Mapping polyprotein cleavage sites in the R78 region of Maize Chlorotic Dwarf Virus

Yujing Zhao, The College of Wooster

Abstract

Maize chlorotic dwarf virus (MCDV) is a maize virus that causes severe stunting, chlorosis of tertiary veins and leaf tearing in corn. It was prevalent in the southeastern United States and was considered the second major corn virus disease in the US. Thus, it is important to understand and characterize the genome and proteome of MCDV. The major Open Reading Frame (ORF) in the single stranded RNA genome of MCDV codes for a 389 kDa polyprotein that is cleaved into smaller functional proteins by the virus-encoded 3C-like protease. The polyprotein is processed into a 78 kDa protein from the N-terminus (R78). Previous studies have suggested that in MCDV-Severe strain (MCDV-S), R78 is further cleaved, with one possible autocatalytic cleavage site (R78 self-cleavage) and a site cleaved by the 3C-like protease (not part of R78). To pinpoint these cleavage sites, we examined the cleavage of R78 using an in vitro system. This study suggested that R78 was cleaved by the 3C-like protease to produce a C-terminal protein around 33 kDa. A higher ratio of protease to R78 and longer incubation time produced more effective cleavage. Experimental results also suggested that R78 might be autocatalytically cleaved into a protein of 60 kDa.