The phosphoprotein (P) and L binding sites reside in the N-terminus of the L subunit of the measles virus RNA polymerase

Çevik, Bayram; Holmes, David E.; Vrotsos, Emmanuel George; Feller, Joyce A.; Smallwood, Sherin; Moyer, Sue A.

doi:10.1016/j.virol.2004.07.002

Cited by 42 publications

(44 citation statements)

References 22 publications

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“…Our subsequent linker insertion scan confirmed these data and, by extending successful epitope insertion to distantly related L proteins, illuminated a conserved Paramyxovirus polymerase organization into at least two distinct sections. In addition, we identified an N-terminal position in MeV L, residue 615 located downstream of the N-terminal L-P and L-L oligomerization domain (8,14), that likewise tolerates enlargement without eliminating L bioactivity. This finding did not extend, however, to acceptance of FLAG or HA epitope tags, which contain a high density of charged or bulky aromatic amino acids (75,76) and are thus expected to be structurally more active than the 10-residue linker used in the primary scan.…”

Section: Discussionmentioning

confidence: 99%

“…These studies highlighted 12 candidate regions in MeV L that were located between CR I and CR VI, received combined propensity scores of Ն4, and were identified by at least two of the algorithms employed. Focusing on the L core, potential interdomain regions located in the N-terminal first 408-amino acid section of MeV L, which mediates both L interaction with the viral P protein and polymerase oligomerization (8,14), and candidates positioned downstream of the LR II/LR III junction (residues 1695-1717 (23, 24)) were not considered for experimental evaluation.…”

Section: In Silico Domain Analysis Of Paramyxovirus L Protein-mentioning

confidence: 99%

“…Of these, the N-terminal sections harboring CR I have been implicated in L oligomerization (8 -10) and/or L interaction with P (8,9,(11)(12)(13)(14), CR III in RNA polymerization (15,16), and CR VI in methyltransferase activity (3,6,17). A conserved GXXT n HR motif in NNSV L CR V, which was first identified in VSV L, is furthermore thought to mediate unusual capping of the viral mRNAs through transfer of 5Ј-monophosphate-mRNA onto GDP (18,19).…”

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confidence: 99%

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Independent Structural Domains in Paramyxovirus Polymerase Protein

Dochow

Krumm

Crowe

et al. 2012

Journal of Biological Chemistry

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Background: RNA-dependent RNA polymerases of nonsegmented negative-strand RNA viruses (Mononegavirales) harbor multiple catalytic activities. Results: Domain screening and trans-complementation of Paramyxovirus polymerase fragments with dimerization tags identifies independent folding-competent domains. Conclusion: Paramyxovirus polymerases harbor at least two independent domains that lack high-affinity interfaces but require molecular compatibility for bioactivity. Significance: First demonstration of Mononegavirales polymerase trans-complementation sets the stage for structural analyses of folding domains.

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Section: Discussionmentioning

confidence: 99%

Section: In Silico Domain Analysis Of Paramyxovirus L Protein-mentioning

confidence: 99%

mentioning

confidence: 99%

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Independent Structural Domains in Paramyxovirus Polymerase Protein

Dochow

Krumm

Crowe

et al. 2012

Journal of Biological Chemistry

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“…However, the data do not eliminate other possible scenarios. For example, the L protein of other paramyxoviruses has been shown to oligomerize (26)(27)(28), raising the possibility that the RdRp can contact the Le 3= end and simultaneously have an active site positioned at or near the NS1 GS signal. Another possibility is that the RdRp is able to bind to nt 3 to 12 and then has some mechanism to scan to the first GS signal without first initiating RNA synthesis.…”

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confidence: 99%

Respiratory Syncytial Virus Polymerase Can Initiate Transcription from Position 3 of the Leader Promoter

