Rotavirus protein NSP3 (NS34) is bound to the 3' end consensus sequence of viral mRNAs in infected cells

Poncet, Didier; Aponte, Carlos; Cohen, Jean

doi:10.1128/jvi.67.6.3159-3165.1993

Cited by 87 publications

(47 citation statements)

References 26 publications

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“…However, deletion of only the terminal C totally abolished binding (Figures 3 and 6), as did the addition of only one C (Table I). Together with our previous localization of the RNA-protein cross-linked site as the last nucleotide of the RNA (Poncet et al, 1993), these observations prompted…”

Section: Minimum Sequence Required For Binding and Cross-linkingmentioning

confidence: 58%

“…Recombinant NSP3A binds to the 3' end of rotaviral mRNAs The recombinant protein NSP3A (rNSP3A) was semipurified from infected Sf9 cells. In non-reducing conditions, the recombinant protein forms multimers, like the natural protein made in rotavirus-infected cells (Mattion et al, 1992;Aponte et al, 1993;Poncet et al, 1993). To determine if the semi-purified rNSP3A was able to bind viral mRNAs, we performed a UV cross-linking immunoprecipitation assay, similar to that used for rotavirusinfected cells (Poncet et al, 1993).…”

Section: Resultsmentioning

confidence: 99%

“…We have previously shown that in group A rotavirusinfected cells, the non-structural protein NSP3 is bound to the 3' conserved sequence of rotavirus mRNAs (Poncet et al, 1993). In the present report we have investigated the RNA sequence needed for binding of NSP3 in vitro.…”

Section: Introductionmentioning

confidence: 86%

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Four nucleotides are the minimal requirement for RNA recognition by rotavirus non-structural protein NSP3.

1994

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The interaction of the group A rotavirus non‐structural protein NSP3 (NSP3A) with RNA has been studied in vitro. Using semi‐purified NSP3A protein expressed by a recombinant baculovirus and in vitro synthesized RNA, we determined by UV cross‐linking and gel retardation assays that NSP3A binds, in a sequence‐specific manner, the consensus sequence (AUGUGACC) present on the 3′ ends of all group A rotavirus mRNAs. Using short oligoribonucleotides, we established that the minimal RNA sequence required for binding of NSP3A is GACC. Modifications of the UGACC oligonucleotide sequence impaired binding of the protein to the RNA. Furthermore, the recombinant NSP3 protein from rotavirus group C showed specificity for the 3′ end consensus sequence (AUGUGGCU) of only group C mRNAs. Sequence analysis of the NSP3 proteins did not reveal significant homologies with other RNA binding proteins, thus the NSP3 proteins of rotaviruses are the prototypes of a new kind of sequence‐specific RNA binding protein.

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Section: Minimum Sequence Required For Binding and Cross-linkingmentioning

confidence: 58%

Section: Resultsmentioning

confidence: 99%

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Four nucleotides are the minimal requirement for RNA recognition by rotavirus non-structural protein NSP3.

1994

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“…The 7th genome segment encodes NSP3 protein [99]. The protein is slightly acidic and binds specifically the conserved tetranucleotide sequence present at the 3 0 end of the rotaviral mRNAs [100]. There is a crystal structure available for NSP3 (PDB ID:1KNZ) {Fig.…”

Section: Non-structural Protein Nsp3mentioning

confidence: 99%

Understanding the penetrance of intrinsic protein disorder in rotavirus proteome

Kumar

Singh

Kumar

et al. 2020

International Journal of Biological Macromolecules

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a b s t r a c tRotavirus is a major cause of severe acute gastroenteritis in the infants and young children. The past decade has evidenced the role of intrinsically disordered proteins/regions (IDPs)/(IDPRs) in viral and other diseases. In general, (IDPs)/(IDPRs) are considered as dynamic conformational ensembles that devoid of a specific 3D structure, being associated with various important biological phenomena. Viruses utilize IDPs/IDPRs to survive in harsh environments, to evade the host immune system, and to highjack and manipulate host cellular proteins. The role of IDPs/IDPRs in Rotavirus biology and pathogenicity are not assessed so far, therefore, we have designed this study to deeply look at the penetrance of intrinsic disorder in rotavirus proteome consisting 12 proteins encoded by 11 segments of viral genome. Also, for all human rotaviral proteins, we have deciphered molecular recognition features (MoRFs), which are disorder based binding sites in proteins. Our study shows the wide spread of intrinsic disorder in several rotavirus proteins, primarily the nonstructural proteins NSP3, NSP4, and NSP5 that are involved in viral replication, translation, viroplasm formation and/or maturation. This study may serve as a primer for understanding the role of IDPs/MoRFs in rotavirus biology, design of alternative therapeutic strategies, and development of disorder-based drugs.

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“…of 5-10 p.f.u./cell. In vivo pulse labelling was performed for 15 min from 0 to 6 h post infection with 0.7 MBq/ml of Trans 35 S-label (ICN: 38 TBq/mmol) as described elsewhere (Poncet et al, 1993).…”

Section: Cells and Virusesmentioning

confidence: 99%

Rotavirus RNA-binding protein NSP3 interacts with eIF4GI and evicts the poly(A) binding protein from eIF4F

et al. 1998

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Most eukaryotic mRNAs contain a 5Ј cap structure and a 3Ј poly(A) sequence that synergistically increase the efficiency of translation. Rotavirus mRNAs are capped, but lack poly(A) sequences. During rotavirus infection, the viral protein NSP3A is bound to the viral mRNAs 3Ј end. We looked for cellular proteins that could interact with NSP3A, using the two-hybrid system in yeast. Screening a CV1 cell cDNA library allowed us to isolate a partial cDNA of the human eukaryotic initiation factor 4GI (eIF4GI). The interaction of NSP3A with eIF4GI was confirmed in rotavirus infected cells by co-immunoprecipitation and in vitro with NSP3A produced in Escherichia coli. In addition, we show that the amount of poly(A) binding protein (PABP) present in eIF4F complexes decreases during rotavirus infection, even though eIF4A and eIF4E remain unaffected. PABP is removed from the eIF4F complex after incubation in vitro with the Cterminal part of NSP3A, but not with its N-terminal part produced in E.coli. These results show that a physical link between the 5Ј and the 3Ј ends of mRNA is necessary for the efficient translation of viral mRNAs and strongly support the closed loop model for the initiation of translation. These results also suggest that NSP3A, by taking the place of PABP on eIF4GI, is responsible for the shut-off of cellular protein synthesis.

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Rotavirus protein NSP3 (NS34) is bound to the 3' end consensus sequence of viral mRNAs in infected cells

Cited by 87 publications

References 26 publications

Four nucleotides are the minimal requirement for RNA recognition by rotavirus non-structural protein NSP3.

Four nucleotides are the minimal requirement for RNA recognition by rotavirus non-structural protein NSP3.

Understanding the penetrance of intrinsic protein disorder in rotavirus proteome

Rotavirus RNA-binding protein NSP3 interacts with eIF4GI and evicts the poly(A) binding protein from eIF4F

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