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Identification of a novel cellular TPR-containing protein, SGT, that interacts with the nonstructural protein NS1 of parvovirus H-1.

ABSTRACT: The nonstructural protein NS1 of autonomous parvoviruses is essential for viral DNA amplification and gene expression and is also the major cytopathic effector of these viruses. NS1 acts as nickase, helicase, and ATPase and upregulates P38-driven transcription of the capsid genes. We report here the identification of a novel cellular protein that interacts with NS1 from parvovirus H-1 and which we termed SGT, for small glutamine-rich tetratricopeptide repeat (TPR)-containing protein. The cDNA encoding full-length SGT was isolated through a two-hybrid screen with, as bait, the truncated NS1dlC69 polypeptide, which lacks the C-terminal transactivation domain of NS1. Full-length NS1 and SGT interacted in the two-hybrid system and in an in vitro interaction assay. Northern blot analysis revealed one major transcript of about 2 kb that was present in all rat tissues investigated. Rat sgt cDNA coded for 314 amino acids, and the protein migrated in sodium dodecyl sulfate-polyacrylamide gel electrophoresis with an apparent molecular mass of 34 kDa. SGT could be detected in both the nucleus and the cytoplasm of rat cells, as determined by indirect immunofluorescence analysis and Western blotting of fractionated cellular extracts with an affinity-purified antiserum raised against recombinant SGT (AC1.1). In H-1 virus-infected rat and human cells, compared to mock-infected controls, differences in the migration of SGT polypeptides were revealed after Western blot analysis of total cellular extracts. Moreover, the transient expression of NS proteins was sufficient to induce SGT modification. These results show that cellular SGT, which we have identified as an NS1-interacting protein, is modified by parvovirus infection as well as NS expression.

SUBMITTER: Cziepluch C

PROVIDER: S-EPMC109644 | biostudies-literature | 1998 May

REPOSITORIES: biostudies-literature

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Identification of a novel cellular TPR-containing protein, SGT, that interacts with the nonstructural protein NS1 of parvovirus H-1.

Cziepluch C C Kordes E E Poirey R R Grewenig A A Rommelaere J J Jauniaux J C JC

Journal of virology 19980501 5

The nonstructural protein NS1 of autonomous parvoviruses is essential for viral DNA amplification and gene expression and is also the major cytopathic effector of these viruses. NS1 acts as nickase, helicase, and ATPase and upregulates P38-driven transcription of the capsid genes. We report here the identification of a novel cellular protein that interacts with NS1 from parvovirus H-1 and which we termed SGT, for small glutamine-rich tetratricopeptide repeat (TPR)-containing protein. The cDNA en ...[more]

PMID: 9557704

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Project description:Dengue virus (DENV) NS1 is a multifunctional protein essential for viral replication. To gain insights into NS1 functions in mosquito cells, the protein interactome of DENV NS1 in C6/36 cells was investigated using a proximity biotinylation system and mass spectrometry. A total of 817 mosquito targets were identified as protein-protein interacting with DENV NS1. Approximately 14% of them coincide with interactomes previously obtained in vertebrate cells, including the oligosaccharide transferase complex, the chaperonin containing TCP-1, vesicle localization, and ribosomal proteins. Notably, other protein pathways not previously reported in vertebrate cells, such as epigenetic regulation and RNA silencing, were also found in the NS1 interactome in mosquito cells. Due to the novel and strong interactions observed for NS1 and the epigenetic regulator DIDO1 (Death-Inducer Obliterator 1), the role of DIDO1 in viral replication was further explored. Interactions between NS1 and DIDO1 were corroborated in infected mosquito cells, by colocalization and proximity ligation assays. Silencing DIDO1 expression results in a significant reduction in DENV and ZIKV replication and progeny production. Comparison of transcription analysis of mock or DENV infected cells silenced for DIDO1 revealed variations in multiple gene expression pathways, including pathways associated with DENV infection such as RNA surveillance, IMD, and Toll. These results suggest that DIDO1 is a host factor involved in the negative modulation of the antiviral response necessary for flavivirus replication in mosquito cells. Our findings uncover novel mechanisms of NS1 to promote DENV and ZIKV replication, and add to the understanding of NS1 as a multifunctional protein. IMPORTANCE Dengue is the most important mosquito-borne viral disease to humans. Dengue virus NS1 is a multifunctional protein essential for replication and modulation of innate immunity. To gain insights into NS1 functions, the protein interactome of dengue virus NS1 in Aedes albopictus cells was investigated using a proximity biotinylation system and mass spectrometry. Several protein pathways, not previously observed in vertebrate cells, such as transcription and epigenetic regulation, were found as part of the NS1 interactome in mosquito cells. Among those, DIDO1 was found to be a necessary host factor for dengue and Zika virus replication in mosquito cells. Transcription analysis of infected mosquito cells silenced for DIDO1 revealed alterations of the IMD and Toll pathways, part of the antiviral response in mosquitoes. The results suggest that DIDO1 is a host factor involved in modulation of the antiviral response and necessary for flavivirus replication.

Dataset Information

Identification of a novel cellular TPR-containing protein, SGT, that interacts with the nonstructural protein NS1 of parvovirus H-1.

Publications

Identification of a novel cellular TPR-containing protein, SGT, that interacts with the nonstructural protein NS1 of parvovirus H-1.

Similar Datasets

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