Wu, Jane Y

2005-01-19

131

Gene Silencing of 4-1BB by RNA Interference Inhibits Acute Rejection in Rats with Liver Transplantation  

PubMed Central

The 4-1BB signal pathway plays a key role in organ transplantation tolerance. In this study, we have investigated the effect of gene silencing of 4-1BB by RNA interference (RNAi) on the acute rejection in rats with liver transplantation. The recombination vector of lentivirus that contains shRNA targeting the 4-1BB gene (LV-sh4-1BB) was constructed. The liver transplantation was performed using the two-cuff technique. Brown-Norway (BN) recipient rats were infected by the recombinant LVs. The results showed that gene silencing of 4-1BB by RNAi downregulated the 4-1BB gene expression of the splenic lymphocytes in vitro, and the splenic lymphocytes isolated from the rats with liver transplantation. LV-sh4-1BB decreased the plasma levels of liver injury markers including AST, ALT, and BIL and also decreased the level of plasma IL-2 and IFN-? in recipient rats with liver transplantation. Lentivirus-mediated delivery of shRNA targeting 4-1BB gene prolonged the survival time of recipient and alleviated the injury of liver morphology in recipient rats with liver transplantation. In conclusion, our results demonstrate that gene silencing of 4-1BB by RNA interference inhibits the acute rejection in rats with liver transplantation. PMID:23484089

Shi, Yang; Hu, Shuqun; Song, Qingwei; Yu, Shengcai; Zhou, Xiaojun; Yin, Jun; Qin, Lei; Qian, Haixin

2013-01-01

132

Ionizing radiation inhibits the PLK cell cycle gene in a G2 checkpoint-dependent manner.  

PubMed

Tumor cell cycle arrest at the cell cycle G2/M boundary after ionizing radiation involves inhibition of the Polo-like kinase 1 (Plk1). We recently found that the mechanism comprised repression of its gene, PLK, mediated by the tumor-suppressor protein BRCA1. In the present study we examined the regulatory responses on PLK and cell cycle phases in breast carcinoma cell lines exposed to various modes of therapeutic irradiation. The tumor cells, harboring different DNA damage checkpoint defects, were irradiated with either a single dose of 8.0 Gy or fractionated doses accumulating to 8.0 Gy. In the BRCA1-/- HCC1937 cell line both radiation regimens caused moderate repression of PLK mRNA expression, whereas the reconstituted wild-type (wt) BRCA1 genotype of the HCC1937/BRCA1wt cell line was associated with significant down-regulation of PLK mRNA expression after irradiation. In contrast to the HCC1937 cell lines, the MCF7/LCC2 cells displayed the characteristic wt TP53 constitution of persistent, radiation-induced CDKN1A mRNA expression (encoding the G1 cell cycle inhibitor p21(Waf1/Cip1/Sdi1)). The regulatory effects on PLK in the MCF7/LCC2 cells, however, were identical to those in the HCC1937/BRCA1wt cell line. Moreover, whereas neither HCC1937 cell line displayed G1/S cell cycle arrest after irradiation but, instead, an apparent accumulation of G2/M-phase cells, the radiation-induced delay at the G1/S boundary seemed to be superior to arrest at the G2/M transition in the MCF7/LCC2 cell line. Since the down-regulation of PLK mRNA expression by ionizing radiation was identical in the wt TP53 MCF7/LCC2 cell line and the TP53-mutated BRCA1-/- HCC1937 cell line reconstituted with wt BRCA1, we conclude that this regulatory effect solely requires an intact G2 checkpoint effector mechanism. PMID:15160994

Ree, Anne Hansen; Bratland, Ase; Solberg Landsverk, Kirsti; Fodstad, Oystein

2004-01-01

133

Transcriptional inhibition of hypertrophic scars by a gene silencer, pyrrole-imidazole polyamide, targeting the TGF-?1 promoter.  

PubMed

Synthetic pyrrole-imidazole (PI) polyamides bind to the minor groove of double-helical DNA with high affinity and specificity, and inhibit the transcription of corresponding genes. We examined the effects of a transforming growth factor (TGF)-?1-targeted PI polyamide (Polyamide) on hypertrophic skin scars in rats. Hypertrophic scars were created dorsally in rats by incisions. FITC-labeled Polyamide was injected to investigate its distribution in the skin. Expression of TGF-?1, connective tissue growth factor (CTGF), collagen type1, and fibronectin mRNAs was evaluated by reverse transcription PCR analysis. The extent of fibrosis and the expression of TGF-?1 were evaluated histologically and immunohistochemically. Polyamide was distributed in almost all nuclei of skin cells. Expression of TGF-?1 mRNA reached a peak at 3 days after skin incision. Expression of CTGF and extracellular matrix mRNAs was increased continuously even after the peak induction of TGF-?1 mRNA. Injection of Polyamide completely inhibited both the development of scars and the induction of growth factors and extracellular matrix mRNAs. The treatment also markedly inhibited fibrotic changes and reduced the numbers of vimentin-positive spindle-shaped fibroblasts. Injection of Polyamide also reduced established hypertrophic scars in rats. Thus, TGF-?1-targeted PI polyamide should be a feasible gene silencer for hypertrophic scars and keloids. PMID:21654833

Washio, Hisayo; Fukuda, Noboru; Matsuda, Hiroyuki; Nagase, Hiroki; Watanabe, Takayoshi; Matsumoto, Yoshiaki; Terui, Tadashi

2011-10-01

134

Monocular inhibition reveals temporal and spatial changes in gene expression in the primary visual cortex of marmoset  

PubMed Central

We investigated the time course of the expression of several activity-dependent genes evoked by visual inputs in the primary visual cortex (V1) in adult marmosets. In order to examine the rapid time course of activity-dependent gene expression, marmosets were first monocularly inactivated by tetrodotoxin (TTX), kept in darkness for two days, and then exposed to various length of light stimulation. Activity-dependent genes including HTR1B, HTR2A, whose activity-dependency were previously reported by us, and well-known immediate early genes (IEGs), c-FOS, ZIF268, and ARC, were examined by in situ hybridization. Using this system, first, we demonstrated the ocular dominance type of gene expression pattern in V1 under this condition. IEGs were expressed in columnar patterns throughout layers II–VI of all the tested monocular marmosets. Second, we showed the regulation of HTR1B and HTR2A expressions by retinal spontaneous activity, because HTR1B and HTR2A mRNA expressions sustained a certain level regardless of visual stimulation and were inhibited by a blockade of the retinal activity with TTX. Third, IEGs dynamically changed its laminar distribution from half an hour to several hours upon a stimulus onset with the unique time course for each gene. The expression patterns of these genes were different in neurons of each layer as well. These results suggest that the regulation of each neuron in the primary visual cortex of marmosets is subjected to different regulation upon the change of activities from retina. It should be related to a highly differentiated laminar structure of marmoset visual systems, reflecting the functions of the activity-dependent gene expression in marmoset V1. PMID:23576954

Nakagami, Yuki; Watakabe, Akiya; Yamamori, Tetsuo

2013-01-01

135

Inhibition of TNF-?-induced MUC5AC mucin gene expression and production by wogonin through the inactivation of NF-?B signaling in airway epithelial cells.  

PubMed

In this study, we investigated whether wogonin significantly affects MUC5AC mucin gene expression and production in human airway epithelial cells. Confluent NCI-H292 cells were pretreated with wogonin for 30 min and then stimulated with tumor necrosis factor-? (TNF-?) for 24 h or the indicated periods. The MUC5AC mucin gene expression and mucin protein production were measured by RT-PCR and ELISA, respectively. We found that incubation of NCI-H292 cells with wogonin significantly inhibited mucin production and down-regulated MUC5AC gene expression induced by TNF-? in a dose-dependent fashion. To elucidate the action mechanism of wogonin, effect of wogonin on TNF-?-induced NF-?B signaling pathway was investigated by western blot analysis. Wogonin inhibited NF-?B activation induced by TNF-?. Inhibition of IKK by wogonin led to the suppression of I?B phosphorylation and degradation, p65 nuclear translocation and NF-?B-regulated gene expression. This, in turn, led to the down-regulation of MUC5AC protein production in NCI-H292 cells. Wogonin also inhibited the gene products involved in cell survival (Bcl-2) and proliferation (cyclooxygenase-2). These results suggest that wogonin inhibits the NF-?B signaling pathway, which may explain its role in the inhibition of MUC5AC mucin gene expression and production. PMID:23463646

Sikder, Md Asaduzzaman; Lee, Hyun Jae; Mia, Md Zakaria; Park, Su Hyun; Ryu, Jiho; Kim, Jang-Hyun; Min, Sang Yeon; Hong, Jang-Hee; Seok, Jeong Ho; Lee, Choong Jae

2014-01-01

136

The mushroom Ganoderma lucidum suppresses breast-to-lung cancer metastasis through the inhibition of pro-invasive genes.  

PubMed

Breast cancer metastasis is one of the major reasons for the high morbidity and mortality of breast cancer patients. In spite of surgical interventions, chemotherapy, radiation therapy and targeted therapy, some patients are considering alternative therapies with herbal/natural products. In the present study, we evaluated a well-characterized extract from the medicinal mushroom Ganoderma lucidum (GLE) for its affects on tumor growth and breast-to-lung cancer metastasis. MDA-MB-231 human breast cancer cells were implanted into the mammary fat pads of nude mice. GLE (100 mg/kg/every other day) was administered to the mice by an oral gavage for 4 weeks, and tumor size was measured using microcalipers. Lung metastases were evaluated by hematoxylin and eosin (H&E) staining. Gene expression in MDA-MB-231 cells was determined by DNA microarray analysis and confirmed by quantitative PCR. Identified genes were silenced by siRNA, and cell migration was determined in Boyden chambers and by wound-healing assay. Although an oral administration of GLE only slightly suppressed the growth of large tumors, the same treatment significantly inhibited the number of breast-to-lung cancer metastases. GLE also downregulated the expression of genes associated with invasive behavior (HRAS, VIL2, S100A4, MCAM, I2PP2A and FN1) in MDA-MB-231 cells. Gene silencing of HRAS, VIL2, S100A4, I2PP2A and FN1 by siRNA suppressed migration of MDA-MB?231 cells. Our study suggests that an oral administration of GLE can inhibit breast-to-lung cancer metastases through the downregulation of genes responsible for cell invasiveness. The anti-metastatic benefits of GLE warrant further clinical studies. PMID:24718855

Loganathan, Jagadish; Jiang, Jiahua; Smith, Amanda; Jedinak, Andrej; Thyagarajan-Sahu, Anita; Sandusky, George E; Nakshatri, Harikrishna; Sliva, Daniel

2014-06-01

137

Reovirus inhibition of cellular DNA synthesis: role of the S1 gene.  

PubMed

Type 3 reovirus inhibits L cell DNA synthesis, whereas type 1 reovirus exerts little or no effect on L cell DNA synthesis. By using recombinant viruses containing both type 1 and type 3 double-standard RNA segments, we determined that one double-stranded RNA segment, the reovirus type 3 S1 double-stranded RNA segment which encodes the viral hemagglutinin, segregates with and is responsible for the capacity of reovirus type 3 to inhibit L cell DNA synthesis. PMID:7241660

Sharpe, A H; Fields, B N

1981-04-01

138

Reovirus inhibition of cellular DNA synthesis: role of the S1 gene.  

PubMed Central

Type 3 reovirus inhibits L cell DNA synthesis, whereas type 1 reovirus exerts little or no effect on L cell DNA synthesis. By using recombinant viruses containing both type 1 and type 3 double-standard RNA segments, we determined that one double-stranded RNA segment, the reovirus type 3 S1 double-stranded RNA segment which encodes the viral hemagglutinin, segregates with and is responsible for the capacity of reovirus type 3 to inhibit L cell DNA synthesis. PMID:7241660

Sharpe, A H; Fields, B N

1981-01-01

139

Theobroxide Treatment Inhibits Wild Fire Disease Occurrence in Nicotiana benthamiana by the Overexpression of Defense-related Genes.  

PubMed

Theobroxide, a novel compound isolated from a fungus Lasiodiplodia theobromae, stimulates potato tuber formation and induces flowering of morning glory by initiating the jasmonic acid synthesis pathway. To elucidate the effect of theobroxide on pathogen resistance in plants, Nicotiana benthamiana plants treated with theobroxide were immediately infiltrated with Pseudomonas syringae pv. tabaci. Exogenous application of theobroxide inhibited development of lesion symptoms, and growth of the bacterial cells was significantly retarded. Semi-quantitative RT-PCRs using the primers of 18 defense-related genes were performed to investigate the molecular mechanisms of resistance. Among the genes, the theobroxide treatment increased the expression of pathogenesis-related protein 1a (PR1a), pathogenesis-related protein 1b (PR1b), glutathione S-transferase (GST), allen oxide cyclase (AOC), and lipoxyganase (LOX). All these data strongly indicate that theobroxide treatment inhibits disease development by faster induction of defense responses, which can be possible by the induction of defense-related genes including PR1a, PR1b, and GST triggered by the elevated jasmonic acid. PMID:25288936

Ahn, Soon Young; Baek, Kwang-Hyun; Moon, Yong Sun; Yun, Hae Keun

2013-03-01

140

Theobroxide Treatment Inhibits Wild Fire Disease Occurrence in Nicotiana benthamiana by the Overexpression of Defense-related Genes  

PubMed Central

Theobroxide, a novel compound isolated from a fungus Lasiodiplodia theobromae, stimulates potato tuber formation and induces flowering of morning glory by initiating the jasmonic acid synthesis pathway. To elucidate the effect of theobroxide on pathogen resistance in plants, Nicotiana benthamiana plants treated with theobroxide were immediately infiltrated with Pseudomonas syringae pv. tabaci. Exogenous application of theobroxide inhibited development of lesion symptoms, and growth of the bacterial cells was significantly retarded. Semi-quantitative RT-PCRs using the primers of 18 defense-related genes were performed to investigate the molecular mechanisms of resistance. Among the genes, the theobroxide treatment increased the expression of pathogenesis-related protein 1a (PR1a), pathogenesis-related protein 1b (PR1b), glutathione S-transferase (GST), allen oxide cyclase (AOC), and lipoxyganase (LOX). All these data strongly indicate that theobroxide treatment inhibits disease development by faster induction of defense responses, which can be possible by the induction of defense-related genes including PR1a, PR1b, and GST triggered by the elevated jasmonic acid. PMID:25288936

Ahn, Soon Young; Baek, Kwang-Hyun; Moon, Yong Sun; Yun, Hae Keun

2013-01-01

141

Ebola Virus Inhibits Induction of Genes by Double-Stranded RNA in Endothelial Cells  

Microsoft Academic Search

Fatal cases of filoviral infection are accompanied by a marked immunosuppression. Endothelial cells play a vital role in the host immune response through the expression of several immunomodulatory genes in addition to the expression of the antiviral genes, 2?,5?-oligoadenylate synthetase [2?-5?(A)N], and the double-stranded RNA (dsRNA)-activated protein kinase (PKR). dsRNA, an intermediate generated during viral replication and gene transcription of

Brian H. Harcourt; Anthony Sanchez; Margaret K. Offermann

1998-01-01

142

A growth factor-repressible gene associated with protein kinase C-mediated inhibition of adipocyte differentiation.  

PubMed

The conversion of determined adipoblasts to fully differentiated adipocytes requires appropriate environmental conditions. A strict dependence on cell confluence and a facilitation by glucocorticoid hormones have previously been described. We have found that agents that are capable of activating protein kinase C, such as basic fibroblast growth factor and phorbol esters, inhibit the differentiation of the adipogenic cell line TA1 without stimulating cell growth. Here we describe the sequence and characterization of a cDNA (clone 5) that detects an RNA, the expression of which is enhanced by glucocorticoids and increasing cell density. In contrast, activators of protein kinase C including basic fibroblast growth factor, phorbol esters, and synthetic diacylglycerols inhibit clone 5 gene expression. It appears that clone 5 expression is closely linked to environmental and hormonal factors that promote the differentiation of adipogenic cells. PMID:2455724

Navre, M; Ringold, G M

1988-07-01

143

Antioxidative dietary compounds modulate gene expression associated with apoptosis, DNA repair, inhibition of cell proliferation and migration.  

PubMed

Many dietary compounds are known to have health benefits owing to their antioxidative and anti-inflammatory properties. To determine the molecular mechanism of these food-derived compounds, we analyzed their effect on various genes related to cell apoptosis, DNA damage and repair, oxidation and inflammation using in vitro cell culture assays. This review further tests the hypothesis proposed previously that downstream products of COX-2 (cyclooxygenase-2) called electrophilic oxo-derivatives induce antioxidant responsive elements (ARE), which leads to cell proliferation under antioxidative conditions. Our findings support this hypothesis and show that cell proliferation was inhibited when COX-2 was down-regulated by polyphenols and polysaccharides. Flattened macrophage morphology was also observed following the induction of cytokine production by polysaccharides extracted from viili, a traditional Nordic fermented dairy product. Coix lacryma-jobi (coix) polysaccharides were found to reduce mitochondrial membrane potential and induce caspase-3- and 9-mediated apoptosis. In contrast, polyphenols from blueberries were involved in the ultraviolet-activated p53/Gadd45/MDM2 DNA repair system by restoring the cell membrane potential. Inhibition of hypoxia-inducible factor-1 by saponin extracts of ginsenoside (Ginsen) and Gynostemma and inhibition of S100A4 by coix polysaccharides inhibited cancer cell migration and invasion. These observations suggest that antioxidants and changes in cell membrane potential are the major driving forces that transfer signals through the cell membrane into the cytosol and nucleus, triggering gene expression, changes in cell proliferation and the induction of apoptosis or DNA repair. PMID:25226533

Wang, Likui; Gao, Shijuan; Jiang, Wei; Luo, Cheng; Xu, Maonian; Bohlin, Lars; Rosendahl, Markus; Huang, Wenlin

2014-01-01

144

Antioxidative Dietary Compounds Modulate Gene Expression Associated with Apoptosis, DNA Repair, Inhibition of Cell Proliferation and Migration  

PubMed Central

Many dietary compounds are known to have health benefits owing to their antioxidative and anti-inflammatory properties. To determine the molecular mechanism of these food-derived compounds, we analyzed their effect on various genes related to cell apoptosis, DNA damage and repair, oxidation and inflammation using in vitro cell culture assays. This review further tests the hypothesis proposed previously that downstream products of COX-2 (cyclooxygenase-2) called electrophilic oxo-derivatives induce antioxidant responsive elements (ARE), which leads to cell proliferation under antioxidative conditions. Our findings support this hypothesis and show that cell proliferation was inhibited when COX-2 was down-regulated by polyphenols and polysaccharides. Flattened macrophage morphology was also observed following the induction of cytokine production by polysaccharides extracted from viili, a traditional Nordic fermented dairy product. Coix lacryma-jobi (coix) polysaccharides were found to reduce mitochondrial membrane potential and induce caspase-3- and 9-mediated apoptosis. In contrast, polyphenols from blueberries were involved in the ultraviolet-activated p53/Gadd45/MDM2 DNA repair system by restoring the cell membrane potential. Inhibition of hypoxia-inducible factor-1 by saponin extracts of ginsenoside (Ginsen) and Gynostemma and inhibition of S100A4 by coix polysaccharides inhibited cancer cell migration and invasion. These observations suggest that antioxidants and changes in cell membrane potential are the major driving forces that transfer signals through the cell membrane into the cytosol and nucleus, triggering gene expression, changes in cell proliferation and the induction of apoptosis or DNA repair. PMID:25226533

Wang, Likui; Gao, Shijuan; Jiang, Wei; Luo, Cheng; Xu, Maonian; Bohlin, Lars; Rosendahl, Markus; Huang, Wenlin

2014-01-01

145

The microRNA159-regulated GAMYB-like genes inhibit growth and promote programmed cell death in Arabidopsis.  

PubMed

The microRNA159 (miR159) family represses the conserved GAMYB-like genes that encode R2R3 MYB domain transcription factors that have been implicated in gibberellin (GA) signaling in anthers and germinating seeds. In Arabidopsis (Arabidopsis thaliana), the two major miR159 family members, miR159a and miR159b, are functionally specific for two GAMYB-like genes, MYB33 and MYB65. These transcription factors have been shown to be involved in anther development, but there are differing reports about their role in the promotion of flowering and little is known about their function in seed germination. To understand the function of this pathway, we identified the genes and processes controlled by these GAMYB-like genes. First, we demonstrate that miR159 completely represses MYB33 and MYB65 in vegetative tissues. We show that GA does not release this repression and that these transcription factors are not required for flowering or growth. By contrast, in the absence of miR159, the deregulation of MYB33 and MYB65 in vegetative tissues up-regulates genes that are highly expressed in the aleurone and GA induced during seed germination. Confirming that these genes are GAMYB-like regulated, their expression was reduced in myb33.myb65.myb101 seeds. Aleurone vacuolation, a GA-mediated programmed cell death process required for germination, was impaired in these seeds. Finally, the deregulation of MYB33 and MYB65 in vegetative tissues inhibits growth by reducing cell proliferation. Therefore, we conclude that miR159 acts as a molecular switch, only permitting the expression of GAMYB-like genes in anthers and seeds. In seeds, these transcription factors participate in GA-induced pathways required for aleurone development and death. PMID:20699403

Alonso-Peral, Maria M; Li, Junyan; Li, Yanjiao; Allen, Robert S; Schnippenkoetter, Wendelin; Ohms, Stephen; White, Rosemary G; Millar, Anthony A

2010-10-01

146

Cryptopleurine Targets NF-?B Pathway, Leading to Inhibition of Gene Products Associated with Cell Survival, Proliferation, Invasion, and Angiogenesis  

PubMed Central

Background Cryptopleurine, a phenanthroquinolizidine alkaloid, was known to exhibit anticancer activity; however, the underlying mechanism is poorly understood. Because the nuclear factor-?B (NF-?B) transcription factors control many physiological processes including inflammation, immunity, and development and progression of cancer, we investigated the effects of cryptopleurine on tumor necrosis factor alpha (TNF-?)-induced NF-?B activation pathway and on the expression of NF-?B-regulated gene products associated with many pathophysiological processes. Methodology and Principal Finding MDA-MB231, MDA-MB435, MCF-7, HEK293, RAW264.7 and Hep3B cells were used to examine cryptopleurine's effect on the NF-?B activation pathway. Major assays were promoter-reporter gene assay, electrophoretic mobility shift assay (EMSA), in vitro immune complex kinase assay, real-time PCR, Western blot analysis, and Matrigel invasion assay. Experiments documenting cell proliferation and apoptosis were analyzed by MTT method and flow cytometry, respectively. The results indicated that cryptopleurine suppressed the NF-?B activation through the inhibition of I?B kinase (IKK) activation, thereby blocking the phosphorylation and degradation of the inhibitor of NF-?B alpha (I?B?) and the nuclear translocation and DNA-binding activity of p65. The suppression of NF-?B by cryptopleurine led to the down-regulation of gene products involved in inflammation, cell survival, proliferation, invasion, and angiogenesis. Conclusions and Significance Our results show that cryptopleurine inhibited NF-?B activation pathway, which leads to inhibition of inflammation, proliferation, and invasion, as well as potentiation of apoptosis. Our findings provide a new insight into the molecular mechanisms and a potential application of cryptopleurine for inflammatory diseases as well as certain cancers associated with abnormal NF-?B activation. PMID:22768286

Cai, Xing Fu; Li, Donghao; Wu, Xue; Nan, Ji Xing; Lee, Jung Joon; Jin, Xuejun

2012-01-01

147

miR-125b inhibits hepatitis B virus expression in vitro through targeting of the SCNN1A gene.  

PubMed

microRNAs (miRNAs) are small noncoding RNAs that modulate gene expression at the posttranscriptional level, playing an important role in many diseases. However, reports concerning the role of miRNA in hepatitis B virus (HBV) infection are limited. miRNA chips were used to investigate miRNA changes during HBV infection in vitro. Bioinformatics analysis was used to explore possible miRNA and target genes during HBV infection. The expression of miR-125b and its potential target gene, sodium channel, non-voltage-gated 1 alpha (SCNN1A), was further analyzed. A total of 136 miRNAs were analyzed in an HBV transient transfection model (HepG2-HBV1.3), and 78 miRNAs were differentially expressed in HepG2.2.15 cells compared with HepG2 cells. miR-125b expression was decreased in both HepG2-HBV1.3 and HepG2.2.15 cells, and ectopic expression of miR-125b inhibited HBV DNA intermediates and secretion of HBsAg and HBeAg. miR-125b also inhibited the mRNA and protein levels of SCNN1A. Using a dual luciferase reporter system, we found that SCNN1A was one of the targets of miR-125b. In this study, we found that miR-125b inhibits HBV expression in vitro by regulating SCNN1A expression. PMID:25173609

Zhang, Zhenzhen; Chen, Juan; He, Yin; Zhan, Xue; Zhao, Ruiqiu; Huang, Yanfeng; Xu, Hongmei; Zhu, Zhaomin; Liu, Quanbo

2014-12-01

148

Inhibition of germination gene expression by Viviparous-1 and ABA during maize kernel development.  

PubMed

Two maize glyoxysomal genes expressed during germination, malate synthase (MS) and isocitrate lyase (ICL), were used to characterize the regulatory roles of the Viviparous-1 (Vp1) regulatory gene and abscisic aicd (ABA) in the induction of embryo quiescence during kernel development. In wild-type maize embryo, MS and ICL transcripts were first detected at 2 (MS) or 3 (ICL) days after germination (DAG), peaked at 5 DAG, and decreased thereafter. By reverse transcriptase-polymerase chain reaction (RT-PCR), the germination-specific genes were amplified in both ABA-insensitive (vp1) and ABA-deficient (vp7 and vp10) mutant embryos at 26 and 33 days after pollination (DAP), but not in wild-type embryos. The repression of these germination-specific genes thus requires the Vp1 gene product and normal levels of ABA to induce embryo quiescence during kernel development. This suggests that a genetic regulatory system exists to prevent vivipary in developing maize embryos. The involvement of the Vp1 gene product and ABA in repressing germination-specific genes complements their previously defined roles in the induction of seed-specific genes such as C1. PMID:9666472

Paek, N C; Lee, B M; Gyu Bai, D; Smith, J D

1998-06-30

149

fMRI Activation during Response Inhibition and Error Processing: The Role of the DAT1 Gene in Typically Developing Adolescents and Those Diagnosed with ADHD  

ERIC Educational Resources Information Center

The DAT1 gene codes for the dopamine transporter, which clears dopamine from the synaptic cleft, and a variant of this gene has previously been associated with compromised response inhibition in both healthy and clinical populations. This variant has also been associated with ADHD, a disorder that is characterised by disturbed dopamine function as…

Braet, Wouter; Johnson, Katherine A.; Tobin, Claire T.; Acheson, Ruth; McDonnell, Caroline; Hawi, Ziarah; Barry, Edwina; Mulligan, Aisling; Gill, Michael; Bellgrove, Mark A.; Robertson, Ian H.; Garavan, Hugh

2011-01-01

150

RECK inhibits stemness gene expression and tumorigenicity of gastric cancer cells by suppressing ADAM-mediated Notch1 activation.  

PubMed

The Reversion-inducing Cysteine-rich Protein with Kazal Motifs (RECK) gene encodes a membrane-anchored glycoprotein that exhibits strong inhibitory activity against various matrix metalloproteinases (MMPs) and a disintegrin and metalloproteinase 10 (ADAM10). RECK functions as a tumor suppressor by inhibiting migration, invasion, and angiogenesis. However, whether RECK can modulate the stem-like phenotypes of cancer cells is not known. In this study, we demonstrate that RECK is down-regulated in gastric cancer cells and is further reduced in CD133-positive cancer stem-like cells. Ectopic expression of RECK induces down-regulation of the expression of stemness genes including Sox2, Oct4, and Nanog and the cancer stem cell marker CD133. Treatment of DAPT (a ?-secretase inhibitor) or TAPI-2 (a hydroxamate-based inhibitor of MMPs, tumor necrosis factor ? converting enzyme and ADAM17) reduces Notch1 shedding and activation which results in attenuation of stemness genes and CD133. Our data show that ADAM10 and ADAM17 are co-pulled down by RECK suggesting a physical interaction between RECK and ADAMs on cell surface. In addition, RECK suppresses sphere formation and sphere size of CD133-positive gastric cancer cells. Overexpression of Notch intracellular domain (NICD) or ADAM17 effectively reverse the inhibitory effect of RECK in CD133-positive cells. More importantly, RECK reduces tumorigenic activity of CD133-positive cells in vivo. Conversely, knockdown of RECK in non-tumorigenic GI2 cells increases stemness and CD133 expression and sphere forming ability. Collectively, these results indicate that RECK represses stemness gene expression and stem-like properties by inhibiting ADAM-mediated Notch1 shedding and activation. PMID:23881612

Hong, Kun-Jing; Wu, Deng-Chyang; Cheng, Kuang-Hung; Chen, Li-Tzong; Hung, Wen-Chun

2014-02-01

151

Molecular Characterization of a Glucose-Inhibited Division Gene, gidA, That Regulates Cytotoxic Enterotoxin of Aeromonas hydrophila  

PubMed Central

By using a mini-transposon, we obtained two mutated strains of a diarrheal isolate, SSU, of Aeromonas hydrophila that exhibited a 50 to 53% reduction in the hemolytic activity and 83 to 87% less cytotoxic activity associated with the cytotoxic enterotoxin (Act). Act is a potent virulence factor of A. hydrophila and has been shown to contribute significantly to the development of both diarrhea and septicemia in animal models. Subsequent cloning and DNA sequence analysis revealed that transposon insertion occurred at different locations in these two mutants within the same 1,890-bp open reading frame for the glucose-inhibited division gene (gidA). A similar reduction in hemolytic (46%) and cytotoxic (81%) activity of Act was noted in the gidA isogenic mutant of A. hydrophila that was generated by marker exchange mutagenesis. Northern blot analysis revealed that the transcription of the cytotoxic enterotoxin gene (act) was not altered in the gidA transposon and isogenic mutants. However, by generating a chromosomal act::alkaline phosphatase gene (phoA) reporter construct, we demonstrated significantly reduced phosphatase activity in these mutants, indicating the effect of glucose-inhibited division (GidA) protein in modulating act gene expression at the translational level. The biological effects of Act in the gidA mutants were restored by complementation. The virulence of the gidA mutants in mice was dramatically reduced compared to the those of the wild-type (WT) and complemented strains of A. hydrophila. The histopathological examination of lungs, in particular, indicated severe congestion, alveolar hemorrhage, and acute inflammatory infiltrate in the interstitial compartment and the alveolar spaces when mice were infected with the WT and complemented strains. Minimal-to-mild changes were noted in the lungs with the gidA mutants. Taken together, our data indicate for the first time that GidA regulates the most-potent virulence factor of A. hydrophila, Act. PMID:14742556

Sha, Jian; Kozlova, E. V.; Fadl, A. A.; Olano, J. P.; Houston, C. W.; Peterson, J. W.; Chopra, A. K.

2004-01-01

152

Short-Chain Fatty Acids Inhibit Growth Hormone and Prolactin Gene Transcription via cAMP/PKA/CREB Signaling Pathway in Dairy Cow Anterior Pituitary Cells  

PubMed Central

Short-chain fatty acids (SCFAs) play a key role in altering carbohydrate and lipid metabolism, influence endocrine pancreas activity, and as a precursor of ruminant milk fat. However, the effect and detailed mechanisms by which SCFAs mediate bovine growth hormone (GH) and prolactin (PRL) gene transcription remain unclear. In this study, we detected the effects of SCFAs (acetate, propionate, and butyrate) on the activity of the cAMP/PKA/CREB signaling pathway, GH, PRL, and Pit-1 gene transcription in dairy cow anterior pituitary cells (DCAPCs). The results showed that SCFAs decreased intracellular cAMP levels and a subsequent reduction in PKA activity. Inhibition of PKA activity decreased CREB phosphorylation, thereby inhibiting GH and PRL gene transcription. Furthermore, PTX blocked SCFAs- inhibited cAMP/PKA/CREB signaling pathway. These data showed that the inhibition of GH and PRL gene transcription induced by SCFAs is mediated by Gi activation and that propionate is more potent than acetate and butyrate in inhibiting GH and PRL gene transcription. In conclusion, this study identifies a biochemical mechanism for the regulation of SCFAs on bovine GH and PRL gene transcription in DCAPCs, which may serve as one of the factors that regulate pituitary function in accordance with dietary intake. PMID:24177567

Wang, Jian-Fa; Fu, Shou-Peng; Li, Su-Nan; Hu, Zhong-Ming; Xue, Wen-Jing; Li, Zhi-Qiang; Huang, Bing-Xu; Lv, Qing-Kang; Liu, Ju-Xiong; Wang, Wei

2013-01-01

153

Therapeutic Effect of Sodium Iodide Symporter Gene Therapy Combined With External Beam Radiotherapy and Targeted Drugs That Inhibit DNA Repair  

PubMed Central

Adenoviral (AdV) transfer of sodium iodide symporter (NIS) gene has translational potential, but relatively low levels of transduction and subsequent radioisotope uptake limit the efficacy of the approach. In previous studies, we showed that combining NIS gene delivery with external beam radiotherapy (EBRT) and DNA damage repair inhibitors increased viral gene expression and radioiodide uptake. Here, we report the therapeutic efficacy of this strategy. An adenovirus expressing NIS from a telomerase promoter (Ad-hTR-NIS) was cytotoxic combined with relatively high-dose (50 µCi) 131I therapy and enhanced the efficacy of EBRT combined with low-dose (10 and 25 µCi) 131I therapy in colorectal and head and neck cancer cells. Combining this approach with ataxia-telangiectasia mutated (ATM) or DNA-dependent protein kinase (DNA-PK) inhibition caused maintenance of double-stranded DNA breaks (DSBs) at 24 hours and increased cytotoxicity on clonogenic assay. When the triplet of NIS-mediated 131I therapy, EBRT, and DNA-PKi was used in vivo, 90% of mice were tumor-free at 5 weeks. Acute radiation toxicity in the EBRT field was not exacerbated. In contrast, DNA-PKi did not enhance the therapeutic efficacy of EBRT plus adenovirus-mediated HSVtk/ganciclovir (GCV). Therefore, combining NIS gene therapy and EBRT represents an ideal strategy to exploit the therapeutic benefits of novel radiosensitizers. PMID:20588260

Hingorani, Mohan; White, Christine L; Zaidi, Shane; Pandha, Hardev S; Melcher, Alan A; Bhide, Shreerang A; Nutting, Christopher M; Syrigos, Konstantinos N; Vile, Richard G; Vassaux, Georges; Harrington, Kevin J

2010-01-01

154

Expression of a reporter gene interrupted by the Candida albicans group I intron is inhibited by base analogs.  

PubMed

We previously reported the identification of an intron (CaLSU) in the 25S ribosomal RNA of some Candida albicans yeast strains. CaLSU was shown to self-splice and has the potential to adopt a secondary structure typical of group I introns. The presence of CaLSU inC. albicans strains correlates with a high degree of susceptibility to base analog antifungal agents, 5-fluorocytosine (5-FC) or 5-fluorouracil (5-FU). Cell death, resulting from addition of base analogs to growing cultures, precluded demonstration of a causal relationship between CaLSU presence and susceptibility to base analogs. In the present study, CaLSU was inserted in a non-essential lacZ reporter gene and expression was examined in Saccharomyces cerevisiae. Different mutations affecting in vitro self-splicing also had similar effects on reporter gene expression in vivo. This indicates that in vivo removal of CaLSU from the reporter gene occurs through the typical self-splicing mechanism of group I introns. Base analogs inhibited expression of the reporter gene product in a concentration-dependent manner upon their addition to the cultures. This supports a model in which disruption of intron secondary structure, consecutive to the incorporation of nucleotide analogs, is a major factor determining the susceptibility of C.albicans cells to base analogs. PMID:9016575

Mercure, S; Cousineau, L; Montplaisir, S; Belhumeur, P; Lemay, G

1997-01-15

155

Adenoviral gene transfer of angiostatic ATF-BPTI inhibits tumour growth  

Microsoft Academic Search

BACKGROUND: The outgrowth of new vessels – angiogenesis – in the tumour mass is considered to be a limiting factor of tumour growth. To inhibit the matrix lysis that is part of the tumour angiogenesis, we employed the chimeric protein mhATF-BPTI, composed of the receptor binding part of the urokinase (ATF) linked to an inhibitor of plasmin (BPTI). METHODS: For

Pierre Lefesvre; Joline Attema; Dirk van Bekkum

2002-01-01

156

The association between 5-HTTLPR gene polymorphism and behavioral inhibition in Chinese toddlers.  

PubMed

As one of the fundamental individual characteristics, behavioral inhibition in early childhood has considerable implications for the development of social, cognitive, and psychological adjustment. The purpose of this study was to examine the relation between the 5-HTTLPR polymorphism and behavioral inhibition in Chinese children using a cross-sectional design. A sample of 263 2-year-old children (134 boys and 129 girls of Han ethnicity; ages ranging from 24 to 26 months) in China participated in the study. Behavioral inhibition was assessed through laboratory observations, and genomic DNA was collected with buccal swabs. The results of analysis of covariance (ANCOVA) indicated that the homozygous short 5-HTTLPR allele was associated with lower levels of behavioral inhibition, which was different from most of the findings based on individuals in Western countries. The results suggest that social and cultural factors may be involved in shaping links between the 5-HTTLPR polymorphism and children's specific behaviors. © 2014 Wiley Periodicals, Inc. Dev Psychobiol 56: 1601-1608, 2014. PMID:25196943

Chen, Xinyin; Zhang, Guangzhen; Liang, Zongbao; Zhang, Minghao; Way, Niobe; Yoshikawa, Hirokazu; Ke, Xiaoyan; Lu, Zuhong; Deng, Huihua

2014-11-01

157

Lost expression of DCC gene in ovarian cancer and its inhibition in ovarian cancer cells  

Microsoft Academic Search

Ovarian cancer is a leading cause of cancer-related women mortality in China. In recent years, the molecular mechanisms involved\\u000a in ovarian carcinoma development and\\/or progression have been intensely studied, and several genes have been identified. Deleted\\u000a in Colorectal Carcinoma (DCC), is an important tumor suppressor gene, which is inactivated in many kinds of tumors, and its function(s) is not clarified.

Liu Meimei; Li Peiling; Li Baoxin; Li Changmin; Zhuang Rujin; Hu Chunjie

2011-01-01

158

The strawberry gene FaGAST affects plant growth through inhibition of cell elongation  

Microsoft Academic Search

The strawberry (Fragaria3ananassa) FaGAST gene encodes a small protein with 12 cysteine residues conserved in the C-terminal region similar to a group of proteins identified in other species with diverse assigned functions such as cell division, elongation, or elongation arrest. This gene is expressed in the fruit receptacle, with two peaks during ripening at the white and the red-ripe stages,

Iraida Amaya; Cristina Castillejo; Jose F. Sanchez-Sevilla; Miguel A. Quesada; Miguel A. Botella; Victoriano Valpuesta

2006-01-01

159

Androgen Inhibits Abdominal Fat Accumulation and Negatively Regulates the PCK1 Gene in Male Chickens  

PubMed Central

Capons are male chickens whose testes have been surgically incised. Capons show a significant increase in fat accumulation compared to intact male chickens. However, while caponization leads to a significant reduction in androgen levels in roosters, little is known about the molecular mechanisms through which androgen status affects lipogenesis in avian species. Therefore, investigation of the influence of androgens on fat accumulation in the chicken will provide insights into this process. In this study, Affymetrix microarray technology was used to analyze the gene expression profiles of livers from capons and intact male chickens because the liver is the major site of lipogenesis in avian species. Through gene ontology, we found that genes involved in hepatic lipogenic biosynthesis were the most highly enriched. Interestingly, among the upregulated genes, the cytosolic form of the phosphoenolpyruvate carboxykinase (PCK1) gene showed the greatest fold change. Additionally, in conjunction with quantitative real-time PCR data, our results suggested that androgen status negatively regulated the PCK1 gene in male chickens. PMID:23544081

Shao, Yonggang; Li, Junying; Ling, Yao; Teng, Kedao; Li, Hongwei; Wu, Changxin

2013-01-01

160

Calcitonin gene-related Peptide inhibits osteolytic factors induced by osteoblast in co-culture system with breast cancer.  

PubMed

Recently, it was found that ?-Calcitonin gene-related peptide (CGRP) was associated with breast cancer metastases, but the role of CGRP in interaction between breast cancer and osteoblast during bone metastases is not clear. Here, we investigated the effect of CGRP on osteoblast in co-culture system with breast cancer. Using a breast cancer-osteoblast co-culture system, we chose MDA-MB-231 for breast cancer and human cell line MG-63 for osteoblast. CGRP was added to this co-culture system. The expression levels of the Runx2, RANK1, and osteoprotegerin (OPG) were analyzed using real-time PCR and western blot. CGRP receptors were investigated by immunofluorescence. We found that breast cancer cells cause osteolysis lesions by upregulating Runx2 expression, decreasing OPG expression, and increasing RANKL expression in osteoblasts. Our data prove that CGRP can regulate osteoclast coupling genes in osteoblast by increasing OPG, and decreasing RANKL and Runx2 expressions in a time-dependent manner; and inhibit those osteolytic factors induced by interaction between breast cancer cells and osteoblast. This inhibition could be abolished by the CGRP antagonist, CGRP8-37. In conclusion, calcitonin receptor-like receptor is the key player for CGRP's effect in this co-culture system. PMID:24853718

Zhao, Hui; Ning, Lin-Lin; Wang, Zhi-Yu; Li, Hong-Tao; Qiao, Dan; Yao, Yang; Qin, Huan-Long

2014-11-01

161

In vitro RNA interference targeting the DNA polymerase gene inhibits orf virus replication in primary ovine fetal turbinate cells.  

