Rubella virus 40S genome RNA specifies a 24S subgenomic mRNA that codes for a precursor to structural proteins

Oker‐Blom, Christian; Ulmanen, Ismo; Kääriäinen, Leevi; Pettersson, Ralf F.

doi:10.1128/jvi.49.2.403-408.1984

Cited by 111 publications

(50 citation statements)

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“…The SPs are synthesized as a precursor polyprotein, which is cleaved into the capsid, E2, and E1 proteins by a host-cell signal peptidase in the lumen of the endoplasmic reticulum. Previous studies have suggested that the capsid protein interacts initially with the viral genomic RNA and is then assembled into the virion with an envelope derived from the host cell membrane containing the E2 and E1 glycoproteins, mainly in the Golgi complex (11)(12)(13)(14). Nucleocapsid formation is a primary function of the capsid protein, but the protein also promotes viral RNA synthesis via an unknown mechanism(s) (15)(16)(17)(18).…”

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Short Self-Interacting N-Terminal Region of Rubella Virus Capsid Protein Is Essential for Cooperative Actions of Capsid and Nonstructural p150 Proteins

Sakata

Otsuki

Okamoto

et al. 2014

J Virol

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Nucleocapsid formation is a primary function of the rubella virus capsid protein, which also promotes viral RNA synthesis via an unknown mechanism. The present study demonstrates that in infected cells, the capsid protein is associated with the nonstructural p150 protein via the short self-interacting N-terminal region of the capsid protein. Mutational analyses indicated that hydrophobic amino acids in this N-terminal region are essential for its N-terminal self-interaction, which is critical for the capsid-p150 association. An analysis based on a subgenomic replicon system demonstrated that the self-interacting N-terminal region of the capsid protein plays a key role in promoting viral gene expression. Analyses using a virus-like particle (VLP) system also showed that the self-interacting N-terminal region of the capsid protein is not essential for VLP production but is critical for VLP infectivity. These results demonstrate that the close cooperative actions of the capsid protein and p150 require the short self-interacting N-terminal region of the capsid protein during the life cycle of the rubella virus. IMPORTANCEThe capsid protein of rubella virus promotes viral RNA replication via an unknown mechanism. This protein interacts with the nonstructural protein p150, but the importance of this interaction is unclear. In this study, we demonstrate that the short N-terminal region of the capsid protein forms a homo-oligomer that is critical for the capsid-p150 interaction. These interactions are required for the viral-gene-expression-promoting activity of the capsid protein, allowing efficient viral growth. These findings provide information about the mechanisms underlying the regulation of rubella virus RNA replication via the cooperative actions of the capsid protein and p150. R ubella virus (RV) is the sole member of the genus Rubivirus in the family Togaviridae. RV is an enveloped, single-stranded, positive-sense RNA virus with a genome of approximately 10 kb. The genome acts as an mRNA and contains three untranslated regions and two open reading frames (ORFs). The ORF at the 5= end encodes two nonstructural proteins (NSPs), p150 and p90, which function in viral RNA replication. The other ORF, at the 3= end, encodes three structural proteins (SPs), capsid, E2, and E1, which are integral components of the virion (1). NSP p150 contains several domains that are conserved among other RNA viruses (2, 3). A putative methyltransferase domain and a protease domain are located at the amino (N) and carboxyl (C) termini, respectively (2, 4, 5). There are also two domains of unknown function, Y and X, between the methyltransferase and protease domains. The putative methyltransferase domain is considered to play a role in capping viral RNA (4). The protease domain cleaves the precursor polyprotein p200 into p150 and p90, and this step is critical in the regulation of viral RNA replication (6-8). NSP p90 has two functional domains, helicase and RNA-dependent RNA polymerase (RdRp) domains, in the N-and C-terminal reg...

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Short Self-Interacting N-Terminal Region of Rubella Virus Capsid Protein Is Essential for Cooperative Actions of Capsid and Nonstructural p150 Proteins

Sakata

Otsuki

Okamoto

et al. 2014

J Virol

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“…RV contains two envelope glycoproteins, E2 and El, both of which are type I membrane proteins, and a capsid protein which associates with the positive-strand genomic RNA in the virion interior (40). The subgenomic RNA which encodes these proteins (41) has been cloned and sequenced (10,13,14). Expression of the cDNAs encoding E2 and E1 together or E1 alone in transfected COS cells resuited in their accumulation in a juxtanuclear location suggestive of the Golgi (18).…”

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The rubella virus E2 and E1 spike glycoproteins are targeted to the Golgi complex.

