The Varicella-Zoster Virus ORFS/L (ORF0) Gene Is Required for Efficient Viral Replication and Contains an Element Involved in DNA Cleavage

Abstract

ABSTRACT The genome of varicella-zoster virus (VZV), a human alphaherpesvirus, consists of two unique regions, unique long (U L ) and unique short (U S ), each of which is flanked by inverted repeats. During replication, four isomers of the viral DNA are generated which are distinguished by the relative orientations of U L and U S . VZV virions predominantly package two isomeric forms of the genome that have a fixed orientation of U L . An open reading frame (ORF) of unknown function, ORFS/L, also referred to as ORF0, is located at the extreme terminus of U L , directly adjacent to the a -like sequences, which are known to be involved in cleavage and packaging of viral DNA. We demonstrate here that the ORFS/L protein localizes to the Golgi network in infected and transfected cells. Furthermore, we were able to demonstrate that deletion of the predicted ORFS/L gene is lethal, while retention of the N-terminal 28 amino acid residues resulted in viable yet replication-impaired virus. The growth defect was only partially attributable to the expression of the ORFS/L product, suggesting that the 5′ region of ORFS/L contains a sequence element crucial for cleavage/packaging of viral DNA. Consequently, mutations introduced into the extreme 5′ terminus of ORFS/L resulted in a defect in DNA cleavage, indicating that the region is indeed involved in the processing of viral DNA. Since the sequence element has no counterpart at the other end of U L , we concluded that our results can provide an explanation for the almost exclusive orientation of the U L seen in packaged VZV DNA.