PubMed

Orf, which is caused by orf virus (ORFV), is distributed worldwide and is endemic in most sheep- and/or goat-raising countries. RNA interference (RNAi) pathways have emerged as important regulators of virus-host cell interactions. In this study, the specific effect of RNAi on the replication of ORFV was explored. The application of RNA interference (RNAi) inhibited the replication of ORFV in cell culture by targeting the ORF025 gene of ORFV, which encodes the viral polymerase. Three small interfering RNA (siRNA) (named siRNA704, siRNA1017 and siRNA1388) were prepared by in vitro transcription. The siRNAs were evaluated for antiviral activity against the ORFV Jilin isolate by the observation of cytopathic effects (CPE), virus titration, and real-time PCR. After 48 h of infection, siRNA704, siRNA1017 and siRNA1388 reduced virus titers by 59- to 199-fold and reduced the level of viral replication by 73-89 %. These results suggest that these three siRNAs can efficiently inhibit ORFV genome replication and infectious virus production. RNAi targeting of the DNA polymerase gene is therefore potentially useful for studying the replication of ORFV and may have potential therapeutic applications. PMID:24178308

Wang, Gaili; He, Wenqi; Song, Deguang; Li, Jida; Bao, Yingfu; Lu, Rongguang; Bi, Jingying; Zhao, Kui; Gao, Feng

2014-05-01

162

BRI3 Inhibits Amyloid Precursor Protein Processing in a Mechanistically Distinct Manner from Its Homologue Dementia Gene BRI2*  

PubMed Central

Alzheimer disease (AD) is characterized by senile plaques, which are mainly composed of ? amyloid (A?) peptides. A? is cleaved off from amyloid precursor protein (APP) with consecutive proteolytic processing: ?-secretase, followed by ?-secretase. Here, we show that BRI3, a member of the BRI gene family that includes the familial British and Danish dementia gene BRI2, interacts with APP and serves as an endogenous negative regulator of A? production. BRI3 colocalizes with APP along neuritis in differentiated N2a cells; endogenous BRI3-APP complexes are readily detectable in mouse brain extract; reducing endogenous BRI3 levels by RNA interference results in increased A? secretion. BRI3 resembles BRI2, because BRI3 overexpression reduces both ?- and ?-APP cleavage. We propose that BRI3 inhibits the various processing of APP by blocking the access of ?- and ?-secretases to APP. However, unlike BRI2, the binding of BRI3 to the ?-secretase cleaved APP C-terminal fragment is negligible and BRI3 does not cause the massive accumulation of this APP fragment, suggesting that, unlike BRI2, BRI3 is a poor ?-cleavage inhibitor. Competitive inhibition of APP processing by BRI3 may provide a new approach to AD therapy and prevention. PMID:19366692

Matsuda, Shuji; Matsuda, Yukiko; D'Adamio, Luciano

2009-01-01

163

RNAi Silencing of the HaHMG-CoA Reductase Gene Inhibits Oviposition in the Helicoverpa armigera Cotton Bollworm  

PubMed Central

RNA interference (RNAi) has considerable promise for developing novel pest control techniques, especially because of the threat of the development of resistance against current strategies. For this purpose, the key is to select pest control genes with the greatest potential for developing effective pest control treatments. The present study demonstrated that the 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMG-CoA reductase; HMGR) gene is a potential target for insect control using RNAi. HMGR is a key enzyme in the mevalonate pathway in insects. A complete cDNA encoding full length HMGR (encoding an 837-aa protein) was cloned from Helicoverpa armigera (Lepidoptera: Noctuidae). The HaHMGR (H. armigera HMGR) knockdown using systemic RNAi in vivo inhibited the fecundity of the females, effectively inhibited ovipostion, and significantly reduced vitellogenin (Vg) mRNA levels. Moreover, the oviposition rate of the female moths was reduced by 98% by silencing HaHMGR compared to the control groups. One-pair experiments showed that both the proportions of valid mating and fecundity were zero. Furthermore, the HaHMGR-silenced females failed to lay eggs (approximate 99% decrease in oviposition) in the semi-field cage performance. The present study demonstrated the potential implications for developing novel pest management strategies using HaHMGR RNAi in the control of H. armigera and other insect pests. PMID:23844078

Wang, Zhijian; Dong, Yongcheng; Desneux, Nicolas; Niu, Changying

2013-01-01

164

Three-dimensionally specific inhibition of DNA repair-related genes by activated KRAS in colon crypt model.  

PubMed

Growth and differentiation of colonic epithelium are regulated in the three-dimensional (3D) physiological architecture, colonic crypt, and deregulation of 3D interactions is involved in tumorigenesis. Cell-based 3D culture systems provide a suitable approach bridging the gap between two-dimensional (2D) culture and animal models. KRAS mutations are found at high frequencies in human colorectal cancer (CRC); however, KRAS-targeted cancer therapy has not been developed. Here, we have established a 3D cell culture model resembling the colonic crypt by use of HKe3 cells, human CRC HCT116 cells disrupted at activated KRAS. In this 3D colonic crypt model, HKe3 cells showed the features of time course-dependent transit-amplifying and terminal-differentiated stages, which are characteristic of normal colonic crypt. On the basis of the features of HCT116 cells, activated KRAS inhibited normal cell polarity and apoptosis in 3D culture. The expression of DNA repair-related tumor suppressor genes including TP53, BRCA1, BRCA2, and EXO-1 was markedly suppressed by activated KRAS in 3D culture but not in 2D culture. These results together suggest that activated KRAS plays critical roles in the accumulation of genetic alterations through inhibition of DNA repair genes and apoptosis and that this 3D culture model will provide a useful tool for investigating the molecular mechanisms of CRC development. PMID:20454511

Tsunoda, Toshiyuki; Takashima, Yasuo; Fujimoto, Takahiro; Koyanagi, Midori; Yoshida, Yasuhiro; Doi, Keiko; Tanaka, Yoko; Kuroki, Masahide; Sasazuki, Takehiko; Shirasawa, Senji

2010-05-01

165

4'-Acetoamido-4-hydroxychalcone, a chalcone derivative, inhibits glioma growth and invasion through regulation of the tropomyosin 1 gene  

SciTech Connect

Research highlights: {yields} 4'-Acetoamido-4-hydroxychalcone (AHC) has anti-cancer property for glioma. {yields} 4'-Acetoamido-4-hydroxychalcone (AHC) increased tropomyosin expreesion through activattion of PKA signaling. {yields} 4'-Acetoamido-4-hydroxychalcone (AHC) inhibits glioma cell migration and invasion. {yields} In vivo administration of 4'-acetoamido-4-hydroxychalcone (AHC) reduced tumor growth. -- Abstract: Chalcones are precursors of flavonoids and have been shown to have anti-cancer activity. Here, we identify the synthetic chalcone derivative 4'-acetoamido-4-hydroxychalcone (AHC) as a potential therapeutic agent for the treatment of glioma. Treatment with AHC reduced glioma cell invasion, migration, and colony formation in a concentration-dependent manner. In addition, AHC inhibited vascular endothelial growth factor-induced migration, invasion, and tube formation in HUVECs. To determine the mechanism underlying the inhibitory effect of AHC on glioma cell invasion and migration, we investigated the effect of AHC on the gene expression change and found that AHC affects actin dynamics in U87MG glioma cells. In actin cytoskeleton regulating system, AHC increased tropomyosin expression and stress fiber formation, probably through activation of PKA. Suppression of tropomyosin expression by siRNA or treatment with the PKA inhibitor H89 reduced the inhibitory effects of AHC on glioma cell invasion and migration. In vivo experiments also showed that AHC inhibited tumor growth in a xenograft mouse tumor model. Together, these data suggest that the synthetic chalcone derivative AHC has potent anti-cancer activity through inhibition of glioma proliferation, invasion, and angiogenesis and is therefore a potential chemotherapeutic candidate for the treatment of glioma.

Ku, Bo Mi [Department of Anatomy and Neurobiology, Institute of Health Sciences, School of Medicine, Gyeongsang National University, Jinju 660-751 (Korea, Republic of)] [Department of Anatomy and Neurobiology, Institute of Health Sciences, School of Medicine, Gyeongsang National University, Jinju 660-751 (Korea, Republic of); Ryu, Hyung Won [Division of Applied Life Science (BK21 Program), EB-NCRC, Institute of Agriculture Life Science, Graduate School of Gyeongsang National University, Jinju 660-701 (Korea, Republic of)] [Division of Applied Life Science (BK21 Program), EB-NCRC, Institute of Agriculture Life Science, Graduate School of Gyeongsang National University, Jinju 660-701 (Korea, Republic of); Lee, Yeon Kyung; Ryu, Jinhyun; Jeong, Joo Yeon; Choi, Jungil [Department of Anatomy and Neurobiology, Institute of Health Sciences, School of Medicine, Gyeongsang National University, Jinju 660-751 (Korea, Republic of)] [Department of Anatomy and Neurobiology, Institute of Health Sciences, School of Medicine, Gyeongsang National University, Jinju 660-751 (Korea, Republic of); Cho, Hee Jun [Department of Microbiology, Research Institute of Life Science, College of Natureal Sciences, Gyeongsang National University, Jinju 660-701 (Korea, Republic of)] [Department of Microbiology, Research Institute of Life Science, College of Natureal Sciences, Gyeongsang National University, Jinju 660-701 (Korea, Republic of); Park, Ki Hun, E-mail: khpark@gnu.ac.kr [Division of Applied Life Science (BK21 Program), EB-NCRC, Institute of Agriculture Life Science, Graduate School of Gyeongsang National University, Jinju 660-701 (Korea, Republic of); Kang, Sang Soo, E-mail: kangss@gnu.ac.kr [Department of Anatomy and Neurobiology, Institute of Health Sciences, School of Medicine, Gyeongsang National University, Jinju 660-751 (Korea, Republic of)

2010-11-19

166

A novel antiestrogen agent Shikonin inhibits estrogen-dependent gene transcription in human breast cancer cells  

Microsoft Academic Search

Shikonin (SK) has been isolated and identified as a key bioactive component in an herbal plant, Shikon (gromwell). In this\\u000a study, we investigated antiestrogen activity of SK in breast cancer cells. In human breast cancer cells, we observed that\\u000a treatment with SK inhibits tumor cell growth in estrogen receptor ? (ER?)-positive, but not ER?-negative breast cancer cells.\\u000a Estrogen-dependent cell growth

Yuan Yao; Qun Zhou

2010-01-01

167

Inhibition of melanogenesis by the extract from Agaricus blazei without affecting iNOS gene expression  

Microsoft Academic Search

Tyrosinase is a key enzyme in melanin synthesis. Owing to enlargement of availability of edible types of mushrooms in food\\u000a medicines, we investigated effects of Agaricus blazei (ABE) on tyrosinase activity using l-tyrosine and l-3, 4-dihydroxyphenylalanine (l-DOPA) as the substrate in normal human epidermal melanocytes (NHEM). ABE inhibited tyrosinase activity similar to arbutin\\u000a and Vitamin C as two whitening agents

Motamed Elsayed Mahmoud; Abd El-Latif Hesham; Yasser Abdel-Galil Ahmed; Mohammed Sayed

2010-01-01

168

Arctigenin from Arctium lappa inhibits interleukin-2 and interferon gene expression in primary human T lymphocytes  

Microsoft Academic Search

Background  \\u000a Arctium lappa (Niubang), a Chinese herbal medicine, is used to treat tissue inflammation. This study investigates the effects of arctigenin (AC),\\u000a isolated from A. lappa, on anti-CD3\\/CD28 Ab-stimulated cell proliferation and cytokine gene expression in primary human T lymphocytes.\\u000a \\u000a \\u000a \\u000a \\u000a Methods  Cell proliferation was determined with enzyme immunoassays and the tritiated thymidine uptake method. Cytokine production\\u000a and gene expression were analyzed

Wei-Jern Tsai; Chu-Ting Chang; Guei-Jane Wang; Tzong-Huei Lee; Shwu-Fen Chang; Shao-Chun Lu; Yuh-Chi Kuo

2011-01-01

169

Inhibition of Chlamydiae by Primary Alcohols Correlates with the Strain-Specific Complement of Plasticity Zone Phospholipase D Genes  

PubMed Central

Members of the genus Chlamydia are obligate intracellular pathogens that have a unique biphasic developmental cycle and interactions with host cells. Many genes that dictate host infection tropism and, putatively, pathogenic manifestations of disease are clustered in a hypervariable region of the genome termed the plasticity zone (PZ). Comparative genomics studies have determined that an uncharacterized family of PZ genes encoding orthologs of eukaryotic and prokaryotic members of the phospholipase D (PLD) enzyme family varies among chlamydiae. Here, we show that the PZ PLD (pzPLD) of Chlamydia trachomatis are transcribed during both normal and persistent infection and that the corresponding PLD proteins are predominately localized in reticulate bodies on the inner leaflet of the inclusion membrane. Further, we show that strains of chlamydiae encoding the pzPLD, but not a strain lacking these genes, are inhibited by primary alcohols, potent PLD inhibitors, during growth in HeLa 229 cells. This inhibitory effect is amplified approximately 5,000-fold during recovery from persistent infection. These findings suggest that the chlamydial pzPLD may be important, strain-specific, pathogenesis factors in vivo. PMID:16368959

Nelson, David E.; Crane, Deborah D.; Taylor, Lacey D.; Dorward, David W.; Goheen, Morgan M.; Caldwell, Harlan D.

2006-01-01

170

Inhibition of Intracellular Antiviral Defense Mechanisms Augments Lentiviral Transduction of Human Natural Killer Cells: Implications for Gene Therapy  

PubMed Central

Abstract Adoptive immunotherapy with genetically modified natural killer (NK) cells is a promising approach for cancer treatment. Yet, optimization of highly efficient and clinically applicable gene transfer protocols for NK cells still presents a challenge. In this study, we aimed at identifying conditions under which optimum lentiviral gene transfer to NK cells can be achieved. Our results demonstrate that stimulation of NK cells with interleukin (IL)-2 and IL-21 supports efficient transduction using a VSV-G pseudotyped lentiviral vector. Moreover, we have identified that inhibition of innate immune receptor signaling greatly enhances transduction efficiency. We were able to boost the efficiency of lentiviral genetic modification on average 3.8-fold using BX795, an inhibitor of the TBK1/IKK? complex acting downstream of RIG-I, MDA-5, and TLR3. We have also observed that the use of BX795 enhances lentiviral transduction efficiency in a number of human and mouse cell lines, indicating a broadly applicable, practical, and safe approach that has the potential of being applicable to various gene therapy protocols. PMID:22779406

Sutlu, Tolga; Nystrom, Sanna; Gilljam, Mari; Stellan, Birgitta; Applequist, Steven E.

2012-01-01

171

Intratumoral decorin gene delivery by AAV vector inhibits brain glioblastomas and prolongs survival of animals by inducing cell differentiation.  

PubMed

Glioblastoma multiforme (GBM) is the most malignant cancer in the central nervous system with poor clinical prognosis. In this study, we investigated the therapeutic effect of an anti-cancer protein, decorin, by delivering it into a xenograft U87MG glioma tumor in the brain of nude mice through an adeno-associated viral (AAV2) gene delivery system. Decorin expression from the AAV vector in vitro inhibited cultured U87MG cell growth by induction of cell differentiation. Intracranial injection of AAV-decorin vector to the glioma-bearing nude mice in vivo significantly suppressed brain tumor growth and prolonged survival when compared to control non-treated mice bearing the same U87MG tumors. Proteomics analysis on protein expression profiles in the U87MG glioma cells after AAV-mediated decorin gene transfer revealed up- and down-regulation of important proteins. Differentially expressed proteins between control and AAV-decorin-transduced cells were identified through MALDI-TOF MS and database mining. We found that a number of important proteins that are involved in apoptosis, transcription, chemotherapy resistance, mitosis, and fatty acid metabolism have been altered as a result of decorin overexpression. These findings offer valuable insight into the mechanisms of the anti-glioblastoma effects of decorin. In addition, AAV-mediated decorin gene delivery warrants further investigation as a potential therapeutic approach for brain tumors. PMID:24625664

Ma, Hsin-I; Hueng, Dueng-Yuan; Shui, Hao-Ai; Han, Jun-Ming; Wang, Chi-Hsien; Lai, Ying-Hsiu; Cheng, Shi-Yuan; Xiao, Xiao; Chen, Ming-Teh; Yang, Yi-Ping

2014-01-01

172

The sodA gene of Haemophilus ducreyi encodes a hydrogen peroxide-inhibitable superoxide dismutase  

Microsoft Academic Search

Haemophilus ducreyi is the etiologic agent of the sexually transmitted disease chancroid, an ulcerative condition implicated in increased HIV transmission. There is increasing evidence for the roles of oxidative stress proteins including superoxide dismutase enzymes in the survival and persistence of pathogenic organisms within the host. The sodA gene of Haemophilus ducreyi was isolated from a genomic plasmid library on

Lani R San Mateo; Kristen L Toffer; Thomas H Kawula

1998-01-01

173

An anti-sense chalcone synthase gene in transgenic plants inhibits flower pigmentation  

Microsoft Academic Search

In most plants flower pigments derive from the flavonoid biosynthesis pathway. Consistent with this pathway in Petunia hybrida the key enzyme in flavonoid synthesis, chalcone synthase, is synthesized in the flower corolla, tube and anthers1. Here we show that constitutive expression of an 'anti-sense' chalcone synthase gene in transgenic petunia and tobacco plants results, with high frequency, in an altered

Alexander R. van der Krol; Peter E. Lenting; Jetty Veenstra; Ingrid M. van der Meer; Ronald E. Koes; Anton G. M. Gerats; Joseph N. M. Mol; Antoine R. Stuitje

1988-01-01

174

Herpes Simplex Virus Gene Products Required for Viral Inhibition of Expression of G1-Phase Functions  

E-print Network

shutoff (vhs) function, DNA replication, and late gene functions were not required for the virus and its synthetic machinery, redirect- ing the cell from synthesis of cellular proteins to synthe- sis for viral DNA synthesis, includ- ing a viral DNA polymerase as well as functional ho- mologs of several S

Knipe, David M.

175

Inhibition of Rev-mediated HIV-1 expression by an RNA binding protein encoded by the interferon-inducible 9-27 gene  

SciTech Connect

Interferon inhibits expression of human immunodeficiency virus type-1 (HIV-1) through unknown mechanisms. A gene inducible by interferon-[alpha] (IFN-[alpha]) and interferon-[gamma] (IFN-[gamma]) was isolated by screening of a human complementary DNA library for proteins binding to the Rev-responsive element (RRE) of HIV-1. The product of this gene, RBP9-27, was shown to bind RNA in vitro and to inhibit HIV-1 expression after transfection into human cells. RBP9-27 primarily inhibited Rev-dependent posttransscriptional steps of viral gene expression. Thus, RBP9-27 is a cellular factor that antagonizes Rev function. These results suggest an inteferon-induced antiviral mechanism operating through the induction of RNA binding proteins such as RBP9-27. Elucidation of RBP9-27 function may lead to a better understanding of the mechanism of interferon action during HIV-1 infection. 29 refs., 4 figs.

Constantoulakis, P.; Campbell, M.; Felber, B.K.; Nasioulas, G.; Afonina, E.; Pavlakis, G.N. (National Cancer Inst., Frederick, MD (United States))

1993-02-26

176

Molecular Cloning, Sequence Analysis, and Expression of the Polygalacturonase-inhibiting Protein (PGIP) Gene in Mulberry  

Microsoft Academic Search

A full-length cDNA sequence encoding polygalacturonase-inhibiting protein (PGIP) from mulberry, which we designated MPGIP (GenBank accession no.: HM044383), was cloned based on mulberry expressed sequence tags (ESTs). Sequence analysis showed\\u000a that the MPGIP is 1,274 base pairs (bp) in length, encoding 333 amino acids with a predicted molecular weight of 37.29 kDa and an isoelectric\\u000a point of 7.25. The expression levels

Dongqing Hu; Ruiqiang Dai; Yuhua Wang; Yinghua Zhang; Zhaoyue Liu; Rongjun Fang; Weiguo Zhao; Long Li; Qiang Lin; Liu Li

177

Human immunodeficiency virus (HIV) type 2-mediated inhibition of HIV type 1: a new approach to gene therapy of HIV-infection.  

PubMed Central

Human immunodeficiency virus (HIV) type 2, the second AIDS-associated human retrovirus, differs from HIV-1 in its natural history, infectivity, and pathogenicity, as well as in details of its genomic structure and molecular behavior. We report here that HIV-2 inhibits the replication of HIV-1 at the molecular level. This inhibition was selective, dose-dependent, and nonreciprocal. The closely related simian immunodeficiency provirus also inhibited HIV-1. The selectivity of inhibition was shown by the observation that HIV-2 did not significantly downmodulate the expression of the unrelated murine leukemia virus; neither did the murine leukemia virus markedly affect HIV-1 or HIV-2 expression. Moreover, while HIV-2 potently inhibited HIV-1, the reverse did not happen, thus identifying yet another and remarkable difference between HIV-1 and HIV-2. Mutational analysis of the HIV-2 genome suggested that the inhibition follows a complex pathway, possibly involving multiple genes and redundant mechanisms. Introduction of inactivating mutations into the structural and regulatory/accessory genes did not render the HIV-2 provirus ineffective. Some of the HIV-2 gene defects, such as that of tat and rev genes, were phenotypically transcomplemented by HIV-1. The HIV-2 proviruses with deletions in the putative packaging signal and defective for virus replication were effective in inducing the suppressive phenotype. Though the exact mechanism remains to be defined, the inhibition appeared to be mainly due to an intracellular molecular event because it could not be explained solely on the basis of cell surface receptor mediated interference. The results support the notion that the inhibition likely occurred at the level of viral RNA, possibly involving competition between viral RNAs for some transcriptional factor essential for virus replication. Induction of a cytokine is another possibility. These findings might be relevant to the clinical-epidemiological data suggesting that infection with HIV-2 may offer some protection against HIV-1 infection. Images Fig. 3 PMID:8633095

Arya, S K; Gallo, R C

1996-01-01

178

Silencing cathepsin S gene expression inhibits growth, invasion and angiogenesis of human hepatocellular carcinoma in vitro.  

PubMed

Cathepsin S (Cat S) plays an important role in tumor invasion and metastasis by its ability to degrade extracellular matrix (ECM). Our previous study suggested there could be a potential association between Cat S and hepatocellular carcinoma (HCC) metastasis. The present study was designed to determine the role of Cat S in HCC cell growth, invasion and angiogenesis, using RNA interference technology. Small interfering RNA (siRNA) sequences for the Cat S gene were synthesized and transfected into human HCC cell line MHCC97-H. The Cat S gene targeted siRNA-mediated knockdown of Cat S expression, leading to potent suppression of MHCC97-H cell proliferation, invasion and angiogenesis. These data suggest that Cat S might be a potential target for HCC therapy. PMID:22796222

Fan, Qi; Wang, Xuedi; Zhang, Hanguang; Li, Chuanwei; Fan, Junhua; Xu, Jing

2012-09-01

179

Cloning and Sequencing of the Gene Encoding Toho-2, a Class A ?-Lactamase Preferentially Inhibited by Tazobactam  

PubMed Central

Escherichia coli TUM1083, which is resistant to ampicillin, carbenicillin, cephaloridine, cephalothin, piperacillin, cefuzonam, and aztreonam while being sensitive to cefoxitin, moxalactam, cefmetazole, ceftazidime, and imipenem, was isolated from the urine of a patient treated with ?-lactam antibiotics. The ?-lactamase (Toho-2) purified from the bacteria hydrolyzed ?-lactam antibiotics such as penicillin G, carbenicillin, cephaloridine, cefoxitin, cefotaxime, ceftazidime, and aztreonam and especially had increased relative hydrolysis rates for cephalothin, cephaloridine, cefotaxime, and ceftizoxime. Different from other extended-spectrum ?-lactamases, Toho-2 was inhibited 16-fold better by the ?-lactamase inhibitor tazobactam than by clavulanic acid. Resistance to ?-lactams was transferred by conjugation from E. coli TUM1083 to E. coli ML4909, and the transferred plasmid was about 54.4 kbp, belonging to the incompatibility group IncFII. The cefotaxime resistance gene for Toho-2 was subcloned from the 54.4-kbp plasmid. The sequence of the gene was determined, and the open reading frame of the gene was found to consist of 981 bases. The nucleotide sequence of the gene (DDBJ accession no. D89862) designated as blatoho was found to have 76.3% identity to class A ?-lactamase CTX-M-2 and 76.2% identity to Toho-1. It has 55.9% identity to SHV-1 ?-lactamase and 47.5% identity to TEM-1 ?-lactamase. Therefore, the newly isolated ?-lactamase designated as Toho-2 produced by E. coli TUM1083 is categorized as an enzyme similar to Toho-1 group ?-lactamases rather than to mutants of TEM or SHV enzymes. According to the amino acid sequence deduced from the DNA sequence, the precursor consisted of 327 amino acid residues. Comparison of Toho-2 with other ?-lactamase (non-Toho-1 group) suggests that the substitutions of threonine for Arg-244 and arginine for Asn-276 are important for the extension of the substrate specificity. PMID:9593147

Ma, Ling; Ishii, Yoshikazu; Ishiguro, Masaji; Matsuzawa, Hiroshi; Yamaguchi, Keizo

1998-01-01

180

Inhibition of protein synthesis stimulates the transcription of human beta-interferon genes in Chinese hamster ovary cells.  

PubMed Central

Using Chinese hamster ovary (CHO) cells transfected with a plasmid carrying the human beta-interferon gene, we find that inhibitors of protein synthesis, in the absence of any other inducer, stimulate the production of interferon RNA; this effect is maintained in cells in which the plasmid sequences have been amplified 25- to 50-fold. Nuclear transcription assays show that a major effect of cycloheximide is to increase the rate of transcription of the interferon gene. This contradicts the generally accepted explanation that inhibitors of protein synthesis augment interferon production by stabilizing interferon mRNA. In addition, we have studied the effects of double stranded RNA [poly(rI) X poly(rC)] on the induction of interferon RNA in the presence and absence of cycloheximide. Our results indicate that poly(rI) X poly(rC) by itself causes a transient increase in interferon RNA; however, in the presence of cycloheximide this effect is prolonged. We do not, however, find an increase in transcription of the interferon gene(s) as an early response to poly(rI) X poly(rC). Finally, we have found that cells treated with cycloheximide or infected with Newcastle disease virus induce large amounts of a secreted 11-kDa protein. This cellular protein is not inducible by poly(rI) X poly(rC). We propose that both interferon and this 11-kDa protein belong to a family of proteins in which production is regulated in a coordinate fashion during viral inhibition of cellular protein synthesis. Images PMID:6330726

Ringold, G M; Dieckmann, B; Vannice, J L; Trahey, M; McCormick, F

1984-01-01

181

Inhibition of protein synthesis stimulates the transcription of human beta-interferon genes in Chinese hamster ovary cells.  

PubMed

Using Chinese hamster ovary (CHO) cells transfected with a plasmid carrying the human beta-interferon gene, we find that inhibitors of protein synthesis, in the absence of any other inducer, stimulate the production of interferon RNA; this effect is maintained in cells in which the plasmid sequences have been amplified 25- to 50-fold. Nuclear transcription assays show that a major effect of cycloheximide is to increase the rate of transcription of the interferon gene. This contradicts the generally accepted explanation that inhibitors of protein synthesis augment interferon production by stabilizing interferon mRNA. In addition, we have studied the effects of double stranded RNA [poly(rI) X poly(rC)] on the induction of interferon RNA in the presence and absence of cycloheximide. Our results indicate that poly(rI) X poly(rC) by itself causes a transient increase in interferon RNA; however, in the presence of cycloheximide this effect is prolonged. We do not, however, find an increase in transcription of the interferon gene(s) as an early response to poly(rI) X poly(rC). Finally, we have found that cells treated with cycloheximide or infected with Newcastle disease virus induce large amounts of a secreted 11-kDa protein. This cellular protein is not inducible by poly(rI) X poly(rC). We propose that both interferon and this 11-kDa protein belong to a family of proteins in which production is regulated in a coordinate fashion during viral inhibition of cellular protein synthesis. PMID:6330726

Ringold, G M; Dieckmann, B; Vannice, J L; Trahey, M; McCormick, F

1984-07-01

182

Selection on Glycine ?-1,3-Endoglucanase Genes Differentially Inhibited by a Phytophthora Glucanase Inhibitor Protein  

PubMed Central

Plant endo-?-1,3-glucanases (EGases) degrade the cell wall polysaccharides of attacking pathogens and release elicitors of additional plant defenses. Isozymes EGaseA and EGaseB of soybean differ in susceptibility to a glucanase inhibitor protein (GIP1) produced by Phytophthora sojae, a major soybean pathogen. EGaseA, the major elicitor-releasing isozyme, is a high-affinity ligand for GIP1, which completely inhibits it, whereas EGaseB is unaffected by GIP1. We tested for departures from neutral evolution on the basis of partial sequences of EGaseA and EGaseB from 20 widespread accessions of Glycine soja (the wild progenitor of soybean), from 4 other Glycine species, and across dicotyledonous plants. G. soja exhibited little intraspecific variation at either locus. Phylogeny-based codon evolution models detected strong evidence of positive selection on Glycine EGaseA and weaker evidence for selection on dicot EGases and Glycine EGaseB. Positively selected peptide sites were identified and located on a structural model of EGase bound to GIP1. Positively selected sites and highly variable sites were found disproportionately within 4.5 Å of bound GIP1. Low variation within G. soja EGases, coupled with positive selection in both Glycine and dicot lineages and the proximity of rapidly evolving sites to GIP1, suggests an arms race involving repeated adaptation to pathogen attack and inhibition. PMID:15545660

Bishop, J. G.; Ripoll, D. R.; Bashir, S.; Damasceno, C. M. B.; Seeds, J. D.; Rose, J. K. C.

2005-01-01

183

Cobalt stimulates HIF-1-dependent but inhibits HIF-2-dependent gene expression in liver cancer cells  

PubMed Central

Hypoxia-inducible factors (HIFs) are transcriptional regulators that mediate the cellular response to low oxygen. Although HIF-1 is usually considered as the principal mediator of hypoxic adaptation, several tissues and different cell types express both HIF-1 and HIF-2 isoforms under hypoxia or when treated with hypoxia mimetic chemicals such as cobalt. However, the similarities or differences between HIF-1 and HIF-2, in terms of their tissue- and inducer-specific activation and function, are not adequately characterized. To address this issue, we investigated the effects of true hypoxia and hypoxia mimetics on HIF-1 and HIF-2 induction and specific gene transcriptional activity in two hepatic cancer cell lines, Huh7 and HepG2. Both hypoxia and cobalt caused rapid induction of both HIF-1? and HIF-2? proteins. Hypoxia induced erythropoietin (EPO) expression and secretion in a HIF-2-dependent way. Surprisingly, however, EPO expression was not induced when cells were treated with cobalt. In agreement, both HIF-1- and HIF-2-dependent promoters (of PGK and SOD2 genes, respectively) were activated by hypoxia while cobalt only activated the HIF-1-dependent PGK promoter. Unlike cobalt, other hypoxia mimetics such as DFO and DMOG activated both types of promoters. Furthermore, cobalt impaired the hypoxic stimulation of HIF-2, but not HIF-1, activity and cobalt-induced HIF-2? interacted poorly with USF-2, a HIF-2-specific co-activator. These data show that, despite similar induction of HIF-1? and HIF-2? protein expression, HIF-1 and HIF-2 specific gene activating functions respond differently to different stimuli and suggest the operation of oxygen-independent and gene- or tissue-specific regulatory mechanisms involving additional transcription factors or co-activators. PMID:23958427

Befani, Christina; Mylonis, Ilias; Gkotinakou, Ioanna-Maria; Georgoulias, Panagiotis; Hu, Cheng-Jun; Simos, George; Liakos, Panagiotis

2013-01-01

184

Flavopiridol inhibits soft tissue sarcoma growth with preferential sensitivity in a CDK4 gene amplified subtype  

Microsoft Academic Search

Introduction: Current chemotherapy for advanced soft tissue sarcoma (STS) has low overall response rates. It has been shown that the CDK4 gene is frequently amplified in some STS subtypes. Here we tested flavopiridol, a pan cyclin dependent kinase inhibitor, with STS subtypes that have amplified (dedifferentiated liposarcoma, DDLS) or non-amplified (malignant peripheral nerve sheath tumor, MPNST) CDK4.Methods: DDLS and MPNST

Elliot B. Sambol; Peter T. Kennealey; Monica Motwani; Jin-Hong Chen; Haider Cheema; Margaret Leversha; Gary K. Schwartz; Samuel Singer

2004-01-01

185

Evolution of the Retroviral Restriction Gene Fv1: Inhibition of Non-MLV Retroviruses  

PubMed Central

Fv1 is the prototypic restriction factor that protects against infection by the murine leukemia virus (MLV). It was first identified in cells that were derived from laboratory mice and was found to be homologous to the gag gene of an endogenous retrovirus (ERV). To understand the evolution of the host restriction gene from its retroviral origins, Fv1s from wild mice were isolated and characterized. Most of these possess intact open reading frames but not all restricted N-, B-, NR-or NB-tropic MLVs, suggesting that other viruses could have played a role in the selection of the gene. The Fv1s from Mus spretus and Mus caroli were found to restrict equine infectious anemia virus (EIAV) and feline foamy virus (FFV) respectively, indicating that Fv1 could have a broader target range than previously thought, including activity against lentiviruses and spumaviruses. Analyses of the Fv1 sequences revealed a number of residues in the C-terminal region that had evolved under positive selection. Four of these selected residues were found to be involved in the novel restriction by mapping studies. These results strengthen the similarities between the two capsid binding restriction factors, Fv1 and TRIM5?, which support the hypothesis that Fv1 defended mice against waves of retroviral infection possibly including non-MLVs as well as MLVs. PMID:24603659

Yap, Melvyn W.; Colbeck, Emily; Ellis, Scott A.; Stoye, Jonathan P.

2014-01-01

186

Evolution of the retroviral restriction gene Fv1: inhibition of non-MLV retroviruses.  

PubMed

Fv1 is the prototypic restriction factor that protects against infection by the murine leukemia virus (MLV). It was first identified in cells that were derived from laboratory mice and was found to be homologous to the gag gene of an endogenous retrovirus (ERV). To understand the evolution of the host restriction gene from its retroviral origins, Fv1s from wild mice were isolated and characterized. Most of these possess intact open reading frames but not all restricted N-, B-, NR-or NB-tropic MLVs, suggesting that other viruses could have played a role in the selection of the gene. The Fv1s from Mus spretus and Mus caroli were found to restrict equine infectious anemia virus (EIAV) and feline foamy virus (FFV) respectively, indicating that Fv1 could have a broader target range than previously thought, including activity against lentiviruses and spumaviruses. Analyses of the Fv1 sequences revealed a number of residues in the C-terminal region that had evolved under positive selection. Four of these selected residues were found to be involved in the novel restriction by mapping studies. These results strengthen the similarities between the two capsid binding restriction factors, Fv1 and TRIM5?, which support the hypothesis that Fv1 defended mice against waves of retroviral infection possibly including non-MLVs as well as MLVs. PMID:24603659

Yap, Melvyn W; Colbeck, Emily; Ellis, Scott A; Stoye, Jonathan P

2014-03-01

187

Inhibition of lung epithelial cell proliferation by hyperoxia. Posttranscriptional regulation of proliferation-related genes.  

PubMed Central

The alveolar surface of the lung is a major target for oxidant injury. After injury, repair of the alveolar epithelium is dependent on the ability of epithelial type 2 (T2) cells to proliferate. The regulation of T2 cell proliferation and the effect of reactive oxygen (O2) species on this lung cell proliferation have not been well defined. To investigate this process we focused on the regulation of two late cell cycle genes, histone and thymidine kinase, in T2 cells and fibroblasts exposed in vitro to varying periods of hyperoxia (95% O2). Hyperoxia for 24 to 48 h arrested cell proliferation in a SV40T-immortalized T2 cell line we have developed and in primary and SV40T-immortalized lung fibroblasts. Despite the cessation of proliferation, histone and TK mRNA continued to be expressed at high levels; mRNA half-lives were markedly prolonged but neither protein was translated. Thus proliferation arrest induced by hyperoxia was associated with posttranscriptional control of at least two late cell cycle-related genes. This form of proliferation arrest is also seen when primary and SV40T-T2 cells but not fibroblasts are serum deprived, suggesting that T2 cells in vitro may be uniquely sensitive to alterations in their redox state and that these alterations in turn affect translational control of a subset of proliferation-related genes. Images PMID:1430207

Clement, A; Edeas, M; Chadelat, K; Brody, J S

1992-01-01

188

18?-Glycyrrhetinic acid inhibits methicillin-resistant Staphylococcus aureus survival and attenuates virulence gene expression.  

PubMed

Methicillin-resistant Staphylococcus aureus (MRSA) has become a major source of infection in hospitals and in the community. Increasing antibiotic resistance in S. aureus strains has created a need for alternative therapies to treat disease. A component of the licorice root Glycyrrhiza spp., 18?-glycyrrhetinic acid (GRA), has been shown to have antiviral, antitumor, and antibacterial activity. This investigation explores the in vitro and in vivo effects of GRA on MRSA pulsed-field gel electrophoresis (PFGE) type USA300. GRA exhibited bactericidal activity at concentrations exceeding 0.223 ?M. Upon exposure of S. aureus to sublytic concentrations of GRA, we observed a reduction in expression of key virulence genes, including saeR and hla. In murine models of skin and soft tissue infection, topical GRA treatment significantly reduced skin lesion size and decreased the expression of saeR and hla genes. Our investigation demonstrates that at high concentrations GRA is bactericidal to MRSA and at sublethal doses it reduces virulence gene expression in S. aureus both in vitro and in vivo. PMID:23114775

Long, Danyelle R; Mead, Julia; Hendricks, Jay M; Hardy, Michele E; Voyich, Jovanka M

2013-01-01

189

Knockdown of DNA methyltransferase-1 inhibits proliferation and derepresses tumor suppressor genes in myeloma cells  

PubMed Central

DNA methyltransferases (including DNMT1, DNMT3A and DNMT3B), catalyze the transfer of methyl groups from S-adenosyl-l-methionine to cytosine position 5; this methylation in promoter regions silences gene expression. In addition, DNMT1 plays a critical role in the maintenance of genomic DNA methylation during DNA replication. In the present study, silencing of DNMT1 with siRNA was performed in RPMI-8226 human multiple myeloma (MM) cells, and the impact on gene methylation status and proliferation of the cells was analyzed. Upon DNMT1 downregulation, proliferation decreased significantly compared with that in the control, non-transfected cells. The expression of B-cell lymphoma 2 and nuclear factor ?B proteins was also significantly reduced. Furthermore, nested methylation-specific polymerase chain reaction revealed that methylation of the tumor suppressor genes, suppressor of cytokine signaling 1 and p16, was significantly reduced upon DNMT1 knockdown. Our results suggest that DNMT1 silencing may be a promising strategy to consider during development of novel MM treatment strategies.

ZHOU, WENWEN; CHEN, HUYING; HONG, XIULI; NIU, XIAOQING; LU, QUANYI

2014-01-01

190

Inhibition of transcription of cytosine-containing DNA in vitro by the alc gene product of bacteriophage T4.  

PubMed Central

The alc gene product (gpalc) of bacteriophage T4 inhibits the transcription of cytosine-containing DNA in vivo. We examined its effect on transcription in vitro by comparing RNA polymerase isolated from Escherichia coli infected with either wild-type T4D+ or alc mutants. A 50 to 60% decline in RNA polymerase activity, measured on phage T7 DNA, was observed by 1 min after infection with either T4D+ or alc mutants; this did not occur when the infecting phage lacked gpalt. In the case of the T4D+ strain but not alc mutants, this was followed by a further decrease. By 5 min after infection the activity of alc mutants was 1.5 to 2.5 times greater than that of the wild type on various cytosine-containing DNA templates, whereas there was little or no difference in activity on T4 HMdC-DNA, in agreement with the in vivo specificity. Effects on transcript initiation and elongation were distinguished by using a T7 phage DNA template. Rifampin challenge, end-labeling with [gamma-32P]ATP, and selective initiation with a dinucleotide all indicate that the decreased in vitro activity of the wild-type polymerase relative to that of the alc mutants was due to inhibition of elongation, not to any difference in initiation rates. Wild-type (but not mutated) gpalc copurified with RNA polymerase on heparin agarose but not in subsequent steps. Immunoprecipitation of modified RNA polymerase also indicated that gpalc was not tightly bound to RNA polymerase intracellularly. It thus appears likely that gpalc inhibits transcript elongation on cytosine-containing DNA by interacting with actively transcribing core polymerase as a complex with the enzyme and cytosine-rich stretches of the template. Images PMID:2185231

Drivdahl, R H; Kutter, E M

1990-01-01

191

Blocking Signaling at the Level of GLI Regulates Downstream Gene Expression and Inhibits Proliferation of Canine Osteosarcoma Cells  

PubMed Central

The Hedgehog-GLI signaling pathway is active in a variety of human malignancies and is known to contribute to the growth and survival of human osteosarcoma cells. In this study, we examined the expression and regulation of GLI transcription factors in multiple canine osteosarcoma cell lines and analyzed the effects of inhibiting GLI with GANT61, a GLI-specific inhibitor. Compared with normal canine osteoblasts, real-time PCR showed that GLI1 and GLI2 were highly expressed in two out of three cell lines and correlated with downstream target gene expression of PTCH1and PAX6. Treatment of canine osteosarcoma cells with GANT61 resulted in decreased expression of GLI1, GLI2, PTCH1, and PAX6. Furthermore, GANT61 inhibited proliferation and colony formation in all three canine osteosarcoma cell lines. The finding that GLI signaling activity is present and active in canine osteosarcoma cells suggests that spontaneously arising osteosarcoma in dogs might serve as a good model for future preclinical testing of GLI inhibitors. PMID:24810746

Shahi, Mehdi Hayat; Holt, Roseline; Rebhun, Robert B.

2014-01-01

192

Blocking signaling at the level of GLI regulates downstream gene expression and inhibits proliferation of canine osteosarcoma cells.  

PubMed

The Hedgehog-GLI signaling pathway is active in a variety of human malignancies and is known to contribute to the growth and survival of human osteosarcoma cells. In this study, we examined the expression and regulation of GLI transcription factors in multiple canine osteosarcoma cell lines and analyzed the effects of inhibiting GLI with GANT61, a GLI-specific inhibitor. Compared with normal canine osteoblasts, real-time PCR showed that GLI1 and GLI2 were highly expressed in two out of three cell lines and correlated with downstream target gene expression of PTCH1and PAX6. Treatment of canine osteosarcoma cells with GANT61 resulted in decreased expression of GLI1, GLI2, PTCH1, and PAX6. Furthermore, GANT61 inhibited proliferation and colony formation in all three canine osteosarcoma cell lines. The finding that GLI signaling activity is present and active in canine osteosarcoma cells suggests that spontaneously arising osteosarcoma in dogs might serve as a good model for future preclinical testing of GLI inhibitors. PMID:24810746

Shahi, Mehdi Hayat; Holt, Roseline; Rebhun, Robert B

2014-01-01

193

Vasopressin inhibits type-I collagen and albumin gene expression in primary cultures of adult rat hepatocytes  

SciTech Connect

The mechanisms that regulate collagen gene expression in hepatic cells are poorly understood. Accelerated Ca2+ fluxes are associated with inhibiting collagen synthesis selectively in human fibroblasts. In suspension cultures of isolated hepatocytes, the Ca2+ agonist vasopressin increases cytosolic levels of free Ca2+. However, whether vasopressin's interactions with plasma membrane V1 receptors attenuate hepatic collagen production is unknown. We investigated this problem by studying vasopressin's effects on collagen synthesis and Ca2+ efflux in long-term primary cultures of differentiated and proliferation-competent adult rat hepatocytes. Twelve-day-old quiescent cultures were exposed to test substances and labeled with (5-3H)proline. Determinations of radioactivity in collagenase-sensitive and collagenase-resistant proteins were used to calculate the relative levels of collagen production. Synthetic (8-arg)vasopressin stimulated 45Ca2+ efflux within 1 min and inhibited hepatocyte collagen production within 3 h by 50%; overall rates of protein synthesis were not affected significantly. In cultures labeled with (35S)methionine, vasopressin also decreased the levels of newly synthesized and secreted albumin, but not fibrinogen, detected in specific immunoprecipitates analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. Northern blot analyses using specific (32P)cDNA probes revealed 70% decreases in hybridizable levels of collagen alpha 1(I) mRNA in hepatocyte cultures treated with either vasopressin or Ca2+ ionophore A23187; hybridizable levels of albumin mRNA also fell approximately 50% following vasopressin treatment.

