Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.48 (transcriptase)
 9,479 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Measles virus (MV) expresses at least 3 proteins from the phosphoprotein (P) cistron. Alternative translation initiation directs synthesis of the C protein from the +1 reading frame, while so-called RNA editing generates a second population of mRNAs which express the V protein from the -1 reading frame which lies within and overlaps the larger P reading frame. While the P protein has been demonstrated to be an essential cofactor for the L protein in the formation of active transcriptase complexes, the functions of the V and C proteins remain unknown. In order to investigate these functions, we have expressed the MV P, V and C proteins as GST fusions in E. coli for affinity purification and use in an in vitro binding assay with other viral and cellular proteins. The P protein was found to interact with L, NP, and with itself. These interactions were mapped to the carboxy-terminal half of the protein which is absent in the V protein. In contrast, both the V and C proteins failed to interact with any other viral proteins, but were each found to interact specifically with one or more cellular proteins. Appropriate aspects of these results were confirmed in vivo using the yeast two-hybrid system. These observations suggest that the V and C proteins may be involved in modulation of the host cellular environment within MV-infected cells. Such activity would be distinct from their previously proposed role in the possible down-regulation of virus-specific RNA transcription and replication.
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PMID:Protein interactions entered into by the measles virus P, V, and C proteins. 857 62

The measles virus RNA-dependent RNA polymerase consists of two virus-encoded subunits, the phosphoprotein (P) and the large (L) protein. The P mRNA also codes for a C protein in the +1 reading frame relative to P. The activities of the measles P and C proteins from the vaccine strain, EdB, a wild-type CM strain, and an SSPE P4 strain were investigated using a CAT reporter minigenome assay. CAT is synthesized following replication and transcription of a DI-CAT minigenome supported by individual P, L, and N plasmids expressed in a mammalian expression system. As measured by CAT activity, CMP1 and P4P1 stimulate transcription and replication four- to six- and six- to eightfold, respectively, better than EdP. There are 10 and 16 amino acid changes in the P protein and three and four changes in C in CMP1 and P4P1, respectively, relative to EdP. By constructing chimeric P genes we showed that mutations throughout P4P1 were required for enhanced polymerase activity, while only mutations in the 5'-terminal portion, encompassing the C ORF, of the CMP1 gene mediated stimulation. Abrogation of C expression from the Ed and CM P genes resulted in an increase in RNA synthesis of twofold for CMP1S and four- to fivefold for EdPS. With the addition of C protein expressed from a separate plasmid that contains only the C ORF, EdC reduces viral RNA synthesis more strongly than CMC. These data suggest that EdC and CMC proteins give a differential inhibition that accounts for most of the differences in RNA synthesis by EdP and CMP1.
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PMID:Mutations in the measles virus C protein that up regulate viral RNA synthesis. 1141 10

Sendai virus encodes an RNA-dependent RNA polymerase which is composed of the L and P proteins. Site-directed mutagenesis of the N terminus of L has identified amino acids important for binding P. Seven of nine mutants in amino acids 1 to 350 of Sendai L lost the ability to bind to Sendai P, although they were still able to bind the viral C protein. Loss of P binding correlated with the loss of all RNA synthesis activities. Two L mutants gave limited P-L complex formation and limited viral transcription and replication.
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PMID:The phosphoprotein (P) binding site resides in the N terminus of the L polymerase subunit of sendai virus. 1186 77

The Sendai virus RNA-dependent RNA polymerase is composed of the L and P proteins. We previously showed that the L protein gives intragenic complementation and forms an oligomer where the L-L interaction site mapped to the N-terminal half of the protein (S. Smallwood et al., 2002, Virology, 00, 000-000). We now show that L oligomerization does not depend on P protein and progressively smaller N-terminal fragments of L from amino acids (aa) 1-1146 through aa 1-174 all bind wild-type L. C-terminal truncations up to aa 424, which bind L, can complement the transcription defect in an L mutant altered at aa 379, although these L truncation mutants do not bind P. The fragment of L comprising aa 1-895, furthermore, acts as a dominant-negative mutant to inhibit transcription of wild-type L. N-terminal deletions of aa 1-189 and aa 1-734 have lost the ability to form the L-L complex as well as the L-P complex, although they still bind C protein. These data are consistent with the L-L interaction site residing in aa 1-174. Site-directed mutations in the N-terminal 347 aa, of L which abolish P binding, do not affect L-L complex formation, so while the L and P binding sites on L are overlapping they are mediated by different amino acids. The N-terminal portions of L with aa 1-424, aa 1-381, and to a lesser extent aa 1-174, can complement the transcription defect in an L mutant altered at aa 77-81, showing their L-L interaction is functional.
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PMID:The L-L oligomerization domain resides at the very N-terminus of the sendai virus L RNA polymerase protein. 1295 19