Hobman

Woodward

Farquhar

1993

The Journal of Cell Biology

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Abstract. Rubella virus (RV) has been reported to bud from intracellular membranes in certain cell types. In this study the intracellular site of targeting of RV envelope E2 and E1 glycoproteins has been investigated in three different cell types (CHO, BHK-21 and Vero cells) transfected with a cDNA encoding the two glycoproteins. By indirect immunofluorescence, E2 and E1 were localized to the Golgi region of all three cell types, and their distribution was disrupted by treatment with BFA or nocodazole. Immunogold labeling demonstrated that E2 and E1 were localized to Golgi cisternae and indicated that the glycoproteins were distributed across the Golgi stack. Analysis of immunoprecipitates obtained from stably transfected CHO cells revealed that E2 and E1 become endo H resistant and undergo sialylation without being transported to the cell surface. Transport of RV glycoproteins to the Golgi complex was relatively slow (tta = 60-90 min). Coprecipitation experiments indicated that E2 and E1 form a heterodimer in the RER. E1 was found to fold much more slowly than E2, suggesting that the delay in transport of the heterodimer to the Golgi may be due to the slow maturation of E1 in the ER. These results indicate that RV glycoproteins behave as integral membrane proteins of the Golgi complex and thus provide a useful model to study targeting and turnover of type I membrane proteins in this organdie.

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“…Rubella virus (RV), the causative agent of German measles, is a positive-strand RNA virus composed of a single genomic RNA (9,757 nucleotides [nt]) beginning with a 5' 7-methyl guanosine cap structure and 3' poly(A) tract (6,31). The RV genome contains two long open reading frames (ORFs), a 5' proximal ORF that encodes the nonstructural proteins (6) and a 3' proximal ORF that encodes the three virus structural proteins (25,30,31,41). In RV-infected cells, a subgenomic RNA is synthesized from which the 3' proximal ORF is expressed (31).…”

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confidence: 99%

“…The RV genome contains two long open reading frames (ORFs), a 5' proximal ORF that encodes the nonstructural proteins (6) and a 3' proximal ORF that encodes the three virus structural proteins (25,30,31,41). In RV-infected cells, a subgenomic RNA is synthesized from which the 3' proximal ORF is expressed (31).…”

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5' sequences of rubella virus RNA stimulate translation of chimeric RNAs and specifically interact with two host-encoded proteins

Pogue

Cao

Singh

et al. 1993

J Virol

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Sequences at the 5' and 3' ends of the rubella virus (RV) genomic RNA can potentially form stable stem-loop (SL) structures that are postulated to be involved in virus replication. We have analyzed the function of these putative SL structures in RNA translation by constructing chimeric chloramphenicol acetyltransferase (CAT) RNAs, flanked either by both 5'and 3'-terminal sequence domains from the RV genome or several deletion derivatives of the same sequences. After in vitro transcription of chimeric RNAs, the translational efficiencies of these RNAs were compared by the rabbit reticulocyte lysate translation system. For in vivo translation studies, the level of CAT activity was measured for chimeric RV/CAT RNAs expressed in transfected cells by the adenovirus major late promoter. Both in vivo and in vitro translation activities of the chimeric RNAs revealed that the presence of 5' and 3' SL sequences of RV RNA, in correct (+) orientation and context [5'(+)SL and 3'(+)SL, respectively] was necessary for efficient translation of chimeric RV/CAT RNAs. The presence of the RV 5'(+)SL sequence had the primary enhancing effect on translation. To identify host proteins which interact with the 5'(+)SL which may be involved in RV RNA translation, RNA gel-shift and UV cross-linking assays were employed. Two host proteins 59 and 52 kDa in size, present in cytosolic extracts from both uninfected and RV-infected cells, specifically interacted with the RV 5'(+)SL RNA. Direct binding comparisons between wild-type and mutant 5'(+)SL RNAs demonstrated that sequences in and around the * Corresponding author. t Present address: Laboratory of Pulmonary and Molecular Immunology, National Heart, Lung, and Blood Institute, Bethesda, MD 20892. sequence termed the internal ribosomal entry site or ribosomal landing pad (8,12,28,33,34). Within this domain of highly complex secondary structure, several structural features, a polypyrimidine tract flanked by two extended SL structures, profoundly influence translation of poliovirus proteins (11). The SL structure present immediately 5' of the polypyrimidine tract must be preserved for both poliovirus translation and 7106

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Rubella virus 40S genome RNA specifies a 24S subgenomic mRNA that codes for a precursor to structural proteins

Cited by 111 publications

References 25 publications

Short Self-Interacting N-Terminal Region of Rubella Virus Capsid Protein Is Essential for Cooperative Actions of Capsid and Nonstructural p150 Proteins

Short Self-Interacting N-Terminal Region of Rubella Virus Capsid Protein Is Essential for Cooperative Actions of Capsid and Nonstructural p150 Proteins

The rubella virus E2 and E1 spike glycoproteins are targeted to the Golgi complex.

5' sequences of rubella virus RNA stimulate translation of chimeric RNAs and specifically interact with two host-encoded proteins

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