Chojkier, M.; Brenner, D.A.; Leffert, H.L.

1989-06-05

194

Acanthoic acid inhibits melanogenesis through tyrosinase downregulation and melanogenic gene expression in B16 melanoma cells.  

PubMed

The aim of this study was to investigate the in vitro inhibitory effects of acanthoic acid (ACAN), isolated from Acanthopanax koreanum, on melanogenesis and its related enzymes such as tyrosinase, tyrosinase-related protein (TRP)-1, and TRP-2 in B16 melanoma cells. We found that ACAN significantly attenuates melanin synthesis and reduces the activity of intracellular tyrosinase, the rate-limiting melanogenic enzyme. Western blot analysis showed that ACAN also decreases tyrosinase, TRP-1, and TRP-2 protein expression. In addition, ACAN significantly decreased the expression of microphthalmia-associated transcription factor (MITF), a key regulator of melanogenesis. These results indicate that ACAN effectively inhibits melanin biosynthesis through down-regulation of MITF and thus could be useful as a new skin-whitening agent. PMID:24354173

Yoon, Weon-Jong; Ham, Young-Min; Yoon, Hun Seok; Lee, Wook-Jae; Lee, Nam Ho; Hyun, Chang-Gu

2013-10-01

195

Aerosol Azacytidine Inhibits Orthotopic Lung Cancers in Mice through Its DNA Demethylation and Gene Reactivation Effects  

PubMed Central

We devised an aerosol based demethylation therapy to achieve therapeutic efficacy in premalignant or in situ lesions of lung cancer, without systemic toxicity. Optimum regimens of aerosolized azacytidine (Aza) were designed and used in orthotopic human non-small cell lung cancer xenograft models. The therapeutic efficacy and toxicity of aerosol Aza were compared with intravenously administered Aza. We observed that 80% of the droplets of the aerosol Aza measured ?0.1–5 microns, which resulted in deposition in the lower bronchial airways. An animal model that phenocopies field carcinogeneisis in humans was developed by intratracheal inoculation of the human lung cancer cells in mice, thus resulting in their distribution throughout the entire airway space. Aerosolized Aza significantly prolonged the survival of mice bearing endo-bronchial lung tumors. The aerosol treatment did not cause any detectable lung toxicity or systemic toxicity. A pre-pharmacokinetic study in mice demonstrated that lung deposition of aerosolized Aza was significantly higher than the intravenous route. Lung tumors were resected after aerosol treatment and the methylation levels of 24 promoters of tumor-suppresser genes related to lung cancer were analyzed. Aerosol Aza significantly reduced the methylation level in 9 of these promoters and reexpressed several genes tested. In conclusion, aerosol Aza at non-cytotoxic doses appears to be effective and results in DNA demethylation and tumor suppressor gene re-expression. The therapeutic index of aerosol Aza is >100-fold higher than that of intravenous Aza. These results provide a preclinical rationale for a phase I clinical trial of aerosol Aza to be initiated at our Institution. PMID:25347303

Qiu, Xuan; Liang, Yuanxin; Sellers, Rani S.; Perez-Soler, Roman; Zou, Yiyu

2014-01-01

196

Acute inhibition of casein kinase 1?/? rapidly delays peripheral clock gene rhythms.  

PubMed

Circadian rhythms are generated through a transcription-translation feedback loop involving clock genes and the casein kinases CSNK1D and CSNK1E. In this study, we investigated the effects of the casein kinase inhibitor PF-670462 (50 mg/kg) on rhythmic expression of clock genes in the liver, pancreas and suprachiasmatic nucleus (SCN) as well as plasma corticosterone, melatonin and running behaviour in rats and compared them to the responses to a 4 h extension of the light phase. PF-670462 acutely phase delayed the rhythmic transcription of Bmal1, Per1, Per2 and Nr1d1 in both liver and pancreas by 4.5 ± 1.3 and 4.5 ± 1.2 h, respectively, 1 day after administration. In the SCN, the rhythm of Nr1d1 and Dbp mRNA expression was delayed by 4.2 and 4 h, respectively. Despite these changes, the time of peak plasma melatonin secretion was not delayed, although the plasma corticosterone rhythm and onset of wheel-running activity were delayed by 2.1 and 1.1 h, respectively. These changes are in contrast to the effects of the 4 h light extension, which resulted in delays in peak expression of the clock genes of less than 1 h and no change in the melatonin or corticosterone rhythms. The ability of the casein kinase inhibitor to bring about large phase shifts in the rhythms of major metabolic target tissues may lead to new drugs being developed to rapidly phase adjust circadian rhythms to alleviate the metabolic impact of shift work. PMID:25245819

Kennaway, D J; Varcoe, T J; Voultsios, A; Salkeld, M D; Rattanatray, L; Boden, M J

2015-01-01

197

Inhibition of Proprotein Convertase SKI-1 Blocks Transcription of Key Extracellular Matrix Genes Regulating Osteoblastic Mineralization*  

PubMed Central

Mineralization, a characteristic phenotypic property of osteoblastic lineage cells, was blocked by 4-(2-aminoethyl) benzenesulfonyl fluoride hydrochloride (AEBSF) and decanoyl-Arg-Arg-Leu-Leu-chloromethyl ketone (dec-RRLL-cmk), inhibitors of SKI-1 (site 1; subtilisin kexin like-1) protease. Because SKI-1 is required for activation of SREBP and CREB (cAMP-response element-binding protein)/ATF family transcription factors, we tested the effect of these inhibitors on gene expression. AEBSF decreased expression of 140 genes by 1.5–3.0-fold including Phex, Dmp1, COL1A1, COL11A1, and fibronectin. Direct comparison of AEBSF and dec-RRLL-cmk, a more specific SKI-1 inhibitor, demonstrated that expression of Phex, Dmp1, COL11A1, and fibronectin was reduced by both, whereas COL1A2 and HMGCS1 were reduced only by AEBSF. AEBSF and dec-RRLL-cmk decreased the nuclear content of SKI-1-activated forms of transcription factors SREBP-1, SREBP-2, and OASIS. In contrast to AEBSF, the actions of dec-RRLL-cmk represent the sum of its direct actions on SKI-1 and indirect actions on caspase-3. Specifically, dec-RRLL-cmk reduced intracellular caspase-3 activity by blocking the formation of activated 19-kDa caspase-3. Conversely, overexpression of SKI-1-activated SREBP-1a and CREB-H in UMR106-01 osteoblastic cells increased the number of mineralized foci and altered their morphology to yield mineralization nodules, respectively. In summary, SKI-1 regulates the activation of transmembrane transcription factor precursors required for expression of key genes required for mineralization of osteoblastic cultures in vitro and bone formation in vivo. Our results indicate that the differentiated phenotype of osteoblastic cells and possibly osteocytes depends upon the non-apoptotic actions of SKI-1. PMID:21075843

Gorski, Jeff P.; Huffman, Nichole T.; Chittur, Sridar; Midura, Ronald J.; Black, Claudine; Oxford, Julie; Seidah, Nabil G.

2011-01-01

198

TBLR1 as an AR coactivator selectively activates AR target genes to inhibit prostate cancer growth  

PubMed Central

Androgen Receptor (AR), a steroid hormone receptor, is critical for prostate cancer growth. However, activation of AR by androgens can also lead to growth suppression and differentiation. Transcriptional cofactors play an important role in this switch between proliferative and anti-proliferative AR target gene programs. TBLR1, a core component of the nuclear receptor corepressor (NCoR) complex, shows both co-repressor and co-activator activities on nuclear receptors, but little is known about its effects on AR and prostate cancer. We characterized TBLR1 as a coactivator of AR in prostate cancer cells and the activation is both phosphorylation and 19S proteosome dependent. We showed that TBLR1 physically interacts with AR and directly occupies the androgen response elements of affected AR target genes in an androgen-dependent manner. TBLR1 is primarily localized in the nucleus in benign prostate cells and nuclear expression is significantly reduced in prostate cancer cells in culture. Similarly, in human tumor samples, the expression of TBLR1 in the nucleus is significantly reduced in the malignant glands compared to the surrounding benign prostatic glands (p<0.005). Stable ectopic expression of nuclear TBLR1 leads to androgen-dependent growth suppression of prostate cancer cells in vitro and in vivo by selective activation of androgen regulated genes associated with differentiation (e.g. KRT18) and growth suppression (e.g. NKX3.1), but not cell proliferation of the prostate. Understanding the molecular switches involved in the transition from AR dependent growth promotion to AR dependent growth suppression will lead to more successful prostate cancer treatments. PMID:24243687

Daniels, Garrett; Li, Yirong; Gellert, Lan Lin; Zhou, Albert; Melamed, Jonathan; Wu, Xinyu; Zhang, Xinming; Zhang, David; Meruelo, Daniel; Logan, Susan K.; Basch, Ross; Lee, Peng

2014-01-01

199

Rad53 Downregulates Mitotic Gene Transcription by Inhibiting the Transcriptional Activator Ndd1  

PubMed Central

The 33 genes in the Saccharomyces cerevisiae mitotic CLB2 transcription cluster have been known to be downregulated by the DNA damage checkpoint for many years. Here, we show that this is mediated by the checkpoint kinase Rad53 and the dedicated transcriptional activator of the cluster, Ndd1. Ndd1 is phosphorylated in response to DNA damage, which blocks recruitment to promoters and leads to the transcriptional downregulation of the CLB2 cluster. Finally, we show that downregulation of Ndd1 is an essential function of Rad53, as a hypomorphic ndd1 allele rescues RAD53 deletion. PMID:24324011

Edenberg, Ellen R.; Vashisht, Ajay; Benanti, Jennifer A.; Wohlschlegel, James

2014-01-01

200

Polygonum cuspidatum, compared with baicalin and berberine, inhibits inducible nitric oxide synthase and cyclooxygenase-2 gene expressions in RAW 264.7 macrophages.  

PubMed

Polygonum cuspidatum water extract (PCWE) was shown to be a potent inhibitor of lipopolysaccharide (LPS)-induced expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). PCWE was compared to baicalin isolated from Scutellaria baicalensis Georgi and berberine of Coptidis rhizoma and Phellodendri cortex, for their effects on LPS-induced nitric oxide (NO) production and iNOS and COX-2 gene expressions in RAW 264.7 macrophages. Both PCWE and the compounds inhibited LPS-induced NO production in a concentration-dependent manner without a cytotoxicity. The decrease in NO production was in parallel with the inhibition of LPS-induced iNOS gene expression by PCWE and the compounds. In contrast, iNOS enzyme activity was not inhibited by PCWE and two agents. In addition, only PCWE inhibited LPS-induced prostaglandin E2 (PGE2) production and COX-2 gene expression without affecting COX-2 enzyme activity, while baicalin or berberine did not. Furthermore, N-nitro-L-arginine (NLA) and N-nitro-L-arginine methyl ester (L-NAME) pretreatment enhanced LPS-induced iNOS protein expression, which was inhibited by these PCWE and two agents, although LPS-induced COX-2 protein expression was not affected by NLA and L-NAME. PCWE inhibited PGE2 production and COX-2 protein expression in NLA/LPS or L-NAME/LPS-co-treated RAW 264.7 cell, however, baicalin or berberine did not. From the results, it was concluded that co-treatment with NOS inhibitors and PCWE effectively blocks acute production of NO and inhibits expression of iNOS and COX-2 genes. PMID:17553752

Kim, Kyung-Woon; Ha, Ki-Tai; Park, Cheol-Soo; Jin, Un-Ho; Chang, Hyen Wook; Lee, In-Seon; Kim, Cheorl-Ho

2007-01-01

201

Predator stress induces behavioral inhibition and amygdala somatostatin receptor 2 gene expression  

PubMed Central

Psychological stressors precipitate and maintain stress-induced psychopathology, and it is likely that altered amygdala function underlies some of the deleterious effects of psychological stress. To understand the mechanisms underlying the linkage between the response to psychological stressors and maladaptive or psychopathological responses, we have focused on amygdala responsivity in animal models employing species-specific psychological stressors. In the present study, we characterized the effects of a 15-min exposure to a natural predator, the ferret, on rat behavior and the expression of the somatostatin family of genes in the amygdala. We examined the somatostatin family of genes because substantial evidence shows that central somatostatin systems are altered in various neuropsychiatric illnesses. We report that rats respond to acute ferret exposure with a significant increase in fearful and anxious behaviors that is accompanied by robust amygdala activation and an increase in somatostatin receptor 2 (sst2) messenger RNA expression within the amygdala and anterior cingulate cortex. These studies are the first to show stress-induced changes in amygdala sst2 expression and may represent one mechanism by which psychological stress is linked to adaptive and maladaptive behavioral responses. PMID:18363859

Nanda, S. A.; Qi, C.; Roseboom, P. H.; Kalin, N. H.

2009-01-01

202

Interferon regulatory factor 1 (IRF-1) mediates cell growth inhibition by transactivation of downstream target genes.  

PubMed Central

Interferon regulatory factor 1 (IRF-1) is a DNA-binding factor which recognizes regulatory elements in the promoters of interferon (IFN)-beta and some IFN-inducible genes. We observed that expression of transfected murine IRF-1 in different mammalian cell lines leads to down-regulation or stop of proliferation depending on the extent of expression. Expression of fusion proteins composed of IRF-1 and the hormone binding domain of the human estrogen receptor does not exhibit IRF-1 activity in the absence of estrogen. However, after estrogen treatment of the cells IFN-beta promoters are activated and the cells stop growing. As shown by expression of IRF-1 mutants both functions of the IRF-1-protein require DNA-binding and transcriptional activation. Since secreted factors including IFNs are not responsible for the anti-proliferative effect of IRF-1 we suggest that IRF-1 may be regarded as a negative regulator of cell growth which acts by activation of down-stream effector genes. Images PMID:8332497

Kirchhoff, S; Schaper, F; Hauser, H

1993-01-01

203

Inducible reduction in pregnancy-associated plasma protein-A gene expression inhibits established atherosclerotic plaque progression in mice.  

PubMed

Pregnancy-associated plasma protein-A (PAPP-A) is a novel zinc metalloproteinase implicated in cardiovascular disease. The aim of this study was to determine whether a reduction in PAPP-A expression in the adult affects the progression of established atherosclerotic plaque. Apolipoprotein E-null mice were fed a high-fat diet for 5 weeks to initiate early-stage plaque development before tamoxifen-inducible, Cre recombinase-mediated excision of the floxed PAPP-A gene. High-fat feeding was continued, and after 10 weeks the aorta and brachiocephalic artery were harvested for atherosclerotic plaque analyses of overall burden and morphology, respectively. An inducible decrease in PAPP-A gene expression significantly inhibited atherosclerotic plaque progression as assessed by a 70% reduction in plaque burden in the aorta (P = .012) without an effect on the elevated circulating levels of cholesterol and triglycerides in this model. Furthermore, this reduction in PAPP-A prevented the development of advanced plaque with necrotic cores and buried fibrous caps in the brachiocephalic artery. These data indicate PAPP-A as a potential target to limit progression of established atherosclerotic plaque. PMID:24506074

Bale, Laurie K; Chakraborty, Suban; Conover, Cheryl A

2014-04-01

204

A novel gene IBF1 is required for the inhibition of brown pigment deposition in rice hull furrows.  

PubMed

The role of flavonoids as the major red, blue, purple and brown pigments in plants has gained these secondary products a great deal of attention over the years. In this study, we characterized a rice inhibitor for brown furrows1 (ibf1) mutant. In the ibf1 mutant, brown pigments specifically accumulate in hull furrows during seed maturation and reach a maximum level in dry seeds. Higher amounts of total flavonoids and anthocyanin in hull may be responsible for the brown pigmentation of ibf1. The IBF1 gene, which encodes a similar kelch repeat-containing F-box protein, was isolated by map-based cloning approach. Real-time RT-PCR and GUS activity assays revealed that IBF1 specifically expressed in reproductive tissues. GFP-IBF1 fusion protein mainly localized in cytoplasm. The expression of some major structural enzymatic genes involved in flavonoids biosynthesis could be up- or down-regulated to some different extent in ibf1 mutant. Our data suggested that IBF1 as a suppressor could inhibit the brown pigmentation of rice hull furrows. PMID:22419106

Shao, Tian; Qian, Qian; Tang, Ding; Chen, Jun; Li, Ming; Cheng, Zhukuan; Luo, Qiong

2012-07-01

205

Systematic screen for mutants resistant to TORC1 inhibition in fission yeast reveals genes involved in cellular ageing and growth.  

PubMed

Target of rapamycin complex 1 (TORC1), which controls growth in response to nutrients, promotes ageing in multiple organisms. The fission yeast Schizosaccharomyces pombe emerges as a valuable genetic model system to study TORC1 function and cellular ageing. Here we exploited the combinatorial action of rapamycin and caffeine, which inhibit fission yeast growth in a TORC1-dependent manner. We screened a deletion library, comprising ?84% of all non-essential fission yeast genes, for drug-resistant mutants. This screen identified 33 genes encoding functions such as transcription, kinases, mitochondrial respiration, biosynthesis, intra-cellular trafficking, and stress response. Among the corresponding mutants, 5 showed shortened and 21 showed increased maximal chronological lifespans; 15 of the latter mutants showed no further lifespan increase with rapamycin and might thus represent key targets downstream of TORC1. We pursued the long-lived sck2 mutant with additional functional analyses, revealing that the Sck2p kinase functions within the TORC1 network and is required for normal cell growth, global protein translation, and ribosomal S6 protein phosphorylation in a nutrient-dependent manner. Notably, slow cell growth was associated with all long-lived mutants while oxidative-stress resistance was not. PMID:24463365

Rallis, Charalampos; López-Maury, Luis; Georgescu, Teodora; Pancaldi, Vera; Bähler, Jürg

2014-01-01

206

Regulation of the grapevine polygalacturonase-inhibiting protein encoding gene: expression pattern, induction profile and promoter analysis.  

PubMed

Regulation of defense in plants is a complex process mediated by various signaling pathways. Promoter analysis of defense-related genes is useful to understand these signaling pathways involved in regulation. To this end, the regulation of the polygalacturonase-inhibiting protein encoding gene from Vitis vinifera L. (Vvpgip1) was analyzed with regard to expression pattern and induction profile as well as the promoter in terms of putative regulatory elements present, core promoter size and the start of transcription. Expression of Vvpgip1 is tissue-specific and developmentally regulated. Vvpgip1 expression was induced in response to auxin, salicylic acid and sugar treatment, wounding and pathogen infection. The start of transcription was mapped to 17 bp upstream of the ATG and the core promoter was mapped to the 137 bp upstream of the ATG. Fructose- and Botrytis responsiveness were identified in the region between positions -3.1 and -1.5 kb. The analyses showed induction in water when the leaves were submersed and this response and the response to wounding mapped to the region between positions -1.1 and -0.1 kb. In silico analyses revealed putative cis-acting elements in these areas that correspond well to the induction stimuli tested. PMID:22932820

Joubert, D Albert; de Lorenzo, Giulia; Vivier, Melané A

2013-03-01

207

Berberine inhibits mouse insulin gene promoter through activation of AMP activated protein kinase and may exert beneficial effect on pancreatic ?-cell.  

PubMed

Berberine is one of the main alkaloids of Rhizoma coptidis, proven to have anti-diabetic potentials through activation of AMP activated protein kinase (AMPK) in liver and muscle. However, the role of berberine on the insulin gene is unknown. Therefore, the effect of berberine on insulin gene transcription was investigated in the present study. Reporter gene assays were used in the mouse ?-cell line NIT-1 to test the effect of berberine on the promoter of mouse insulin gene Ins2. The mRNA and protein levels of insulin were also detected. Diet induced glucose intolerant mice were used to explore the effect of berberine on blood glucose homeostasis and insulin resistance in vivo. The insulin content in islet was semi-quantified by an image analysis software in the immunohistochemistry sections. The results revealed that berberine caused a reversible concentration-dependent inhibition of insulin gene transcription in NIT-1 cells which showed a significant difference from the long term used AMPK activator metformin. Such inhibition on insulin promoter resulted in the reduction of mRNA and protein of insulin. Furthermore, the inhibition of insulin promoter was totally abolished by AMPK inhibitor Compound C. Berberine significantly improved insulin resistance and glucose intolerance of mice. Likewise, insulin content in islets of berberine treated mice was also decreased. Thus, the insulin gene represents a novel target of AMPK that may contribute to the action of berberine in type 2 diabetes mellitus. PMID:22955013

Shen, Ning; Huan, Yi; Shen, Zhu-fang

2012-11-01

208

?-Amylase inhibitor-1 gene from Phaseolus vulgaris expressed in Coffea arabica plants inhibits ?-amylases from the coffee berry borer pest  

PubMed Central

Background Coffee is an important crop and is crucial to the economy of many developing countries, generating around US$70 billion per year. There are 115 species in the Coffea genus, but only two, C. arabica and C. canephora, are commercially cultivated. Coffee plants are attacked by many pathogens and insect-pests, which affect not only the production of coffee but also its grain quality, reducing the commercial value of the product. The main insect-pest, the coffee berry borer (Hypotheneumus hampei), is responsible for worldwide annual losses of around US$500 million. The coffee berry borer exclusively damages the coffee berries, and it is mainly controlled by organochlorine insecticides that are both toxic and carcinogenic. Unfortunately, natural resistance in the genus Coffea to H. hampei has not been documented. To overcome these problems, biotechnological strategies can be used to introduce an ?-amylase inhibitor gene (?-AI1), which confers resistance against the coffee berry borer insect-pest, into C. arabica plants. Results We transformed C. arabica with the ?-amylase inhibitor-1 gene (?-AI1) from the common bean, Phaseolus vulgaris, under control of the seed-specific phytohemagglutinin promoter (PHA-L). The presence of the ?-AI1 gene in six regenerated transgenic T1 coffee plants was identified by PCR and Southern blotting. Immunoblotting and ELISA experiments using antibodies against ?-AI1 inhibitor showed a maximum ?-AI1 concentration of 0.29% in crude seed extracts. Inhibitory in vitro assays of the ?-AI1 protein against H. hampei ?-amylases in transgenic seed extracts showed up to 88% inhibition of enzyme activity. Conclusions This is the first report showing the production of transgenic coffee plants with the biotechnological potential to control the coffee berry borer, the most important insect-pest of crop coffee. PMID:20565807

2010-01-01

209

Overexpression of the vimentin gene in transgenic mice inhibits normal lens cell differentiation  

PubMed Central

To investigate the role of the intermediate filament protein vimentin in the normal differentiation and morphogenesis of the eye lens fiber cells, we generated transgenic mice bearing multiple copies of the chicken vimentin gene. In most cases, the vimentin transgene was overexpressed in the lenses of these animals, reaching up to 10 times the endogenous levels. This high expression of vimentin interfered very strongly with the normal differentiation of the lens fibers. The normal fiber cell denucleation and elongation processes were impaired and the animals developed pronounced cataracts, followed by extensive lens degeneration. The age of appearance and extent of these abnormalities in the different transgenic lines were directly related to the vimentin level. Electron microscopic analysis revealed that the accumulated transgenic protein forms normal intermediate filaments. PMID:2793935

1989-01-01

210

Inhibition of Experimental Liver Cirrhosis in Mice by Telomerase Gene Delivery  

NASA Astrophysics Data System (ADS)

Accelerated telomere loss has been proposed to be a factor leading to end-stage organ failure in chronic diseases of high cellular turnover such as liver cirrhosis. To test this hypothesis directly, telomerase-deficient mice, null for the essential telomerase RNA (mTR) gene, were subjected to genetic, surgical, and chemical ablation of the liver. Telomere dysfunction was associated with defects in liver regeneration and accelerated the development of liver cirrhosis in response to chronic liver injury. Adenoviral delivery of mTR into the livers of mTR-/- mice with short dysfunctional telomeres restored telomerase activity and telomere function, alleviated cirrhotic pathology, and improved liver function. These studies indicate that telomere dysfunction contributes to chronic diseases of continual cellular loss-replacement and encourage the evaluation of ``telomerase therapy'' for such diseases.

Rudolph, Karl Lenhard; Chang, Sandy; Millard, Melissa; Schreiber-Agus, Nicole; DePinho, Ronald A.

2000-02-01

211

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

PubMed Central

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

Durham, Paul L.

2011-01-01

212

BZP, a novel serum-responsive zinc finger protein that inhibits gene transcription.  

PubMed Central

We report the fortuitous isolation of cDNA clones encoding a novel zinc finger DNA-binding protein termed BZP. The protein encoded is 114 kDa and contains eight zinc finger motifs, seven of which are present in two clusters at opposite ends of the molecule. Both finger clusters bound to the 9-bp sequence AAAGGTGCA with apparent Kds of approximately 2.5 nM. Two of the finger motifs within the amino- and carboxy-terminal finger clusters share 63% amino acid identity. BZP inhibited transcription of the herpes simplex virus thymidine kinase promoter when copies of the 9-bp target motif were linked in cis, suggesting that it functions as a transcriptional repressor. BZP mRNA and immunoreactivity were detected in several established cell lines but were most abundant in hamster insulinoma (HIT) cells, the parental source of the cDNAs. In mouse tissues, BZP mRNA and immunoreactivity were identified in cells of the endocrine pancreas, anterior pituitary, and central nervous system. Interestingly, in HIT cells proliferating in culture, BZP immunoreactivity was predominately nuclear in location, whereas it was usually located in the cytoplasm in most neural and neuroendocrine tissues. Serum deprivation of HIT cells caused BZP immunoreactivity to become predominantly cytoplasmic in location and attenuated its inhibitory effect on transcription, thereby suggesting that the both the subcellular location and the function of this protein are modulated by factors in serum. Images PMID:7935395

Franklin, A J; Jetton, T L; Shelton, K D; Magnuson, M A

1994-01-01

213

Inhibition of neurite outgrowth and alteration of cytoskeletal gene expression by sodium arsenite.  

PubMed

Arsenic compounds that are often found in drinking water increase the risk of developmental brain disorders. In this study, we performed live imaging analyses of Neuro-2a cells expressing SCAT3, a caspase-3 cleavage peptide sequence linking two fluorescent proteins; enhanced cyan fluorescence protein (ECFP) and Venus, to determine whether sodium arsenite (NaAsO(2); 0, 1, 5, or 10 ?M) affects both neurite outgrowth and/or induces apoptosis with the same doses and in the same cell cultures. We observed that the area ratio of neurite to cell body in SCAT3-expressing cells was significantly reduced by 5 and 10 ?M NaAsO(2), but not by 1 ?M, although the emission ratio of ECFP to Venus, an endpoint of caspase-3 activity, was not changed. However, cytological assay using apoptotic and necrotic markers resulted in that apoptosis, but not necrosis, was significantly induced in Neuro-2a cells when NaAsO(2) exposure continued after the significant effects of NaAsO(2) on neurite outgrowth were found by live imaging. These results suggested that neurite outgrowth was suppressed by NaAsO(2) prior to NaAsO(2)-induced apoptosis. Next, we examined the effects of NaAsO(2) on cytoskeletal gene expression in Neuro-2a cells. NaAsO(2) increased the mRNA levels of the light and medium subunits of neurofilament and decreased the mRNA levels of tau and tubulin in a dose-dependent manner; no significant effect was found in the mRNA levels of the heavy subunit of neurofilament, microtubule-associated protein 2, or actin. The changes in cytoskeletal gene expression are likely responsible for the inhibitory effects of NaAsO(2) on neurite outgrowth. PMID:23022324

Aung, Kyaw Htet; Kurihara, Ryohei; Nakashima, Shizuka; Maekawa, Fumihiko; Nohara, Keiko; Kobayashi, Tetsuya; Tsukahara, Shinji

2013-01-01

214

A new group of phage anti-CRISPR genes inhibits the type I-E CRISPR-Cas system of Pseudomonas aeruginosa.  

PubMed

CRISPR-Cas systems are one of the most widespread phage resistance mechanisms in prokaryotes. Our lab recently identified the first examples of phage-borne anti-CRISPR genes that encode protein inhibitors of the type I-F CRISPR-Cas system of Pseudomonas aeruginosa. A key question arising from this work was whether there are other types of anti-CRISPR genes. In the current work, we address this question by demonstrating that some of the same phages carrying type I-F anti-CRISPR genes also possess genes that mediate inhibition of the type I-E CRISPR-Cas system of P. aeruginosa. We have discovered four distinct families of these type I-E anti-CRISPR genes. These genes do not inhibit the type I-F CRISPR-Cas system of P. aeruginosa or the type I-E system of Escherichia coli. Type I-E and I-F anti-CRISPR genes are located at the same position in the genomes of a large group of related P. aeruginosa phages, yet they are found in a variety of combinations and arrangements. We have also identified functional anti-CRISPR genes within nonprophage Pseudomonas genomic regions that are likely mobile genetic elements. This work emphasizes the potential importance of anti-CRISPR genes in phage evolution and lateral gene transfer and supports the hypothesis that more undiscovered families of anti-CRISPR genes exist. Finally, we provide the first demonstration that the type I-E CRISPR-Cas system of P. aeruginosa is naturally active without genetic manipulation, which contrasts with E. coli and other previously characterized I-E systems. IMPORTANCE The CRISPR-Cas system is an adaptive immune system possessed by the majority of prokaryotic organisms to combat potentially harmful foreign genetic elements. This study reports the discovery of bacteriophage-encoded anti-CRISPR genes that mediate inhibition of a well-studied subtype of CRISPR-Cas system. The four families of anti-CRISPR genes described here, which comprise only the second group of anti-CRISPR genes to be identified, encode small proteins that bear no sequence similarity to previously studied phage or bacterial proteins. Anti-CRISPR genes represent a newly discovered and intriguing facet of the ongoing evolutionary competition between phages and their bacterial hosts. PMID:24736222

Pawluk, April; Bondy-Denomy, Joseph; Cheung, Vivian H W; Maxwell, Karen L; Davidson, Alan R

2014-01-01

215

Antisense inhibition of gene expression and growth in gram-negative bacteria by cell-penetrating peptide conjugates of peptide nucleic acids targeted to rpoD gene.  

PubMed

Gram-negative bacteria (GNB) cause common and severe hospital- and community-acquired infections with a high incidence of multidrug resistance (MDR) and mortality. The emergence and spread of MDR-GNB strains limit therapeutic options and highlight the need to develop new therapeutic strategies. In this study, the peptide (RXR)(4)XB- and (KFF)(3)K-conjugated peptide nucleic acids (PPNAs) were developed to target rpoD, which encodes an RNA polymerase primary ?(70) that is thought to be essential for bacterial growth. Their antimicrobial activities were tested against different clinical isolates of MDR-GNB in vitro and in infection models. The (RXR)(4)XB- and (KFF)(3)K- conjugated PNAs were bactericidal against different strains of MDR-GNB in concentration-dependent and sequence-selective manner, whereas a PPNA with a scrambled base sequence had no effect on growth. Among tested PPNAs, (RXR)(4)XB conjugate PPNA06 showed more potent and broad spectrum inhibition in multidrug-resistant Escherichia coli, Salmonella enterica, Klebsiella pneumoniae, and Shigella flexneri in vitro and in vivo. The results were associated with suppression of rpoD mRNA and ?(70) expression, as well as ?(70) downstream regulated genes including ftsZ, mazF, prfB, rpoS, seqA, turfB and ygjD. The treatment of PPNA06 on mono- or multiple MDR-GBN infected human gastric mucosal epithelial cells demonstrated the complete inhibition on bacterial growth and no influence on morphology and growth of human cells. Also, PPNA06 did not show the induction of antibiotic resistance as compared with classical antibiotics in GNB. These findings firstly demonstrate that rpoD is potential target for developing antisense antibiotics, and indicate that peptide conjugates of anti-rpoD PNA are active against GNBs in vitro and in vivo. Our results offer a feasible strategy for treating MDR-GNB infections. PMID:22000398

Bai, Hui; You, Yu; Yan, Hua; Meng, Jingru; Xue, Xiaoyan; Hou, Zheng; Zhou, Ying; Ma, Xue; Sang, Guojun; Luo, Xiaoxing

2012-01-01

216

A New Group of Phage Anti-CRISPR Genes Inhibits the Type I-E CRISPR-Cas System of Pseudomonas aeruginosa  

PubMed Central

ABSTRACT CRISPR-Cas systems are one of the most widespread phage resistance mechanisms in prokaryotes. Our lab recently identified the first examples of phage-borne anti-CRISPR genes that encode protein inhibitors of the type I-F CRISPR-Cas system of Pseudomonas aeruginosa. A key question arising from this work was whether there are other types of anti-CRISPR genes. In the current work, we address this question by demonstrating that some of the same phages carrying type I-F anti-CRISPR genes also possess genes that mediate inhibition of the type I-E CRISPR-Cas system of P. aeruginosa. We have discovered four distinct families of these type I-E anti-CRISPR genes. These genes do not inhibit the type I-F CRISPR-Cas system of P. aeruginosa or the type I-E system of Escherichia coli. Type I-E and I-F anti-CRISPR genes are located at the same position in the genomes of a large group of related P. aeruginosa phages, yet they are found in a variety of combinations and arrangements. We have also identified functional anti-CRISPR genes within nonprophage Pseudomonas genomic regions that are likely mobile genetic elements. This work emphasizes the potential importance of anti-CRISPR genes in phage evolution and lateral gene transfer and supports the hypothesis that more undiscovered families of anti-CRISPR genes exist. Finally, we provide the first demonstration that the type I-E CRISPR-Cas system of P. aeruginosa is naturally active without genetic manipulation, which contrasts with E. coli and other previously characterized I-E systems. PMID:24736222

Pawluk, April; Bondy-Denomy, Joseph; Cheung, Vivian H. W.; Maxwell, Karen L.; Davidson, Alan R.

2014-01-01

217

Inhibition of storage pathology in prenatal CLN5-deficient sheep neural cultures by lentiviral gene therapy.  

PubMed

The neuronal ceroid lipofuscinoses (NCLs, Batten disease) are inherited neurodegenerative lysosomal storage diseases caused by mutations in several different genes. Mutations in CLN5 cause a variant late-infantile human disease and some cases of juvenile and adult clinical disease. NCLs also occur in animals, and a flock of New Zealand Borderdale sheep with a CLN5 splice-site mutation has been developed for model studies. Dissociated mixed neural cells from CLN5-deficient foetal sheep brains contained no obvious storage bodies at plating but these accumulated rapidly in culture, mainly in microglial cells and also in neurons and astrocytes. Accumulation was very obvious after a week, as monitored by fluorescent microscopy and immunostaining for subunit c of mitochondrial ATP synthase. Photography at intervals revealed the dynamic nature of the cultures and a flow of storage bodies between cells, specifically the phagocytosis of storage-body containing cells by microglia and incorporation of the storage bodies into the host cells. No storage was observed in cultured control cells. Transduction of cell cultures with a lentiviral vector expressing a C-terminal Myc tagged CLN5 resulted in secretion of post-translationally glycosylated and processed CLN5. Transduction of CLN5-deficient cultures with this construct rapidly reversed storage body accumulation, to less than half in only six days. These results show that storage body accumulation is reversible with enzyme correction and support the use of these cultures for testing of therapeutics prior to whole animal studies. PMID:24269732

Hughes, Stephanie M; Hope, Katie M; Xu, Janet Boyu; Mitchell, Nadia L; Palmer, David N

2014-02-01

218

Resveratrol Inhibits Sodium/Iodide Symporter Gene Expression and Function in Rat Thyroid Cells  

PubMed Central

Resveratrol is a polyphenol found in grapes and berries that has antioxidant, antiproliferative and anti-inflammatory properties. For these reasons, it is available as a dietary supplement, and it is under investigation in several clinical trials. Few data are available regarding the effects of resveratrol on thyroid function. A previous study showed that resveratrol transiently increases iodide influx in FRTL-5 rat thyroid cells. Indeed, this increase arises after short treatment times (6–12 h), and no further effects are seen after 24 h. The aim of the present study was to investigate the effects of resveratrol on iodide uptake and sodium/iodide symporter expression in thyroid cells after longer times of treatment. For this purpose, the effects of resveratrol were evaluated both in vitro and in vivo using the rat thyroid FRTL-5 cell line and Sprague-Dawley rats, respectively. In FRTL-5 cells, resveratrol decreased the sodium/iodide symporter RNA and protein expression as a function of time. Furthermore, resveratrol decreased cellular iodide uptake after 48 h of treatment. The inhibitory effect of resveratrol on iodide uptake was confirmed in vivo in Sprague-Dawley rats. This study demonstrates that with longer-term treatment, resveratrol is an inhibitor of sodium/iodide symporter gene expression and function in the thyroid. These data suggest that resveratrol can act as a thyroid disruptor, which indicates the need for caution as a supplement and in therapeutic use. PMID:25251397

Giuliani, Cesidio; Bucci, Ines; Di Santo, Serena; Rossi, Cosmo; Grassadonia, Antonino; Mariotti, Marianna; Piantelli, Mauro; Monaco, Fabrizio; Napolitano, Giorgio

2014-01-01

219

Metformin inhibits food intake and neuropeptide Y gene expression in the hypothalamus  

PubMed Central

Metformin may reduce food intake and body weight, but the anorexigenic effects of metformin are still poorly understood. In this study, Sprague-Dawley rats were administered a single intracere-broventricular dose of metformin and compound C, in a broader attempt to investigate the regula-tory effects of metformin on food intake and to explore the possible mechanism. Results showed that central administration of metformin significantly reduced food intake and body weight gain, par-ticularly after 4 hours. A reduction of neuropeptide Y expression and induction of AMP-activated protein kinase phosphorylation in the hypothalamus were also observed 4 hours after metformin administration, which could be reversed by compound C, a commonly-used antagonist of AMP-activated protein kinase. Furthermore, metformin also improved lipid metabolism by reducing plasma low-density lipoprotein. Our findings suggest that under normal physiological conditions, central regulation of appetite by metformin is related to a decrease in neuropeptide Y gene expres-sion, and that the activation of AMP-activated protein kinase may simply be a response to the anorexigenic effect of metformin. PMID:25206548

Duan, Yale; Zhang, Rui; Zhang, Min; Sun, Lijuan; Dong, Suzhen; Wang, Gang; Zhang, Jun; Zhao, Zheng

2013-01-01

220

Screening effective short interfering RNA/short hairpin RNA for inhibition of human astrovirus ORF2 gene expression in cultured cells.  

PubMed

In this study, we have evaluated four different 21-nt duplexes of small interfering RNA (siRNA-469, siRNA-852, siRNA-1802 and siRNA-1806) that specifically target the ORF2 gene of human astrovirus (HAstV) in inhibiting HAstV capsid protein expression in transfected BHK-21 cells. Furthermore, fluorescence analysis, real-time quantitative PCR (RT-qPCR) and western blot assays showed that pGPU6/GFP/Neo-shRNA inhibits ORF2 gene expression in Caco2 cells. The results indicate that siRNA/shRNA-469 and siRNA/shRNA-1802 can interfere with capsid protein expression in cell culture, and this provides a powerful tool for the study of HAstV gene functions and the biological properties of the capsid protein. PMID:24162825

Zhao, Wei; Niu, Ke; Liu, Wen-Hui; Zhao, Jian; Jin, Yi-Ming; Sui, Ting-Ting

2014-05-01

221

Characterization of the polydnaviral 'T. rostrale virus' (TrV) gene family: TrV1 expression inhibits in vitro cell proliferation.  

PubMed

Tranosema rostrale ichnovirus (TrIV) is a polydnavirus (PDV) transmitted by the endoparasitic wasp T. rostrale to its host Choristoneura fumiferana during oviposition. PDV genes are expressed in infected caterpillars, causing physiological disturbances that promote the survival of the developing endoparasite. The previously sequenced genome of TrIV contains ~86 genes organized in multigene families and distributed on multiple segments of circular dsDNA. Among these, the 'T. rostrale virus' (TrV) family comprises seven genes that are absent in other PDV genomes examined to date and whose function(s) remain(s) unknown. Here, we initiated a functional analysis of the TrV family using qPCR, transfection and RNAi approaches. TrV family genes were weakly expressed in wasp ovaries, but some displayed high transcript abundance in parasitized caterpillars. Whilst TrV1 was the most highly transcribed TrV gene in infected caterpillars, transcript levels for TrV5 and TrV6 were nearly undetectable, indicating that they may be pseudogenes. Temporal and tissue-specific patterns of transcript abundance were similar for all expressed TrV family genes, indicative of an apparent lack of difference in function or tissue specificity. Infection of Cf-203 and Sf-21 insect cells with TrIV led to a dose-dependent inhibition of cell proliferation with no sign of apoptosis. Whilst similar inhibition was observed following transfection of cells with a cloned genome segment carrying the TrV1 gene, RNA interference targeting TrV1 largely restored cell growth in TrIV-infected cells, indicating that TrV1 expression was responsible for the observed inhibition. We suggest that TrV genes may contribute to host developmental disruption by interfering with host-cell proliferation during parasitism. PMID:23343630

Djoumad, Abdelmadjid; Dallaire, Fréderic; Lucarotti, Christopher J; Cusson, Michel

2013-05-01

222

CK2 Phosphorylates and Inhibits TAp73 Tumor Suppressor Function to Promote Expression of Cancer Stem Cell Genes and Phenotype in Head and Neck Cancer.  