The complete genome of mandarin fish Siniperca chuatsi rhabdovirus (SCRV) was cloned and sequenced. It comprises 11,545 nucleotides and contains five genes encoding the nucleoprotein N, the phosphoprotein P, the matrix protein M, the glycoprotein G, and the RNA-dependent RNA polymerase protein L. At the 3' and 5' termini of SCRV genome, leader and trailer sequences show inverse complementarity. The N, P, M and G proteins share the highest sequence identities (ranging from 14.8 to 41.5%) with the respective proteins of rhabdovirus 903/87, the L protein has the highest identity with those of vesiculoviruses, especially with Chandipura virus (44.7%). Phylogenetic analysis of L proteins showed that SCRV clustered with spring vireamia of carp virus (SVCV) and was most closely related to viruses in the genus Vesiculovirus. In addition, an overlapping open reading frame (ORF) predicted to encode a protein similar to vesicular stomatitis virus C protein is present within the P gene of SCRV. Furthermore, an unoverlapping small ORF downstream of M ORF within M gene is predicted (tentatively called orf4). Therefore, the genomic organization of SCRV can be proposed as 3' leader-N-P/C-M-(orf4)-G-L-trailer 5'. Orf4 transcription or translation products could not be detected by northern or Western blot, respectively, though one similar mRNA band to M mRNA was found. This is the first report on one small unoverlapping ORF in M gene of a fish rhabdovirus.
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PMID:Genomic sequence of mandarin fish rhabdovirus with an unusual small non-transcriptional ORF. 1806 57

The Nipah virus (NiV) phosphoprotein (P) gene encodes the C, P, V, and W proteins. P, V, and W, have in common an amino-terminal domain sufficient to bind STAT1, inhibiting its interferon (IFN)-induced tyrosine phosphorylation. P is also essential for RNA-dependent RNA polymerase function. C is encoded by an alternate open reading frame (ORF) within the common amino-terminal domain. Mutations within residues 81 to 113 of P impaired its polymerase cofactor function, as assessed by a minireplicon assay, but these mutants retained STAT1 inhibitory function. Mutations within the residue 114 to 140 region were identified that abrogated interaction with and inhibition of STAT1 by P, V, and W without disrupting P polymerase cofactor function. Recombinant NiVs were then generated. A G121E mutation, which abrogated inhibition of STAT1, was introduced into a C protein knockout background (C(ko)) because the mutation would otherwise also alter the overlapping C ORF. In cell culture, relative to the wild-type virus, the C(ko) mutation proved attenuating but the G121E mutant virus replicated identically to the C(ko) virus. In cells infected with the wild-type and C(ko) viruses, STAT1 was nuclear despite the absence of tyrosine phosphorylation. This latter observation mirrors what has been seen in cells expressing NiV W. In the G121E mutant virus-infected cells, STAT1 was not phosphorylated and was cytoplasmic in the absence of IFN stimulation but became tyrosine phosphorylated and nuclear following IFN addition. These data demonstrate that the gene for NiV P encodes functions that sequester inactive STAT1 in the nucleus, preventing its activation and suggest that the W protein is the dominant inhibitor of STAT1 in NiV-infected cells.
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PMID:Nipah virus sequesters inactive STAT1 in the nucleus via a P gene-encoded mechanism. 1951 82

Measles virus (MeV), like all viruses of the order Mononegavirales, utilizes a complex consisting of genomic RNA, nucleoprotein, the RNA-dependent RNA polymerase, and a polymerase cofactor, the phosphoprotein (P), for transcription and replication. We previously showed that a recombinant MeV that does not express another viral protein, C, has severe transcription and replication deficiencies, including a steeper transcription gradient than the parental virus and generation of defective interfering RNA. This virus is attenuated in vitro and in vivo However, how the C protein operates and whether it is a component of the replication complex remained unclear. Here, we show that C associates with the ribonucleocapsid and forms a complex that can be purified by immunoprecipitation or ultracentrifugation. In the presence of detergent, the C protein is retained on purified ribonucleocapsids less efficiently than the P protein and the polymerase. The C protein is recruited to the ribonucleocapsid through its interaction with the P protein, as shown by immunofluorescence microscopy of cells expressing different combinations of viral proteins and by split luciferase complementation assays. Forty amino-terminal C protein residues are dispensable for the interaction with P, and the carboxyl-terminal half of P is sufficient for the interaction with C. Thus, the C protein, rather than being an "accessory" protein as qualified in textbooks so far, is a ribonucleocapsid-associated protein that interacts with P, thereby increasing replication accuracy and processivity of the polymerase complex.IMPORTANCE Replication of negative-strand RNA viruses relies on two components: a helical ribonucleocapsid and an RNA-dependent RNA polymerase composed of a catalytic subunit, the L protein, and a cofactor, the P protein. We show that the measles virus (MeV) C protein is an additional component of the replication complex. We provide evidence that the C protein is recruited to the ribonucleocapsid by the P protein and map the interacting segments of both C and P proteins. We conclude that the primary function of MeV C is to improve polymerase processivity and accuracy, rather than uniquely to antagonize the type I interferon response. Since most viruses of the Paramyxoviridae family express C proteins, their primary function may be conserved.
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PMID:The C Protein Is Recruited to Measles Virus Ribonucleocapsids by the Phosphoprotein. 3174 90