Function and structure of RNA polymerase from vesicular stomatitis virus

Area of Science:

• Virology
• Molecular Biology
• Biochemistry

Background:

• Vesicular stomatitis virus (VSV) possesses an RNA-dependent RNA polymerase (RdRp) essential for its replication.
• Understanding the composition and function of viral polymerases is crucial for developing antiviral strategies.

Purpose of the Study:

• To isolate and characterize the RdRp from VSV.
• To elucidate the subunit composition and enzymatic activities of the purified VSV RdRp.

Main Methods:

• Purification of VSV RdRp using cesium sulfate and cesium chloride density gradients, followed by glycerol gradient centrifugation or ion-exchange chromatography.
• Analysis of enzyme subunits and determination of activity based on RNA and poly(A) synthesis.

Main Results:

• A homogeneous preparation of VSV RdRp was obtained, containing L (large protein) and NS proteins.
• Multiple enzyme species were identified with varying L:NS molar ratios; the most active form contained equimolar amounts of L and NS, suggesting a (L)1(NS)1 protomer structure.
• The purified RdRp catalyzed both RNA synthesis and the addition of poly(A) sequences to the 3' termini of RNA products.

Conclusions:

• The VSV RdRp is composed of L and NS proteins, with an active protomer structure of (L)1(NS)1.
• The VSV RdRp exhibits a novel dual function, catalyzing both RNA transcription and post-transcriptional polyadenylation.
• This finding suggests the viral transcriptase itself modifies mRNA by adding poly(A) tails, a mechanism requiring further investigation.