PubMed

Cancer stem cells (CSC) and genes have been linked to cancer development and therapeutic resistance, but the signaling mechanisms regulating CSC genes and phenotype are incompletely understood. CK2 has emerged as a key signal serine/threonine kinase that modulates diverse signal cascades regulating cell fate and growth. We previously showed that CK2 is often aberrantly expressed and activated in head and neck squamous cell carcinomas (HNSCC), concomitantly with mutant (mt) tumor suppressor TP53, and inactivation of its family member, TAp73. Unexpectedly, we observed that classical stem cell genes Nanog, Sox2, and Oct4, are overexpressed in HNSCC with inactivated TAp73 and mtTP53. However, the potential relationship between CK2, TAp73 inactivation, and CSC phenotype is unknown. We reveal that inhibition of CK2 by pharmacologic inhibitors or siRNA inhibits the expression of CSC genes and side population (SP), while enhancing TAp73 mRNA and protein expression. Conversely, CK2 inhibitor attenuation of CSC protein expression and the SP by was abrogated by TAp73 siRNA. Bioinformatic analysis uncovered a single predicted CK2 threonine phosphorylation site (T27) within the N-terminal transactivation domain of TAp73. Nuclear CK2 and TAp73 interaction, confirmed by co-immunoprecipitation, was attenuated by CK2 inhibitor, or a T27A point-mutation of this predicted CK2 threonine phospho-acceptor site of TAp73. Further, T27A mutation attenuated phosphorylation, while enhancing TAp73 function in repressing CSC gene expression and SP cells. A new CK2 inhibitor, CX-4945, inhibited CSC related SP cells, clonogenic survival, and spheroid formation. Our study unveils a novel regulatory mechanism whereby aberrant CK2 signaling inhibits TAp73 to promote the expression of CSC genes and phenotype. PMID:25379016

Lu, Hai; Yan, Carol; Quan, Xin Xin; Yang, Xinping; Zhang, Jialing; Bian, Yansong; Chen, Zhong; Van Waes, Carter

2014-10-01

223

CK2 Phosphorylates and Inhibits TAp73 Tumor Suppressor Function to Promote Expression of Cancer Stem Cell Genes and Phenotype in Head and Neck Cancer12  

PubMed Central

Cancer stem cells (CSC) and genes have been linked to cancer development and therapeutic resistance, but the signaling mechanisms regulating CSC genes and phenotype are incompletely understood. CK2 has emerged as a key signal serine/threonine kinase that modulates diverse signal cascades regulating cell fate and growth. We previously showed that CK2 is often aberrantly expressed and activated in head and neck squamous cell carcinomas (HNSCC), concomitantly with mutant (mt) tumor suppressor TP53, and inactivation of its family member, TAp73. Unexpectedly, we observed that classical stem cell genes Nanog, Sox2, and Oct4, are overexpressed in HNSCC with inactivated TAp73 and mtTP53. However, the potential relationship between CK2, TAp73 inactivation, and CSC phenotype is unknown. We reveal that inhibition of CK2 by pharmacologic inhibitors or siRNA inhibits the expression of CSC genes and side population (SP), while enhancing TAp73 mRNA and protein expression. Conversely, CK2 inhibitor attenuation of CSC protein expression and the SP by was abrogated by TAp73 siRNA. Bioinformatic analysis uncovered a single predicted CK2 threonine phosphorylation site (T27) within the N-terminal transactivation domain of TAp73. Nuclear CK2 and TAp73 interaction, confirmed by co-immunoprecipitation, was attenuated by CK2 inhibitor, or a T27A point-mutation of this predicted CK2 threonine phospho-acceptor site of TAp73. Further, T27A mutation attenuated phosphorylation, while enhancing TAp73 function in repressing CSC gene expression and SP cells. A new CK2 inhibitor, CX-4945, inhibited CSC related SP cells, clonogenic survival, and spheroid formation. Our study unveils a novel regulatory mechanism whereby aberrant CK2 signaling inhibits TAp73 to promote the expression of CSC genes and phenotype. PMID:25379016

Lu, Hai; Yan, Carol; Quan, Xin Xin; Yang, Xinping; Zhang, Jialing; Bian, Yansong; Chen, Zhong; Van Waes, Carter

2014-01-01

224

Overexpression of the PLAP-1 gene inhibits the differentiation of BMSCs into osteoblast-like cells.  

PubMed

Periodontal ligament-associated protein-1 (PLAP-1) is a newly discovered member of the extracellular matrix family of proteins known as proteoglycans and is a negative regulator that plays a crucial role in the homeostasis of periodontal tissues. It can protect the periodontal ligament from excessive osteogenesis. However, the molecular mechanisms of PLAP-1 during osteogenic differentiation and osteogenesis remain unclear. In this study, we constructed a PLAP-1 recombinant retroviral plasmid vector named pBABE-hygro-PLAP-1. We transfected this plasmid into rat bone marrow stromal cells (rBMSCs) to obtain a stable cell line with overexpression of PLAP-1 to verify whether PLAP-1 also acts as an inhibitory factor in rBMSCs during bone mineralization. A rBMSC line stably overexpressing PLAP-1 was established successfully as determined by the mRNA levels of PLAP-1, which were measured by real time-qPCR (RT-qPCR), and protein expression, which was measured by immunocytochemistry and western blot analysis. At the same time, a Cell Counting Kit-8 assay did not reveal any statistically significant changes in the transfected cells (P > 0.05). Then, mineral-inducing cultures were performed, and mineralized nodules were observed at weeks 2, 3 and 4 under a microscope. Alizarin Red (Sigma) staining was performed at 4 week to illustrate calcium accumulation. The mineralized nodules in the PLAP-1-transfected rBMSC group were fewer than those in the control groups. The time span of the formation of the mineralized nodules was prolonged. Meanwhile, osteogenic genes were also detected in the mineral-inducing cells by RT-qPCR. An RT-qPCR analysis demonstrated that the levels of the osteoblast markers of rBMSCs that were transfected with pBABE-hygro-PLAP-1, including Runx2, Osterix, alkaline phosphatase, bone sialoprotein and osteocalcin, were lower than those in the non-transfected rBMSCs and rBMSCs that were transfected with empty vector (P < 0.01). These results suggest that PLAP-1 has an inhibitory function in rBMSCs when they differentiate into osteoblast-like cells. PMID:25038933

Sun, Jing; Zhang, Ting; Zhang, Panpan; Lv, Linlin; Wang, Yanzhi; Zhang, Jing; Li, Shu

2014-10-01

225

Upregulation of CRMP4, a new prostate cancer metastasis suppressor gene, inhibits tumor growth in a nude mouse intratibial injection model.  

PubMed

Prostate cancer, the most commonly diagnosed male cancer in North America, has a high incidence of bone metastasis. Our previous study showed collapsin response mediator protein 4 (CRMP4) gene inhibited prostate cancer migration and invasion. In this study, we investigated whether overexpression of CRMP4 gene in prostate cancer cells inhibit tumor bone metastasis. The stable prostate cancer cells overexpressing the CRMP4 gene were constructed using lentivirus infection. Prostate cancer bone metastasis nude mouse model was built though orthotopic prostate implantation, intracardiac injection and intratibial injection with CRMP4 overexpress and control cancer cells. Small animal PET/CT scanning results showed no difference of bone metastatic capacity in orthotopic and intracardiac injection models between CRMP4 overexpression and control group, while CRMP4 overexpression inhibited tumor growth in the intratibial injection model. Moreover, our in vitro study showed CRMP4 overexpression downregulates the Neuropilin1 (NRP1) expression and upregulate the Noggin expression. Immunohistochemical staining of the hind limbs of intratibial injection model was confirmed with cytological experiments. Taken together, our research indicated CRMP4 inhibits prostate cancer cells growth in the nude mouse bone microenvironment and this effect may relate with regulation of NRP1 and Noggin expression. PMID:25338524

Zhou, Wei; Xie, Peigen; Pang, Mao; Yang, Bu; Fang, Youqiang; Shu, Tao; Liu, Chang; Wang, Xuan; Zhang, Liangming; Li, Shangfu; Rong, Limin

2015-01-01

226

Antisense inhibition of a pectate lyase gene supports a role for pectin depolymerization in strawberry fruit softening  

PubMed Central

Cell wall disassembly in softening fruits is a complex process involving the cumulative action of many families of wall-modifying proteins on interconnected polysaccharide matrices. One strategy to elucidate the in vivo substrates of specific enzymes and their relative importance and contribution to wall modification is to suppress their expression in transgenic fruit. It has been reported previously that inhibiting the expression of pectate lyase genes by antisense technology in strawberry (Fragaria×ananassa Duch.) fruit resulted in prolonged fruit firmness. This suggested that pectin depolymerization might make a more important contribution to strawberry fruit softening than is often stated. In this present study, three independent transgenic lines were identified exhibiting a greater than 90% reduction in pectate lyase transcript abundance. Analyses of sequential cell wall extracts from the transgenic and control fruit collectively showed clear quantitative and qualitative differences in the extractability and molecular masses of populations of pectin polymers. Wall extracts from transgenic fruits showed a reduction in pectin solubility and decreased depolymerization of more tightly bound polyuronides. Additional patterns of differential extraction of other wall-associated pectin subclasses were apparent, particularly in the sodium carbonate- and chelator-soluble polymers. In addition, microscopic studies revealed that the typical ripening-associated loss of cell–cell adhesion was substantially reduced in the transgenic fruits. These results indicate that pectate lyase plays an important degradative role in the primary wall and middle lamella in ripening strawberry fruit, and should be included in synergistic models of cell wall disassembly. PMID:18522930

Santiago-Domenech, Nieves; Jimenez-Bemudez, Silvia; Matas, Antonio J.; Rose, Jocelyn K. C.; Munoz-Blanco, Juan; Mercado, Jose A.; Quesada, Miguel A.

2008-01-01

227

Highly potent dUTPase inhibition by a bacterial repressor protein reveals a novel mechanism for gene expression control.  

PubMed

Transfer of phage-related pathogenicity islands of Staphylococcus aureus (SaPI-s) was recently reported to be activated by helper phage dUTPases. This is a novel function for dUTPases otherwise involved in preservation of genomic integrity by sanitizing the dNTP pool. Here we investigated the molecular mechanism of the dUTPase-induced gene expression control using direct techniques. The expression of SaPI transfer initiating proteins is repressed by proteins called Stl. We found that ?11 helper phage dUTPase eliminates SaPIbov1 Stl binding to its cognate DNA by binding tightly to Stl protein. We also show that dUTPase enzymatic activity is strongly inhibited in the dUTPase:Stl complex and that the dUTPase:dUTP complex is inaccessible to the Stl repressor. Our results disprove the previously proposed G-protein-like mechanism of SaPI transfer activation. We propose that the transfer only occurs if dUTP is cleared from the nucleotide pool, a condition promoting genomic stability of the virulence elements. PMID:25274731

Szabó, Judit E; Németh, Veronika; Papp-Kádár, Veronika; Nyíri, Kinga; Leveles, Ibolya; Bendes, Abris Á; Zagyva, Imre; Róna, Gergely; Pálinkás, Hajnalka L; Besztercei, Balázs; Ozohanics, Olivér; Vékey, Károly; Liliom, Károly; Tóth, Judit; Vértessy, Beáta G

2014-10-29

228

Gene therapy using genetically modified lymphocytes targeting VEGFR-2 inhibits the growth of vascularized syngenic tumors in mice  

PubMed Central

Immunotherapies based on adoptive cell transfer are highly effective in the treatment of metastatic melanoma, but the use of this approach in other cancer histologies has been hampered by the identification of appropriate target molecules. Immunologic approaches targeting tumor vasculature provide a means for the therapy of multiple solid tumor types. We developed a method to target tumor vasculature, using genetically redirected syngeneic or autologous T cells. Mouse and human T cells were engineered to express a chimeric antigen receptor (CAR) targeted against VEGFR-2, which is overexpressed in tumor vasculature and is responsible for VEGF-mediated tumor progression and metastasis. Mouse and human T cells expressing the relevant VEGFR-2 CARs mediated specific immune responses against VEGFR-2 protein as well as VEGFR-2–expressing cells in vitro. A single dose of VEGFR-2 CAR-engineered mouse T cells plus exogenous IL-2 significantly inhibited the growth of 5 different types of established, vascularized syngeneic tumors in 2 different strains of mice and prolonged the survival of mice. T cells transduced with VEGFR-2 CAR showed durable and increased tumor infiltration, correlating with their antitumor effect. This approach provides a potential method for the gene therapy of a variety of human cancers. PMID:20978347

Chinnasamy, Dhanalakshmi; Yu, Zhiya; Theoret, Marc R.; Zhao, Yangbing; Shrimali, Rajeev K.; Morgan, Richard A.; Feldman, Steven A.; Restifo, Nicholas P.; Rosenberg, Steven A.

2010-01-01

229

Orally delivered thioketal nanoparticles loaded with TNF-?-siRNA target inflammation and inhibit gene expression in the intestines  

NASA Astrophysics Data System (ADS)

Small interfering RNAs (siRNAs) directed against proinflammatory cytokines have the potential to treat numerous diseases associated with intestinal inflammation; however, the side-effects caused by the systemic depletion of cytokines demands that the delivery of cytokine-targeted siRNAs be localized to diseased intestinal tissues. Although various delivery vehicles have been developed to orally deliver therapeutics to intestinal tissue, none of these strategies has demonstrated the ability to protect siRNA from the harsh environment of the gastrointestinal tract and target its delivery to inflamed intestinal tissue. Here, we present a delivery vehicle for siRNA, termed thioketal nanoparticles (TKNs), that can localize orally delivered siRNA to sites of intestinal inflammation, and thus inhibit gene expression in inflamed intestinal tissue. TKNs are formulated from a polymer, poly-(1,4-phenyleneacetone dimethylene thioketal), that degrades selectively in response to reactive oxygen species (ROS). Therefore, when delivered orally, TKNs release siRNA in response to the abnormally high levels of ROS specific to sites of intestinal inflammation. Using a murine model of ulcerative colitis, we demonstrate that orally administered TKNs loaded with siRNA against the proinflammatory cytokine tumour necrosis factor-alpha (TNF-?) diminish TNF-? messenger RNA levels in the colon and protect mice from ulcerative colitis.

Wilson, D. Scott; Dalmasso, Guillaume; Wang, Lixin; Sitaraman, Shanthi V.; Merlin, Didier; Murthy, Niren

2010-11-01

230

Strong inhibition of fimbrial 3 subunit gene transcription by a novel downstream repressive element in Bordetella pertussis.  

PubMed

The Bvg-regulated promoters for the fimbrial subunit genes fim2 and fim3 of Bordetella pertussis behave differently from each other both in vivo and in vitro. In vivo?Pfim2 is significantly stronger than Pfim3 , even though predictions based on the DNA sequences of BvgA-binding motifs and core promoter elements would indicate the opposite. In vitro?Pfim3 demonstrated robust BvgA?P-dependent transcriptional activation, while none was seen with Pfim2 . This apparent contradiction was investigated further. By swapping sequence elements we created a number of hybrid promoters and assayed their strength in vivo. We found that, while Pfim3 promoter elements upstream of the +1 transcriptional start site do indeed direct Bvg-activated transcription more efficiently than those of Pfim2 , the overall promoter strength of Pfim3 ?in vivo is reduced due to sequences downstream of +1 that inhibit transcription more than 250-fold. This element, the DRE (downstream repressive element), was mapped to the 15 bp immediately downstream of the Pfim3 +1. Placing the DRE in different promoter contexts indicated that its activity was not specific to fim promoters, or even to Bvg-regulated promoters. However it does appear to be specific to Bordetella species in that it did not function in Escherichia coli. PMID:24963821

Chen, Qing; Boulanger, Alice; Hinton, Deborah M; Stibitz, Scott

2014-08-01

231

Polycomb group gene BMI1 controls invasion of medulloblastoma cells and inhibits BMP-regulated cell adhesion  

PubMed Central

Background Medulloblastoma is the most common intracranial childhood malignancy and a genetically heterogeneous disease. Despite recent advances, current therapeutic approaches are still associated with high morbidity and mortality. Recent molecular profiling has suggested the stratification of medulloblastoma from one single disease into four distinct subgroups namely: WNT Group (best prognosis), SHH Group (intermediate prognosis), Group 3 (worst prognosis) and Group 4 (intermediate prognosis). BMI1 is a Polycomb group repressor complex gene overexpressed across medulloblastoma subgroups but most significantly in Group 4 tumours. Bone morphogenetic proteins are morphogens belonging to TGF-? superfamily of growth factors, known to inhibit medulloblastoma cell proliferation and induce apoptosis. Results Here we demonstrate that human medulloblastoma of Group 4 characterised by the greatest overexpression of BMI1, also display deregulation of cell adhesion molecules. We show that BMI1 controls intraparenchymal invasion in a novel xenograft model of human MB of Group 4, while in vitro assays highlight that cell adhesion and motility are controlled by BMI1 in a BMP dependent manner. Conclusions BMI1 controls MB cell migration and invasion through repression of the BMP pathway, raising the possibility that BMI1 could be used as a biomarker to identify groups of patients who may benefit from a treatment with BMP agonists. PMID:24460684

2014-01-01

232

Gene expression analysis reveals inhibition of radiation-induced TGF?-signaling by hyperbaric oxygen therapy in mouse salivary glands.  

PubMed

A side effect of radiation therapy in the head and neck region is injury to surrounding healthy tissues such as irreversible impaired function of the salivary glands. Hyperbaric oxygen therapy (HBOT) is clinically used to treat radiation-induced damage but its mechanism of action is largely unknown. In this study, we investigated the molecular pathways that are affected by HBOT in mouse salivary glands two weeks after radiation therapy by microarray analysis. Interestingly, HBOT led to significant attenuation of the radiation-induced expression of a set of genes and upstream regulators that are involved in processes such as fibrosis and tissue regeneration. Our data suggest that the TGF?-pathway, which is involved in radiation-induced fibrosis and chronic loss of function after radiation therapy, is affected by HBOT. On the longer term, HBOT reduced the expression of the fibrosis-associated factor ?-smooth muscle actin in irradiated salivary glands. This study highlights the potential of HBOT to inhibit the TGF?-pathway in irradiated salivary glands and to restrain consequential radiation induced tissue injury. PMID:24849810

Spiegelberg, Linda; Swagemakers, Sigrid M A; Van Ijcken, Wilfred F J; Oole, Edwin; Wolvius, Eppo B; Essers, Jeroen; Braks, Joanna A M

2014-01-01

233

Gene Expression Analysis Reveals Inhibition of Radiation-Induced TGF?-Signaling by Hyperbaric Oxygen Therapy in Mouse Salivary Glands  

PubMed Central

A side effect of radiation therapy in the head and neck region is injury to surrounding healthy tissues such as irreversible impaired function of the salivary glands. Hyperbaric oxygen therapy (HBOT) is clinically used to treat radiation-induced damage but its mechanism of action is largely unknown. In this study, we investigated the molecular pathways that are affected by HBOT in mouse salivary glands two weeks after radiation therapy by microarray analysis. Interestingly, HBOT led to significant attenuation of the radiation-induced expression of a set of genes and upstream regulators that are involved in processes such as fibrosis and tissue regeneration. Our data suggest that the TGF?-pathway, which is involved in radiation-induced fibrosis and chronic loss of function after radiation therapy, is affected by HBOT. On the longer term, HBOT reduced the expression of the fibrosis-associated factor ?-smooth muscle actin in irradiated salivary glands. This study highlights the potential of HBOT to inhibit the TGF?-pathway in irradiated salivary glands and to restrain consequential radiation induced tissue injury. PMID:24849810

Spiegelberg, Linda; Swagemakers, Sigrid MA; van IJcken, Wilfred FJ; Oole, Edwin; Wolvius, Eppo B; Essers, Jeroen; Braks, Joanna AM

2014-01-01

234

Inhibition of CD44 gene expression in human skin models, using self-delivery short interfering RNA administered by dissolvable microneedle arrays.  

PubMed

Treatment of skin disorders with short interfering RNA (siRNA)-based therapeutics requires the development of effective delivery methodologies that reach target cells in affected tissues. Successful delivery of functional siRNA to the epidermis requires (1) crossing the stratum corneum, (2) transfer across the keratinocyte membrane, followed by (3) incorporation into the RNA-induced silencing complex. We have previously demonstrated that treatment with microneedle arrays loaded with self-delivery siRNA (sd-siRNA) can achieve inhibition of reporter gene expression in a transgenic mouse model. Furthermore, treatment of human cultured epidermal equivalents with sd-siRNA resulted in inhibition of target gene expression. Here, we demonstrate inhibition of CD44, a gene that is uniformly expressed throughout the epidermis, by sd-siRNA both in vitro (cultured human epidermal skin equivalents) and in vivo (full-thickness human skin equivalents xenografted on immunocompromised mice). Treatment of human skin equivalents with CD44 sd-siRNA markedly decreased CD44 mRNA levels, which led to a reduction of the target protein as confirmed by immunodetection in epidermal equivalent sections with a CD44-specific antibody. Taken together, these results demonstrate that sd-siRNA, delivered by microneedle arrays, can reduce expression of a targeted endogenous gene in a human skin xenograft model. PMID:22480249

Lara, Maria Fernanda; González-González, Emilio; Speaker, Tycho J; Hickerson, Robyn P; Leake, Devin; Milstone, Leonard M; Contag, Christopher H; Kaspar, Roger L

2012-08-01

235

Salicylic acid inhibits gibberellin-induced alpha-amylase expression and seed germination via a pathway involving an abscisic-acid-inducible WRKY gene.  

PubMed

It is well known that abscisic acid (ABA) antagonizes gibberellin (GA)-promoted seed germination. Recent circumstantial evidence suggests that salicylic acid (SA) also inhibits seed germination in maize and Arabidopsis. Our study shows that SA blocks barley seed germination in a dosage dependent manner. As an initial effort to addressing the mechanism controlling the crosstalk of SA, GA and ABA signaling in barley, we studied the regulation of alpha-amylases by SA and a WRKY gene whose expression is modulated by these hormones. Assays of alpha-amylase activity reveal that GA-induced alpha-amylase production in aleurone cells is inhibited by bioactive SA, but not its analogs, 3-hydroxybenzoic acid and 4-hydroxybenzoic acid. This inhibitory effect is unlikely due to repressing alpha-amylase secretion or inhibiting alpha-amylase enzyme activities. Northern blot analyses indicate that SA suppresses GA-induced expression of a barley low pI alpha-amylase gene (Amy32b). Because our previous data indicate that ABA-inducible and GA-suppressible WRKY genes inhibit the expression of alpha-amylase genes in rice, we studied the steady state mRNA levels of a barley WRKY gene, HvWRKY38. The expression of HvWRKY38 in barley aleurone cells is down-regulated by GA, but up-regulated by SA and ABA. However, the regulation of HvWRKY38 by SA appears to be different from that of ABA in term of the kinetics and levels of induction. Over-expression of HvWRKY38 in aleurone cells by particle bombardment blocks GA induction of the Amy32b promoter reporter construct (Amy32b-GUS). Therefore, HvWRKY38 might serve as a converging node of SA and ABA signal pathways involved in suppressing GA-induced seed germination. PMID:17390108

Xie, Zhen; Zhang, Zhong-Lin; Hanzlik, Shane; Cook, Everett; Shen, Qingxi J

2007-06-01

236

Superiority of Combined Phosphodiesterase PDE3/PDE4 Inhibition over PDE4 Inhibition Alone on Glucocorticoid- and Long-Acting ?2-Adrenoceptor Agonist-Induced Gene Expression in Human Airway Epithelial Cells.  

PubMed

Glucocorticoids, also known as corticosteroids, induce effector gene transcription as a part of their anti-inflammatory mechanisms of action. Such genomic effects can be significantly enhanced by long-acting ?2-adrenoceptor agonists (LABAs) and may contribute to the clinical superiority of inhaled corticosteroid (ICS)/LABA combinations in asthma and chronic obstructive pulmonary disease (COPD) over ICSs alone. Using models of cAMP- and glucocorticoid-induced transcription in human bronchial epithelial BEAS-2B cells, we show that combining inhibitors of phosphodiesterase (PDE) 3 and PDE4 provides greater benefits compared with inhibiting either PDE alone. In respect to cAMP-dependent transcription, inhibitors of PDE3 (siguazodan, cilostazol) and PDE4 (rolipram, GSK256066, roflumilast N-oxide) each sensitized to the LABA, formoterol. This effect was magnified by dual PDE3 and PDE4 inhibition. Siguazodan plus rolipram was also more effective at inducing cAMP-dependent transcription than either inhibitor alone. Conversely, the concentration-response curve describing the enhancement of dexamethasone-induced, glucocorticoid response element-dependent transcription by formoterol was displaced to the left by PDE4, but not PDE3, inhibition. Overall, similar effects were described for bona fide genes, including RGS2, CD200, and CRISPLD2. Importantly, the combination of siguazodan plus rolipram prolonged the duration of gene expression induced by formoterol, dexamethasone, or dexamethasone plus formoterol. This was most apparent for RGS2, a bronchoprotective gene that may also reduce the proinflammatory effects of constrictor mediators. Collectively, these data provide a rationale for the use of PDE3 and PDE4 inhibitors in the treatment of COPD and asthma where they may enhance, sensitize, and prolong the effects of LABA/ICS combination therapies. PMID:25324049

BinMahfouz, Hawazen; Borthakur, Bibhusana; Yan, Dong; George, Tresa; Giembycz, Mark A; Newton, Robert

2015-01-01

237

Human natural killer cell line modified with a chimeric immunoglobulin T-cell receptor gene leads to tumor growth inhibition in vivo.  

PubMed

The gene transfer of tumor-specific chimeric immunoglobulin T-cell receptors (cIgTCRs) combining antibody-like specificity with the effector cell function could be an attractive tool in immunotherapy. In this study, we directed the human natural killer (NK) cell line YT to tumor cells by gene transfer of a cIgTCR with specificity against the human carcinoembryonic antigen (CEA). The cIgTCR was constructed of a CEA-specific humanized single-chain Fv antibody fragment fused to the IgG1 Fc domain and the CD3 zeta chain. YT cells were transfected with the cIgTCR gene by electroporation and cIgTCR-expressing cells were enriched by immunoaffinity purification. cIgTCR-expressing YT cells specifically lysed CEA(+) colon carcinoma cell lines, which were resistant to the parental YT cell line. The lysis was not inhibited in the presence of soluble CEA. Receptor gene-modified YT cells retained their CEA-specific cytolytic activity after gamma-irradiation in vitro and inhibited the tumor growth in vivo after adoptive transfer into NOD/SCID mice. This gene-modified NK cell line available in unlimited source might be useful in clinical immunotherapy of CEA(+) cancer. PMID:11960290

Schirrmann, Thomas; Pecher, Gabriele

2002-04-01

238

Molecular mechanism of inhibition of estrogen-induced cathepsin D gene expression by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in MCF-7 cells  

SciTech Connect

This report describes how 17{beta}-estradiol (E2) induces cathepsin D gene expression, but is inhibited by the aryl hydrocarbon receptor by disruption of the estrogen receptor/pBC12/S1/pac plasmid complex by interaction with an overlapping xenobiotic responsive element. It was also determined that 2,3,7,8-tetrachlorobenzo-p-dioxin (TCDD) alone does not affect cathepsin D gene expression but can together with E2 to affect the rate of transcription and levels of immunoreactive protein. 85 refs., 6 figs., 2 tabs.

Krishnan, V.; Porter, W.; Santostefano, M.; Wang, Xiahong [Texas A& M Univ., College Station, TX (United States)] [and others

1995-12-01

239

Glucocorticoids selectively inhibit the transcription of the interleukin 1 beta gene and decrease the stability of interleukin 1 beta mRNA.  

PubMed Central

Transcription of the interleukin 1 beta (IL-1 beta) gene was studied by mRNA hybridization with a cDNA probe in the human promonocytic cell line U-937. Phorbol ester and lipopolysaccharide increased the steady-state level of IL-1 beta mRNA. Glucocorticoids markedly decreased IL-1 beta mRNA levels by two mechanisms. Transcription of the IL-1 gene was inhibited, as shown by in vitro transcription assays with nuclei isolated from glucocorticoid-treated cells. Moreover, kinetic analyses and pulse-labeling of mRNAs showed that glucocorticoids selectively decrease the stability of IL-1 beta mRNA, without affecting the stability of beta-actin and FOS mRNAs. Inhibition of the formation and effects IL-1 is a mechanism by which glucocorticoids can exert antiinflammatory and immunosuppressive effects. Images PMID:3257575

Lee, S W; Tsou, A P; Chan, H; Thomas, J; Petrie, K; Eugui, E M; Allison, A C

1988-01-01

240

Glucocorticoid inhibition of SP-A gene expression in lung type II cells is mediated via the TTF-1-binding element.  

PubMed

Induction of surfactant protein-A (SP-A) gene expression in fetal lung type II cells by cAMP and IL-1 is mediated by increased binding of thyroid transcription factor-1 (TTF-1) and NF-B proteins p50 and p65 to the TTF-1-binding element (TBE) at -183 bp. In type II cell transfections, dexamethasone (Dex) markedly inhibits cAMP-induced expression of rabbit SP-A:human growth hormone (hGH) fusion genes containing as little as 300 bp of the SP-A 5'-flanking sequence. Dex inhibition is blocked by RU-486, suggesting a role of the glucocorticoid receptor (GR). The present study was undertaken to define the mechanisms for GR inhibition of SP-A expression. Cotransfection of primary cultures of type II cells with a GR expression vector abrogated cAMP induction of SP-A promoter activity while, at the same time, causing a 60-fold induction of cotransfected mouse mammary tumor virus (MMTV) promoter. In lung cells transfected with a fusion gene containing three TBEs fused to the basal SP-A promoter, Dex prevented the stimulatory effect of IL-1 on TTF-1 induction of SP-A promoter activity, suggesting that the GR inhibits SP-A promoter activity through the TBE. In gel shift assays using nuclear extracts from human fetal type II cells cultured in the absence or presence of cAMP, Dex markedly reduced binding of nuclear proteins to the TBE and blocked the stimulatory effect of cAMP on TBE-binding activity. Our finding that Dex increased expression of the NF-kappaB inhibitory partner IkappaB-alpha suggests that the decrease in TBE-binding activity may be caused, in part, by GR inhibition of NF-kappaB interaction with this site. PMID:14633512

Alcorn, Joseph L; Islam, Kazi N; Young, Pampee P; Mendelson, Carole R

2004-04-01

241

mda-7/IL-24 expression inhibits breast cancer through upregulation of growth arrest-specific gene 3 (gas3) and disruption of ?1 integrin function.  

PubMed

Melanoma differentiation-associated gene (MDA)-7)/interleukin (IL)-24, a member of the IL-10 family of cytokines, inhibits growth of various human cancer cells, yet the underlying mechanism is largely unknown. Here, we report that mda-7/IL-24 efficiently suppresses the development of rat mammary tumors in vivo. Microarray analysis for genes differentially expressed in rat mammary tumor cells overexpressing MDA-7/IL-24 compared with those that do not express this cytokine identified growth arrest-specific gene-3 (gas3) as a target for mda-7/IL-24. Upregulation of gas3 by mda-7/IL-24 was STAT3 dependent. Induction of gas3 inhibited attachment and proliferation of tumor cells in vitro and in vivo by inhibiting the interaction of ?1 integrin with fibronectin. A mutated GAS3, which is unable to bind ?1 integrin, was also unable to inhibit fibronectin-mediated attachment and cell growth both in adherent and suspension cultures, suggesting that GAS3 exerts its effects through interaction with and regulation of ?1 integrin. Thus, mda-7/IL-24 inhibits breast cancer growth, at least in part, through upregulation of GAS3 and disruption of ?1 integrin function. Importantly, the expression of the mda-7/IL-24 receptor, IL-20R1, is highly correlated with GAS3 expression in human breast cancer (P = 1.02 × 10(-9)), and the incidence of metastases is significantly reduced in patients with HER2(+) breast cancer expressing high-levels of IL-20R1. Together, our results identify a novel MDA-7/IL-24-GAS3-?1integrin-fibronectin signaling pathway that suppresses breast cancer growth and can be targeted for therapy. PMID:23468528

Li, You-Jun; Liu, Guodong; Li, Yanmei; Vecchiarelli-Federico, Laura M; Liu, Jeff C; Zacksenhaus, Eldad; Shan, Sze W; Yang, Burton B; Li, Qi; Dash, Rupesh; Fisher, Paul B; Archer, Michael C; Ben-David, Yaacov

2013-06-01

242

Calreticulin inhibits glucocorticoid– but not cAMP–sensitive expression of tyrosine aminotransferase gene in cultured McA–RH7777 hepatocytes  

Microsoft Academic Search

Calreticulin is a ubiquitously expressed Ca2+ binding protein of the endoplasmic reticulum which inhibits DNA binding and transcriptional activation by steroid hormone receptors. In this study the effects of calreticulin on tyrosine aminotransferase (TAT) gene expression in cultured McA–RH7777 hepatocytes was investigated. McA–RH7777 cells were stably transfected with calreticulin expression vector to generate cells overexpressing the protein. The transcriptional activity

Kimberly Burns; Michal Opas; Marek Michalak

1997-01-01

243

Dynamin inhibition interferes with inflammasome activation and cytokine gene expression in Streptococcus pyogenes-infected human macrophages.  

PubMed

In the present study, we have analysed the ability of Streptococcus pyogenes [Group A streptococcus (GAS)] to activate the NACHT-domain-, leucine-rich repeat- and PYD-containing protein 3 (NALP3) inflammasome complex in human monocyte-derived macrophages and the molecules and signalling pathways involved in GAS-induced inflammatory responses. We focused upon analysing the impact of dynamin-dependent endocytosis and the role of major streptococcal virulence factors streptolysin O (SLO) and streptolysin S (SLS) in the immune responses induced by GAS. These virulence factors are involved in immune evasion by forming pores in host cell membranes, and aid the bacteria to escape from the endosome-lysosome pathway. We analysed cytokine gene expression in human primary macrophages after stimulation with live or inactivated wild-type GAS as well as with live SLO and SLS defective bacteria. Interleukin (IL)-1?, IL-10, tumour necrosis factor (TNF)-? and chemokine (C-X-C motif) ligand (CXCL)-10 cytokines were produced after bacterial stimulation in a dose-dependent manner and no differences in cytokine levels were seen between live, inactivated or mutant bacteria. These data suggest that streptolysins or other secreted bacterial products are not required for the inflammatory responses induced by GAS. Our data indicate that inhibition of dynamin-dependent endocytosis in macrophages attenuates the induction of IL-1?, TNF-?, interferon (IFN)-? and CXCL-10 mRNAs. We also observed that pro-IL-1? protein was expressed and efficiently cleaved into mature-IL-1? via inflammasome activation after bacterial stimulation. Furthermore, we demonstrate that multiple signalling pathways are involved in GAS-stimulated inflammatory responses in human macrophages. PMID:25079511

Latvala, S; Mäkelä, S M; Miettinen, M; Charpentier, E; Julkunen, I

2014-11-01

244

Inhibition of Gene Expression of Organic Cation/Carnitine Transporter and Antioxidant Enzymes in Oxazaphosphorines-Induced Acute Cardiomyopathic Rat Models  

PubMed Central

It is well documented that high therapeutic doses of oxazaphosphorines, cyclophosphamide (CP) and ifosfamide (IFO), are associated with cardiomyopathy. This study investigated whether oxazaphosphorines alter the expression of organic cation/carnitine transporter (OCTN2) and antioxidant genes and if so, whether these alterations contribute to CP and IFO-induced cardiotoxicity. Adult male Wistar albino rats were assigned to one of six treatment groups namely, control, L carnitine, CP, IFO, CP plus L carnitine and IFO plus L carnitine. In cardiac and kidney tissues, CP and IFO significantly decreased mRNA and protein expression of OCTN2. Oxazaphosphorines significantly increased serum acyl-carnitine/free carnitine ratio and urinary carnitine excretion and significantly decreased total carnitine in cardiac tissues. Interestingly, carnitine supplementation completely reversed the biochemical and gene expression changes-induced by oxazaphosphorines to the control values, except OCTN2 expression remained inhibited by IFO. Data from this study suggest that: (1) Oxazaphosphorines decreased myocardial carnitine content following the inhibition of OCTN2 mRNA and protein expression in cardiac tissues. (2) Oxazaphosphorine therapy increased urinary loss of carnitine secondary to the inhibition of OCTN2 mRNA and protein expression in proximal tubules of the kidney. (3) Carnitine supplementation attenuates CP but not IFO-induced inhibition of OCTN2 mRNA and protein expression in heart and kidney tissues. PMID:22701146

Sayed-Ahmed, Mohamed M.; Aldelemy, Meshan Lafi; Hafez, Mohamed M.; Al-Shabanah, Othman A.

2012-01-01

245

Disruption of HDAC4/N-CoR complex by histone deacetylase inhibitors leads to inhibition of IL-2 gene expression.  

PubMed

Previous studies have shown that HDAC inhibitors selectively inhibit IL-2 gene expression, but the mechanism of this inhibition remains to be elucidated. It was recently reported that HDAC4, a component of the nuclear hormone receptor corepressor (N-CoR) complex, associates with the IL-2 promoter via the transcription factor myocyte enhancer factor 2 (MEF2). We therefore focused on the role of HDAC4/N-CoR complex in the transcriptional regulation of IL-2. Four approaches were used to characterize this role and to investigate the relation between the regulatory function of HDAC4/N-CoR complex and HDAC4-enzymatic activity: (i) HDAC4 silencing by RNA interference, (ii) overexpression of N-CoR repression domain 3 (RD3), (iii) overexpression of HDAC4 point mutants, and (iv) treatment with HDAC inhibitors. Here, we report that HDAC4 plays an essential role in IL-2 promoter activation, and that the formation of the HDAC4/N-CoR complex, which is closely related to HDAC4-enzymatic activity, might be involved in HDAC inhibitor-mediated inhibition of IL-2 gene expression. These observations indicate that the selective inhibition of HDAC4 or the interaction of HDAC4 with N-CoR is likely a potential target for the development of novel immunosuppressants. PMID:17559812

Matsuoka, Hideaki; Fujimura, Takao; Hayashi, Masako; Matsuda, Kaori; Ishii, Yoshinori; Aramori, Ichiro; Mutoh, Seitaro

2007-08-01

246

ATF4-dependent regulation of the JMJD3 gene during amino acid deprivation can be rescued in Atf4-deficient cells by inhibition of deacetylation.  

PubMed

Following amino acid deprivation, the amino acid response (AAR) induces transcription from specific genes through a collection of signaling mechanisms, including the GCN2-eIF2-ATF4 pathway. The present report documents that the histone demethylase JMJD3 is an activating transcription factor 4 (ATF4)-dependent target gene. The JMJD3 gene contains two AAR-induced promoter activities and chromatin immunoprecipitation (ChIP) analysis showed that the AAR leads to enhanced ATF4 recruitment to the C/EBP-ATF response element (CARE) upstream of Promoter-1. AAR-induced histone modifications across the JMJD3 gene locus occur upon ATF4 binding. Jmjd3 transcription is not induced in Atf4-knock-out cells, but the AAR-dependent activation was rescued by inhibition of histone deacetylation with trichostatin A (TSA). The TSA rescue of AAR activation in the absence of Atf4 also occurred for the Atf3 and C/EBP homology protein (Chop) genes, but not for the asparagine synthetase gene. ChIP analysis of the Jmjd3, Atf3, and Chop genes in Atf4 knock-out cells documented that activation of the AAR in the presence of TSA led to specific changes in acetylation of histone H4. The results suggest that a primary function of ATF4 is to recruit histone acetyltransferase activity to a sub-set of AAR target genes. Thus, absolute binding of ATF4 to these particular genes is not required and no ATF4 interaction with the general transcription machinery is necessary. The data are consistent with the hypothesis that ATF4 functions as a pioneer factor to alter chromatin structure and thus, enhance transcription in a gene-specific manner. PMID:22955275

Shan, Jixiu; Fu, Lingchen; Balasubramanian, Mukundh N; Anthony, Tracy; Kilberg, Michael S

2012-10-19

247

Phorbol esters selectively and reversibly inhibit a subset of myofibrillar genes responsible for the ongoing differentiation program of chick skeletal myotubes.  

PubMed Central

Phorbol esters selectively and reversibly disassemble the contractile apparatus of cultured skeletal muscle as well as inhibit the synthesis of many contractile proteins without inhibiting that of housekeeping proteins. We now demonstrate that phorbol esters reversibly decrease the mRNA levels of at least six myofibrillar genes: myosin heavy chain, myosin light chain 1/3, myosin light chain 2, cardiac and skeletal alpha-actin, and skeletal troponin T. The steady-state message levels decrease 50- to 100-fold after 48 h of exposure to phorbol esters. These decreases can be attributed at least in part to decreases in transcription rates. For at least two genes, cardiac and skeletal alpha-actin, some of the decreases are the result of increased mRNA turnover. In contrast, the cardiac troponin T steady-state message level does not change, and its transcription rate decreases only transiently upon exposure to phorbol esters. Phorbol esters do not decrease the expression of the housekeeping genes, alpha-tubulin, beta-actin, and gamma-actin. Phorbol esters do not decrease the steady-state message levels of MyoD1, a gene known to be important in the activation of many skeletal muscle-specific genes. Cycloheximide blocks the phorbol ester-induced decreases in transcription, message stability, and the resulting steady-state message level but does not block the tetradecanoyl phorbol acetate-induced rapid disassembly of the I-Z-I complexes. These results suggests a common mechanism for the regulation of many myofibrillar genes independent of MyoD1 mRNA levels, independent of housekeeping genes, but dependent on protein synthesis. Images PMID:1875933

Choi, J K; Holtzer, S; Chacko, S A; Lin, Z X; Hoffman, R K; Holtzer, H

1991-01-01

248

Guava Leaf Extract Inhibits Quorum-Sensing and Chromobacterium violaceum Induced Lysis of Human Hepatoma Cells: Whole Transcriptome Analysis Reveals Differential Gene Expression  

PubMed Central

Quorum sensing (QS) is a process mediated via small molecules termed autoinducers (AI) that allow bacteria to respond and adjust according to the cell population density by altering the expression of multitudinous genes. Since QS governs numerous bioprocesses in bacteria, including virulence, its inhibition promises to be an ideal target for the development of novel therapeutics. We found that the aqueous leaf extract of Psidium guajava (GLE) exhibited anti-QS properties as evidenced by inhibition of violacein production in Chromobacterium violaceum and swarming motility of Pseudomonas aeruginosa. The gram-negative bacterium, C. violaceum is a rare pathogen with high mortality rate. In this study, perhaps for the first time, we identified the target genes of GLE in C. violaceum MTCC 2656 by whole transcriptome analysis on Ion Torrent. Our data revealed that GLE significantly down-regulated 816 genes at least three fold, with p value?0.01, which comprises 19% of the C. violaceum MTCC 2656 genome. These genes were distributed throughout the genome and were associated with virulence, motility and other cellular processes, many of which have been described as quorum regulated in C. violaceum and other gram negative bacteria. Interestingly, GLE did not affect the growth of the bacteria. However, consistent with the gene expression pattern, GLE treated C. violaceum cells were restrained from causing lysis of human hepatoma cell line, HepG2, indicating a positive relationship between the QS-regulated genes and pathogenicity. Overall, our study proposes GLE as a QS inhibitor (QSI) with the ability to attenuate virulence without affecting growth. To the best of our knowledge, this is the first report which provides with a plausible set of candidate genes regulated by the QS system in the neglected pathogen C. violaceum. PMID:25229331

Tiwary, Bipransh Kumar; Kumar, Anoop

2014-01-01

249

A 122.5-kilobase deletion of the P gene underlies the high prevalence of oculocutaneous albinism type 2 in the Navajo population.  