Tremaglio

Noton

Deflubé

et al. 2013

J Virol

103

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The mechanisms by which the respiratory syncytial virus (RSV) RNA-dependent RNA polymerase (RdRp) initiates mRNA transcription and RNA replication are poorly understood. A previous study, using an RSV minigenome, suggested that the leader (Le) promoter region at the 3= end of the genome has two initiation sites, one at position ؉1, opposite the 3= terminal nucleotide of the genome, and a second site at position ؉3, at a sequence that closely resembles the gene start (GS) signal of the RSV L gene. In this study, we show that the ؉3 initiation site of the Le is utilized with apparently high frequency in RSV-infected cells and yields small RNA transcripts that are heterogeneous in length but mostly approximately 25 nucleotides (nt) long. Experiments with an in vitro assay in which RSV RNA synthesis was reconstituted using purified RdRp and an RNA oligonucleotide showed that nt 1 to 14 of the Le promoter were sufficient to signal initiation from ؉3 and that the RdRp could access the ؉3 initiation site without prior initiation at ؉1. In a minigenome assay, nucleotide substitutions within the Le to increase its similarity to a GS signal resulted in more-efficient elongation of the RNA initiated from position ؉3 and a reduction in RNA initiated from the NS1 gene start signal at ؉45. Taken together, these data suggest a new model for initiation of sequential transcription of the RSV genes, whereby the RdRp initiates the process from a gene start-like sequence at position ؉3 of the Le. R espiratory syncytial virus (RSV) is the leading cause of pneumonia and viral deaths in infants worldwide and is increasingly recognized as a significant pathogen of the elderly and immunocompromised (1). Treatment for RSV infection is limited to supportive care, and at present there are no approved vaccines. RSV is a member of the family Paramyxoviridae in the order Mononegavirales, the nonsegmented negative-strand (NNS) RNA viruses. Like other viruses in this order, the RSV genome serves as the template for mRNA transcription and genome replication, carried out by the viral RNA-dependent RNA polymerase (RdRp) (reviewed in reference 2).The RSV genome is 15.2 kb and encodes mRNAs from 10 sequentially arranged genes (reviewed in reference 3). Two short extragenic regions border the genome, a 3= 44-nt leader region (Le) and a 5= 155-nt trailer (Tr) region (4, 5). The Le region contains promoter sequences that are required for transcription of all RSV genes and genome replication (4). Transcription occurs in an obligatorily sequential, polarized manner to generate 10 subgenomic mRNAs (6). During this process, the RdRp is controlled by cis-acting gene start (GS) and gene end (GE) sequences that flank each of the genes (7, 8). The GS sequences direct initiation of RNA synthesis. They are highly conserved in RSV, with 9 of 10 GS sequences being identical and the tenth having only slight differences (9). In related viruses, the GS signals have been shown to be multifunctional, directing the RdRp to cap and methylate the nascent transcript (10-12...

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“…Moreover, the SSPE virus-associated mutations in their study were different from those observed for SSPE-Kobe-1 in our study. Cevik et al (11) recently identified a number of mutations in an N-terminal region of the L protein that affected interaction with the P protein. Our present study revealed that SSPE-Kobe-1 did not possess any of those mutations.…”

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confidence: 99%

Full‐Length Sequence Analysis of Subacute Sclerosing Panencephalitis (SSPE) Virus, a Mutant of Measles Virus, Isolated from Brain Tissues of a Patient Shortly after Onset of SSPE

Hotta

Nihei

Abe

et al. 2006

Microbiology and Immunology

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Subacute sclerosing panencephalitis (SSPE) virus, a measles virus (MeV) mutant, was isolated from brain tissues of a patient shortly after the clinical onset, and the entire viral genome was sequenced. The virus, named SSPE‐Kobe‐1, formed syncytia on B95a and Vero/SLAM cells without producing cell‐free infectious virus particles, which is characteristic of SSPE virus. Phylogenetic analysis classified SSPE‐Kobe‐1 into genotype D3. When compared with an MeV field isolate of the same genotype (Ich‐B strain), SSPE‐Kobe‐1 exhibited mutation rates of 0.8–1.6% at the nucleotide level in each of the protein‐coding regions of the viral genome. It is noteworthy that the mutation rate of the M gene (1.2%) of SSPE‐Kobe‐1 was considerably lower than for other SSPE virus strains reported so far, but that the majority of the mutations (75%) were the uridine‐to‐cytidine biased hypermutation characteristic of the SSPE virus M gene. At the amino acid level, the viral proteins, such as N, P., C, V, M., F, H and L proteins, had point‐mutations on 3, 7, 1, 4, 3, 9, 8 and 14 residues, respectively, compared with the Ich‐B strain. In addition, the F and H proteins had mutated C‐termini due to single‐point mutations near or at the stop codons. Two of the three mutations in the M protein were Leu‐to‐Pro mutations, which are likely to affect the conformation and, therefore, the function of the protein. Because of the relatively small number of mutations, SSPE‐Kobe‐1 would be a useful tool to study genetic evolution of SSPE virus.

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The phosphoprotein (P) and L binding sites reside in the N-terminus of the L subunit of the measles virus RNA polymerase

Cited by 42 publications

References 22 publications

Independent Structural Domains in Paramyxovirus Polymerase Protein

Independent Structural Domains in Paramyxovirus Polymerase Protein

Respiratory Syncytial Virus Polymerase Can Initiate Transcription from Position 3 of the Leader Promoter

Full‐Length Sequence Analysis of Subacute Sclerosing Panencephalitis (SSPE) Virus, a Mutant of Measles Virus, Isolated from Brain Tissues of a Patient Shortly after Onset of SSPE

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