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Coding assignments of the genome of adult diarrhea rotavirus
1Viral Gastroenteritis Unit, Centers for Disease Control, Atlanta, Georgia.
Abstract:
Adult diarrhea rotavirus (ADRV) has caused epidemics of diarrhea in China since 1982 and remains the only group B rotavirus associated with widespread disease in humans. We recently characterized the proteins of ADRV and have now proceeded to identify the gene segments encoding each protein. Viral RNA transcripts were synthesized in vitro with the endogenous viral RNA polymerase and separated by electrophoresis in agarose. The individual transcripts were translated in a cell-free system using nuclease-treated rabbit reticulocyte lysates. The translation products were compared with polypeptides found in purified virus and were characterized by SDS-PAGE, immunoprecipitation, and Western blot analysis using antisera to double- and single-shelled virions, virus cores, and monoclonal antibodies. Furthermore, individual RNA transcripts were hybridized to total dsRNA to determine their genomic origin. Based on this analysis, the core polypeptides VP1, VP2 and VP3 are encoded by segments 1, 2, and 3, respectively. The main polypeptides in the inner capsid, VP6, and the outer capsid, VP4 and VP7, are encoded by segments 6, 4, and 8 respectively. Segments 5, 7, and 9 code for 60, 45, and 30 kDa nonstructural polypeptides. Two other nonstructural polypeptides (24 and 25 kDa) are derived from gene segment 11. Gene segment 10 codes for a 26 kDa polypeptide that is precipitated with serum to ADRV and may be a structural protein VP9. With this exception, gene coding assignments of ADRV are comparable to those of the group A rotaviruses. Our results have clear implications for further work in cloning, sequencing, and expression genes of ADRV and can provide direction towards understanding the origin and the evolution of this virus.
Insights
Adult diarrhea rotavirus (ADRV) gene segments were identified, mapping viral proteins to specific genes. This research clarifies ADRV
Area of Science:
- Virology
- Molecular Biology
- Genomics
Background:
- Adult diarrhea rotavirus (ADRV), a Group B rotavirus, has caused human diarrhea epidemics in China since 1982.
- Previous characterization of ADRV proteins necessitated identifying the specific gene segments encoding them.
Purpose of the Study:
- To identify the gene segments responsible for encoding each protein in Adult diarrhea rotavirus (ADRV).
- To establish the genetic basis of ADRV protein structure and function for future research.
Main Methods:
- In vitro synthesis of viral RNA transcripts using endogenous viral RNA polymerase.
- Separation of transcripts via agarose electrophoresis and cell-free translation.
- Characterization of translation products using SDS-PAGE, immunoprecipitation, Western blot, and RNA-dsRNA hybridization.
Main Results:
- Core polypeptides VP1, VP2, and VP3 are encoded by gene segments 1, 2, and 3, respectively.
- Capsid proteins VP6, VP4, and VP7 are encoded by segments 6, 4, and 8.
- Segments 5, 7, 9, and 11 encode nonstructural polypeptides of varying molecular weights (60, 45, 30, 24/25 kDa).
- Gene segment 10 encodes a 26 kDa polypeptide, potentially VP9.
Conclusions:
- Gene coding assignments for ADRV are largely comparable to Group A rotaviruses.
- These findings provide a foundation for cloning, sequencing, and expressing ADRV genes.
- The study offers insights into the origin and evolution of ADRV.