PubMed

Oculocutaneous albinism (OCA) is a genetically heterogeneous disorder. There are four known types of OCA: OCA1-OCA4. The clinical manifestations of all types of OCA include skin and hair hypopigmentation and visual impairment. Although there are a few documented observations of high frequency of albinism among Native Americans, including the Hopi, Zuni, Kuna, Jemez, Laguna, San Juan, and Navajo, no causative molecular defect has been previously reported. In the present study, we show that albinism in one Native American population, the Navajo, is caused by a LINE-mediated 122.5-kilobase deletion of the P gene, thus demonstrating that albinism in this population is OCA2. This deletion appears to be Navajo specific, because this allele was not detected in 34 other individuals with albinism who listed other Native American origins, nor has it been reported in any other ethnic group. The molecular characterization of this deletion allele allowed us to design a three-primer polymerase chain reaction system to estimate the carrier frequency in the Navajo population by screening 134 unrelated normally pigmented Navajos. The carrier frequency was found to be approximately 4.5%. The estimated prevalence of OCA2 in Navajos is between approximately 1 per 1,500 and 1 per 2,000. We further estimate that this mutation originated 400-1,000 years ago from a single founder. PMID:12469324

Yi, Zanhua; Garrison, Nanibaa'; Cohen-Barak, Orit; Karafet, Tatiana M; King, Richard A; Erickson, Robert P; Hammer, Michael F; Brilliant, Murray H

2003-01-01

250

A 122.5-Kilobase Deletion of the P Gene Underlies the High Prevalence of Oculocutaneous Albinism Type 2 in the Navajo Population  

PubMed Central

Oculocutaneous albinism (OCA) is a genetically heterogeneous disorder. There are four known types of OCA: OCA1–OCA4. The clinical manifestations of all types of OCA include skin and hair hypopigmentation and visual impairment. Although there are a few documented observations of high frequency of albinism among Native Americans, including the Hopi, Zuni, Kuna, Jemez, Laguna, San Juan, and Navajo, no causative molecular defect has been previously reported. In the present study, we show that albinism in one Native American population, the Navajo, is caused by a LINE-mediated 122.5-kilobase deletion of the P gene, thus demonstrating that albinism in this population is OCA2. This deletion appears to be Navajo specific, because this allele was not detected in 34 other individuals with albinism who listed other Native American origins, nor has it been reported in any other ethnic group. The molecular characterization of this deletion allele allowed us to design a three-primer polymerase chain reaction system to estimate the carrier frequency in the Navajo population by screening 134 unrelated normally pigmented Navajos. The carrier frequency was found to be ?4.5%. The estimated prevalence of OCA2 in Navajos is between ?1 per 1,500 and 1 per 2,000. We further estimate that this mutation originated 400–1,000 years ago from a single founder. PMID:12469324

Yi, Zanhua; Garrison, Nanibaa'; Cohen-Barak, Orit; Karafet, Tatiana M.; King, Richard A.; Erickson, Robert P.; Hammer, Michael F.; Brilliant, Murray H.

2003-01-01

251

Mechanisms of hormonal regulation of CAD gene expression and inhibition by Aryl hydrocarbon receptor agonist in human breast cancer cells  

E-print Network

The CAD gene is trifunctional and expresses carbamoylphosphate synthetase/aspartate carbamyltransferase/dihydroorotase, which are required for pyrimidine biosynthesis. CAD gene activities are induced in MCF-7 human breast cancer cells, and treatment...

Khan, Shaheen Munawar Ali

2007-04-25

252

Wild-type p53 inhibits pro-invasive properties of TGF-?3 in breast cancer, in part through regulation of EPHB2, a new TGF-? target gene.  

PubMed

The p53 tumor suppressor protein is primarily known for its important role in tumor suppression. In addition, p53 affects tumor cell migration, invasion, and epithelial-mesenchymal transition (EMT); processes also regulated by the transforming growth factor-? (TGF-?) signaling pathway. Here, we investigated the role of p53 in breast tumor cell invasion, migration, and EMT and examined the interplay of p53 with TGF-?3 in these processes. MCF-10A1 and MCF-10CA1a breast cancer cells were treated with Nutlin-3 and TGF-?3, and the effects on tumor cell migration and invasion were studied in transwell and 3D spheroid invasion assays. The effects of Nutlin-3 and TGF-?3 on EMT were examined in NMuMG cells. To identify genes involved in TGF-?-induced invasion that are modulated by p53, a Human Tumor Metastasis-specific RT-PCR array was performed. Verification of EPHB2 regulation by TGF-?3 and p53 was performed on breast cancer tumor cell lines. We demonstrate that p53 inhibits basal and TGF-?3-induced invasion, migration, and EMT in normal breast epithelial and breast cancer cells. Pharmacological activation of p53 inhibited induction of several TGF-?3 targets involved in TGF-?3-induced tumor cell invasion, i.e., matrix metallo proteinase (MMP)2, MMP9, and integrin ? 3 . The ephrin-type B receptor 2 (EPHB2) gene was identified as a new TGF-? target important for TGF-?3-mediated invasion and migration, whose transcriptional activation by TGF-?3 is also inhibited by p53. The results show an intricate interplay between p53 and TGF-?3 whereby p53 inhibits the TGF-?3-induced expression of genes, e.g., EPHB2, to impede tumor cell invasion and migration. PMID:25257729

Lam, Suzanne; Wiercinska, Eliza; Teunisse, Amina F A S; Lodder, Kirsten; Ten Dijke, Peter; Jochemsen, Aart G

2014-11-01

253

Structure-activity relationship studies of naphthol AS-E and its derivatives as anticancer agents by inhibiting CREB-mediated gene transcription  

PubMed Central

CREB (cyclic AMP-response element binding protein) is a downstream transcription factor of a multitude of signaling pathways emanating from receptor tyrosine kinases or G-protein coupled receptors. CREB is not activated until it is phosphorylated at Ser133 and its subsequent binding to CREB-binding protein (CBP) through kinase-inducible domain (KID) in CREB and KID-interacting (KIX) domain in CBP. Tumor tissues from various organs present higher level of expression and activation of CREB. Thus CREB has been proposed as a promising cancer drug target. We previously described naphthol AS-E (1a) as a small molecule inhibitor of CREB-mediated gene transcription in living cells. Here we report the structure–activity relationship (SAR) studies of 1a by modifying the appendant phenyl ring. All the compounds were evaluated for in vitro inhibition of KIX–KID interaction, cellular inhibition of CREB-mediated gene transcription and inhibition of proliferation of four cancer cell lines (A549, MCF-7, MDA-MB-231 and MDA-MB-468). SAR indicated that a small and electron-withdrawing group was preferred at the para-position for KIX–KID interaction inhibition. Compound 1a was selected for further biological characterization and it was found that 1a down-regulated the expression of endogenous CREB target genes. Expression of a constitutively active CREB mutant, VP16-CREB in MCF-7 cells rendered the cells resistant to 1a, suggesting that CREB was critical in mediating its anticancer activity. Furthermore, 1a was not toxic to normal human cells. Collectively, these data support that 1a represents a structural template for further development into potential cancer therapeutics with a novel mechanism of action. PMID:23102993

Li, Bingbing X.; Yamanaka, Kinrin; Xiao, Xiangshu

2012-01-01

254

Grb7 Upregulation Is a Molecular Adaptation to HER2 Signaling Inhibition Due to Removal of Akt-Mediated Gene Repression  

PubMed Central

The efficacy of anti-HER2 therapeutics, such as lapatinib and trastuzumab, is limited by primary and acquired resistance. Cellular adaptations that allow breast cancer cell to survive prolonged HER2 inhibition include de-repression of the transcription factor FOXO3A with consequent estrogen receptor activation, and/or increased HER3 signaling. Here, we used low-density arrays, quantitative PCR, and western blotting to determine how HER2 signaling inhibition with lapatinib or PI3K inhibitors affects the expression of genes involved in breast cancer metastatic spread and overall prognosis. Retroviral transgenesis was used to express constitutively active forms of Akt in the HER2+ breast cancer cell line SKBR3, and Grb7 in MCF7 cells. Specific gene silencing was obtained by siRNAs transfection. A murine BT474 xenograft cancer model was used to assess the effect of lapatinib on gene expression in vivo. We found that lapatinib induces upregulation of Grb7, an adaptor protein involved in receptor tyrosine kinase signaling and promoting cell survival and cell migration. Grb7 upregulation induced by lapatinib was found to occur in cancer cells in vitro and in vivo. We demonstrate that Grb7 upregulation is recreated by PI3K inhibitors while being prevented by constitutively active Akt. Thus, Grb7 is repressed by PI3K signaling and lapatinib-mediated Akt inhibition is responsible for Grb7 de-repression. Finally, we show that Grb7 removal by RNA-interference reduces breast cancer cell viability and increases the activity of lapatinib. In conclusion, Grb7 upregulation is a potentially adverse consequence of HER2 signaling inhibition. Preventing Grb7 accumulation and/or its interaction with receptor tyrosine kinases may increase the benefit of HER2-targeting drugs. PMID:20126311

Nencioni, Alessio; Cea, Michele; Garuti, Anna; Passalacqua, Mario; Raffaghello, Lizzia; Soncini, Debora; Moran, Eva; Zoppoli, Gabriele; Pistoia, Vito; Patrone, Franco; Ballestrero, Alberto

2010-01-01

255

Caprylic acid and medium-chain triglycerides inhibit IL-8 gene transcription in Caco-2 cells: comparison with the potent histone deacetylase inhibitor trichostatin A  

PubMed Central

Medium-chain triglyceride (MCT) is often administered to patients with Crohn's disease (CD) or short-bowel syndrome. However, little is known about the effects of medium-chain fatty acids (MCFAs) and MCT on intestinal inflammation. In this study we examined whether caprylic acid, one of the MCFAs, and MCT suppress IL-8 secretion by differentiated Caco-2 cells.We found for the first time that caprylic acid and MCT suppress IL-8 secretion by Caco-2 cells at the transcriptional level when precultured together for 24 h. We also tried to clarify the mechanism of IL-8 gene inhibition by examining the activation of NF-?B and other transcription factors by electrophoretic mobility shift assay (EMSA), and found that caprylic acid did not modulate their activation.The result of dual-luciferase assay using Caco-2 cells transfected with IL-8 promoter/luciferase reporter plasmid revealed that caprylic acid inhibited the activation of IL-8 promoter.Similar results were observed when cells were precultured with the well-known potent histone deacetylase inhibitor trichostatin A (TSA).We examined the state of H4 acetylation in IL-8 promoter using the technique known as chromatin immunoprecipitation (Chr-IP). TSA rapidly induced H4 acetylation in IL-8 promoter chromatin, whereas caprylic acid did not. These results suggest that the inhibition of IL-8 gene transcription induced by caprylic acid and TSA does not necessarily require the marked suppression of transcription factors, and the mechanism of inhibition of IL-8 gene transcription may be different between caprylic acid and TSA. PMID:12010777

Hoshimoto, Aihiro; Suzuki, Yasuo; Katsuno, Tatsuro; Nakajima, Hiroshi; Saito, Yasushi

2002-01-01

256

NCYM, a Cis-Antisense Gene of MYCN, Encodes a De Novo Evolved Protein That Inhibits GSK3? Resulting in the Stabilization of MYCN in Human Neuroblastomas  

PubMed Central

The rearrangement of pre-existing genes has long been thought of as the major mode of new gene generation. Recently, de novo gene birth from non-genic DNA was found to be an alternative mechanism to generate novel protein-coding genes. However, its functional role in human disease remains largely unknown. Here we show that NCYM, a cis-antisense gene of the MYCN oncogene, initially thought to be a large non-coding RNA, encodes a de novo evolved protein regulating the pathogenesis of human cancers, particularly neuroblastoma. The NCYM gene is evolutionally conserved only in the taxonomic group containing humans and chimpanzees. In primary human neuroblastomas, NCYM is 100% co-amplified and co-expressed with MYCN, and NCYM mRNA expression is associated with poor clinical outcome. MYCN directly transactivates both NCYM and MYCN mRNA, whereas NCYM stabilizes MYCN protein by inhibiting the activity of GSK3?, a kinase that promotes MYCN degradation. In contrast to MYCN transgenic mice, neuroblastomas in MYCN/NCYM double transgenic mice were frequently accompanied by distant metastases, behavior reminiscent of human neuroblastomas with MYCN amplification. The NCYM protein also interacts with GSK3?, thereby stabilizing the MYCN protein in the tumors of the MYCN/NCYM double transgenic mice. Thus, these results suggest that GSK3? inhibition by NCYM stabilizes the MYCN protein both in vitro and in vivo. Furthermore, the survival of MYCN transgenic mice bearing neuroblastoma was improved by treatment with NVP-BEZ235, a dual PI3K/mTOR inhibitor shown to destabilize MYCN via GSK3? activation. In contrast, tumors caused in MYCN/NCYM double transgenic mice showed chemo-resistance to the drug. Collectively, our results show that NCYM is the first de novo evolved protein known to act as an oncopromoting factor in human cancer, and suggest that de novo evolved proteins may functionally characterize human disease. PMID:24391509

Suenaga, Yusuke; Islam, S. M. Rafiqul; Alagu, Jennifer; Kaneko, Yoshiki; Kato, Mamoru; Tanaka, Yukichi; Kawana, Hidetada; Hossain, Shamim; Matsumoto, Daisuke; Yamamoto, Mami; Shoji, Wataru; Itami, Makiko; Shibata, Tatsuhiro; Nakamura, Yohko; Ohira, Miki; Haraguchi, Seiki; Takatori, Atsushi; Nakagawara, Akira

2014-01-01

257

Inhibition of ?-Defensin Gene Expression in Airway Epithelial Cells by Low Doses of Residual Oil Fly Ash is Mediated by Vanadium  

PubMed Central

Poor ambient air quality is associated with increased morbidity and mortality, including respiratory infections. However, its effects on various host-defense mechanisms are poorly understood. This study utilized an in vitro model to study the effect of particulate matter (PM2.5) on one antimicrobial mechanism of host defense in the airway, ?-defensin-2 and its bovine homologue, tracheal antimicrobial peptide (TAP) induction in response to lipopolysaccharide (LPS) and IL-1?. Our model utilized cultured primary bovine tracheal epithelial (BTE) cells and the human alveolar type II epithelial cell line, A549, treated with 0–20 ?g/cm2 residual oil fly ash (ROFA) for 6 h. The cells were then washed and stimulated for 18 h with 100 ng/ml LPS or for 6 h with 100 ng/ml IL-1?. ROFA inhibited the LPS-induced increase in TAP mRNA and protein without inducing significant cytotoxicity. As little as 2.5 ?g/cm2 of ROFA inhibited LPS-induced TAP gene expression by 30%. The inhibitory activity was associated with the soluble fraction and not the washed particle. The activity in the leachate was attributed to vanadium, but not nickel or iron. SiO2 and TiO2 were utilized as controls and did not inhibit LPS induction of TAP gene expression in BTE. ROFA also inhibited the increase of IL-1?–induced human ?-defensin-2, a homologue of TAP, in A549 cells. The results show that ROFA, V2O5, and VOSO4 inhibit the ability of airway epithelial cells to respond to inflammatory stimuli at low, physiologically relevant doses and suggest that exposure to these agents could result in an impairment of defense against airborne pathogens. PMID:16641320

Klein-Patel, Marcia E.; Diamond, Gill; Boniotto, Michele; Saad, Sherif; Ryan, Lisa K.

2007-01-01

258

© 2010 Molecular Vision Spectrum of candidate gene mutations associated with Indian familial oculocutaneous and ocular albinism  

E-print Network

Purpose: Albinism is a group of genetic disorders, showing a broad spectrum of different phenotypes. The purpose of this study was to screen known candidate genes for oculocutaneous albinism (OCA) and ocular albinism (OA) mutations in Indian patients. Methods: Blood samples were collected from 23 probands and 13 affected family members from 23 genetically unrelated Indian families (22 diagnosed as OCA and 1 diagnosed as OA) and analyzed by bidirectional DNA sequencing of the classic OCA genes — tyrosinase (TYR, or oculocutaneous albinism IA), pink eyed dilution (P; or oculocutaneous albinism II (OCA2]), tyrosinase-related protein 1 (TYRP1), solute carrier family 45, member 2 (SLC45A2; or membrane-associated transporter protein [MATP])—and the OA1 gene, G protein-coupled receptor 143 (GPR143). Results: Three missense mutations, c. 715 C>T (R239W), c. 896 G>A (R299H), c.1255 G>A (G419R), and one termination c. 832 C>T (R278X), were identified in TYR, as well as one novel mutation, c.1453 G>A (G485R) in P. One novel single nucleotide polymorphism (SNP) was identified in both TYR and P; few reported SNPs were identified. The G>A base substitution caused relatively conservative amino acid changes, which altered glycine to arginine residues within the topological domain. The novel OCA2 mutation was not present in 100 control samples. This study identified two probands carrying mutations alone, 16 probands carrying SNPs alone, 4 probands carrying both mutations and SNPs and only one proband carrying neither mutations nor SNPs.

Kathirvel Renugadevi; Asim Kumar Sil; Vijayalakshmi Perumalsamy; Periasamy Sundaresan

259

Heat shock selectively inhibits ribosomal RNA gene transcription and down-regulates E1BF/Ku in mouse lymphosarcoma cells.  

PubMed

The effect of heat shock on RNA polymerase I (pol I)-directed transcription of the rRNA gene was studied in S-100 extract derived from mouse lymphosarcoma cells, and by in vivo labelling of rRNA. Exposure of cells to 42 degrees C for 2 h resulted in complete inhibition of rRNA synthesis in vivo. Pol I transcription was inhibited by 50% within 2 h of heat shock and was abolished after 3 h exposure at 42 degrees C. Under this condition, the core-promoter-binding activity of the factor (CPBF) that modulates pol I transcription was unaffected. In contrast, the promoter-binding activity of enhancer-1-binding factor, a protein related to the Ku autoantigen, which is involved in pol I transcription initiation, was reduced by 50 and 90% after 2 and 3 h of heat shock respectively. Western-blot analysis with antibodies specific for the two subunits of Ku protein showed the absence of p72 subunit after 3 h of heat shock. Under this condition, pol II transcription from the adenovirus major late promoter and pol III transcription of 5 S RNA gene remained unaffected. Mixing experiments ruled out the possibility that the inhibition of transcription was due to activation of nucleases or other inhibitors. This is the first report to show selective down-regulation of pol I transcription in vitro by heat shock and of the potential involvement of a pol I transcription factor in this process. PMID:8760351

Ghoshal, K; Jacob, S T

1996-08-01

260

St. John's Wort increases brain serotonin synthesis by inhibiting hepatic tryptophan 2, 3 dioxygenase activity and its gene expression in stressed rats.  

PubMed

We aimed to investigate the effects of herbal St. John's Wort (SJW) on transcriptional regulation of hepatic tryptophan 2, 3 - dioxygenase (TDO) enzyme activity and brain regional serotonin (5-HT) levels in rats exposed to forced swim test (FST). TDO mRNA expression was quantified using real-time reverse transcription polymerase chain (RT-PCR) reaction and brain regional indoleamines were determined by high performance liquid chromatography coupled to fluorescence detector. Behavioral analysis shows significant reduction in immobility time in SJW (500mg/kg/ml) administered rats. It was found that pretreatment of SJW to rats did not prevent stress-induced elevation in plasma corticosterone levels however it increases serotonin synthesis by virtue of inhibiting hepatic TDO enzyme activity and its gene expression, ascertaining the notion that there exists an inverse relationship between hepatic TDO enzyme activity and brain 5-HT. The drug also decreases serotonin turnover in all the brain areas (hypothalamus, hippocampus amygdala) in stressed rats endorsing its monoamine oxidase inhibition property. Inhibition of TDO enzyme activity and its gene expression by the drug provides new insights for the development of therapeutic interventions for stress related mental illnesses. PMID:25176236

Bano, Samina; Ara, Iffat; Saboohi, Kausar; Moattar, Tariq; Chaoudhry, Bushra

2014-09-01

261

Hexokinase-mediated sugar signaling controls expression of the calcineurin B-like interacting protein kinase 15 gene and is perturbed by oxidative phosphorylation inhibition.  

PubMed

Calcineurin B-like (CBL) interacting protein kinase 15 (CIPK15) is a newly identified positive regulator which is critical to directing the O(2) deficiency signal to the sugar signaling cascade as part of Amy3D (representative Amy3 gene) regulation in rice. It is located upstream and probably contributes to reserve mobilization under anoxia. In isolated starving embryos, the temporal pattern of accumulation of CIPK15 transcripts and leaky suppression of this gene suggests that factors other than CIPK15 may also be involved in the regulation of Amy3D expression. Probing of a variety of sugars and sugar analogs has shown that hexokinase mediates the sugar regulation of CIPK15. For example, hexokinase substrates, such as mannose, 2-deoxyglucose, and other metabolizable sugars, repressed CIPK15 expression, whereas 3-O-methylglucose and 6-deoxyglucose did not. By using glucosamine, a hexokinase inhibitor, to release glucose-dependent CIPK15 suppression, we confirmed that hexokinase mediates regulation of this gene. Chemical inhibitors of mitochondrial electron transfer, proton separation or ATP synthase also effectively abolished sugar-induced repression of CIPK15. This type of interference, the release from glucose-induced repression of gene expression by inhibition of oxidative phosphorylation, was previously identified for the Amy3D gene, which suggests that hexokinase-mediated sugar signaling may be coordinated with the cellular energy status. Analysis of a transgenic rice cell line harboring the GUS reporter gene under the control of the CIPK15 promoter, and transient expression assay for 3' UTR of the CIPK15 gene indicate that sugar regulation of the rice CIPK15 gene is likely mediated by 2548-bp 5'-flanking region, with no additional post-transcriptional control. PMID:22796010

Yim, Hui-Kyeong; Lim, Mi-Na; Lee, Sung-Eun; Lim, Jun; Lee, Yew; Hwang, Yong-Sic

2012-10-15

262

Herpes simplex virus 1 late gene expression is preferentially inhibited during infection of the TAF250 mutant ts13 cell line.  

PubMed

A key component of the polymerase II transcription machinery is the transcription factor TFIID, a complex that contains the TATA-box binding protein and several (10-12) associated factors designated as TAFs (TBP-associated factors). ts13 cells, which contain a temperature-sensitive mutant in TAF250, the largest subunit of TFIID, exhibit promoter-specific defects in gene expression at the nonpermissive temperature, suggesting that individual TAFs are required for transcription of specific subsets of eukaryotic genes. Herpes simplex virus 1 (HSV-1) does not replicate in ts13 cells at the nonpermissive temperature, but the point at which the replicative process is blocked is not known. We used the TAF250 defect in ts13 cells to investigate the role of TAF250 in the expression of HSV-1 genes of each temporal class. At a low m.o.i., expression of most immediate-early mRNAs is reduced at the nonpermissive temperature, and consequently, there is little expression of early genes and no viral DNA replication. In contrast, at high m.o.i., expression of immediate-early genes is unaffected by the TAF250 defect and is not dependent on de novo viral protein synthesis. Early genes and early proteins are produced under these conditions, and viral DNA replication ensues, albeit at somewhat reduced levels. In contrast, late gene expression and late protein synthesis are severely restricted, even in the presence of appreciable viral DNA replication. Thus the lack of late protein synthesis is responsible for the inability of HSV-1 to replicate in ts13 cells at the nonpermissive temperature. Further, it appears that late viral gene expression may be preferentially inhibited by the TAF250 mutation in ts13 cells. PMID:10772991

Dhar, S; Weir, J P

2000-04-25

263

Metformin inhibits epithelial-mesenchymal transition in prostate cancer cells: involvement of the tumor suppressor miR30a and its target gene SOX4.  

PubMed

Tumor metastasis is the leading cause of mortality and morbidity of prostate cancer (PCa) patients. Epithelial-mesenchymal transition (EMT) plays a critical role in cancer progression and metastasis. Recent evidence suggested that diabetic patients treated with metformin have lower PCa risk and better prognosis. This study was aimed to investigate the effects of metformin on EMT in PCa cells and the possible microRNA (miRNA)-based mechanisms. MiRNAs have been shown to regulate various processes of cancer metastasis. We herein showed that metformin significantly inhibits proliferation of Vcap and PC-3 cells, induces G0/G1 cell cycle arrest and inhibits invasiveness and motility capacity of Vcap cells. Metformin could inhibit TGF-?-induced EMT in Vcap cells, as manifested by inhibition of the increase of N-cadherin (p=0.013), Vimentin (p=0.002) and the decrease of E-cadherin (p=0.0023) and ?-catenin (p=0.034) at mRNA and protein levels. Notably, we demonstrated significant upregulation of miR30a levels by metformin (P<0.05) and further experiments indicated that miR30a significantly inhibits proliferation and EMT process of Vcap cells. Interestingly, we identified that SOX4, a previously reported oncogenic transcriptional factor and modulator of EMT, is a direct target gene of miR30a. Finally, we screened the expression of miR30a and SOX4 in 84 PCa cases with radical prostatectomy. Of note, SOX4 overexpression is significantly associated with decreased levels of miR30a in PCa cases. In all, our study suggested that inhibition of EMT by metformin in PCa cells may involve upregulation of miR30a and downregulation of SOX4. PMID:25201727

Zhang, Jing; Shen, Chengwu; Wang, Lin; Ma, Quanping; Xia, Pingtian; Qi, Mei; Yang, Muyi; Han, Bo

2014-09-26

264

The EWS/FLI1 oncogenic protein inhibits expression of the Wnt inhibitor DICKKOPF-1 gene and antagonizes beta-catenin/TCF-mediated transcription.  

PubMed

Tumours of the Ewing family, which comprise Ewing's sarcoma and peripheral primitive neuroectodermal tumours, are highly aggressive and mostly affect children and adolescents. They are characterized by chromosomal translocations leading to the generation of fusion proteins between EWS (or very rarely FUS) and members of the E-twenty-six (ETS) family of transcription factors that are capable of transforming cells. EWS/FLI1, the most frequent fusion, is thought to cause transformation through activation or repression of specific target genes. We present evidence demonstrating that the Wnt inhibitor and beta-catenin/T-cell factor (TCF)-responsive gene DICKKOPF-1 (DKK-1) is a transcriptional target of EWS/FLI1, which can inhibit both basal and beta-catenin-induced transactivation of the DKK-1 promoter. Moreover, our data indicate that EWS/FLI1 has a more general effect on beta-catenin/TCF-mediated transcription since it can block transactivation of a consensus beta-catenin/TCF reporter construct. Consistently, Ewing tumour cells expressing different EWS/ETS translocations cannot engage beta-catenin/TCF-dependent transcription, whereas silencing of EWS/FLI1 restores beta-catenin responsiveness in A673 and RD-ES Ewing tumour cells. Accordingly, gene set enrichment analysis shows that beta-catenin/TCF target genes are significantly enriched among genes downregulated by EWS/FLI1 in the Ewing cell line A673. Mechanistically, the inhibitory effect of EWS/FLI1 can be overcome by a constitutively active TCF4 protein (TCF4-VP16). Moreover, EWS/FLI1 binds lymphoid enhancer factor 1, a TCF family member, and interferes with its binding to beta-catenin, which could explain its negative effect on beta-catenin/TCF-mediated transcription. Our results show that EWS/FLI1 inhibits both DKK-1 expression as well as beta-catenin/TCF-dependent transcription, which could contribute to progression of tumours of the Ewing family. PMID:20019092

Navarro, Diego; Agra, Noelia; Pestaña, Angel; Alonso, Javier; González-Sancho, José M

2010-03-01

265

Transcriptional up-regulation of antioxidant genes by PPAR? inhibits angiotensin II-induced premature senescence in vascular smooth muscle cells.  

PubMed

This study evaluated peroxisome proliferator-activated receptor (PPAR) ? as a potential target for therapeutic intervention in Ang II-induced senescence in human vascular smooth muscle cells (hVSMCs). Activation of PPAR? by GW501516, a specific agonist of PPAR?, significantly inhibited the Ang II-induced premature senescence of hVSMCs. Agonist-activated PPAR? suppressed the generation of Ang II-triggered reactive oxygen species (ROS) with a concomitant reduction in DNA damage. Notably, GW501516 up-regulated the expression of antioxidant genes, such as glutathione peroxidase 1, thioredoxin 1, manganese superoxide dismutase and heme oxygenase 1. siRNA-mediated down-regulation of these antioxidant genes almost completely abolished the effects of GW501516 on ROS production and premature senescence in hVSMCs treated with Ang II. Taken together, the enhanced transcription of antioxidant genes is responsible for the PPAR?-mediated inhibition of premature senescence through sequestration of ROS in hVSMCs treated with Ang II. PMID:21352808

Kim, Hyo Jung; Ham, Sun Ah; Paek, Kyung Shin; Hwang, Jung Seok; Jung, Si Young; Kim, Min Young; Jin, Hanna; Kang, Eun Sil; Woo, Im Sun; Kim, Hye Jung; Lee, Jae Heun; Chang, Ki Churl; Han, Chang Woo; Seo, Han Geuk

2011-03-25

266

Ethyl acetate extract from Jiedu Xiaozheng Yin inhibits the proliferation of human hepatocellular carcinoma cells by suppressing polycomb gene product Bmi1 and Wnt/?-catenin signaling.  

PubMed

Jiedu Xiaozheng Yin (JXY) is a Chinese herbal decoction used to treat hepatocellular carcinoma (HCC). Previous studies have demonstrated that JXY can inhibit HCC cell proliferation via induction of G0/G1 phase arrest. In this study, we investigated whether the inhibitory effect of JXY on HCC cells is associated with the inhibition of the Wnt/??catenin pathway and the polycomb gene product Bmi1. Ethyl acetate extract from JXY (EE-JXY) was prepared. Methyl thiazolyl tetrazolium (MTT) and colony formation assays were used to measure cell proliferation. Immunofluorescence was used to analyze the expression and location of ?-catenin and Bmi1. Immunohistochemistry was used to examine the expression of proliferating cell nuclear antigen (PCNA), c-myc and cyclin D1. ?-catenin, Bmi1, c-myc, cyclin D1 and p16INK4A mRNA levels were detected by RT-PCR. The results demonstrated that EE-JXY inhibited the expression of PCNA, c-myc, cyclin D1 and Bmi1, and upregulated the expression of p16INK4A. We also found that EE-JXY could facilitate ?-catenin translocation from the cytoplasm and nuclei to the cytomembrane. Finally, suppression of cell proliferation and expression of Bmi1 and Wnt/?-catenin by EE-JXY was confirmed in a mouse xenograft model of HCC. Thus, EE-JXY can inhibit the proliferation of HCC partially via suppression of the Bmi1 and Wnt/?-catenin signaling pathways. PMID:25333742

Chen, Xu-Zheng; Cao, Zhi-Yun; Li, Jin-Nong; Hu, Hai-Xia; Zhang, You-Quan; Huang, Yun-Mei; Liu, Zhi-Zhen; Hu, Dan; Liao, Lian-Ming; Du, Jian

2014-12-01

267

Tandemly Duplicated Arabidopsis Genes That Encode Polygalacturonase-Inhibiting Proteins Are Regulated Coordinately by Different Signal Transduction Pathways in Response to Fungal Infection  

PubMed Central

Polygalacturonase-inhibiting proteins (PGIPs) are plant proteins that counteract fungal polygalacturonases, which are important virulence factors. Like many other plant defense proteins, PGIPs are encoded by gene families, but the roles of individual genes in these families are poorly understood. Here, we show that in Arabidopsis, two tandemly duplicated PGIP genes are upregulated coordinately in response to Botrytis cinerea infection, but through separate signal transduction pathways. AtPGIP2 expression is mediated by jasmonate and requires COI1 and JAR1, whereas AtPGIP1 expression is upregulated strongly by oligogalacturonides but is unaffected by salicylic acid, jasmonate, or ethylene. Both AtPGIP1 and AtPGIP2 encode functional inhibitors of polygalacturonase from Botrytis, and their overexpression in Arabidopsis significantly reduces Botrytis disease symptoms. Therefore, gene duplication followed by the divergence of promoter regions may result in different modes of regulation of similar defensive proteins, thereby enhancing the likelihood of defense gene activation during pathogen infection. PMID:12509524

Ferrari, Simone; Vairo, Donatella; Ausubel, Frederick M.; Cervone, Felice; De Lorenzo, Giulia

2003-01-01

268

Breast Tumors with Elevated Expression of 1q Candidate Genes Confer Poor Clinical Outcome and Sensitivity to Ras/PI3K Inhibition  

PubMed Central

Genomic aberrations are common in cancers and the long arm of chromosome 1 is known for its frequent amplifications in breast cancer. However, the key candidate genes of 1q, and their contribution in breast cancer pathogenesis remain unexplored. We have analyzed the gene expression profiles of 1635 breast tumor samples using meta-analysis based approach and identified clinically significant candidates from chromosome 1q. Seven candidate genes including exonuclease 1 (EXO1) are consistently over expressed in breast tumors, specifically in high grade and aggressive breast tumors with poor clinical outcome. We derived a EXO1 co-expression module from the mRNA profiles of breast tumors which comprises 1q candidate genes and their co-expressed genes. By integrative functional genomics investigation, we identified the involvement of EGFR, RAS, PI3K / AKT, MYC, E2F signaling in the regulation of these selected 1q genes in breast tumors and breast cancer cell lines. Expression of EXO1 module was found as indicative of elevated cell proliferation, genomic instability, activated RAS/AKT/MYC/E2F1 signaling pathways and loss of p53 activity in breast tumors. mRNA–drug connectivity analysis indicates inhibition of RAS/PI3K as a possible targeted therapeutic approach for the patients with activated EXO1 module in breast tumors. Thus, we identified seven 1q candidate genes strongly associated with the poor survival of breast cancer patients and identified the possibility of targeting them with EGFR/RAS/PI3K inhibitors. PMID:24147022

Viveka Thangaraj, Soundara; Periasamy, Jayaprakash; Bhaskar Rao, Divya; Barnabas, Georgina D.; Raghavan, Swetha; Ganesan, Kumaresan

2013-01-01

269

Inhibition of interferon gene activation by death-effector domain-containing proteins from the molluscum contagiosum virus.  

PubMed

Apoptosis, NF-?B activation, and IRF3 activation are a triad of intrinsic immune responses that play crucial roles in the pathogenesis of infectious diseases, cancer, and autoimmunity. FLIPs are a family of viral and cellular proteins initially found to inhibit apoptosis and more recently to either up- or down-regulate NF-?B. As such, a broad role for FLIPs in disease regulation is postulated, but exactly how a FLIP performs such multifunctional roles remains to be established. Here we examine FLIPs (MC159 and MC160) encoded by the molluscum contagiosum virus, a dermatotropic poxvirus causing skin infections common in children and immunocompromised individuals, to better understand their roles in viral pathogenesis. While studying their molecular mechanisms responsible for NF-?B inhibition, we discovered that each protein inhibited IRF3-controlled luciferase activity, identifying a unique function for FLIPs. MC159 and MC160 each inhibited TBK1 phosphorylation, confirming this unique function. Surprisingly, MC159 coimmunoprecipitated with TBK1 and IKK? but MC160 did not, suggesting that these homologs use distinct molecular mechanisms to inhibit IRF3 activation. Equally surprising was the finding that the FLIP regions necessary for TBK1 inhibition were distinct from those MC159 or MC160 regions previously defined to inhibit NF-?B or apoptosis. These data reveal previously unappreciated complexities of FLIPs, and that subtle differences within the conserved regions of FLIPs possess distinct molecular and structural fingerprints that define crucial differences in biological activities. A future comparison of mechanistic differences between viral FLIP proteins can provide new means of precisely manipulating distinct aspects of intrinsic immune responses. PMID:24379396

Randall, Crystal M H; Biswas, Sunetra; Selen, Catherine V; Shisler, Joanna L

2014-01-14

270

Inhibition of interferon gene activation by death-effector domain-containing proteins from the molluscum contagiosum virus  

PubMed Central

Apoptosis, NF-?B activation, and IRF3 activation are a triad of intrinsic immune responses that play crucial roles in the pathogenesis of infectious diseases, cancer, and autoimmunity. FLIPs are a family of viral and cellular proteins initially found to inhibit apoptosis and more recently to either up- or down-regulate NF-?B. As such, a broad role for FLIPs in disease regulation is postulated, but exactly how a FLIP performs such multifunctional roles remains to be established. Here we examine FLIPs (MC159 and MC160) encoded by the molluscum contagiosum virus, a dermatotropic poxvirus causing skin infections common in children and immunocompromised individuals, to better understand their roles in viral pathogenesis. While studying their molecular mechanisms responsible for NF-?B inhibition, we discovered that each protein inhibited IRF3-controlled luciferase activity, identifying a unique function for FLIPs. MC159 and MC160 each inhibited TBK1 phosphorylation, confirming this unique function. Surprisingly, MC159 coimmunoprecipitated with TBK1 and IKK? but MC160 did not, suggesting that these homologs use distinct molecular mechanisms to inhibit IRF3 activation. Equally surprising was the finding that the FLIP regions necessary for TBK1 inhibition were distinct from those MC159 or MC160 regions previously defined to inhibit NF-?B or apoptosis. These data reveal previously unappreciated complexities of FLIPs, and that subtle differences within the conserved regions of FLIPs possess distinct molecular and structural fingerprints that define crucial differences in biological activities. A future comparison of mechanistic differences between viral FLIP proteins can provide new means of precisely manipulating distinct aspects of intrinsic immune responses. PMID:24379396

Randall, Crystal M. H.; Biswas, Sunetra; Selen, Catherine V.; Shisler, Joanna L.

2014-01-01

271

Novel Dominant-Negative Mutation Within the Six Domain of the Conserved Eye Specification Gene sine oculis Inhibits Eye Development in Drosophila  

PubMed Central

The development of the compound eye of Drosophila is controlled, in part, by the concerted actions of several nuclear proteins that form an intricate regulatory system. One member of this network is sine oculis (so), the founding member of the Six gene family. Mutations within so affect the entire visual system, including the compound eye. The vertebrate homologs Six3 and Six6 also appear to play crucial roles in retinal formation. Mutations in Six3 inhibit retinal formation in chickens and fish, whereas those in Six6 are the underlying cause of bilateral anophthalmia in humans. Together, these phenotypes suggest a conserved role for the Six genes in eye development. In this report, we describe the effects of a dominant-negative mutation of sine oculis on the development of the compound eye of Drosophila. The mutation resides within the Six domain and may have implications for eye development and disease. PMID:15704100

Roederer, Kristin; Cozy, Loralyn; Anderson, Jason; Kumar, Justin P.

2009-01-01

272

Targeted gene delivery in tumor xenografts by the combination of ultrasound-targeted microbubble destruction and polyethylenimine to inhibit survivin gene expression and induce apoptosis  

PubMed Central

Background Noninvasive and tissue-specific technologies of gene transfection would be valuable in clinical gene therapy. This present study was designed to determine whether it could enhance gene transfection in vivo by the combination of ultrasound-targeted microbubble destruction (UTMD) with polyethylenimine (PEI) in tumor xenografts, and illuminate the effects of gene silencing and apoptosis induction with short hairpin RNA (shRNA) interference therapy targeting human survivin by this novel technique. Methods Two different expression vectors (pCMV-LUC and pSIREN) were incubated with PEI to prepare cationic complexes (PEI/DNA) and confirmed by the gel retardation assay. Human cervical carcinoma (Hela) tumors were planted subcutaneously in both flanks of nude mice. Tumor-bearing mice were administered by tail vein with PBS, plasmid, plasmid and SonoVue microbubble, PEI/DNA and SonoVue microbubble. One tumor was exposed to ultrasound irradiation, while the other served as control. The feasibility of targeted delivery and tissue specificity facilitated by UTMD and PEI were investigated. Moreover, immunohistochemistry analyses about gene silencing and apoptosis induction were detected. Results Electrophoresis experiment revealed that PEI could condense DNA efficiently. The application of UTMD significantly increases the tissue transfection. Both expression vectors showed that gene expressions were present in all sections of tumors that received ultrasound exposure but not in control tumors. More importantly, the increases in transgene expression were related to UTMD with the presence of PEI significantly. Silencing of the survivin gene could induce apoptosis effectively by downregulating survivin and bcl-2 expression, also cause up-regulation of bax and caspase-3 expression. Conclusions This noninvasive, novel combination of UTMD with PEI could enhance targeted gene delivery and gene expression in tumor xenografts at intravenous administration effectively without causing any apparently adverse effect, and might be a promising candidate for gene therapy. Silencing of survivin gene expression with shRNA could be facilitated by this non-viral technique, and lead to significant cell apoptosis. PMID:21092274

2010-01-01

273

Inhibition of all-trans retinoic acid on MDM2 gene expression in astrocytoma cell line SHG-44  

Microsoft Academic Search

Objective  To investigate the impact of all-trans retinoic acid (ATRA) on MDM2 gene expression in astrocytoma cell line SHG-44, and to provide basic data for further research on the progression mechanism\\u000a and gene therapy of human astrocytoma.\\u000a \\u000a \\u000a \\u000a Methods  The differential expressions of MDM2 gene and protein in SHG-44 cells were detected by cDNA microarray and Western blot, respectively, before and after treatment

Yi Zeng; Zhong Yang; Xiao-Dong Long; Chao You

2008-01-01

274

RB2/p130 gene-enhanced expression down-regulates vascular endothelial growth factor expression and inhibits angiogenesis in vivo.  

