Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.6 (RNA polymerase)
34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The genome of influenza A virus consists of eight single-stranded RNA molecules of negative polarity. Their replication and transcription take place in the nucleus of infected cells using ribonucleoprotein complexes (RNPs) as templates. Two of the viral transcripts, those generated by RNPs 7 and 8, can be spliced and lead to two alternative protein products (M1 and M2, NS1 and NEP/NS2, respectively). Previous studies have shown that when expressed from cDNA, NS1 protein alters the splicing and transport of RNA polymerase II-driven transcripts. Here we used a transient replication/transcription system, in which RNP 8 is replicated and transcribed by recombinant RNA and proteins, to study the splicing and nucleo-cytoplasmic transport of true viral transcripts. Our results show that the encoded NS1 protein inhibits the splicing of the collinear transcript. This regulation is mediated by the N-terminal region of the protein but does not involve its RNA-binding activity. We also show that NS1 protein preferentially blocks the nucleo-cytoplasmic transport of the collinear RNP 8 transcript in an RNA-binding dependent manner. These results rule out previous models to explain the regulation of mRNA processing and transport by NS1 and underlines the relevance of NS1 protein in the control of virus gene expression.
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PMID:Mutation analysis of a recombinant NS replicon shows that influenza virus NS1 protein blocks the splicing and nucleo-cytoplasmic transport of its own viral mRNA. 1748 45

The influenza virus RNA polymerase transcribes the negative-sense viral RNA segments (vRNA) into mRNA and replicates them via complementary RNA (cRNA) intermediates into more copies of vRNA. It is not clear how the relative amounts of the three RNA products, mRNA, cRNA and vRNA, are regulated during the viral life cycle. We found that in viral ribonucleoprotein (vRNP) reconstitution assays involving only the minimal components required for viral transcription and replication (the RNA polymerase, the nucleoprotein and a vRNA template), the relative levels of accumulation of RNA products differed from those observed in infected cells, suggesting a regulatory role for additional viral proteins. Expression of the viral NS2/NEP protein in RNP reconstitution assays affected viral RNA levels by reducing the accumulation of transcription products and increasing the accumulation of replication products to more closely resemble those found during viral infection. This effect was functionally conserved in influenza A and B viruses and was influenza-virus-type-specific, demonstrating that the NS2/NEP protein changes RNA levels by specific alteration of the viral transcription and replication machinery, rather than through an indirect effect on the host cell. Although NS2/NEP has been shown previously to play a role in the nucleocytoplasmic export of viral RNPs, deletion of the nuclear export sequence region that is required for its transport function did not affect the ability of the protein to regulate RNA levels. A role for the NS2/NEP protein in the regulation of influenza virus transcription and replication that is independent of its viral RNP export function is proposed.
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PMID:NS2/NEP protein regulates transcription and replication of the influenza virus RNA genome. 1926 57

RNA segment 8 (NS) of influenza A virus encodes two proteins. The NS1 protein is translated from the unspliced primary mRNA transcript, whereas the second protein encoded by this segment, NS2/NEP, is translated from a spliced mRNA. Splicing of influenza NS1 mRNA is thought to be regulated so that the levels of NS2 spliced transcripts are approximately 10 % of total NS mRNA. Regulation of splicing of the NS1 mRNA has been studied at length, and a number of often-contradictory control mechanisms have been proposed. In this study, we used (32)P-labelled gene-specific primers to investigate influenza A NS1 mRNA splicing regulation. It was found that the efficiency of splicing of NS1 mRNA was maintained at similar levels in both virus infection and ribonucleoprotein-reconstitution assays, and NS2 mRNA comprised approximately 15 % of total NS mRNA in both assays. The effect of NS1 protein expression on the accumulation of viral NS2 mRNA and spliced cellular beta-globin mRNA was analysed, and it was found that NS1 protein expression reduced spliced beta-globin mRNA levels, but had no effect on the accumulation of NS2 mRNA. We conclude that the NS1 protein specifically inhibits the accumulation of cellular RNA polymerase II-driven mRNAs, but does not affect the splicing of its own viral NS1 mRNA.
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PMID:Splicing of influenza A virus NS1 mRNA is independent of the viral NS1 protein. 2051 56

