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Polyadenylate in the virion RNA of mouse hepatitis virus
Abstract:
Mouse hepatitis (MH) virus was grown in SR-CDF1-DBT, a mouse cell line, and purified by ammonium sulfate precipitation and by density gradient centrifugation. Extraction of RNA from purified virions with 1% SDS and sedimentation analysis of the RNA revealed a major 50S component and two minor components. Treatment of virions with phenol/chloroform also produced the 50S component, although its yield was lower. MH virion RNA can bind to a poly(U)-fiberglass filter, indicating that MH virion RNA contains poly(A). A poly(A)-like fragment was isolated by digestion with ribonuclease A [EC 3.1.4.22] and T1 [EC 3.1.4.8] and by DEAE-Sephadex column chromatography. Analysis of the fragment for base composition showed it to be an adenine-rich material. Its chain length was about 90 nucleotides, as determined by ion-exchange chromatography and gel electrophoresis.
Insights
Mouse hepatitis virus (MHV) RNA was isolated and analyzed. The study found MHV virion RNA contains a poly(A) tail, a crucial finding for understanding viral replication.
Area of Science:
- Virology
- Molecular Biology
- Biochemistry
Background:
- Mouse hepatitis virus (MHV) is a significant pathogen in mice.
- Understanding the structure of MHV is crucial for developing antiviral strategies.
Purpose of the Study:
- To characterize the RNA component of purified MHV virions.
- To determine if MHV virion RNA contains a polyadenylated (poly(A)) tail.
Main Methods:
- Cell culture of MHV in SR-CDF1-DBT cells.
- Purification of virions using ammonium sulfate precipitation and density gradient centrifugation.
- RNA extraction, sedimentation analysis, poly(U)-fiberglass filter binding assay, enzymatic digestion, and chromatographic/electrophoretic analysis.
Main Results:
- A major 50S RNA component was identified in purified MHV virions.
- MHV virion RNA demonstrated binding to a poly(U)-fiberglass filter, indicating the presence of poly(A).
- A poly(A)-like fragment of approximately 90 nucleotides was isolated and characterized as adenine-rich.
Conclusions:
- MHV virion RNA possesses a poly(A) tail.
- The presence of poly(A) in MHV RNA has implications for viral RNA replication and stability.