PubMed

Angiogenesis is an essential step in the progression of tumor formation and development. The switch to an angiogenetic phenotype can occur as a distinct step before progression to a neoplastic phenotype and is linked to genetic changes such as mutations in key cell cycle regulatory genes. The pathogenesis of the angiogenetic phenotype may involve the inactivation of tumor suppressor genes such as the "guardian of the genome," p53, and the cyclin-dependent kinase inhibitor p16. Retinoblastoma family member RB2/p130 encodes a cell cycle regulatory protein and has been found mutated in different tumor types. Overexpression of RB2/p130 not only suppresses tumor formation in nude mice but also causes regression of established tumor grafts, suggesting that RB2/p130 may modulate the angiogenetic balance. We found that induction of RB2/p130 expression using a tetracycline-regulated gene expression system as well as retroviral and adenoviral-mediated gene delivery inhibited angiogenesis in vivo. This correlated with pRb2/p130-mediated down-regulation of vascular endothelial growth factor protein expression both in vitro and in vivo. PMID:11212232

Claudio, P P; Stiegler, P; Howard, C M; Bellan, C; Minimo, C; Tosi, G M; Rak, J; Kovatich, A; De Fazio, P; Micheli, P; Caputi, M; Leoncini, L; Kerbel, R; Giordano, G G; Giordano, A

2001-01-15

275

Inhibition of systemic onset of post-transcriptional gene silencing by non-toxic concentrations of cadmium  

E-print Network

of cadmium Shoko Ueki and Vitaly Citovsky* Department of Biochemistry and Cell Biology, State University±plant interactions. To better understand this process, the heavy metal cadmium was identi®ed as a speci®c inhibitor in two different PTGS systems, constitutive and inducible. The pattern of cadmium-induced inhibition

Citovsky, Vitaly

276

In vivo gene transfer: prevention of neointima formation by inhibition of mitogen-activated protein kinase kinase  

Microsoft Academic Search

Background The mitogen- activated protein kinase kinase (MAPKK) is a protein downstream ras which is rapidly activated in cells stimulated with various extracellular signals. These proteins are believed to play a pivotal role in integrating and transmitting transmembrane signals required for cell growth. Methods and Results To study the effect of inhibition of MAPKK on smooth muscle cell (SMC) prolifera-

C. Indolfi; E. V. Avvedimento; A. Rapacciuolo; G. Esposito; E. Di Lorenzo; A. Leccia; A. Pisani; A. Chieffo; A. Coppola; M. Chiariello

1997-01-01

277

Targeted nonviral gene-based inhibition of G?i/o-mediated vagal signaling in the posterior left atrium decreases vagal-induced atrial fibrillation  

PubMed Central

BACKGROUND Pharmacologic and ablative therapies for atrial fibrillation (AF) have suboptimal efficacy. Newer gene-based approaches that target specific mechanisms underlying AF are likely to be more efficacious in treating AF. Parasympathetic signaling appears to be an important contributor to AF substrate. OBJECTIVE The purpose of this study was to develop a nonviral gene-based strategy to selectively inhibit vagal signaling in the left atrium and thereby suppress vagal-induced AF. METHODS In eight dogs, plasmid DNA vectors (minigenes) expressing G?i C-terminal peptide (G?ictp) was injected in the posterior left atrium either alone or in combination with minigene expressing G?octp, followed by electroporation. In five control dogs, minigene expressing scrambled peptide (G?Rctp) was injected. Vagal- and carbachol-induced left atrial effective refractory periods (ERPs), AF inducibility, and G?i/octp expression were assessed 3 days following minigene delivery. RESULTS Vagal stimulation- and carbachol-induced effective refractory period shortening and AF inducibility were significantly attenuated in atria receiving a G?i2ctp-expressing minigene and were nearly eliminated in atria receiving both G?i2ctp- and G?o1ctp-expressing minigenes. CONCLUSION Inhibition of both Gi and Go proteins is necessary to abrogate vagal-induced AF in the left atrium and can be achieved via constitutive expression of G?i/octps expressed by nonviral plasmid vectors delivered to the posterior left atrium. PMID:21689540

Aistrup, Gary L.; Cokic, Ivan; Ng, Jason; Gordon, David; Koduri, Hemanth; Browne, Suzanne; Arapi, Dorina; Segon, Yogita; Goldstein, Jacob; Angulo, Abigail; Wasserstrom, J. Andrew; Goldberger, Jeffrey J.; Kadish, Alan H.; Arora, Rishi

2012-01-01

278

Effective inhibition of human cytomegalovirus gene expression and replication by a ribozyme derived from the catalytic RNA subunit of RNase P from Escherichia coli  

PubMed Central

A sequence-specific ribozyme (M1GS RNA) derived from the catalytic RNA subunit of RNase P from Escherichia coli was used to target the overlapping exon 3 region of the mRNAs encoding the major transcription regulatory proteins IE1 and IE2 of human cytomegalovirus. A reduction of more than 80% in the expression levels of IE1 and IE2 and a reduction of about 150-fold in viral growth were observed in human cells that stably expressed the ribozyme. In contrast, a reduction of less than 10% in the IE1/IE2 expression and viral growth was observed in cells that either did not express the ribozyme or produced a “disabled” ribozyme that carried mutations that abolished its catalytic activity. Examination of the expression of several other viral early and late genes in the cells that expressed the M1GS ribozyme further revealed an overall reduction of at least 80% in their expression. These results are consistent with the notion that the antiviral effects in these cells are due to the fact that the ribozyme specifically inhibits the expression of IE1 and IE2 and, consequently, abolishes the expression of viral early and late genes as well as viral growth. Our study is the first, to our knowledge, to use M1GS ribozyme for inhibiting human cytomegalovirus replication and demonstrates the utility of this ribozyme for antiviral applications. PMID:10811889

Trang, Phong; Lee, Manfred; Nepomuceno, Edward; Kim, Joe; Zhu, Hua; Liu, Fenyong

2000-01-01

279

(-)-Xanthatin up-regulation of the GADD45? tumor suppressor gene in MDA-MB-231 breast cancer cells: Role of topoisomerase II? inhibition and reactive oxygen species  

PubMed Central

Previously, we reported that (?)-xanthatin, a naturally occurring xanthanolide present in the Cocklebur plant, exhibits potent anti-proliferative effects on human breast cancer cells, accompanied by an induction of the growth arrest and DNA damage-inducible gene 45? (GADD45?), recognized recently as a novel tumor suppressor gene. However, the mechanisms mediating this activation were unknown. Topoisomerase II? (Topo II?) inhibition has been reported to produce a cell death response accompanied by an atypical DNA laddering fragmentation profile, similar to that noted previously for (–)-xanthatin. Therefore we hypothesized that (?)-xanthatin’s GADD45? activation was mediated through the Topo II? pathway. Here, we identify that (?)-xanthatin does function as a catalytic inhibitor of Topo II?, promoting DNA damage. In addition, reactive oxygen species (ROS) were elevated in cells treated with this agent. Mechanistically, it was determined that the induced levels of GADD45? mRNA resulting from (?)-xanthatin exposures were stabilized by coordinately produced ROS, and that the consequent induction of GADD45? mRNA, GADD45? protein and ROS generation were abrogated by co-treatment with N-acetyl-l-cysteine. Taken together, the data support the concept that Topo II? inhibition by (?)-xanthatin is a trigger that stimulates expression of DNA damage-inducible GADD45? mRNA and that concomitantly produced ROS act downstream to further enhance the GADD45? mRNA/GADD45? protein induction process, resulting in breast cancer cell death. PMID:23313378

Takeda, Shuso; Noguchi, Momoko; Matsuo, Kazumasa; Yamaguchi, Yasuhiro; Kudo, Taichi; Nishimura, Hajime; Okamoto, Yoshiko; Amamoto, Toshiaki; Shindo, Mitsuru; Omiecinski, Curtis J.; Aramaki, Hironori

2014-01-01

280

Short-hairpin RNA-mediated Heat shock protein 90 gene silencing inhibits human breast cancer cell growth in vitro and in vivo  

SciTech Connect

Highlights: Black-Right-Pointing-Pointer Hsp90 is over-expressed in human breast cancer. Black-Right-Pointing-Pointer The shRNA-mediated gene silencing of Hsp90 resulted in inhibition of cell growth. Black-Right-Pointing-Pointer Akt and NF-kB were down-regulation after transfection due to Hsp90 silencing. Black-Right-Pointing-Pointer The tumor growth ratio was decline due to Hsp90 silencing. Black-Right-Pointing-Pointer The PCNA expression was down-regulation due to Hsp90 silencing. -- Abstract: Hsp90 interacts with proteins that mediate signaling pathways involved in the regulation of essential processes such as proliferation, cell cycle control, angiogenesis and apoptosis. Hsp90 inhibition is therefore an attractive strategy for blocking abnormal pathways that are crucial for cancer cell growth. In the present study, the role of Hsp90 in human breast cancer MCF-7 cells was examined by stably silencing Hsp90 gene expression with an Hsp90-silencing vector (Hsp90-shRNA). RT-PCR and Western blot analyses showed that Hsp90-shRNA specifically and markedly down-regulated Hsp90 mRNA and protein expression. NF-kB and Akt protein levels were down-regulated in Hsp90-shRNA transfected cells, indicating that Hsp90 knockout caused a reduction of survival factors and induced apoptosis. Treatment with Hsp90-shRNA significantly increased apoptotic cell death and caused cell cycle arrest in the G1/S phase in MCF-7 cells, as shown by flow cytometry. Silencing of Hsp90 also reduced cell viability, as determined by MTT assay. In vivo experiments showed that MCF-7 cells stably transfected with Hsp90-shRNA grew slowly in nude mice as compared with control groups. In summary, the Hsp90-shRNA specifically silenced the Hsp90 gene, and inhibited MCF-7 cell growth in vitro and in vivo. Possible molecular mechanisms underlying the effects of Hsp90-shRNA include the degradation of Hsp90 breast cancer-related client proteins, the inhibition of survival signals and the upregulation of apoptotic pathways. shRNA-mediated interference may have potential therapeutic utility in human breast cancer.

Zuo, Keqiang [Department of General Surgery, Shanghai 10th People's Hospital, Tongji University School of Medicine, Shanghai 200072 (China)] [Department of General Surgery, Shanghai 10th People's Hospital, Tongji University School of Medicine, Shanghai 200072 (China); Li, Dan [Department of Nuclear Medicine, Shanghai 10th People's Hospital, Tongji University School of Medicine, Shanghai 200072 (China)] [Department of Nuclear Medicine, Shanghai 10th People's Hospital, Tongji University School of Medicine, Shanghai 200072 (China); Pulli, Benjamin [Center for Systems Biology, Massachusetts General Hospital, Harvard Medical School, 185 Cambridge Street, Boston, MA 02114 (United States)] [Center for Systems Biology, Massachusetts General Hospital, Harvard Medical School, 185 Cambridge Street, Boston, MA 02114 (United States); Yu, Fei; Cai, Haidong; Yuan, Xueyu [Department of Nuclear Medicine, Shanghai 10th People's Hospital, Tongji University School of Medicine, Shanghai 200072 (China)] [Department of Nuclear Medicine, Shanghai 10th People's Hospital, Tongji University School of Medicine, Shanghai 200072 (China); Zhang, Xiaoping, E-mail: zxpsibs@163.com [Department of Nuclear Medicine, Shanghai 10th People's Hospital, Tongji University School of Medicine, Shanghai 200072 (China)] [Department of Nuclear Medicine, Shanghai 10th People's Hospital, Tongji University School of Medicine, Shanghai 200072 (China); Lv, Zhongwei, E-mail: heyixue163@163.com [Department of Nuclear Medicine, Shanghai 10th People's Hospital, Tongji University School of Medicine, Shanghai 200072 (China)] [Department of Nuclear Medicine, Shanghai 10th People's Hospital, Tongji University School of Medicine, Shanghai 200072 (China)

2012-05-04

281

Curcumin-induced DNA damage and inhibited DNA repair genes expressions in mouse-rat hybrid retina ganglion cells (N18).  

PubMed

Curcumin is reported to be a potent inhibitor of the initiation and promotion of many cancer cells. We investigated to examine whether or not curcumin induce DNA damage in mouse-rat hybrid retina ganglion cell line N18 cells. The Comet assay showed that incubation of N18 cells with 10, 25 and 30 microM of curcumin led to a longer DNA migration smear (Comet tail). The DNA gel electrophoresis showed that 20 microM of curcumin for 24 and 48 h treatment induced DNA damage and fragments in N18 cells. The real time PCR analysis showed that 20 microM of curcumin for 48 h treatment decreased ATM, ATR, BRCA1, 14-3-3sigma, DNA-PK and MGMT mRNA, and ATM and MGMT mRNA expression were inhibited in a time-dependent manner. Our results indicate that curcumin caused DNA damage and inhibited DNA repair genes which may be the factors for curcumin-inhibited cell growth. PMID:19263217

Lu, Hsu-Feng; Yang, Jai-Sing; Lai, Kuang-Chi; Hsu, Shu-Chun; Hsueh, Shu-Ching; Chen, Yuan-Liang; Chiang, Jo-Hua; Lu, Chi-Cheng; Lo, Chyi; Yang, Mei-Due; Chung, Jing-Gung

2009-08-01

282

Induction of mammary differentiation by mammary-derived growth inhibitor-related gene that interacts with an omega-3 fatty acid on growth inhibition of breast cancer cells.  

PubMed

We previously identified and characterized a novel tumor growth inhibitor and a fatty acid-binding protein in human mammary gland and named it the mammary-derived growth inhibitor-related gene (MRG). Here, the effects of MRG on mammary gland differentiation and its interaction with omega-3 polyunsaturated fatty acids (omega-3 PUFAs) on growth inhibition were investigated. MRG protein expression was associated with human mammary gland differentiation, with the highest expression observed in the differentiated alveolar mammary epithelial cells from the lactating gland. Overexpression of MRG in human breast cancer cells induced differentiation with changes in cellular morphology and a significant increase in the production of lipid droplets. Treatment of mouse mammary gland in organ culture with MRG protein resulted in a differentiated morphology and stimulation of beta-casein expression. Treatment of human breast cancer cells with the omega-3 PUFA docosahexaenoic acid resulted in a differential growth inhibition proportional to their MRG expression. MRG-transfected cells or MRG protein treated cells were much more sensitive to docosahexaenoic acid-induced growth inhibition than MRG-negative or untreated control cells. Our results suggest that MRG is a candidate mediator of the differentiating effect of pregnancy on breast epithelial cells and may play a major role in omega-3 PUFA-mediated tumor suppression. PMID:11103817

Wang, M; Liu, Y E; Ni, J; Aygun, B; Goldberg, I D; Shi, Y E

2000-11-15

283

Inhibiting eukaryotic transcription  

PubMed Central

This review first discusses ways in which we can evaluate transcription inhibition, describe changes in nuclear structure due to transcription inhibition, and report on genes that are paradoxically stimulated by transcription inhibition. Next, it summarizes the characteristics and mechanisms of commonly used inhibitors: ?-amanitin is highly selective for RNAP II and RNAP III but its action is slow, actinomycin D is fast but its selectivity is poor, CDK9 inhibitors such as DRB and flavopiridol are fast and reversible but many genes escape transcription inhibition. New compounds, such as triptolide, are fast and selective and able to completely arrest transcription by triggering rapid degradation of RNAP II. PMID:21922053

2011-01-01

284

Transcriptional and Posttranscriptional Regulation of Cytokine Gene Expression in HIV-1 Antigen-Specific CD8+ T Cells That Mediate Virus Inhibition  

PubMed Central

ABSTRACT The ability of CD8+ T cells to effectively limit HIV-1 replication and block HIV-1 acquisition is determined by the capacity to rapidly respond to HIV-1 antigens. Understanding both the functional properties and regulation of an effective CD8+ response would enable better evaluation of T cell-directed vaccine strategies and may inform the design of new therapies. We assessed the antigen specificity, cytokine signature, and mechanisms that regulate antiviral gene expression in CD8+ T cells from a cohort of HIV-1-infected virus controllers (VCs) (<5,000 HIV-1 RNA copies/ml and CD4+ lymphocyte counts of >400 cells/?l) capable of soluble inhibition of HIV-1. Gag p24 and Nef CD8+ T cell-specific soluble virus inhibition was common among the VCs and correlated with substantial increases in the abundance of mRNAs encoding the antiviral cytokines macrophage inflammatory proteins MIP-1?, MIP-1?P (CCL3L1), and MIP-1?; granulocyte-macrophage colony-stimulating factor (GM-CSF); lymphotactin (XCL1); tumor necrosis factor receptor superfamily member 9 (TNFRSF9); and gamma interferon (IFN-?). The induction of several of these mRNAs was driven through a coordinated response of both increased transcription and stabilization of mRNA, which together accounted for the observed increase in mRNA abundance. This coordinated response allows rapid and robust induction of mRNA messages that can enhance the CD8+ T cells' ability to inhibit virus upon antigen encounter. IMPORTANCE We show that mRNA stability, in addition to transcription, is key in regulating the direct anti-HIV-1 function of antigen-specific memory CD8+ T cells. Regulation at the level of RNA helps enable rapid recall of memory CD8+ T cell effector functions for HIV-1 inhibition. By uncovering and understanding the mechanisms employed by CD8+ T cell subsets with antigen-specific anti-HIV-1 activity, we can identify new strategies for comprehensive identification of other important antiviral genes. This will, in turn, enhance our ability to inhibit virus replication by informing both cure strategies and HIV-1 vaccine designs that aim to reduce transmission and can aid in blocking HIV-1 acquisition. PMID:24899193

Payne, Tamika L.; Blackinton, Jeff; Frisbee, Alyse; Pickeral, Joy; Sawant, Sheetal; Vandergrift, Nathan A.; Freel, Stephanie A.; Ferrari, Guido; Keene, Jack D.

2014-01-01

285

The E3L and K3L vaccinia virus gene products stimulate translation through inhibition of the double-stranded RNA-dependent protein kinase by different mechanisms.  

PubMed Central

Vaccinia virus has evolved multiple mechanisms to counteract the interferon-induced antiviral host cell response. Recently, two vaccinia virus gene products were shown to interfere with the activity of the double-stranded RNA-dependent protein kinase (PKR): the K3L gene product and the E3L gene product. We have evaluated the efficiency by which these gene products inhibit PKR and whether they act in a synergistic manner. The effects of the two vaccinia virus gene products were compared in an in vivo system in which translation of a reporter gene (dihydrofolate reductase or eukaryotic translation initiation factor 2 alpha [eIF-2 alpha]) was inhibited because of the localized activation of PKR. In this system, the E3L gene product, and to a lesser extent the K3L gene product, potentiated translation of the reporter gene and inhibited eIF-2 alpha phosphorylation. Analysis in vitro demonstrated that the E3L gene product inhibited PKR approximately 50- to 100-fold more efficiently than the K3L gene product. However, further studies demonstrated that the mechanism of action of these two inhibitors was different. Whereas the E3L inhibitor interfered with the binding of the kinase to double-stranded RNA, the K3L inhibitor did not. We propose that the K3L inhibitor acts through its homology to eIF-2 alpha to interfere with the interaction of eIF-2 alpha with PKR. The two inhibitors did not display a synergistic effect on translation or eIF-2 alpha phosphorylation. In addition, neither K3L nor E3L expression detectably altered cellular protein synthesis. Images PMID:8094759

Davies, M V; Chang, H W; Jacobs, B L; Kaufman, R J

1993-01-01

286

Histone deacetylase inhibition induces long-lasting changes in maternal behavior and gene expression in female mice.  

PubMed

In many species, including mice, maternal responsiveness is experience-dependent and permanent, lasting for long periods (months to years). We have shown that after brief exposures to pups, virgin female mice continue to respond maternally toward pups for at least one month. Administration of a histone deacetylase inhibitor (HDACi) reduces the amount of maternal experience required to affect maternal behavior and gene expression. In this set of studies, we examined the epigenetic mechanisms that underlie these motivated behaviors. We assessed whether the effects of HDACi persisted 1 month after the initial experience (in the absence of continued pup experience or HDACi treatment) and whether the maintenance of maternal memory was associated with stable changes in gene expression. Using chromatin immunoprecipitation, we examined whether Esr2 and Oxt gene expression might be mediated by recruitment of the histone acetyltransferase cAMP response element binding protein (CBP) to their promoter regions after maternal memory consolidation. We report that HDACi treatment induced long-lasting changes in maternal responsiveness. Maternal learning was associated with increased recruitment of CBP to the Esr2 and Oxt gene promoters during the consolidation of maternal memory as well as a persistent increase in estrogen receptor-? (Esr2) mRNA and decreased expression of the de novo DNA methyltransferase Dnmt3a within the medial preoptic area. The consolidation of the maternal experience may involve the CBP recruitment and stable changes in gene expression, which maintain increased maternal responsiveness for long periods of time. PMID:24932804

Stolzenberg, Danielle S; Stevens, Jacqueline S; Rissman, Emilie F

2014-09-01

287

Inhibition of HIV1 gene expression by Ciclopirox and Deferiprone, drugs that prevent hypusination of eukaryotic initiation factor 5A  

Microsoft Academic Search

BACKGROUND: Eukaryotic translation initiation factor eIF5A has been implicated in HIV-1 replication. This protein contains the apparently unique amino acid hypusine that is formed by the post-translational modification of a lysine residue catalyzed by deoxyhypusine synthase and deoxyhypusine hydroxylase (DOHH). DOHH activity is inhibited by two clinically used drugs, the topical fungicide ciclopirox and the systemic medicinal iron chelator deferiprone.

Mainul Hoque; Hartmut M Hanauske-Abel; Paul Palumbo; Deepti Saxena; Darlene D'Alliessi Gandolfi; Myung Hee Park; Tsafi Pe'ery; Michael B Mathews

2009-01-01

288

Factor inhibiting HIF limits the expression of hypoxia-inducible genes in podocytes and distal tubular cells  

Microsoft Academic Search

The two hypoxia-inducible factors (HIF-1? and HIF-2?) are transcription factors that regulate the response to hypoxia. Recently, the factor inhibiting HIF (FIH1) was identified as a molecular oxygen-dependent dioxygenase that blunts the transcriptional activity of HIF and has also been implicated in HIF-dependent and -independent hypoxia responses. Interestingly, HIF accumulation in the kidney has been shown to confer renal protection

Johannes Schödel; Daniela Bohr; Bernd Klanke; Gunnar Schley; Ursula Schlötzer-Schrehardt; Christina Warnecke; Armin Kurtz; Kerstin Amann; Kai-Uwe Eckardt; Carsten Willam

2010-01-01

289

HIPK2 inhibits both MDM2 gene and protein by, respectively, p53-dependent and independent regulations  

Microsoft Academic Search

We address here the involvement of the homeodomain-interacting protein kinase 2 (HIPK2)\\/p53 complex on MDM2 regulation following apoptotic DNA damage. Our results provide a plausible transcriptional (p53-dependent) and posttranscriptional (p53-independent) double mechanism by which HIPK2 accomplishes MDM2 downmodulation. First, in wtp53-carrying cells HIPK2-dependent p53Ser46 phosphorylation selectively inhibits MDM2 at transcriptional level. Secondly, HIPK2 interacts with MDM2 in vitro and in

Valeria Di Stefano; Marina Mattiussi; Ada Sacchi; Gabriella D’Orazi

2005-01-01

290

Genes  

NSDL National Science Digital Library

Illustration of the placement of genes in a chromosome. A gene can be defined as a region of DNA that controls a hereditary characteristic. It usually corresponds to a sequence used in the production of a specific protein or RNA. A gene carries biological information in a form that must be copied and transmitted from each cell to all its progeny. This includes the entire functional unit: coding DNA sequences, non-coding regulatory DNA sequences, and introns. Genes can be as short as 1000 base pairs or as long as several hundred thousand base pairs. It can even be carried by more than one chromosome. The estimate for the number of genes in humans has decreased as our knowledge has increased. As of 2001, humans are thought to have between 30,000 and 40,000 genes.

Excellence, Access

2005-03-12

291

Cold exposure inhibits hypothalamic Kiss-1 gene expression, serum leptin concentration, and delays reproductive development in male Brandt's vole (Lasiopodomys brandtii)  

NASA Astrophysics Data System (ADS)

Cold commonly affects growth and reproductive development in small mammals. Here, we test the hypothesis that low ambient temperature will affect growth and puberty onset, associated with altered hypothalamic Kiss-1 gene expression and serum leptin concentration in wild rodents. Male Brandt's voles (Lasiopodomys brandtii) were exposed to cold (4 ± 1 °C) and warm (23 ± 1 °C) conditions from the birth and sacrificed on different developmental stages (day 26, day 40, day 60, and day 90, respectively). Brandt's voles increased the thermogenic capacity of brown adipose tissue, mobilized body fat, decreased serum leptin levels, and delayed the reproductive development especially on day 40 in the cold condition. They increased food intake to compensate for the high energy demands in the cold. The hypothalamic Kiss-1 gene expression on day 26 was decreased, associated with lower wet testis mass and testis testosterone concentration on day 40, in the cold-exposed voles compared to that in the warm. Serum leptin was positively correlated with body fat, testis mass, and testosterone concentration. These data suggested that cold exposure inhibited hypothalamic Kiss-1 gene expression during the early stage of development, decreased serum leptin concentration, and delayed reproductive development in male Brandt's voles.

Zhang, Qiang; Lin, Yi; Zhang, Xue-Ying; Wang, De-Hua

2014-08-01

292

The BEL1-type homeobox gene SH5 induces seed shattering by enhancing abscission-zone development and inhibiting lignin biosynthesis.  

PubMed

Seed shattering is an important trait that influences grain yield. A major controlling quantitative trait locus in rice is qSH1. Although the degree of shattering is correlated with the level of expression of qSH1, some qSH1-defective cultivars display moderate shattering while others show a non-shattering phenotype. Os05 g38120 (SH5) on chromosome 5 is highly homologous to qSH1. Although we detected SH5 transcripts in various organs, this gene was highly expressed at the abscission zone (AZ) in the pedicels. When expression of this gene was suppressed in easy-shattering 'Kasalath', development of the AZ was reduced and thereby so was seed loss. By contrast, the extent of shattering, as well as AZ development, was greatly enhanced in moderate-shattering 'Dongjin' rice when SH5 was overexpressed. Likewise, overexpression of SH5 in the non-shattering 'Ilpum' led to an increase in seed shattering because lignin levels were decreased in the basal region of spikelets in the absence of development of an AZ. We also determined that two shattering-related genes, SHAT1 and Sh4, which are necessary for proper formation of an AZ, were induced by SH5. Based on these observations, we propose that SH5 modulates seed shattering by enhancing AZ development and inhibiting lignin biosynthesis. PMID:24923192

Yoon, Jinmi; Cho, Lae-Hyeon; Kim, Song Lim; Choi, Heebak; Koh, Hee-Jong; An, Gynheung

2014-09-01

293

Adenovirus-mediated kallikrein gene transfer inhibits neointima formation via increased production of nitric oxide in rat artery  

Microsoft Academic Search

Tissue kallikrein cleaves kininogen substrate to produce vasoactive kinin peptides that have been implicated to play a role . in the proliferation of vascular smooth muscle cells VSMC . In order to explore potential roles of the kallikrein-kinin system in vascular biology, we evaluated the effects of adenovirus-mediated kallikrein gene delivery on neointima formation in balloon-injured rat artery. Infection of

Hideyuki Murakami; Robert Q. Miao; Lee Chao; Julie Chao

1999-01-01

294

Histone post-translational modifications induced by histone deacetylase inhibition in transcriptional control units of NIS gene.  

PubMed

Histone post-translational modifications (HPTMs) play a major role in control of gene transcription. Among them, histone acetylation and methylation have been extensively investigated. Histone acetylation at different residues is generally associated to active gene transcription. In contrast, histone methylation can be associated either to transcriptional activation or repression, depending primarily on the histone residue that is subjected to the modification. Herein, effects of the histone deacetylase inhibitor SAHA on the sodium-iodide symporter (NIS) gene expression were investigated in breast cancer cells (MDA157 and MDA468). SAHA treatment induces high increase of NIS mRNA levels in MDA468 cells (300-fold), but moderate increase in MDA157 cells (fivefold). Histone H3 HPTMs (acetylation and methylations) on transcriptional units of NIS gene were investigated in these cell lines upon SAHA treatment. Our data indicate that HPTMs, particularly the H3 lysine 27 trimethylation, may operate in contrast to current models that relate epigenetic modifications with transcriptional activity. PMID:24844212

Baldan, Federica; Lavarone, Elisa; Di Loreto, Carla; Filetti, Sebastiano; Russo, Diego; Damante, Giuseppe; Puppin, Cinzia

2014-08-01

295

Defective Mitochondrial Gene Expression Results in Reactive Oxygen Species-Mediated Inhibition of Respiration and Reduction of Yeast Life Span  

Microsoft Academic Search

Mitochondrial dysfunction causes numerous human diseases and is widely believed to be involved in aging. However, mechanisms through which compromised mitochondrial gene expression elicits the reported variety of cellular defects remain unclear. The amino-terminal domain (ATD) of yeast mitochondrial RNA polymerase is required to couple transcription to translation during expression of mitochondrial DNA-encoded oxidative phosphorylation subunits. Here we report that

Nicholas D. Bonawitz; Matthew S. Rodeheffer; Gerald S. Shadel

2006-01-01

296

TET2 Inhibits Differentiation of Embryonic Stem Cells but Does Not Overcome Methylation-Induced Gene Silencing  

PubMed Central

TET2 is a methylcytosine dioxygenase that is frequently mutated in myeloid malignancies, notably myelodysplasia and acute myeloid leukemia. TET2 catalyses the conversion of 5?-methylcytosine to 5?-hydroxymethylcytosine within DNA and has been implicated in the process of genomic demethylation. However, the mechanism by which TET2 loss of function results in hematopoietic dysplasia and leukemogenesis is poorly understood. Here, we show that TET2 is expressed in undifferentiated embryonic stem cells and that its knockdown results in reduction of 5?-hydroxymethylcytosine in genomic DNA. We also present DNA methylation data from bone marrow samples obtained from patients with TET2-mutated myelodysplasia. Based on these findings, we sought to identify the role of TET2 in regulating pluripotency and differentiation. We show that overexpression of TET2 in a stably integrated transgene leads to increased alkaline phosphatase expression in differentiating ES cells and impaired differentiation in methylcellulose culture. We speculate that this effect is due to TET2-mediated expression of stem cell genes in ES cells via hydroxylation of 5?-methylcytosines at key promoter sequences within genomic DNA. This leads to relative hypomethylation of gene promoters as 5?-hydroxymethylcytosine is not a substrate for DNMT1-mediated maintenance methylation. We sought to test this hypothesis by cotransfecting the TET2 gene with methylated reporter genes. The results of these experiments are presented. PMID:25276435

2014-01-01

297

Peroxisome Proliferator-Activated Receptor Gamma Activators Inhibit Gene Expression and Migration in Human Vascular Smooth Muscle Cells  

Microsoft Academic Search

Migration of vascular smooth muscle cells (VSMCs) plays an important role in atherogenesis and restenosis after arterial interventions. The expression of matrix metalloproteinases (MMPs), particularly MMP-9, contributes to VSMC migration. This process requires degradation of basal laminae and other components of the arterial extracellular matrix. Peroxisome proliferator-activated receptors (PPARs), members of the nuclear receptor family, regulate gene expression after activation

Nikolaus Marx; Uwe Schonbeck; Mitchell A. Lazar; Peter Libby; Jorge Plutzky

298

Exploring signatures of positive selection in pigmentation candidate genes in populations of East Asian ancestry  

PubMed Central

Background Currently, there is very limited knowledge about the genes involved in normal pigmentation variation in East Asian populations. We carried out a genome-wide scan of signatures of positive selection using the 1000 Genomes Phase I dataset, in order to identify pigmentation genes showing putative signatures of selective sweeps in East Asia. We applied a broad range of methods to detect signatures of selection including: 1) Tests designed to identify deviations of the Site Frequency Spectrum (SFS) from neutral expectations (Tajima’s D, Fay and Wu’s H and Fu and Li’s D* and F*), 2) Tests focused on the identification of high-frequency haplotypes with extended linkage disequilibrium (iHS and Rsb) and 3) Tests based on genetic differentiation between populations (LSBL). Based on the results obtained from a genome wide analysis of 25 kb windows, we constructed an empirical distribution for each statistic across all windows, and identified pigmentation genes that are outliers in the distribution. Results Our tests identified twenty genes that are relevant for pigmentation biology. Of these, eight genes (ATRN, EDAR, KLHL7, MITF, OCA2, TH, TMEM33 and TRPM1,) were extreme outliers (top 0.1% of the empirical distribution) for at least one statistic, and twelve genes (ADAM17, BNC2, CTSD, DCT, EGFR, LYST, MC1R, MLPH, OPRM1, PDIA6, PMEL (SILV) and TYRP1) were in the top 1% of the empirical distribution for at least one statistic. Additionally, eight of these genes (BNC2, EGFR, LYST, MC1R, OCA2, OPRM1, PMEL (SILV) and TYRP1) have been associated with pigmentary traits in association studies. Conclusions We identified a number of putative pigmentation genes showing extremely unusual patterns of genetic variation in East Asia. Most of these genes are outliers for different tests and/or different populations, and have already been described in previous scans for positive selection, providing strong support to the hypothesis that recent selective sweeps left a signature in these regions. However, it will be necessary to carry out association and functional studies to demonstrate the implication of these genes in normal pigmentation variation. PMID:23848512

2013-01-01

299

Overexpression of D-Xylose Reductase (xyl1) Gene and Antisense Inhibition of D-Xylulokinase (xyiH) Gene Increase Xylitol Production in Trichoderma reesei  

PubMed Central

T. reesei is an efficient cellulase producer and biomass degrader. To improve xylitol production in Trichoderma reesei strains by genetic engineering, two approaches were used in this study. First, the presumptive D-xylulokinase gene in T. reesei (xyiH), which has high homology to known fungi D-xylulokinase genes, was silenced by transformation of T. reesei QM9414 strain with an antisense construct to create strain S6-2-2. The expression of the xyiH gene in the transformed strain S6-2-2 decreased at the mRNA level, and D-xylulokinase activity decreased after 48?h of incubation. This led to an increase in xylitol production from undetectable levels in wild-type T. reesei QM9414 to 8.6?mM in S6-2-2. The T. reesei ?xdh is a xylose dehydrogenase knockout strain with increased xylitol production compared to the wild-type T. reesei QM9414 (22.8?mM versus undetectable). The copy number of the xylose reductase gene (xyl1) in T. reesei ?xdh strain was increased by genetic engineering to create a new strain ?9-5-1. The ?9-5-1 strain showed a higher xyl1 expression and a higher yield of xylose reductase, and xylitol production was increased from 22.8?mM to 24.8?mM. Two novel strains S6-2-2 and ?9-5-1 are capable of producing higher yields of xylitol. T. reesei has great potential in the industrial production of xylitol. PMID:25013760

Hong, Yuanyuan; Dashtban, Mehdi; Kepka, Greg; Chen, Sanfeng; Qin, Wensheng

2014-01-01

300

Gene Disruption of the Calcium Channel Orai1 Results in Inhibition of Osteoclast and Osteoblast Differentiation and Impairs Skeletal Development  

PubMed Central

Calcium signaling plays a central role in the regulation of bone cells, though uncertainty remains with regard to the channels involved. In previous studies, we determined that the calcium channel Orai1 was required for the formation of multinucleated osteoclasts in vitro. To define the skeletal functions of calcium release-activated calcium currents, we compared mice with targeted deletion of the calcium channel Orai1 to wild-type littermate controls, and examined differentiation and function of osteoblast and osteoclast precursors in vitro with and without Orai1 inhibition. Consistent with in vitro findings, Orai1?/? mice lacked multinucleated osteoclasts. Yet they did not develop osteopetrosis. Mononuclear cells expressing osteoclast products were found in Orai1?/? mice, and in vitro studies showed significantly reduced, but not absent, mineral resorption by the mononuclear osteoclast-like cells that form in culture from peripheral blood monocytic cells when Orai1 is inhibited. More prominent in Orai1?/? mice was a decrease in bone with retention of fetal cartilage. Micro-computed tomography showed reduced cortical ossification and thinned trabeculae in Orai1?/? animals compared to controls; bone deposition was markedly decreased in the knock-out. This suggested a previously unrecognized role for Orai1 within osteoblasts. Analysis of osteoblasts and precursors in Orai1?/? and control mice showed a significant decrease in alkaline phosphatase-expressing osteoblasts. In vitro studies confirmed that inhibiting Orai1 activity impaired differentiation and function of human osteoblasts, supporting a critical function for Orai1 in osteoblasts, in addition to its role as a regulator of osteoclast formation. PMID:22546867

Robinson, Lisa J.; Mancarella, Salvatore; Songsawad, Duangrat; Tourkova, Irina L.; Barnett, John B.; Gill, Donald L.; Soboloff, Jonathan; Blair, Harry C.

2012-01-01

301

Inhibition of gene expression of carnitine palmitoyltransferase I and heart fatty acid binding protein in cyclophosphamide and ifosfamide-induced acute cardiotoxic rat models.  

PubMed

This study investigated whether cyclophosphamide (CP) and ifosfamide (IFO) therapy alters the expression of the key genes engaged in long-chain fatty acid (LCFA) oxidation outside rat heart mitochondria, and if so, whether these alterations should be viewed as a mechanism during CP- and IFO-induced cardiotoxicity. Adult male Wistar albino rats were assigned to one of the six treatment groups: Rats in group 1 (control) and group 2 (L-carnitine) were injected intraperitoneal (i.p.) with normal saline and L-carnitine (200 mg/kg/day), respectively, for 10 successive days. Animals in group 3 (CP group) were injected i.p. with normal saline for 5 days before and 5 days after a single dose of CP (200 mg/kg, i.p.). Rats in group 4 (IFO group) received normal saline for 5 successive days followed by IFO (50 mg/kg/day, i.p.) for 5 successive days. Rats in group 5 (CP-carnitine supplemented) were given the same doses of L-carnitine as group 2 for 5 days before and 5 days after a single dose of CP as group 3. Rats in group 6 (IFO-carnitine supplemented) were given the same doses of L-carnitine as group 2 for 5 days before and 5 days concomitant with IFO as group 4. Immediately, after the last dose of the treatment protocol, blood samples were withdrawn and animals were killed for biochemical, histopathological and gene expression studies. Treatment with CP and IFO significantly decreased expression of heart fatty acid binding protein (H-FABP) and carnitine palmitoyltransferase I (CPT I) genes in cardiac tissues. Moreover, CP but not IFO significantly increased acetyl-CoA carboxylase2 mRNA expression. Conversely, IFO but not CP significantly decreased mRNA expression of malonyl-CoA decarboxylase. Both CP and IFO significantly increased serum lactate dehydrogenase, creatine kinase isoenzyme MB and malonyl-CoA content and histopathological lesions in cardiac tissues. Interestingly, carnitine supplementation completely reversed all the biochemical, histopathological and gene expression changes induced by CP and IFO to the control values, except CPT I mRNA, and protein expression remained inhibited by IFO. Data from the current study suggest, for the first time, that (1) CP and IFO therapy is associated with the inhibition of the expression of H-FABP and CPT I genes in cardiac tissues with the consequent inhibition of mitochondrial transport and oxidation of LCFA. (2) The progressive increase in cardiotoxicity enzymatic indices and the decrease in H-FABP and CPT I expression may point to the possible contribution of these genes to CP- and IFO-induced cardiotoxicity. PMID:24469765

Sayed-Ahmed, Mohamed M; Aldelemy, Meshan L; Al-Shabanah, Othman A; Hafez, Mohamed M; Al-Hosaini, Khaled A; Al-Harbi, Naif O; Al-Sharary, Shakir D; Al-Harbi, Mohamed M

2014-09-01

302

Axl gene knockdown inhibits the metastasis properties of hepatocellular carcinoma via PI3K/Akt-PAK1 signal pathway.  

PubMed

The objective of this study is to clarify the possible role and mechanism of Axl in the tumorigenicity and metastasis process of hepatocellular carcinoma. The mRNA and protein expression levels of Axl in MHCC97-H and MHCC97-L cell lines were evaluated by real-time PCR and Western blot analysis. The key factor of phosphatidylinositol-3-kinase (PI3K)/Akt-p21-activated kinases-1 (PAK1) signaling pathway was studied after Axl expression was downregulated by shRNA. Finally, we analyzed the expression status of Axl protein expression in hepatocellular carcinoma tissues and its relationship with the prognosis of hepatocellular carcinoma. Axl was observed to be higher expressed in MHCC97-H cell lines compared to MHCC97-L cell lines. The downregulation of Axl in MHCC97-H cell lines resulted in the inhibition of the invasion ability of MHCC97-H cells both in vitro and in vivo. Interestingly, blocking PI3K/Akt signaling pathway by LY294002 or Akt siRNA could remarkably inhibit the PAK1 activation and cell invasion. Finally, the Axl protein expression was positively correlated with differentiation, lymph node metastasis, and clinical stage in patients with hepatocellular carcinoma patients (all P < 0.01). These findings suggest that Axl can also regulate the metastasis process of hepatocellular carcinoma and may serve as a new prognostic marker and therapeutic target for treating hepatocellular carcinoma metastasis. PMID:24347489

Xu, Jingchao; Jia, Li; Ma, Hongye; Li, Yanping; Ma, Zhenhai; Zhao, Yongfu

2014-04-01

303

In Vivo Gene Therapy with Interleukin12 Inhibits Primary Vascular Tumor Growth and Induces Apoptosis in a Mouse Model1  

Microsoft Academic Search

Interleukin-12 is proposed to have anti-neoplastic activity on the basis of both its anti-angiogenic and immunologic effects. Gene gun therapy with interleukin-12 cDNA into the peritumoral area of immunocompetent 129\\/J mice with life-threatening primary vascular tumors reduced tumor volume 7.5-fold and almost tripled the duration of mouse survival, in contrast with luciferase-bombarded control mice. Epidermal expression of mouse interleukin-12 elevated

Chong Wang; M. Eugenia Quevedo; Brian J. Lannutti; Kenneth B. Gordon; Danqing Guo; Wenn Sun; Amy S. Paller

1999-01-01

304

Renal Pro-Inflammatory Cytokine Gene Expression in Diabetic Nephropathy: Effect of Angiotensin-Converting Enzyme Inhibition and Pentoxifylline Administration  

Microsoft Academic Search

Background: Recent studies have shown a role for inflammation in the pathogenesis of diabetic nephropathy (DN). Tumor necrosis factor (TNF)-?, interleukin (IL)-1 and IL-6 are cytokines with a prevalent pro-inflammatory activity. Our objective was to study the renal gene expression of TNF-?, IL-1 and IL-6 in DN and their relationship with renal damage assessed by urinary albumin excretion (UAE). In

Juan F. Navarro; Francisco J. Milena; Carmen Mora; Candelaria León; Javier García

2006-01-01

305

TcpH Influences Virulence Gene Expression in Vibrio cholerae by Inhibiting Degradation of the Transcription Activator TcpP  

Microsoft Academic Search

Expression of toxT, the transcription activator of cholera toxin and pilus production in Vibrio cholerae ,i s the consequence of a complex cascade of regulatory events that culminates in activation of the toxT promoter by TcpP and ToxR, two membrane-localized transcription factors. Both are encoded in operons with genes whose products, TcpH and ToxS, which are also membrane localized, are

Nancy A. Beck; Eric S. Krukonis; Victor J. DiRita

2004-01-01

306

Histone Deacetylase 5 Is Not a p53 Target Gene, But Its Overexpression Inhibits Tumor Cell Growth and Induces Apoptosis  

Microsoft Academic Search

p53 tumor suppressor is activated by phosphorylation and acetylation on DNA damage. One of unknown p53 early transcripts was identified to be histone deacetylase-5 (HDAC5). We tested a hypothesis that HDAC5 is a p53 down-stream target gene that on induction by p53 inactivates p53 by removal of acetyl group in p53 molecule, thus functioning as an auto- regulatory negative feedback

Yuanhui Huang; Mingjia Tan; Mark Gosink; Kevin K. W. Wang; Yi Sun

2002-01-01

307

High Temperature Inhibits Ascorbate Recycling and Light Stimulation of the Ascorbate Pool in Tomato despite Increased Expression of Biosynthesis Genes  

PubMed Central

Understanding how the fruit microclimate affects ascorbate (AsA) biosynthesis, oxidation and recycling is a great challenge in improving fruit nutritional quality. For this purpose, tomatoes at breaker stage were harvested and placed in controlled environment conditions at different temperatures (12, 17, 23, 27 and 31°C) and irradiance regimes (darkness or 150 µmol m-2 s-1). Fruit pericarp tissue was used to assay ascorbate, glutathione, enzymes related to oxidative stress and the AsA/glutathione cycle and follow the expression of genes coding for 5 enzymes of the AsA biosynthesis pathway (GME, VTC2, GPP, L-GalDH, GLDH). The AsA pool size in pericarp tissue was significantly higher under light at temperatures below 27°C. In addition, light promoted glutathione accumulation at low and high temperatures. At 12°C, increased AsA content was correlated with the enhanced expression of all genes of the biosynthesis pathway studied, combined with higher DHAR and MDHAR activities and increased enzymatic activities related to oxidative stress (CAT and APX). In contrast, at 31°C, MDHAR and GR activities were significantly reduced under light indicating that enzymes of the AsA/glutathione cycle may limit AsA recycling and pool size in fruit pericarp, despite enhanced expression of genes coding for AsA biosynthesis enzymes. In conclusion, this study confirms the important role of fruit microclimate in the regulation of fruit pericarp AsA content, as under oxidative conditions (12°C, light) total fruit pericarp AsA content increased up to 71%. Moreover, it reveals that light and temperature interact to regulate both AsA biosynthesis gene expression in tomato fruits and AsA oxidation and recycling. PMID:24367665

Massot, Capucine; Bancel, Doriane; Lopez Lauri, Felicie; Truffault, Vincent; Baldet, Pierre; Stevens, Rebecca; Gautier, Helene

2013-01-01

308

Inhibition of cell proliferation and induction of apoptosis by oleanane triterpenoid (CDDO-Me) in pancreatic cancer cells is associated with the suppression of hTERT gene expression and its telomerase activity  

SciTech Connect

Highlights: Black-Right-Pointing-Pointer CDDO-Me inhibits hTERT gene expression. Black-Right-Pointing-Pointer CDDO-Me inhibits hTERT protein expression. Black-Right-Pointing-Pointer CDDO-Me inhibits hTERT telomerase activity. Black-Right-Pointing-Pointer CDDO-Me inhibits hTERT regulatory proteins. -- Abstract: Methyl-2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oate (CDDO-Me) is a multifunctional oleanane synthetic triterpenoid with potent anti-inflammatory and antitumorigenic properties. The mechanisms of the antisurvival and apoptosis-inducing activities of CDDO-Me and related derivatives of oleanolic acid have been defined; however, to date, no study has been carried out on the effect of CDDOs on human telomerase reverse transcriptase (hTERT) gene or telomerase activity. Here we report for the first time that inhibition of cell proliferation and induction of apoptosis by CDDO-Me in pancreatic cancer cell lines is associated with the inhibition of hTERT gene expression, hTERT telomerase activity and a number of proteins that regulate hTERT expression and activity. Furthermore, abrogation or overexpression of hTERT protein altered the susceptibility of tumor cells to CDDO-Me. These findings suggest that telomerase (hTERT) is a relevant target of CDDO-Me in pancreatic cancer cells.