Arabidopsis seed formation is coupled with two plastid differentiation processes. Chloroplast formation starts during embryogenesis and ends with the maturation phase. It is followed by chloroplast dedifferentiation/degeneration that starts at the end of the maturation phase and leads to the presence of small non-photosynthetic plastids in dry seeds. We have analysed mRNA and protein levels of nucleus- and plastid-encoded (NEP and PEP) components of the plastid transcriptional machinery, mRNA and protein levels of some plastid RNA polymerase target genes, changes in plastid transcriptome profiles and mRNA and protein levels of some selected nucleus-encoded plastid-related genes in developing seeds during embryogenesis, maturation and desiccation. As expected, most of the mRNAs and proteins increase in abundance during maturation and decrease during desiccation, when plastids dedifferentiate/degenerate. In contrast, mRNAs and proteins of components of the plastid transcriptional apparatus do not decrease or even still increase during the period of plastid dedifferentiation. Results suggest that proteins of the plastid transcriptional machinery are specifically protected from degradation during the desiccation period and conserved in dry seeds to allow immediate regain of plastid transcriptional activity during stratification/germination. In addition, results reveal accumulation and storage of mRNAs coding for RNA polymerase components and sigma factors in dry seeds. They should provide immediately-to-use templates for translation on cytoplasmic ribosomes in order to enhance RNA polymerase protein levels and to provide regulatory proteins for stored PEP to guaranty efficient plastid genome transcription during germination.
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PMID:Plastid gene expression during chloroplast differentiation and dedifferentiation into non-photosynthetic plastids during seed formation. 2349 53

Influenza A viruses are major pathogens in humans and in animals, whose genome consists of eight single-stranded RNA segments of negative polarity. Viral mRNAs are synthesized by the viral RNA-dependent RNA polymerase in the nucleus of infected cells, in close association with the cellular transcriptional machinery. Two proteins essential for viral multiplication, the exportin NS2/NEP and the ion channel protein M2, are produced by splicing of the NS1 and M1 mRNAs, respectively. Here we identify two human spliceosomal factors, RED and SMU1, that control the expression of NS2/NEP and are required for efficient viral multiplication. We provide several lines of evidence that in infected cells, the hetero-trimeric viral polymerase recruits a complex formed by RED and SMU1 through interaction with its PB2 and PB1 subunits. We demonstrate that the splicing of the NS1 viral mRNA is specifically affected in cells depleted of RED or SMU1, leading to a decreased production of the spliced mRNA species NS2, and to a reduced NS2/NS1 protein ratio. In agreement with the exportin function of NS2, these defects impair the transport of newly synthesized viral ribonucleoproteins from the nucleus to the cytoplasm, and strongly reduce the production of infectious influenza virions. Overall, our results unravel a new mechanism of viral subversion of the cellular splicing machinery, by establishing that the human splicing factors RED and SMU1 act jointly as key regulators of influenza virus gene expression. In addition, our data point to a central role of the viral RNA polymerase in coupling transcription and alternative splicing of the viral mRNAs.
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PMID:Recruitment of RED-SMU1 complex by Influenza A Virus RNA polymerase to control Viral mRNA splicing. 2494 53

Plastid genes are transcribed by two types of RNA polymerase in angiosperms: the bacterial type plastid-encoded RNA polymerase (PEP) and one (RPOTp in monocots) or two (RPOTp and RPOTmp in dicots) nuclear-encoded RNA polymerase(s) (NEP). PEP is a bacterial-type multisubunit enzyme composed of core subunits (coded for by the plastid rpoA, rpoB, rpoC1 and rpoC2 genes) and additional protein factors (sigma factors and polymerase associated protein, PAPs) encoded in the nuclear genome. Sigma factors are required by PEP for promoter recognition. Six different sigma factors are used by PEP in Arabidopsis plastids. NEP activity is represented by phage-type RNA polymerases. Only one NEP subunit has been identified, which bears the catalytic activity. NEP and PEP use different promoters. Many plastid genes have both PEP and NEP promoters. PEP dominates in the transcription of photosynthesis genes. Intriguingly, rpoB belongs to the few genes transcribed exclusively by NEP. Both NEP and PEP are active in non-green plastids and in chloroplasts at all stages of development. The transcriptional activity of NEP and PEP is affected by endogenous and exogenous factors. This article is part of a Special Issue entitled: Chloroplast Biogenesis.
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PMID:Chloroplast RNA polymerases: Role in chloroplast biogenesis. 2568 May 13