Deeb, Dorrah; Gao, Xiaohua; Liu, Yongbo [Department of Surgery, Henry Ford Health System, Detroit, MI (United States)] [Department of Surgery, Henry Ford Health System, Detroit, MI (United States); Kim, Sahn-Ho [Department of Urology, Henry Ford Health System, Detroit, MI (United States)] [Department of Urology, Henry Ford Health System, Detroit, MI (United States); Pindolia, Kirit R. [Department of Medical Genetics, Henry Ford Health System, Detroit, MI (United States)] [Department of Medical Genetics, Henry Ford Health System, Detroit, MI (United States); Arbab, Ali S. [Department of Radiology, Henry Ford Health System, Detroit, MI (United States)] [Department of Radiology, Henry Ford Health System, Detroit, MI (United States); Gautam, Subhash C., E-mail: sgautam1@hfhs.org [Department of Surgery, Henry Ford Health System, Detroit, MI (United States)

2012-06-15

309

Inhibition of Inflammatory Gene Expression in Keratinocytes Using a Composition Containing Carnitine, Thioctic Acid and Saw Palmetto Extract  

PubMed Central

Chronic inflammation of the hair follicle (HF) is considered a contributing factor in the pathogenesis of androgenetic alopecia (AGA). Previously, we clinically tested liposterolic extract of Serenoa repens (LSESr) and its glycoside, ?-sitosterol, in subjects with AGA and showed a highly positive response to treatment. In this study, we sought to determine whether blockade of inflammation using a composition containing LSESr as well as two anti-inflammatory agents (carnitine and thioctic acid) could alter the expression of molecular markers of inflammation in a well-established in vitro system. Using a well-validated assay representative of HF keratinocytes, specifically, stimulation of cultured human keratinocyte cells in vitro, we measured changes in gene expression of a spectrum of well-known inflammatory markers. Lipopolysaccharide (LPS) provided an inflammatory stimulus. In particular, we found that the composition effectively suppressed LPS-activated gene expression of chemokines, including CCL17, CXCL6 and LTB(4) associated with pathways involved in inflammation and apoptosis. Our data support the hypothesis that the test compound exhibits anti-inflammatory characteristics in a well-established in vitro assay representing HF keratinocyte gene expression. These findings suggest that 5-alpha reductase inhibitors combined with blockade of inflammatory processes could represent a novel two-pronged approach in the treatment of AGA with improved efficacy over current modalities. PMID:19692448

Chittur, Sridar; Parr, Brian; Marcovici, Geno

2011-01-01

310

Overexpression of the m4 and malpha genes of the E(spl)-complex antagonizes notch mediated lateral inhibition.  

PubMed

Intercellular signalling mediated by Notch proteins is crucial to many cell fate decisions in metazoans. Its profound effects on cell fate and proliferation require that a complex set of responses involving positive and negative signal transducers be orchestrated around each instance of signalling. In Drosophila the basic-helix-loop-helix (bHLH) repressor encoding genes of the E(spl) locus are induced by Notch signalling and mediate some of its effects, such as suppression of neural fate. Here we report on a novel family of Notch responsive genes, whose products appear to act as antagonists of the Notch signal in the process of adult sensory organ precursor singularization. They, too, reside in the E(spl) locus and comprise transcription units E(spl) m4 and E(spl) malpha. Overexpression of these genes causes downregulation of E(spl) bHLH expression accompanied by cell autonomous overcommitment of sensory organ precursors and tufting of bristles. Interestingly, negative regulation of the Notch pathway by overexpression of E(spl) m4 and malpha is specific to the process of sensory organ precursor singularization and does not impinge on other instances of Notch signalling. PMID:10446264

Apidianakis, Y; Nagel, A C; Chalkiadaki, A; Preiss, A; Delidakis, C

1999-08-01

311

Inhibition of the Nrf2 transcription factor by the alkaloid trigonelline renders pancreatic cancer cells more susceptible to apoptosis through decreased proteasomal gene expression and proteasome activity.  

PubMed

Evidence accumulates that the transcription factor nuclear factor E2-related factor 2 (Nrf2) has an essential role in cancer development and chemoresistance, thus pointing to its potential as an anticancer target and undermining its suitability in chemoprevention. Through the induction of cytoprotective and proteasomal genes, Nrf2 confers apoptosis protection in tumor cells, and inhibiting Nrf2 would therefore be an efficient strategy in anticancer therapy. In the present study, pancreatic carcinoma cell lines (Panc1, Colo357 and MiaPaca2) and H6c7 pancreatic duct cells were analyzed for the Nrf2-inhibitory effect of the coffee alkaloid trigonelline (trig), as well as for its impact on Nrf2-dependent proteasome activity and resistance to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and anticancer drug-induced apoptosis. Chemoresistant Panc1 and Colo357 cells exhibit high constitutive Nrf2 activity, whereas chemosensitive MiaPaca2 and H6c7 cells display little basal but strong tert-butylhydroquinone (tBHQ)-inducible Nrf2 activity and drug resistance. Trig efficiently decreased basal and tBHQ-induced Nrf2 activity in all cell lines, an effect relying on a reduced nuclear accumulation of the Nrf2 protein. Along with Nrf2 inhibition, trig blocked the Nrf2-dependent expression of proteasomal genes (for example, s5a/psmd4 and ?5/psma5) and reduced proteasome activity in all cell lines tested. These blocking effects were absent after treatment with Nrf2 siRNA, a condition in which proteasomal gene expression and proteasome activity were already decreased, whereas siRNA against the related transcription factor Nrf1 did not affect proteasome activity and the inhibitory effect of trig. Depending on both Nrf2 and proteasomal gene expression, the sensitivity of all cell lines to anticancer drugs and TRAIL-induced apoptosis was enhanced by trig. Moreover, greater antitumor responses toward anticancer drug treatment were observed in tumor-bearing mice when receiving trig. In conclusion, representing an efficient Nrf2 inhibitor capable of blocking Nrf2-dependent proteasome activity and thereby apoptosis protection in pancreatic cancer cells, trig might be beneficial in improving anticancer therapy. PMID:23108405

Arlt, A; Sebens, S; Krebs, S; Geismann, C; Grossmann, M; Kruse, M-L; Schreiber, S; Schäfer, H

2013-10-01

312

Inhibition of tomato (Solanum lycopersicum L.) root growth by cyanamide is due to altered cell division, phytohormone balance and expansin gene expression.  

PubMed

Cyanamide (CA) has been reported as a natural compound produced by hairy vetch (Vicia villosa Roth.) and it was shown also to be an allelochemical, responsible for strong allelopathic potential in this species. CA phytotoxicity has been demonstrated on various plant species, but to date little is known about its mode of action at cellular level. Treatment of tomato (Solanum lycopersicum L.) roots with CA (1.2 mM) resulted in inhibition of growth accompanied by alterations in cell division, and imbalance of plant hormone (ethylene and auxin) homeostasis. Moreover, the phytotoxic effect of CA was also manifested by modifications in expansin gene expression, especially in expansins responsible for cell wall remodeling after the cytokinesis (LeEXPA9, LeEXPA18). Based on these results the phytotoxic activity of CA on growth of roots of tomato seedlings is likely due to alterations associated with cell division. PMID:22847024

Soltys, Dorota; Rudzi?ska-Langwald, Anna; Gniazdowska, Agnieszka; Wi?niewska, Anita; Bogatek, Renata

2012-11-01

313

Reproductive Development Modulates Gene Expression and Metabolite Levels with Possible Feedback Inhibition of Artemisinin in Artemisia annua1[C][W][OA  

PubMed Central

The relationship between the transition to budding and flowering in Artemisia annua and the production of the antimalarial sesquiterpene, artemisinin (AN), the dynamics of artemisinic metabolite changes, AN-related transcriptional changes, and plant and trichome developmental changes were measured. Maximum production of AN occurs during full flower stage within floral tissues, but that changes in the leafy bracts and nonbolt leaves as the plant shifts from budding to full flower. Expression levels of early pathway genes known to be involved in isopentenyl diphosphate and farnesyl diphosphate biosynthesis leading to AN were not immediately positively correlated with either AN or its precursors. However, we found that the later AN pathway genes, amorpha-4,11-diene synthase (ADS) and the cytochrome P450, CYP71AV1 (CYP), were more highly correlated with AN’s immediate precursor, dihydroartemisinic acid, within all leaf tissues tested. In addition, leaf trichome formation throughout the developmental phases of the plant also appears to be more complex than originally thought. Trichome changes correlated closely with the levels of AN but not its precursors. Differences were observed in trichome densities that are dependent both on developmental stage (vegetative, budding, and flowering) and on position (upper and lower leaf tissue). AN levels declined significantly as plants matured, as did ADS and CYP transcripts. Spraying leaves with AN or artemisinic acid inhibited CYP transcription; artemisinic acid also inhibited ADS transcription. These data allow us to present a novel model for the differential control of AN biosynthesis as it relates to developmental stage and trichome maturation and collapse. PMID:20724645

Arsenault, Patrick R.; Vail, Daniel; Wobbe, Kristin K.; Erickson, Karen; Weathers, Pamela J.

2010-01-01

314

Co-Inoculation with Rhizobia and AMF Inhibited Soybean Red Crown Rot: From Field Study to Plant Defense-Related Gene Expression Analysis  

PubMed Central

Background Soybean red crown rot is a major soil-borne disease all over the world, which severely affects soybean production. Efficient and sustainable methods are strongly desired to control the soil-borne diseases. Principal Findings We firstly investigated the disease incidence and index of soybean red crown rot under different phosphorus (P) additions in field and found that the natural inoculation of rhizobia and arbuscular mycorrhizal fungi (AMF) could affect soybean red crown rot, particularly without P addition. Further studies in sand culture experiments showed that inoculation with rhizobia or AMF significantly decreased severity and incidence of soybean red crown rot, especially for co-inoculation with rhizobia and AMF at low P. The root colony forming unit (CFU) decreased over 50% when inoculated by rhizobia and/or AMF at low P. However, P addition only enhanced CFU when inoculated with AMF. Furthermore, root exudates of soybean inoculated with rhizobia and/or AMF significantly inhibited pathogen growth and reproduction. Quantitative RT-PCR results indicated that the transcripts of the most tested pathogen defense-related (PR) genes in roots were significantly increased by rhizobium and/or AMF inoculation. Among them, PR2, PR3, PR4 and PR10 reached the highest level with co-inoculation of rhizobium and AMF. Conclusions Our results indicated that inoculation with rhizobia and AMF could directly inhibit pathogen growth and reproduction, and activate the plant overall defense system through increasing PR gene expressions. Combined with optimal P fertilization, inoculation with rhizobia and AMF could be considered as an efficient method to control soybean red crown rot in acid soils. PMID:22442737

Gao, Xiang; Lu, Xing; Wu, Man; Zhang, Haiyan; Pan, Ruqian; Tian, Jiang; Li, Shuxian; Liao, Hong

2012-01-01

315

Inhibition of HIV-1 gene expression by novel macrophage-tropic DNA enzymes targeted to cleave HIV-1 TAT/Rev RNA.  

PubMed Central

Many regions of the HIV-1 genome have been targeted in earlier studies by RNA-cleaving DNA enzymes possessing the 10-23 catalytic motif, and efficient inhibition of HIV-1 gene expression was reported. All these studies employed charged synthetic lipids to introduce the catalytic DNA into the mammalian cells, which severely limits its practical application and usefulness in vivo. Taking advantage of the ability of G residues to interact directly with the scavenger receptors on the macrophages, we synthesized a DNA enzyme 5970 that contained 10 G residues at the 3' end. With the aim of improving the intracellular stability of the DNA enzyme 5970, we added two short stretches of stem-loop structures that were 12 bases long on either side of the DNA enzyme 5970. DNA enzyme 5970 without the poly-G tracts cleaved the synthetic RNA of HIV-1 TAT/Rev, two important regulatory proteins of HIV, very efficiently in a sequence-specific manner. Addition of 10 G residues at the 3' end of the DNA enzyme affected the cleavage efficiency only marginally whereas the same DNA enzyme with stem-loop structures on either end was significantly less efficient. The DNA enzyme with the poly-G tract at its 3' end was taken up specifically by a human macrophage-specific cell line directly in the absence of Lipofectin and was also able to inhibit HIV-1 gene expression in a transient-expression system as well as when challenged with the virus. The potential applications of these novel macrophage-tropic DNA enzymes are discussed. PMID:11415445

Unwalla, H; Banerjea, A C

2001-01-01

316

Inhibition of Lipolysis in the Novel Transgenic Quail Model Overexpressing G0/G1 Switch Gene 2 in the Adipose Tissue during Feed Restriction  

PubMed Central

In addition to the issue of obesity in humans, the production of low-fat meat from domestic animals is important in the agricultural industry to satisfy consumer demand. Understanding the regulation of lipolysis in adipose tissue could advance our knowledge to potentially solve both issues. Although the G0/G1 switch gene 2 (G0S2) was recently identified as an inhibitor of adipose triglyceride lipase (ATGL) in vitro, its role in vivo has not been fully clarified. This study was conducted to investigate the role of G0S2 gene in vivo by using two independent transgenic quail lines during different energy conditions. Unexpectedly, G0S2 overexpression had a negligible effect on plasma NEFA concentration, fat cell size and fat pad weight under ad libitum feeding condition when adipose lipolytic activity is minimal. A two-week feed restriction in non-transgenic quail expectedly caused increased plasma NEFA concentration and dramatically reduced fat cell size and fat pad weight. Contrary, G0S2 overexpression under a feed restriction resulted in a significantly less elevation of plasma NEFA concentration and smaller reductions in fat pad weights and fat cell size compared to non-transgenic quail, demonstrating inhibition of lipolysis and resistance to loss of fat by G0S2. Excessive G0S2 inhibits lipolysis in vivo during active lipolytic conditions, such as food restriction and fasting, suggesting G0S2 as a potential target for treatment of obesity. In addition, transgenic quail are novel models for studying lipid metabolism and mechanisms of obesity. PMID:24964090

Shin, Sangsu; Choi, Young Min; Han, Jae Yong; Lee, Kichoon

2014-01-01

317

Sequence-specific inhibition of gene expression by a novel antisense oligodeoxynucleotide phosphorothioate directed against a nonregulatory region of the human immunodeficiency virus type 1 genome.  

PubMed Central

Previous studies have demonstrated that oligodeoxynucleotide phosphorothioates complementary to human immunodeficiency virus type 1 (HIV-1) RNA are more nuclease resistant and are effective inhibitors of HIV-1 replication than their unmodified counterpart. In this study, antisense oligodeoxynucleotide sequences were evaluated for therapeutic potential in the treatment of HIV infections. The use of HIV-infected lymphocytes to test the efficacy of a drug is very complex, and therefore it is difficult to draw conclusions about the mechanism. We used a COS-like Monkey kidney cell line (CMT3) stably transfected with plasmids pCMVgagpol-rre-r (containing gag and pol genes) and pCMVrev (containing the rev gene of HIV-1), derived from cDNA clone BH10, as a model. A biologically active provirus that transcribes and translates their nucleotide sequences into viral proteins p24, p39/41, p55, and p160 was generated. Sequence-specific and dose-dependent inhibition of HIV-1 viral protein synthesis and significant inhibition at the mRNA level were demonstrated by antisense construct GPI2A, directed against a nonregulatory region of the HIV-1 genome. Also, our studies demonstrated enhancement of the antisense effect through encapsulation in a cationic lipid preparation. The observed attenuation of HIV-1 mRNA levels suggests that, at least in part, the mechanism of action of GPI2A was at the transcript level. Further studies have also shown antiviral activity of this construct as determined by the reverse transcriptase assay using acutely and chronically infected cells of lymphoid origin (H9 cells). Toxicological studies involving cell growth characteristics, colony-forming ability, effects on cellular proteins, specific activities of labeled proteins, and DNA synthesis in cell culture showed no cytotoxic effects of GPI2A. PMID:7853519

Anazodo, M I; Wainberg, M A; Friesen, A D; Wright, J A

1995-01-01

318

Recombinant lentivirus targeting the pleotrophin gene reduces pleotrophin protein expression in pancreatic cancer cells and inhibits neurite outgrowth of dorsal root ganglion neurons.  

PubMed

The objectives of the present study were to construct the recombinant primate lentivirus?short hairpin RNA-pleiotrophin (pLV-shRNA-PTN) vector, to investigate the silencing effect of pLV-shRNA-PTN on PTN expression in MIA PaCa-2 cells and to observe the inhibition efficiency of pLV-shRNA?PTN on neurite outgrowth from dorsal root ganglion (DRG) neurons in vitro. The construction procedure for recombinant lentivirus pLV-shRNA-PTN has been described previously. In the present study, pLV-shRNA?PTN was used to infect MIA PaCa-2 pancreatic cancer cells and the efficiency of the knockdown of the PTN gene on day 7 following infection was analyzed using western blotting. The morphological changes in the cultured DRG neurons were observed by monoculture of DRG neurons and co-culture with MIA PaCa-2 cells in vitro. The recombinant lentivirus pLV-shRNA?PTN was successfully constructed. The western blot analysis showed that the inhibition rates of PTN expression were 46, 80, 20 and 21%, respectively, following pLV-shRNA?PTN-A, B, C and D infection. pLV-shRNA-PTN?B showed the highest knockdown efficiency. DRG neurons co-cultured with infected MIA PaCa-2 cells were decreased in size when compared with the control, and there was a significant decrease in the number and length of neurites. The results suggest that efficient and specific knockdown of PTN in MIA PaCa-2 pancreatic cancer cells and the subsequent reduction in PTN expression results in the inhibition of neurite outgrowth from DRG neurons. PMID:24469464

Yao, Jun; Li, Wen-Yao; Li, Shuo-Guo; Feng, Xiao-Shan; Gao, She-Gan

2014-03-01

319

MicroRNA-124 Suppresses the Transactivation of Nuclear Factor of Activated T Cells by Targeting Multiple Genes and Inhibits the Proliferation of Pulmonary Artery Smooth Muscle Cells*  

PubMed Central

Abnormal proliferation and phenotypic modulation of pulmonary artery smooth muscle cells (PASMC) contributes to the pathogenesis of numerous cardiovascular disorders, including pulmonary arterial hypertension (PAH). The nuclear factor of activated T cells (NFAT) signaling pathway is linked to PASMC proliferation and PAH. MicroRNAs (miRNAs) are small non-coding RNAs that function in diverse biological processes. To systemically identify the specific miRNAs that regulate the NFAT pathway, a human primary miRNA library was applied for cell-based high throughput screening with the NFAT luciferase reporter system. Eight miRNAs were found to modulate NFAT activity efficiently. Of them, miR-124 robustly inhibited NFAT reporter activity and decreased both the dephosphorylation and the nuclear translocation of NFAT. miR-124 also inhibited NFAT-dependent transcription of IL-2 in Jurkat T cells. miR-124 exerted its effects by targeting multiple genes, including a known component of the NFAT pathway, NFATc1, and two new regulators of NFAT signaling, CAMTA1 (calmodulin-binding transcription activator 1) and PTBP1 (polypyrimidine tract-binding protein 1). Physiologically, miR-124 was down-regulated by hypoxia in human PASMC, consistent with the activation of NFAT during this process. Down-regulation of miR-124 was also observed in 3-week hypoxia-treated mouse lungs. Furthermore, the overexpression of miR-124 not only inhibited human PASMC proliferation but also maintained its differentiated phenotype by repressing the NFAT pathway. Taken together, our data provide the first evidence that miR-124 acts as an inhibitor of the NFAT pathway. Down-regulation of miR-124 in hypoxia-treated PASMC and its antiproliferative and prodifferentiation effects imply a potential value for miR-124 in the treatment of PAH. PMID:23853098

Kang, Kang; Peng, Xiao; Zhang, Xiaoying; Wang, Yuna; Zhang, Lishu; Gao, Li; Weng, Tingting; Zhang, Honghao; Ramchandran, Ramaswamy; Raj, J. Usha; Gou, Deming; Liu, Lin

2013-01-01

320

Glycyrrhizin Exerts Antioxidative Effects in H5N1 Influenza A Virus-Infected Cells and Inhibits Virus Replication and Pro-Inflammatory Gene Expression  

PubMed Central

Glycyrrhizin is known to exert antiviral and anti-inflammatory effects. Here, the effects of an approved parenteral glycyrrhizin preparation (Stronger Neo-Minophafen C) were investigated on highly pathogenic influenza A H5N1 virus replication, H5N1-induced apoptosis, and H5N1-induced pro-inflammatory responses in lung epithelial (A549) cells. Therapeutic glycyrrhizin concentrations substantially inhibited H5N1-induced expression of the pro-inflammatory molecules CXCL10, interleukin 6, CCL2, and CCL5 (effective glycyrrhizin concentrations 25 to 50 µg/ml) but interfered with H5N1 replication and H5N1-induced apoptosis to a lesser extent (effective glycyrrhizin concentrations 100 µg/ml or higher). Glycyrrhizin also diminished monocyte migration towards supernatants of H5N1-infected A549 cells. The mechanism by which glycyrrhizin interferes with H5N1 replication and H5N1-induced pro-inflammatory gene expression includes inhibition of H5N1-induced formation of reactive oxygen species and (in turn) reduced activation of NF?B, JNK, and p38, redox-sensitive signalling events known to be relevant for influenza A virus replication. Therefore, glycyrrhizin may complement the arsenal of potential drugs for the treatment of H5N1 disease. PMID:21611183

Michaelis, Martin; Geiler, Janina; Naczk, Patrizia; Sithisarn, Patchima; Leutz, Anke; Doerr, Hans Wilhelm; Cinatl, Jindrich

2011-01-01

321

New-generation taxoid SB-T-1214 inhibits stem cell-related gene expression in 3D cancer spheroids induced by purified colon tumor-initiating cells  

PubMed Central

Background Growing evidence suggests that the majority of tumors are organized hierarchically, comprising a population of tumor-initiating, or cancer stem cells (CSCs) responsible for tumor development, maintenance and resistance to drugs. Previously we have shown that the CD133high/CD44high fraction of colon cancer cells is different from their bulk counterparts at the functional, morphological and genomic levels. In contrast to the majority of colon cancer cells expressing moderate levels of CD133, CD44 and CD166, cells with a high combined expression of CD133 and CD44 possessed several characteristic stem cell features, including profound self-renewal capacity in vivo and in vitro, and the ability to give rise to different cell phenotypes. The present study was undertaken for two aims: a) to determine stem cell-related genomic characteristics of floating 3D multicellular spheroids induced by CD133high/CD44high colon cancer cells; and b) to evaluate CSC-specific alterations induced by new-generation taxoid SB-T-1214. Results Selected CSC phenotype was isolated from three independent invasive colon cancer cell lines, HCT116, HT29 and DLD-1. A stem cell-specific PCR array assay (SABiosciences) revealed that colonospheres induced by purified CD133high/CD44high expressing cells display profound up-regulation of stem cell-related genes in comparison with their bulk counterparts. The FACS analysis has shown that the 3D colonospheres contained some minority cell populations with high levels of expression of Oct4, Sox2, Nanog and c-Myc, which are essential for stem cell pluripotency and self-renewal. Single administration of the SB-T-1214 at concentration 100 nM-1 ?M for 48 hr not only induced growth inhibition and apoptotic cell death in these three types of colon cancer spheroids in 3D culture, but also mediated massive inhibition of the stem cell-related genes and significant down-regulation of the pluripotency gene expression. PCR array and FACS data were confirmed with western blotting. Importantly, viable cells that survived this treatment regimen were no longer able to induce secondary floating spheroids and exhibited significant morphological abnormalities. Conclusions We report here that a new-generation taxoid SB-T-1214 possesses significant activity against colon cancer spheroids induced by and enriched with drug resistant tumorigenic CD133high/CD44high cells and efficiently inhibited expression of the majority of stem cell-related genes. Our data indicates that the previously observed long-term efficacy of SB-T-1214 against drug resistant colon tumors in vivo may be explained by the down-regulation of multiple stem cell-related genes in the tumorigenic cell population, in addition to its known efficacy as a mitotic poison against proliferating cancer cells. PMID:20630067

2010-01-01

322

Inhibition of hepatitis B virus gene expression by small interfering RNAs targeting cccDNA and X antigen.  

PubMed

To test the possible inhibition of hepatitis B virus (HBV) replication and expression by small interfering RNAs (siRNAs) targeting simultaneously covalenthy closed circular DNA (dnacccDNA) and X antigen, corresponding recombinant plasmids were transfected into HepG2.2.15 cells and the levels of cccDNA, HBXAg, HBcAg, and HBeAg were assayed at various times post transfection. As expected, the single siRNAs showed marked inhibitory effects but their combination was even more efficient. These results provide a new insight into the development of a potential anti-HBV strategy of enhancing the efficacy of individual antivirals and overcoming the high mutation rate of HBV. PMID:22404609

Xie, Q; Zhang, S; Wang, W; Li, Y M; Du, T; Su, X L; Wei, Y Q; Deng, H X

2012-01-01

323

Inhibition of Glucose-Stimulated Insulin Secretion by KCNJ15, a Newly Identified Susceptibility Gene for Type 2 Diabetes  

PubMed Central

Potassium inwardly rectifying channel, subfamily J, member 15 (KCNJ15) is a type 2 diabetes–associated risk gene, and Kcnj15 overexpression suppresses insulin secretion in rat insulinoma (INS1) cells. The aim of the current study was to characterize the role of Kcnj15 by knockdown of this gene in vitro and in vivo. Human islet cells were used to determine the expression of KCNJ15. Expression of KCNJ15 mRNA in islets was higher in subjects with type 2 diabetes. In INS1 cells, Kcnj15 expression was induced by high glucose–containing medium. Regulation of Kcnj15 by glucose and its effect on insulin secretion were analyzed in INS1 cells and in normal mice and diabetic mice by the inactivation of Kcnj15 using small interfering RNA. Knockdown of Kcnj15 increased the insulin secretion in vitro and in vivo. KCNJ15 and Ca2+-sensing receptor (CsR) interact in the kidney. Binding of Kcnj15 with CsR was also detected in INS1 cells. In conclusion, downregulation of Kcnj15 leads to increased insulin secretion in vitro and in vivo. The mechanism to regulate insulin secretion involves KCNJ15 and CsR. PMID:22566534

Okamoto, Koji; Iwasaki, Naoko; Doi, Kent; Noiri, Eisei; Iwamoto, Yasuhiko; Uchigata, Yasuko; Fujita, Toshiro; Tokunaga, Katsushi

2012-01-01

324

Adenovirus-mediated gene transfer of endostatin in vivo results in high level of transgene expression and inhibition of tumor growth and metastases  

NASA Astrophysics Data System (ADS)

Inhibition of angiogenesis has been shown to be an effective strategy in cancer therapy in mice. However, its widespread application has been hampered by difficulties in the large-scale production of the antiangiogenic proteins. This limitation may be resolved by in vivo delivery and expression of the antiangiogenic genes. We have constructed a recombinant adenovirus that expresses murine endostatin that is biologically active both in vitro, as determined in endothelial cell proliferation assays, and in vivo, by suppression of angiogenesis induced by vascular endothelial growth factor 165. Persistent high serum levels of endostatin (605-1740 ng/ml; mean, 936 ng/ml) were achieved after systemic administration of the vector to nude mice, which resulted in significant reduction of the growth rates and the volumes of JC breast carcinoma and Lewis lung carcinoma (P < 0.001 and P < 0.05, respectively). In addition, the endostatin vector treatment completely prevented the formation of pulmonary micrometastases in Lewis lung carcinoma (P = 0.0001). Immunohistochemical staining of the tumors demonstrated a decreased number of blood vessels in the treatment group versus the controls. In conclusion, the present study clearly demonstrates the potential of vector-mediated antiangiogenic gene therapy as a component in cancer therapy.

Sauter, Bernhard V.; Martinet, Olivier; Zhang, Wei-Jian; Mandeli, John; Woo, Savio L. C.

2000-04-01

325

EGCG protects endothelial cells against PCB 126-induced inflammation through inhibition of AhR and induction of Nrf2-regulated genes  

SciTech Connect

Tea flavonoids such as epigallocatechin gallate (EGCG) protect against vascular diseases such as atherosclerosis via their antioxidant and anti-inflammatory functions. Persistent and widespread environmental pollutants, including polychlorinated biphenyls (PCB), can induce oxidative stress and inflammation in vascular endothelial cells. Even though PCBs are no longer produced, they are still detected in human blood and tissues and thus considered a risk for vascular dysfunction. We hypothesized that EGCG can protect endothelial cells against PCB-induced cell damage via its antioxidant and anti-inflammatory properties. To test this hypothesis, primary vascular endothelial cells were pretreated with EGCG, followed by exposure to the coplanar PCB 126. Exposure to PCB 126 significantly increased cytochrome P450 1A1 (Cyp1A1) mRNA and protein expression and superoxide production, events which were significantly attenuated following pretreatment with EGCG. Similarly, EGCG also reduced DNA binding of NF-?B and downstream expression of inflammatory markers such as monocyte chemotactic protein-1 (MCP-1) and vascular cell adhesion protein-1 (VCAM-1) after PCB exposure. Furthermore, EGCG decreased endogenous or base-line levels of Cyp1A1, MCP-1 and VCAM-1 in endothelial cells. Most of all, treatment of EGCG upregulated expression of NF-E2-related factor 2 (Nrf2)-controlled antioxidant genes, including glutathione S transferase (GST) and NAD(P)H:quinone oxidoreductase 1 (NQO1), in a dose-dependent manner. In contrast, silencing of Nrf2 increased Cyp1A1, MCP-1 and VCAM-1 and decreased GST and NQO1 expression, respectively. These data suggest that EGCG can inhibit AhR regulated genes and induce Nrf2-regulated antioxidant enzymes, thus providing protection against PCB-induced inflammatory responses in endothelial cells. -- Highlights: ? PCBs cause endothelial inflammation and subsequent atherosclerosis. ? Nutrition can modulate toxicity by environmental pollutants. ? We demonstrated that EGCG can decrease PCB-induced inflammation. ? EGCG protection was via inhibition of AhR and induction of Nrf2 regulatory genes.

Han, Sung Gu [Superfund Research Program, University of Kentucky, Lexington, KY 40536 (United States) [Superfund Research Program, University of Kentucky, Lexington, KY 40536 (United States); Department of Animal and Food Sciences, College of Agriculture, University of Kentucky, Lexington, KY 40536 (United States); Han, Seong-Su [Department of Pathology, College of Medicine, University of Iowa, Iowa City, IA 52242 (United States)] [Department of Pathology, College of Medicine, University of Iowa, Iowa City, IA 52242 (United States); Toborek, Michal [Department of Neurosurgery, University of Kentucky, Lexington, KY 40536 (United States)] [Department of Neurosurgery, University of Kentucky, Lexington, KY 40536 (United States); Hennig, Bernhard, E-mail: bhennig@uky.edu [Superfund Research Program, University of Kentucky, Lexington, KY 40536 (United States) [Superfund Research Program, University of Kentucky, Lexington, KY 40536 (United States); Department of Animal and Food Sciences, College of Agriculture, University of Kentucky, Lexington, KY 40536 (United States)

2012-06-01

326

Inhibition of Annexin A2 gene transcription is a promising molecular target for hepatoma cell proliferation and metastasis  

PubMed Central

Hepatocyte Annexin A2 (ANXA2) expression is associated with the progression and metastasis of hepatocellular carcinoma (HCC). Circulating ANXA2 levels in HCC patients are significantly higher compared with that of patients with benign liver disease. ANXA2 levels have been found to correlate with hepatitis B virus infection, extrahepatic metastasis and portal vein thrombus. By contrast, ANXA2 levels do not correlate with tumour size and AFP levels. However, the underlying mechanisms of ANXA2 remain obscure. The results of the current study identified that abnormalities in hepatic ANXA2 expression were localised to the cell membrane and cytoplasm of HCC tissues and mainly in the cytoplasm of para-cancerous tissues. ANXA2 was overexpressed in MHCC97-H cells which have high metastatic potential. Following specific ANXA2-small hairpin RNA (shRNA) transfection in vitro, ANXA-2 was effectively inhibited and the S phase ratio of cells was 27.76%, compared with 36.14% in mock-treated cells. In addition, the invading cell ratio was reduced in the shRNA-treated group (52.16%) compared with the mock-treated group (86.14%). The growth and volume of xenograft tumours in vivo was significantly suppressed (P<0.05) in the shRNA group compared with that of the mock group, indicating that ANXA2 may be a novel and useful target for elucidating molecular mechanisms involving the proliferation and metastasis of HCC. PMID:24348815

DONG, ZHIZHEN; YAO, MIN; ZHANG, HAIJIAN; WANG, LI; HUANG, HUA; YAN, MEIJUAN; WU, WEI; YAO, DENGFU

2014-01-01

327

Inhibition of the expression of penicillin resistance in Streptococcus pneumoniae by inactivation of cell wall muropeptide branching genes  

PubMed Central

Penicillin-resistant strains of Streptococcus pneumoniae contain low affinity penicillin-binding proteins and often also produce abnormal indirectly crosslinked cell walls. However the relationship between cell wall abnormality and penicillin resistance has remained obscure. We now show that the genome of S. pneumoniae contains an operon composed of two genes (murM and murN) that encode enzymes involved with the biosynthesis of branched structured cell wall muropeptides. The sequences of murMN were compared in two strains: the penicillin-susceptible strain R36A producing the species-specific pneumococcal cell wall peptidoglycan in which branched stem peptides are rare, and the highly penicillin-resistant transformant strain Pen6, the cell wall of which is enriched for branched-structured stem peptides. The two strains carried different murM alleles: murM of the penicillin-resistant strain Pen6 had a “mosaic” structure encoding a protein that was only 86.5% identical to the product of murM identified in the isogenic penicillin-susceptible strain R36A. Mutants of R36A and Pen6 in which the murMN operon was interrupted by insertion-duplication mutagenesis produced peptidoglycan from which all branched muropeptide components were missing. The insertional mutant of Pen6 carried a pbp2x gene with the same “mosaic” sequence found in Pen6. On the other hand, inactivation of murMN in strain Pen6 and other resistant strains caused a virtually complete loss of penicillin resistance. Our observations indicate that the capacity to produce branched cell wall precursors plays a critical role in the expression of penicillin resistance in S. pneumoniae. PMID:10759563

Filipe, Sergio R.; Tomasz, Alexander

2000-01-01

328

S-adenosylmethionine decarboxylase gene expression in rat hepatoma cells: regulation by insulin and by inhibition of protein synthesis.  

PubMed Central

We have investigated expression of the S-adenosylmethionine decarboxylase (AdoMetDC) gene in H4-II-E rat hepatoma cells treated with growth factors (epidermal growth factor and transforming growth factor beta 1) and inducers (cAMP and insulin). Treatment with insulin caused a marked increase in both RNA level and enzyme activity. The stability of AdoMetDC mRNA was not altered by insulin treatment: the accumulation of mRNA in hepatoma cells therefore seems to be due to an increase in the transcription rate. Cycloheximide was found to be a strong inducer of AdoMetDC mRNA transcription and the effects of insulin and cycloheximide were additive, suggesting that they increase expression by separate mechanisms. Chloramphenicol acetyltransferase assays in rat hepatoma cells using 5' flanking regions of different lengths revealed that the promoter region extending 337 bp upstream from the transcription start site contains elements involved in insulin response. PMID:8645217

Soininen, T; Liisanantti, M K; Pajunen, A E

1996-01-01

329

Inhibition of AMPK and Krebs cycle gene expression drives metabolic remodeling of Pten-deficient preneoplastic thyroid cells  

PubMed Central

Rapidly proliferating and neoplastically transformed cells generate the energy required to support rapid cell division by increasing glycolysis and decreasing flux through the oxidative phosphorylation pathway (OXPHOS), usually without alterations in mitochondrial function. In contrast, little is known of the metabolic alterations, if any, which occur in cells harboring mutations that prime their neoplastic transformation. To address this question, we used a Pten-deficient mouse model to examine thyroid cells where a mild hyperplasia progresses slowly to follicular thyroid carcinoma. Using this model, we report that constitutive PI3K activation caused by PTEN deficiency in non-transformed thyrocytes results in a global down-regulation of Krebs cycle and OXPHOS gene expression, defective mitochondria, reduced respiration and an enhancement in compensatory glycolysis. We found that this process does not involve any of the pathways classically associated with the Warburg effect. Moreover, this process was independent of proliferation but contributed directly to thyroid hyperplasia. Our findings define a novel metabolic switch to glycolysis driven by PI3K-dependent AMPK inactivation with a consequent repression in the expression of key metabolic transcription regulators. PMID:23796563

Antico Arciuch, Valeria G.; Russo, Marika A.; Kang, Kristy S.; Di Cristofano, Antonio

2013-01-01

330

Inhibition of lung metastasis by chemokine CCL17-mediated in vivo silencing of genes in CCR4+ Tregs  

PubMed Central

Despite significant attractiveness of anti-sense oligonucleotide/RNAi technology, its clinical application has been precluded by a lack of methods for targeted delivery and transduction of primary immune cells in vivo. Here, we devised a chemokine CCL17-based strategy (TARC-arp) that transiently silences expression of genes in immune cells by delivering inhibitory oligonucleotides via their chemokine receptors. In modeling studies using mice with established 4T1.2 breast cancer, we show that IL10 produced by CCR4+ cells, in particular FoxP3+ regulatory T cells (Tregs), plays an important role in lung metastasis. As such, TARC-arp-mediated silencing of IL-10 or FoxP3 in CCR4+ Tregs is sufficient to block lung metastasis. Thus, we provide a simple solution that circumvents the problems of RNAi use in vivo, indicating that a disease outcome can be successfully controlled by delivering inhibitory oligonucleotides with chemokines to inactivate a selective subset of immune cells. PMID:23603860

Biragyn, Arya; Bodogai, Monica; Olkhanud, Purevdorj B.; Denny-Brown, Sinan R.; Puri, Nitin; Ayukawa, Koichi; Kanegasaki, Shiro; Hogaboam, Cory M.; Wejksza, Katarzyna; Lee-Chang, Catalina

2013-01-01

331

Inhibition of AMPK and Krebs cycle gene expression drives metabolic remodeling of Pten-deficient preneoplastic thyroid cells.  

PubMed

Rapidly proliferating and neoplastically transformed cells generate the energy required to support rapid cell division by increasing glycolysis and decreasing flux through the oxidative phosphorylation (OXPHOS) pathway, usually without alterations in mitochondrial function. In contrast, little is known of the metabolic alterations, if any, which occur in cells harboring mutations that prime their neoplastic transformation. To address this question, we used a Pten-deficient mouse model to examine thyroid cells where a mild hyperplasia progresses slowly to follicular thyroid carcinoma. Using this model, we report that constitutive phosphoinositide 3-kinase (PI3K) activation caused by PTEN deficiency in nontransformed thyrocytes results in a global downregulation of Krebs cycle and OXPHOS gene expression, defective mitochondria, reduced respiration, and an enhancement in compensatory glycolysis. We found that this process does not involve any of the pathways classically associated with the Warburg effect. Moreover, this process was independent of proliferation but contributed directly to thyroid hyperplasia. Our findings define a novel metabolic switch to glycolysis driven by PI3K-dependent AMPK inactivation with a consequent repression in the expression of key metabolic transcription regulators. PMID:23796563

Antico Arciuch, Valeria G; Russo, Marika A; Kang, Kristy S; Di Cristofano, Antonio

2013-09-01

332

Water-stress-induced inhibition of ?-tubulin gene expression during growth, and its implications for reproductive success in rice.  

PubMed

A drought-suppressed cDNA (RiP-3), encoding a putative ?-tubulin protein was isolated from rice panicle at pollen-mother-cell meiosis stage. Analysis of its deduced amino acid sequence showed all the typical structural motifs for plant ?-tubulins. The expression of ?-tubulin transcripts was observed in all the reproductive organs of rice panicle, and in 5- or 15-day old seedlings, but not in mature leaves. Expression levels were positively correlated with the regions and periods of high growth, and the transcript level declined in parallel with drought-induced reduction in growth rates in all tissues examined. Immunoblot analysis of proteins separated by SDS-PAGE with anti-?-tubulin monoclonal antibody showed that the level of protein paralleled the changes in the transcript abundance in these organs. In situ immunolocalization of the ?-tubulin protein in sections of the basal part of 5-day old seedlings showed that the highest levels of the protein were associated with the fastest growing leaf whorls, and the protein level declined upon a brief episode of water stress. Given the known critical role of tubulin in cell division and elongation, the results indicate that the expression of ?-tubulin gene may be part of the events that suppress panicle elongation during water deficit, which is in turn a suspected cause of male reproductive failure and yield reduction in rice. PMID:24814750

Sheoran, Inder S; Koonjul, Priyum; Attieh, Jihad; Saini, Hargurdeep S

2014-07-01

333

Isolation of a wheat (Triticum aestivum L.) mutant in ABA 8?-hydroxylase gene: effect of reduced ABA catabolism on germination inhibition under field condition  

PubMed Central

Pre-harvest sprouting, the germination of mature seeds on the mother plant under moist condition, is a serious problem in cereals. To investigate the effect of reduced abscisic acid (ABA) catabolism on germination in hexaploid wheat (Triticum aestivum L.), we cloned the wheat ABA 8?-hydroxyase gene which was highly expressed during seed development (TaABA8?OH1) and screened for mutations that lead to reduced ABA catabolism. In a screen for natural variation, one insertion mutation in exon 5 of TaABA8?OH1 on the D genome (TaABA8?OH1-D) was identified in Japanese cultivars including ‘Tamaizumi’. However, a single mutation in TaABA8?OH1-D had no clear effect on germination inhibition in double haploid lines. In a screen for a mutation, one deletion mutant lacking the entire TaABA8?OH1 on the A genome (TaABA8?OH1-A), TM1833, was identified from gamma-ray irradiation lines of ‘Tamaizumi’. TM1833 (a double mutant in TaABA8?OH1-A and TaABA8?OH1-D) showed lower TaABA8?OH1 expression, higher ABA content in embryos during seed development under field condition and lower germination than those in ‘Tamaizumi’ (a single mutant in TaABA8?OH1-D). These results indicate that reduced ABA catabolism through mutations in TaABA8?OH1 may be effective in germination inhibition in field-grown wheat. PMID:23641187

Chono, Makiko; Matsunaka, Hitoshi; Seki, Masako; Fujita, Masaya; Kiribuchi-Otobe, Chikako; Oda, Shunsuke; Kojima, Hisayo; Kobayashi, Daisuke; Kawakami, Naoto

2013-01-01

334

Isolation of a wheat (Triticum aestivum L.) mutant in ABA 8'-hydroxylase gene: effect of reduced ABA catabolism on germination inhibition under field condition.  

PubMed

Pre-harvest sprouting, the germination of mature seeds on the mother plant under moist condition, is a serious problem in cereals. To investigate the effect of reduced abscisic acid (ABA) catabolism on germination in hexaploid wheat (Triticum aestivum L.), we cloned the wheat ABA 8'-hydroxyase gene which was highly expressed during seed development (TaABA8'OH1) and screened for mutations that lead to reduced ABA catabolism. In a screen for natural variation, one insertion mutation in exon 5 of TaABA8'OH1 on the D genome (TaABA8'OH1-D) was identified in Japanese cultivars including 'Tamaizumi'. However, a single mutation in TaABA8'OH1-D had no clear effect on germination inhibition in double haploid lines. In a screen for a mutation, one deletion mutant lacking the entire TaABA8'OH1 on the A genome (TaABA8'OH1-A), TM1833, was identified from gamma-ray irradiation lines of 'Tamaizumi'. TM1833 (a double mutant in TaABA8'OH1-A and TaABA8'OH1-D) showed lower TaABA8'OH1 expression, higher ABA content in embryos during seed development under field condition and lower germination than those in 'Tamaizumi' (a single mutant in TaABA8'OH1-D). These results indicate that reduced ABA catabolism through mutations in TaABA8'OH1 may be effective in germination inhibition in field-grown wheat. PMID:23641187

Chono, Makiko; Matsunaka, Hitoshi; Seki, Masako; Fujita, Masaya; Kiribuchi-Otobe, Chikako; Oda, Shunsuke; Kojima, Hisayo; Kobayashi, Daisuke; Kawakami, Naoto

2013-03-01

335

A Novel Rice Cytochrome P450 Gene, CYP72A31, Confers Tolerance to Acetolactate Synthase-Inhibiting Herbicides in Rice and Arabidopsis.  

PubMed

Target-site and non-target-site herbicide tolerance are caused by the prevention of herbicide binding to the target enzyme and the reduction to a nonlethal dose of herbicide reaching the target enzyme, respectively. There is little information on the molecular mechanisms involved in non-target-site herbicide tolerance, although it poses the greater threat in the evolution of herbicide-resistant weeds and could potentially be useful for the production of herbicide-tolerant crops because it is often involved in tolerance to multiherbicides. Bispyribac sodium (BS) is an herbicide that inhibits the activity of acetolactate synthase. Rice (Oryza sativa) of the indica variety show BS tolerance, while japonica rice varieties are BS sensitive. Map-based cloning and complementation tests revealed that a novel cytochrome P450 monooxygenase, CYP72A31, is involved in BS tolerance. Interestingly, BS tolerance was correlated with CYP72A31 messenger RNA levels in transgenic plants of rice and Arabidopsis (Arabidopsis thaliana). Moreover, Arabidopsis overexpressing CYP72A31 showed tolerance to bensulfuron-methyl (BSM), which belongs to a different class of acetolactate synthase-inhibiting herbicides, suggesting that CYP72A31 can metabolize BS and BSM to a compound with reduced phytotoxicity. On the other hand, we showed that the cytochrome P450 monooxygenase CYP81A6, which has been reported to confer BSM tolerance, is barely involved, if at all, in BS tolerance, suggesting that the CYP72A31 enzyme has different herbicide specificities compared with CYP81A6. Thus, the CYP72A31 gene is a potentially useful genetic resource in the fields of weed control, herbicide development, and molecular breeding in a broad range of crop species. PMID:24406793

Saika, Hiroaki; Horita, Junko; Taguchi-Shiobara, Fumio; Nonaka, Satoko; Nishizawa-Yokoi, Ayako; Iwakami, Satoshi; Hori, Kiyosumi; Matsumoto, Takashi; Tanaka, Tsuyoshi; Itoh, Takeshi; Yano, Masahiro; Kaku, Koichiro; Shimizu, Tsutomu; Toki, Seiichi

2014-11-01

336

Human Papillomavirus 16 Oncoprotein E7 Stimulates UBF1-Mediated rDNA Gene Transcription, Inhibiting a p53-Independent Activity of p14ARF  

PubMed Central

High-risk human papillomavirus oncoproteins E6 and E7 play a major role in HPV-related cancers. One of the main functions of E7 is the degradation of pRb, while E6 promotes the degradation of p53, inactivating the p14ARF-p53 pathway. pRb and p14ARF can repress ribosomal DNA (rDNA) transcription in part by targeting the Upstream Binding Factor 1 (UBF1), a key factor in the activation of RNA polymerase I machinery. We showed, through ectopic expression and siRNA silencing of p14ARF and/or E7, that E7 stimulates UBF1-mediated rDNA gene transcription, partly because of increased levels of phosphorylated UBF1, preventing the inhibitory function of p14ARF. Unexpectedly, activation of rDNA gene transcription was higher in cells co-expressing p14ARF and E7, compared to cells expressing E7 alone. We did not find a difference in P-UBF1 levels that could explain this data. However, p14ARF expression induced E7 to accumulate into the nucleolus, where rDNA transcription takes place, providing an opportunity for E7 to interact with nucleolar proteins involved in this process. GST-pull down and co-immunoprecipitation assays showed interactions between p14ARF, UBF1 and E7, although p14ARF and E7 are not able to directly interact. Co-expression of a pRb-binding-deficient mutant (E7C24G) and p14ARF resulted in EC24G nucleolar accumulation, but not in a significant higher activation of rDNA transcription, suggesting that the inactivation of pRb is involved in this phenomenon. Thus, p14ARF fails to prevent E7-mediated UBF1 phosphorylation, but could facilitate nucleolar pRb inactivation by targeting E7 to the nucleolus. While others have reported that p19ARF, the mouse homologue of p14ARF, inhibits some functions of E7, we showed that E7 inhibits a p53-independent function of p14ARF. These results point to a mutually functional interaction between p14ARF and E7 that might partly explain why the sustained p14ARF expression observed in most cervical pre-malignant lesions and malignancies may be ineffective. PMID:24798431

Dichamp, Isabelle; Seite, Paule; Agius, Gerard; Barbarin, Alice; Beby-Defaux, Agnes

2014-01-01

337

Human papillomavirus 16 oncoprotein E7 stimulates UBF1-mediated rDNA gene transcription, inhibiting a p53-independent activity of p14ARF.  

PubMed

High-risk human papillomavirus oncoproteins E6 and E7 play a major role in HPV-related cancers. One of the main functions of E7 is the degradation of pRb, while E6 promotes the degradation of p53, inactivating the p14ARF-p53 pathway. pRb and p14ARF can repress ribosomal DNA (rDNA) transcription in part by targeting the Upstream Binding Factor 1 (UBF1), a key factor in the activation of RNA polymerase I machinery. We showed, through ectopic expression and siRNA silencing of p14ARF and/or E7, that E7 stimulates UBF1-mediated rDNA gene transcription, partly because of increased levels of phosphorylated UBF1, preventing the inhibitory function of p14ARF. Unexpectedly, activation of rDNA gene transcription was higher in cells co-expressing p14ARF and E7, compared to cells expressing E7 alone. We did not find a difference in P-UBF1 levels that could explain this data. However, p14ARF expression induced E7 to accumulate into the nucleolus, where rDNA transcription takes place, providing an opportunity for E7 to interact with nucleolar proteins involved in this process. GST-pull down and co-immunoprecipitation assays showed interactions between p14ARF, UBF1 and E7, although p14ARF and E7 are not able to directly interact. Co-expression of a pRb-binding-deficient mutant (E7C24G) and p14ARF resulted in EC24G nucleolar accumulation, but not in a significant higher activation of rDNA transcription, suggesting that the inactivation of pRb is involved in this phenomenon. Thus, p14ARF fails to prevent E7-mediated UBF1 phosphorylation, but could facilitate nucleolar pRb inactivation by targeting E7 to the nucleolus. While others have reported that p19ARF, the mouse homologue of p14ARF, inhibits some functions of E7, we showed that E7 inhibits a p53-independent function of p14ARF. These results point to a mutually functional interaction between p14ARF and E7 that might partly explain why the sustained p14ARF expression observed in most cervical pre-malignant lesions and malignancies may be ineffective. PMID:24798431

Dichamp, Isabelle; Séité, Paule; Agius, Gérard; Barbarin, Alice; Beby-Defaux, Agnès

2014-01-01

338

The metastasis suppressor, N-myc downstream-regulated gene 1 (NDRG1), inhibits stress-induced autophagy in cancer cells.  

PubMed

N-myc downstream regulated gene 1 (NDRG1) is a potent metastasis suppressor with an undefined role in the stress response. Autophagy is a pro-survival pathway and can be regulated via the protein kinase-like endoplasmic reticulum kinase (PERK)/eIF2?-mediated endoplasmic reticulum (ER) stress pathway. Hence, we investigated the role of NDRG1 in stress-induced autophagy as a mechanism of inhibiting metastasis via the induction of apoptosis. As thiosemicarbazone chelators induce stress and up-regulate NDRG1 to inhibit metastasis, we studied their effects on the ER stress response and autophagy. This was important to assess, as little is understood regarding the role of the stress induced by iron depletion and its role in autophagy. We observed that the chelator, di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone (Dp44mT), which forms redox-active iron and copper complexes, effectively induced ER stress as shown by activation of the PERK/eIF2? pathway. Dp44mT also increased the expression of the autophagic marker, LC3-II, and this was dependent on activation of the PERK/eIF2? axis, as silencing PERK prevented LC3-II accumulation. The effect of Dp44mT on LC3-II expression was at least partially due to iron-depletion, as this effect was also demonstrated with the classical iron chelator, desferrioxamine (DFO), and was not observed for the DFO-iron complex. NDRG1 overexpression also inhibited basal autophagic initiation and the ER stress-mediated autophagic pathway via suppression of the PERK/eIF2? axis. Moreover, NDRG1-mediated suppression of the pro-survival autophagic pathway probably plays a role in its anti-metastatic effects by inducing apoptosis. In fact, multiple pro-apoptotic markers were increased, whereas anti-apoptotic Bcl-2 was decreased upon NDRG1 overexpression. This study demonstrates the role of NDRG1 as an autophagic inhibitor that is important for understanding its mechanism of action. PMID:24532803

Sahni, Sumit; Bae, Dong-Hun; Lane, Darius J R; Kovacevic, Zaklina; Kalinowski, Danuta S; Jansson, Patric J; Richardson, Des R

2014-04-01

339

The Niemann-Pick C1 gene is downregulated in livers of C57BL/6J mice by dietary fatty acids, but not dietary cholesterol, through feedback inhibition of the SREBP pathway.  

PubMed

The Niemann-Pick C1 (NPC1) gene is associated with human obesity. Mouse models with decreased Npc1 gene dosage are susceptible to weight gain when fed a high-fat diet, but not a low-fat diet, consistent with an Npc1 gene-diet interaction. The objectives of this study were to define regulation of the Npc1 gene and to investigate the Npc1 gene-diet interaction responsible for weight gain. The experimental design involved feeding C57BL/6J male mice a low-fat diet (with 0.00, 0.10, or 1.00% cholesterol) or a high-fat diet (with 0.02% cholesterol) until 30 wk to determine regulation of the Npc1 gene in liver. The key results showed that the Npc1 gene was downregulated by dietary fatty acids (54%, P = 0.022), but not by dietary cholesterol, through feedback inhibition of the sterol regulatory element-binding protein (SREBP) pathway. However, the dietary fatty acids secondarily increased liver cholesterol, which also inhibits the SREBP pathway. Similarly, the Npc1 gene was downregulated in peritoneal fibroblasts isolated from C57BL/6J weanling male mice not exposed to the experimental diets and incubated in media supplemented with purified oleic acid (37%, P = 0.038) but not in media supplemented with purified cholesterol. These results are important because they suggest a novel mechanism for the interaction of fatty acids with the Npc1 gene to influence energy balance and to promote weight gain. Moreover, the responsiveness of the Npc1 gene to fatty acids is consistent with studies that suggest that the encoded NPC1 protein has a physiologic role in regulating both cholesterol and fatty acid metabolism. PMID:22990467

Jelinek, David; Castillo, Joseph J; Richardson, Lisa M; Luo, Li; Heidenreich, Randall A; Garver, William S

2012-11-01

340

Nocistatin inhibits pregnant rat uterine contractions in vitro: roles of calcitonin gene-related peptide and calcium-dependent potassium channel.  

PubMed

The endogenous neuropeptide nociceptin/orphanin FQ, translated from the prepronociceptin gene, exerts a contraction-inhibitory effect on the rat uterus. As nocistatin has been reported to cause functional antagonism of the pro-nociceptive effects of nociceptin, we set out to investigate its effects on the pregnant rat uterus and to elucidate its signalling pathway. The expression of prepronociceptin mRNA in the uterus and nocistatin levels in the uterus and the plasma were confirmed by RT-PCR and radioimmunoassay. The uterine levels of prepronociceptin mRNA and nocistatin were significantly increased by the last day of pregnancy, while the plasma nocistatin levels remained unchanged. In the isolated organ bath studies nocistatin inhibited the prostaglandin- and the KCl-evoked contractions in the uterus dose-dependently. This latter effect was decreased by preincubation with capsaicin. Incubation with calcitonin gene-related peptide after capsaicin treatment caused an elevation in the contraction-inhibitory effect of nocistatin. The effect of nocistatin was also decreased by the Ca(2+)-dependent K(+) channel inhibitor paxilline, against spontaneous uterine contractions. Nociceptin potentiated the action of nocistatin. Naloxone decreased the effect of nocistatin administered either alone or in combination with nociceptin. In Ca(2+)-poor environment, this effect of naloxone was suspended. Enzyme immunoassay for the uterine intracellular cAMP levels partially confirmed the results of in vitro contractility studies. We conclude that nocistatin, generated locally in the uterus, exerts an inhibitory effect, the mechanism being mediated in part by Ca(2+)-dependent K(+) channels, the elevation of cAMP levels and sensory neuropeptides. PMID:23792038

Deák, Beáta H; Klukovits, Anna; Tekes, Kornélia; Ducza, Eszter; Falkay, George; Gáspár, Róbert

2013-08-15

341

Pheromone exposure influences preoptic arginine vasotocin gene expression and inhibits social approach behavior in response to rivals, but not potential mates  

PubMed Central

The nonapeptides arginine vasotocin (AVT) and vasopressin (AVP) mediate a variety of social behaviors in vertebrates. However, the effects of these peptides on behavior can vary considerably both between and within species. AVT, in particular, stimulates aggressive and courtship responses typical of dominant males in several species, although it can also inhibit social interactions in some cases. Such differential effects may depend upon AVT influences within brain circuits that differ among species or between males that adopt alternative reproductive phenotypes and/or upon the differential activation of those circuits in different social contexts. However, to date, very little is known about how social stimuli that promote alternative behavioral responses influence AVT circuits within the brain. To address this issue, we exposed adult male goldfish to androstenedione (AD), a pheromonal signal that is released by both males and females during the breeding season, and measured social approach responses of males towards same- and other-sex individuals before and after AD exposure. In a second experiment, we measured AD-induced AVT gene expression using in situ hybridization. We found that brief exposure to AD induces social avoidance in response to rival males, but does not affect the level of sociality exhibited in response to sexually receptive females. Exposure to AD also increases AVT gene expression in the preoptic area of male goldfish, particularly in the parvocellular population of the preoptic nucleus. Together, these data suggest that AD is part of a social signaling system that induces social withdrawal specifically during male-male interactions by activating AVT neurons. PMID:23712040

Mangiamele, Lisa A.; Keeney, Alex D.T.; D'Agostino, Erin N.; Thompson, Richmond R.

2013-01-01

342

EGCG protects endothelial cells against PCB 126-induced inflammation through inhibition of AhR and induction of Nrf2-regulated genes.  

PubMed

Tea flavonoids such as epigallocatechin gallate (EGCG) protect against vascular diseases such as atherosclerosis via their antioxidant and anti-inflammatory functions. Persistent and widespread environmental pollutants, including polychlorinated biphenyls (PCB), can induce oxidative stress and inflammation in vascular endothelial cells. Even though PCBs are no longer produced, they are still detected in human blood and tissues and thus considered a risk for vascular dysfunction. We hypothesized that EGCG can protect endothelial cells against PCB-induced cell damage via its antioxidant and anti-inflammatory properties. To test this hypothesis, primary vascular endothelial cells were pretreated with EGCG, followed by exposure to the coplanar PCB 126. Exposure to PCB 126 significantly increased cytochrome P450 1A1 (Cyp1A1) mRNA and protein expression and superoxide production, events which were significantly attenuated following pretreatment with EGCG. Similarly, EGCG also reduced DNA binding of NF-?B and downstream expression of inflammatory markers such as monocyte chemotactic protein-1 (MCP-1) and vascular cell adhesion protein-1 (VCAM-1) after PCB exposure. Furthermore, EGCG decreased endogenous or base-line levels of Cyp1A1, MCP-1 and VCAM-1 in endothelial cells. Most of all, treatment of EGCG upregulated expression of NF-E2-related factor 2 (Nrf2)-controlled antioxidant genes, including glutathione S transferase (GST) and NAD(P)H:quinone oxidoreductase 1 (NQO1), in a dose-dependent manner. In contrast, silencing of Nrf2 increased Cyp1A1, MCP-1 and VCAM-1 and decreased GST and NQO1 expression, respectively. These data suggest that EGCG can inhibit AhR regulated genes and induce Nrf2-regulated antioxidant enzymes, thus providing protection against PCB-induced inflammatory responses in endothelial cells. PMID:22521609

Han, Sung Gu; Han, Seong-Su; Toborek, Michal; Hennig, Bernhard

2012-06-01

343

Vitamin E Conditionally Inhibits Atherosclerosis in ApoE Knockout Mice by Anti-oxidation and Regulation of Vasculature Gene Expressions.  

PubMed

Lipid deposition in artery walls is implied in the pathogenesis of atherosclerosis and imbalance between uptake and efflux of cholesterol favors the deposition. We investigated the effect of vitamin E with the same dose and duration on the different stages of atherosclerosis in Apolipoprotein E knockout (ApoE KO) mice and explored the potential mechanisms. The results showed that the ApoE KO mouse spontaneously develops atherosclerosis in an age-dependent manner from 14 to 46 weeks on the regular chow. Vitamin E (100 mg/kg) supplementation to ApoE KO mice at 6, 14, and 22 weeks for 8 weeks significantly reduced the atherosclerotic lesion area by 41, 29 and 19 % respectively compared to the age-matched control mice; however had no significant effect on the lesion when given at 30 and 38 weeks. In addition, vitamin E supplemented at the ages from 6 to 30 weeks decreased the contents of serum oxLDL and TBARS without affecting the TC and TAG contents in serum and liver. Furthermore, vitamin E supplemented at 6, 14 and 22 weeks down-regulated vasculature mRNA expressions of scavenger receptor CD36 and up-regulated mRNA expressions of PPAR?, LXR? and ABCA1 which are involved in reverse cholesterol transportation; however had no significant effects on these genes when given at 30 and 38 weeks. In conclusion, vitamin E with same dose and duration inhibits the early but not advanced atherosclerotic lesion in ApoE KO mice by anti-oxidation and regulation of mRNA expression of genes involved in cholesterol uptake and efflux, which favors the improvement of atherosclerosis. PMID:25385496

Tang, Futian; Lu, Meili; Zhang, Suping; Mei, Meng; Wang, Tieqiao; Liu, Peiqing; Wang, Hongxin

2014-12-01

344

Identification of a mutation in the tyrosinase related protein 1 (TRP1) gene associated with brown oculocutaneous albinism (OCA3)  

SciTech Connect

The genes responsible for the two most common types of human oculocutaneous albinism (OCA) have been identified. Mutations of the tyrosinase gene (chromosome 11q14-21) produce OCA1, and mutations of the P gene (chromosome 15q11.2-13) produce OCA2. Another type of OCA known as brown OCA or OCA3 is found commonly in the African and African-American population. OCA3 is characterized by light brown skin and hair with the ocular features of albinism and represents the third most frequent type of OCA. We previously identified dizygotic African-American twin boys who were discordant for OCA3. Melanocytes from the affected twin produced brown melanin and contained no detectable TRP1 protein. We have now characterized the TRP1 gene from the affected twin. The human TRP1 gene, homologous to the murine brown locus, contains 8 exons and maps to chromosome 9p23. Using PCR amplification of each exon coupled with SSCP analysis and direct DNA sequencing, we found the affected twin to homozygous for a single bp deletion in exon 6. The deletion removes a G in codon 368 leading to a premature stop at codon 384. We also identified a Tsp509 polymorphism in the 3{prime} UTR. We conclude that mutations of the TRP1 gene are responsible for brown OCA or OCA3, making this the third major OCA gene identified in humans.

Wildenberg, S.C.; Oetting, W.S.; Fryer, J.P. [Univ. of Minnesota, Minneapolis, MN (United States)] [and others

1994-09-01

345

Cloning and functional analysis of three genes encoding polygalacturonase-inhibiting proteins from Capsicum annuum and transgenic CaPGIP1 in tobacco in relation to increased resistance to two fungal pathogens.  

PubMed

Polygalacturonase-inhibiting proteins (PGIPs) are plant cell wall glycoproteins that can inhibit fungal endopolygalacturonases (PGs). The PGIPs directly reduce the aggressive potential of PGs. Here, we isolated and functionally characterized three members of the pepper (Capsicum annuum) PGIP gene family. Each was up-regulated at a different time following stimulation of the pepper leaves by Phytophthora capcisi and abiotic stresses including salicylic acid, methyl jasmonate, abscisic acid, wounding and cold treatment. Purified recombinant proteins individually inhibited activity of PGs produced by Alternaria alternata and Colletotrichum nicotianae, respectively, and virus-induced gene silencing in pepper conferred enhanced susceptibility to P. capsici. Because three PGIP genes acted similarily in conferring resistance to infection by P. capsici, and because individually purified proteins showed consistent inhibition against PG activity of both pathogens, CaPGIP1 was selected for manipulating transgenic tobacco. The crude proteins from transgenic tobacco exhibited distinct enhanced resistance to PG activity of both fungi. Moreover, the transgenic tobacco showed effective resistance to infection and a significant reduction in the number of infection sites, number of lesions and average size of lesions in the leaves. All results suggest that CaPGIPs may be involved in plant defense response and play an important role in a plant's resistance to disease. PMID:23334855

Wang, Xiuju; Zhu, Xiaoping; Tooley, Paul; Zhang, Xiuguo

2013-03-01

346

Trichostatin A induces mesenchymal-like morphological change and gene expression but inhibits migration and colony formation in human cancer cells.  

PubMed

Histone deacetylases (HDACs) are enzymes that catalyze the removal of acetyl from lysine residues in histones and other proteins, which results in gene transcriptional repression and subsequent changes in signaling events. HDACs inhibitors (HDACIs) have been used to reverse the aberrant epigenetic changes associated with cancer. However, the effects of HDACIs on epithelial-mesenchymal transition (EMT) in human cancer cells remain unclear. EMT is a fundamental process governing morphogenesis in multicellular organisms and promotes cancer invasion and metastasis. In this study, human cancer cells were treated with the HDACI trichostatin A (TSA). TSA was found to induce mesenchymal?like morphological changes in BGC-823 human gastric cancer and MCF-7 breast cancer cells, and increase the expression levels of the mesenchymal markers Vimentin and Twist. However, the expression levels of the epithelial cell marker E-cadherin were also increased in response to TSA treatment, while cell migration was reduced by TSA. Furthermore, TSA decreased cancer cell colony formation in BGC-823 and MCF-7 cells, and led to the deregulation of ?-catenin, a critical signaling molecule involved in EMT. In conclusion, the results suggested that TSA exhibits dual functions in EMT induction and inhibition in human cancer cells, but the detailed mechanisms require further investigation. PMID:25269990

Han, Rong-Fei; Li, Kai; Yang, Zi-Shan; Chen, Zhi-Guo; Yang, Wan-Cai

2014-12-01

347

Notch1 signaling inhibits growth of EC109 esophageal carcinoma cells through downmodulation of HPV18 E6/E7 gene expression  

PubMed Central

Aim: To investigate the role of the Notch1 signaling pathway in growth arrest of an esophageal carcinoma cell line (EC109) in vitro and the mechanism involved. Methods: An intracellular domain of Notch1 (ICN) was transfected into cultured EC109 cells by lipofectamine transfection. Subsequently, the proliferation of the transfected cells was measured by an MTT assay. Cell cycle distribution was analyzed by flow cytometry. Human papillomavirus type 18 (HPV18) E6/E7 mRNA expression was detected by RT-PCR, and p53 protein expression was detected by Western blot. Results: Activation of Notch1 signaling resulted in inhibition of EC109 cell proliferation with the induction of G2/M arrest, downmodulation of HPV18 E6/E7 gene expression, and upregulation of p53 expression. Conclusion: Repression of HPV18 E6/E7 expression by Notch1 signaling results in the activation of p53-mediated pathways with concomitant growth suppression of HPV18-positive EC109 cells. PMID:19122673

Zhang, Ke-jie; Lu, Quan-yi; Niu, Xiao-qing; Zhang, Peng; Zhao, Jiang-ning; Wang, Zhao; Hu, Jia-sheng; Li, Pu; Liu, Wen-li

2009-01-01

348

Signal-transducing mechanisms of ketamine-caused inhibition of interleukin-1{beta} gene expression in lipopolysaccharide-stimulated murine macrophage-like Raw 264.7 cells  

SciTech Connect

Ketamine may affect the host immunity. Interleukin-1{beta} (IL-1{beta}), IL-6, and tumor necrosis factor-{alpha} (TNF-{alpha}) are pivotal cytokines produced by macrophages. This study aimed to evaluate the effects of ketamine on the regulation of inflammatory cytokine gene expression, especially IL-1{beta}, in lipopolysaccharide (LPS)-activated murine macrophage-like Raw 264.7 cells and its possible signal-transducing mechanisms. Administration of Raw 264.7 cells with a therapeutic concentration of ketamine (100 {mu}M), LPS, or a combination of ketamine and LPS for 1, 6, and 24 h was not cytotoxic to macrophages. Exposure to 100 {mu}M ketamine decreased the binding affinity of LPS and LPS-binding protein but did not affect LPS-induced RNA and protein synthesis of TLR4. Treatment with LPS significantly increased IL-1{beta}, IL-6, and TNF-{alpha} gene expressions in Raw 264.7 cells. Ketamine at a clinically relevant concentration did not affect the synthesis of these inflammatory cytokines, but significantly decreased LPS-caused increases in these cytokines. Immunoblot analyses, an electrophoretic mobility shift assay, and a reporter luciferase activity assay revealed that ketamine significantly decreased LPS-induced translocation and DNA binding activity of nuclear factor-kappa B (NF{kappa}B). Administration of LPS sequentially increased the phosphorylations of Ras, Raf, MEK1/2, ERK1/2, and IKK. However, a therapeutic concentration of ketamine alleviated such augmentations. Application of toll-like receptor 4 (TLR4) small interfering (si)RNA reduced cellular TLR4 amounts and ameliorated LPS-induced RAS activation and IL-1{beta} synthesis. Co-treatment with ketamine and TLR4 siRNA synergistically ameliorated LPS-caused enhancement of IL-1{beta} production. Results of this study show that a therapeutic concentration of ketamine can inhibit gene expression of IL-1{beta} possibly through suppressing TLR4-mediated signal-transducing phosphorylations of Ras, Raf, MEK1/2, ERK1/2, and IKK and subsequent translocation and transactivation of NF{kappa}B.

Chen, T.-L. [Department of Anesthesiology, Taipei Medical University, Taipei, Taiwan (China); Chang, C.-C. [Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan (China); Lin, Y.-L. [Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan (China); Drug Abuse Research Center and Cell Physiology and Molecular Image Research Center, Taipei Medical University-Wan Fang Hospital, Taipei, Taiwan (China); Ueng, Y.-F. [Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan (China); National Research Institute of Chinese Medicine, Taipei, Taiwan (China); Chen, R.-M. [Graduate Institute of Medical Sciences, College of Medicine, Taipei Medical University, Taipei, Taiwan (China); Drug Abuse Research Center and Cell Physiology and Molecular Image Research Center, Taipei Medical University-Wan Fang Hospital, Taipei, Taiwan (China); Department of Anesthesiology, Taipei Medical University-Wan Fang Hospital, Taipei, Taiwan (China)], E-mail: rmchen@tmu.edu.tw

2009-10-01

349

Inhibition of Irvingia gabonensis seed extract (OB131) on adipogenesis as mediated via down regulation of the PPARgamma and Leptin genes and up-regulation of the adiponectin gene  

PubMed Central

Background Endeavors to manage obesity have been heavily reliant on controlling energy intake and expenditure equilibrium, but have failed to curtail the overweight and obesity epidemic. This dynamic equilibrium is more complex than originally postulated and is influenced by lifestyle, calorie and nutrient intake, reward cravings and satiation, energy metabolism, stress response capabilities, immune metabolism and genetics. Fat metabolism is an important indicator of how efficiently and to what extent these factors are competently integrating. We investigated whether an Irvingia gabonensis seed extract (IGOB131) would provide a more beneficial comprehensive approach influencing multiple mechanisms and specifically PPAR gamma, leptin and adiponectin gene expressions, important in anti-obesity strategies. Methods Using murine 3T3-L1 adipocytes as a model for adipose cell biology research, the effects of IGOB131 were investigated on PPAR gamma, adiponectin, and leptin. These adipocytes were harvested 8 days after the initiation of differentiation and treated with 0 to 250 microM of IGOB131 for 12 and 24 h at 37 degree C in a humidified 5 percent CO2 incubator. The relative expression of PPAR gamma, adiponectin, and leptin in 3T3-L1 adipocytes was quantified densitometrically using the software LabWorks 4.5, and calculated according to the reference bands of beta-actin. Results The IGOB131 significantly inhibited adipogenesis in adipocytes. The effect appears to be mediated through the down-regulated expression of adipogenic transcription factors (PPAR gamma) [P less than 0.05] and adipocyte-specific proteins (leptin) [P less than 0.05], and by up-regulated expression of adiponectin [P less than 0.05]. Conclusion IGOB131 may play an important multifaceted role in the control of adipogenesis and have further implications in in-vivo anti obesity effects by targeting the PPAR gamma gene, a known contributory factor to obesity in humans. PMID:19014517

Oben, Julius E; Ngondi, Judith L; Blum, Kenneth

2008-01-01

350

Curcumin blocks migration and invasion of mouse-rat hybrid retina ganglion cells (N18) through the inhibition of MMP-2, -9, FAK, Rho A and Rock-1 gene expression.  

PubMed

Cancer metastasis involves multiple processes which may complicate clinical management and even lead to death. Matrix metalloproteinases (MMPs) play an important role in cancer cell invasion, metastasis and angiogenesis, depending on whether agents can inhibit MMPs which could lead to inhibition of the migration and invasion of cancer cells. Curcumin, the active constituent of the dietary spice turmeric, has potential for the prevention and therapy of cancer. However, there is no study to address the effects of curcumin on migration and invasion of mouse-rat hybrid retina ganglion cells (N18). This is the first study to explore the anti-migration and -invasion of curcumin in mouse-rat hybrid retina ganglion cells (N18) in vitro. Curcumin exerted a dose- and time-dependent inhibitory effect on the invasion and migration of N18 cells in vitro. Results from Western blotting showed that curcumin inhibited the protein levels of PKC, FAK, NF-kappaB p65 and Rho A leading to the inhibition of ERK1/2, MKK7, COX-2 and ROCK1, respectively, finally causing the inhibition of MMP-2 and -9 for the inhibition of migration and invasion of N18 cells. Moreover, this action was involved in the inhibition of gene expression of MMP-2 and -7, FAK, ROCK1 and Rho A. Overall, the above data show that the anticancer effect of curcumin also exists for the inhibition of migration and invasion in N18 cells, and that curcumin may be a powerful candidate for developing preventive agents for cancer metastasis. PMID:20127004

Lin, Hui-Ju; Su, Chin-Cheng; Lu, Hsu-Feng; Yang, Jai-Sing; Hsu, Shu-Chun; Ip, Siu-Wan; Wu, Jia-Jiuan; Li, Yu-Ching; Ho, Chin-Chin; Wu, Chih-Chung; Chung, Jing-Gung

2010-03-01

351

Molt regulation in green and red color morphs of the crab Carcinus maenas: gene expression of molt-inhibiting hormone signaling components.  

PubMed

In decapod crustaceans, regulation of molting is controlled by the X-organ/sinus gland complex in the eyestalks. The complex secretes molt-inhibiting hormone (MIH), which suppresses production of ecdysteroids by the Y-organ (YO). MIH signaling involves nitric oxide and cGMP in the YO, which expresses nitric oxide synthase (NOS) and NO-sensitive guanylyl cyclase (GC-I). Molting can generally be induced by eyestalk ablation (ESA), which removes the primary source of MIH, or by multiple leg autotomy (MLA). In our work on Carcinus maenas, however, ESA has limited effects on hemolymph ecdysteroid titers and animals remain in intermolt at 7 days post-ESA, suggesting that adults are refractory to molt induction techniques. Consequently, the effects of ESA and MLA on molting and YO gene expression in C. maenas green and red color morphotypes were determined at intermediate (16 and 24 days) and long-term (~90 days) intervals. In intermediate-interval experiments, ESA of intermolt animals caused transient twofold to fourfold increases in hemolymph ecdysteroid titers during the first 2 weeks. In intermolt animals, long-term ESA increased hemolymph ecdysteroid titers fourfold to fivefold by 28 days post treatment, but there was no late premolt peak (>400 pg ?l(-1)) characteristic of late premolt animals and animals did not molt by 90 days post-ESA. There was no effect of ESA or MLA on the expression of Cm-elongation factor 2 (EF2), Cm-NOS, the beta subunit of GC-I (Cm-GC-I?), a membrane receptor GC (Cm-GC-II) and a soluble NO-insensitive GC (Cm-GC-III) in green morphs. Red morphs were affected by prolonged ESA and MLA treatments, as indicated by large decreases in Cm-EF2, Cm-GC-II and Cm-GC-III mRNA levels. ESA accelerated the transition of green morphs to the red phenotype in intermolt animals. ESA delayed molting in premolt green morphs, whereas intact and MLA animals molted by 30 days post treatment. There were significant effects on YO gene expression in intact animals: Cm-GC-I? mRNA increased during premolt and Cm-GC-III mRNA decreased during premolt and increased during postmolt. Cm-MIH transcripts were detected in eyestalk ganglia, the brain and the thoracic ganglion from green intermolt animals, suggesing that MIH in the brain and thoracic ganglion prevents molt induction in green ESA animals. PMID:24198255

Abuhagr, Ali M; Blindert, Jennifer L; Nimitkul, Sukkrit; Zander, Ian A; Labere, Stefan M; Chang, Sharon A; Maclea, Kyle S; Chang, Ernest S; Mykles, Donald L

2014-03-01

352

Inhibition of Myo6 gene expression by co?expression of a mutant of transcription factor POU4F3 (BRN?3C) in hair cells.  

PubMed

An eight?base pair (bp) deletion in the Pou4f3 gene in hair cells is associated with DFNA15, a hereditary form of hearing loss. To explore the pathological mechanisms underlying the development of DFNA15, the effect of the mutation in Pou4f3 on the activity of the myosin VI (Myo6) promoter, was investigated. The upstream regulatory sequence of Myo6 (2625 bp), consisting of an 1899 bp upstream sequence and a 727 bp intron 1 sequence, was amplified using polymerase chain reaction and subcloned into the pGL3?Basic vector expressing firefly luciferase. For verification of inserted fragments, plasmids were subjected to restriction analysis and then sequenced. HEK293T human embryonic kidney cells were transiently transfected with renilla luciferase?thymidine kinase vectors expressing Renilla luciferase and the Myo6 promoter?driven firefly luciferase expressing vectors along with pIRES2?enhanced green fluorescent protein (EGFP)?Pou4f3 (expressing wild?type Pou4f3) or pIRES2?EGFP?Pou4f3 (expressing the truncation mutant of Pou4f3). The relative luciferase activities were measured to determine the activity of the Myo6 promoter. The Myo6 promoter activity was not affected by co?expression of wild?type Pou4f3, as indicated by the comparable relative luciferase activities in the presence of the pIRES2?EGFP?Pou4f3 and the empty control vectors. However, co?expression of mutated Pou4f3 significantly inhibited the activity of the Myo6 promoter to almost half of that of the control (P<0.001). The data suggests that mutated Pou4f3 has a negative role in the promoter activity of Myo6, and by extension, the expression of myosin VI, and this may be an underlying mechanism of DFNA15 hearing loss. PMID:24535414

Ma, Deng-Bin; Chen, Jie; Xia, Yang; Zhu, Guang-Jie; Ma, Xiao-Feng; Zhou, Han; Gu, Ya-Jun; Yu, Chen-Jie; Zhu, Min-Sheng; Qian, Xiao-Yun; Gao, Xia

2014-04-01

353

Inhibition of CBF/NF-Y mediated transcription activation arrests cells at G2/M phase and suppresses expression of genes activated at G2/M phase of the cell cycle  

PubMed Central

Previous studies showed that binding of the CBF/NF-Y (CBF) transcription factor to cellular promoters is essential for cell proliferation. This observation prompted us to investigate the function of CBF in relation to cell cycle progression and in cell-cycle-regulated transcription. In this study, we used a tetracycline-inducible adenoviral vector to express a truncated CBF-B subunit, Bdbd, lacking a transcription activation domain in various mammalian cell lines. The Bdbd polypeptide interacts with cellular CBF-A/CBF-C and binds to promoters containing CBF-binding sites. Interestingly, expression of Bdbd in various mammalian cells resulted in the inhibition of cell proliferation and specific cell cycle arrest at G2/M phase. Gene expression analysis demonstrated that the expression of Bdbd strongly suppressed cell cycle-dependent transcription activation of Cyclin B1, Aurora A and CDK1 genes, key regulators for cell cycle progression at G2/M phase. Chromatin immunoprecipitation analysis showed that Bdbd significantly inhibited binding of TATA-binding protein, TBP to both Cyclin B1 and Aurora A promoters, but did not inhibit binding of E2F3 activator to Cyclin B1 promoter. This study suggested that the activation domain of CBF-B plays an essential role in the transcription activation of Cyclin B1 and Aurora A genes at G2/M phase, thus regulating cell cycle progression at G2/M phase. PMID:17098936

Hu, Qianghua; Lu, Jing-Fang; Luo, Rong; Sen, Subrata; Maity, Sankar N.

2006-01-01

354

The C. elegans class A synthetic multivulva genes inhibit ectopic RAS-mediated vulval development by tightly restricting expression of lin-3 EGF  

E-print Network

The class A and B synthetic multivulva (synMuv) genes of C. elegans redundantly antagonize an EGF/Ras pathway to prevent ectopic vulval induction. The class B synMuv genes encode many proteins known to remodel chromatin ...

Saffer, Adam M

2011-01-01

355

Cellular and Gene Expression Responses Involved in the Rapid Growth Inhibition of Human Cancer Cells by RNA Interference-mediated Depletion of Telomerase RNA  

Microsoft Academic Search

Inhibition of the up-regulated telomerase activity in cancer cells has previously been shown to slow cell growth but only after prior telomere shortening. Previ- ously, we have reported that, unexpectedly, a hairpin short interfering RNA specifically targeting human te- lomerase RNA rapidly inhibits the growth of human cancer cells independently of p53 or telomere length and without bulk telomere shortening

Shang Li; Julia Crothers; Christopher M. Haqq; Elizabeth H. Blackburn

2005-01-01

356

Suicide HSVtk Gene Delivery by Neurotensin-Polyplex Nanoparticles via the Bloodstream and GCV Treatment Specifically Inhibit the Growth of Human MDA-MB-231 Triple Negative Breast Cancer Tumors Xenografted in Athymic Mice  

PubMed Central

The human breast adenocarcinoma cell line MDA-MB-231 has the triple-negative breast cancer (TNBC) phenotype, which is an aggressive subtype with no specific treatment. MDA-MB-231 cells express neurotensin receptor type 1 (NTSR1), which makes these cells an attractive target of therapeutic genes that are delivered by the neurotensin (NTS)-polyplex nanocarrier via the bloodstream. We addressed the relevance of this strategy for TNBC treatment using NTS-polyplex nanoparticles harboring the herpes simplex virus thymidine kinase (HSVtk) suicide gene and its complementary prodrug ganciclovir (GCV). The reporter gene encoding green fluorescent protein (GFP) was used as a control. NTS-polyplex successfully transfected both genes in cultured MDA-MB-231 cells. The transfection was demonstrated pharmacologically to be dependent on activation of NTSR1. The expression of HSVtk gene decreased cell viability by 49% (P<0.0001) and induced apoptosis in cultured MDA-MB-231 cells after complementary GCV treatment. In the MDA-MB-231 xenograft model, NTS-polyplex nanoparticles carrying either the HSVtk gene or GFP gene were injected into the tumors or via the bloodstream. Both routes of administration allowed the NTS-polyplex nanoparticles to reach and transfect tumorous cells. HSVtk expression and GCV led to apoptosis, as shown by the presence of cleaved caspase-3 and Apostain immunoreactivity, and significantly inhibited the tumor growth (55–60%) (P<0.001). At the end of the experiment, the weight of tumors transfected with the HSVtk gene was 55% less than that of control tumors (P<0.05). The intravenous transfection did not induce apoptosis in peripheral organs. Our results offer a promising gene therapy for TNBC using the NTS-polyplex nanocarrier. PMID:24824754

Castillo-Rodriguez, Rosa A.; Arango-Rodriguez, Martha L.; Escobedo, Lourdes; Hernandez-Baltazar, Daniel; Gompel, Anne

2014-01-01