Functional chloroplast generation depends on the precise coordination of gene expression between the plastid and the nucleus and is essential for plant growth and development. In this study, a rice (Oryza sativa) mutant that exhibited albino and seedling-lethal phenotypes was isolated from a⁶⁰Co-irradiated rice population. The mutant gene was identified as an ortholog of the Arabidopsis plastid transcriptionally active chromosome protein 2 (pTAC2) gene, and the mutant strain was designated osptac2. Sequence and transcription analyses showed that OspTAC2 encodes a putative chloroplast protein consisting of 10 pentratricopeptide repeat (PPR) domains and a C-terminal small MutS-related (SMR) domain. Cytological observations via microscopy showed that the OspTAC2-green fluorescent fusion protein is localized in the chloroplasts. Transmission electron microscopy revealed that the chloroplast of the osptac2 mutant lacks an organized thylakoid membrane. The transcript levels of all investigated PEP (plastid-encoded RNA polymerase)-dependent genes were dramatically reduced in the osptac2 mutant, whereas the transcript levels of NEP (nuclear-encoded polymerase)-dependent genes were increased. These results suggest that OspTAC2 plays a critical role in chloroplast development and indicate that the molecular function of the OspTAC2 gene is conserved in rice and Arabidopsis.
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PMID:OspTAC2 encodes a pentatricopeptide repeat protein and regulates rice chloroplast development. 2776 Jul 23

In higher plant chloroplasts, the plastid-encoded RNA polymerase (PEP) consists of four catalytic subunits and numerous nuclear-encoded accessory proteins, including pTAC10, an S1-domain-containing protein. In this study, pTAC10 knockout lines were characterized. Two ptac10 mutants had an albino phenotype and severely impaired chloroplast development. The pTAC10 genomic sequence fused to a four-tandem MYC tag driven by its own promoter functionally complemented the ptac10-1 mutant phenotype. pTAC10 was present in both the chloroplast stroma and thylakoids. Two-dimensional blue native polyacrylamide gel electrophoresis (BN-PAGE), and immunoblotting assays showed that pTAC10:MYC co-migrates with one of the PEP core subunits, RpoB. A comprehensive investigation of the plastid gene expression profiles by quantitative RT-PCR revealed that, compared with wild-type plants, the abundance of PEP-dependent plastid transcripts is severely decreased in the ptac10-1 mutant, while the amount of plastid transcripts exclusively transcribed by NEP either barely changes or even increases. RNA blot analysis confirmed that PEP-dependent chloroplast transcripts, including psaB, psbA and rbcL, substantially decrease in the ptac10-1 mutant. Immunoblotting showed reduced accumulation of most chloroplast proteins in the ptac10 mutants. These data indicate the essential role of pTAC10 in plastid gene expression and plastid development. pTAC10 interacts with chloroplast-targeted casein kinase 2 (cpCK2) in vitro and in vivo and can be phosphorylated by Arabidopsis cpCK2 in vitro at sites Ser95, Ser396 and Ser434. RNA-EMSA assays showed that pTAC10 is able to bind to the psbA, atpE and accD transcripts, suggesting a non-specific RNA-binding activity of pTAC10. The RNA affinity of pTAC10 was enhanced by phosphorylation and decreased by the amino acid substitution Ser434-Ala of pTAC10. These data show that pTAC10 is essential for plastid gene expression in Arabidopsis and that it can be phosphorylated by cpCK2.
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PMID:pTAC10, an S1-domain-containing component of the transcriptionally active chromosome complex, is essential for plastid gene expression in Arabidopsis thaliana and is phosphorylated by chloroplast-targeted casein kinase II. 2933 Jul 2

The chloroplast is an essential photosynthetic apparatus that is more sensitive to low temperatures than other organelles. Sigma factors were revealed regulating specific gene expression for maintaining photosynthetic efficiency and adapting to physiological and environmental conditions. However, the regulatory mechanisms of SIG genes supporting chloroplast development under low temperature in rice have not yet been reported. Here, we uncovered the essential role of OsSIG2A in rice chloroplast development at low temperatures by a newly reported thermo-sensitive chlorophyll deficient 12 (tcd12) mutant, which exhibited albino leaves with decreased chlorophyll content and malformed chloroplasts at seedling stage under low temperature. OsSIG2A is a typical chloroplast-localised RNA polymerase sigma factor, and constitutively expresses in different rice tissues, especially for young leaves and stems. Moreover, the transcription level of both PEP- and NEP- dependent genes, which are necessary for chloroplast development at early leaf development stage, was greatly affected in the tcd12 mutant under low temperature. Taken together, our findings indicate that OsSIG2A is required for early chloroplast differentiation under low temperatures by regulating plastid genes expression.
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PMID:OsSIG2A is required for chloroplast development in rice (Oryza sativa L.) at low temperature by regulating plastid genes expression. 3104 2


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