Spliced mRNA encoding the murine cytomegalovirus chemokine homolog predicts a beta chemokine of novel structure

A viral mRNA of the late kinetic class expressed by murine cytomegalovirus (MCMV) contains an open reading frame (ORF) whose predicted protein, design ated MCK-1, has homology to beta chemokines (M. R. MacDonald, X.-Y. Li, and H, W. Virgin IV, J, Virol, 71:1671-1678, 1997), The present study analyzed further the structure of the transcript in infected fibroblast cells. A sp licing event removed the MCK-1 stop codon, bringing a downstream ORF into f rame with the chemokine homolog and demonstrating that the MCK-1 ORF was an exon of a larger gene. The predicted 31.4-kDa protein, designated MCK-2, c ontains a putative amino-terminal signal sequence and a beta chemokine doma in, followed by a carboxyl-terminal domain without significant homology to known proteins. Quantitative analysis of mRNA forms in MCMV-infected fibrob last cells at late times after infection indicated that the viral chemokine RNA was predominantly spliced. There was no evidence for expression of RNA encoding either MCK-1 or MCK-2 at immediate early or early times after inf ection with MCMV. Monoclonal antibodies generated against bacterially expre ssed MCK-2 recognized multiple proteins in the range of similar to 30 to si milar to 45 kDa in Western blot analysis of MCK-2. expressed in transfected COS cells. The monoclonal antibodies immunoprecipitated a similar group of proteins in transfected COS cells metabolically labeled with radioactive c ysteine, Radiolabelled protein of apparent higher molecular;lr mass was imm unoprecipitated from culture medium overlying the transfected cells, sugges ting that posttranslationally modified MCK-2 can be secreted. Two proteins with apparent molecular mass suggestive of posttranslational modification w ere detected by Western blot analysis of cells harvested at late times afte r infection with MCMV. These studies show that MCMV encodes and expresses a beta chemokine homolog a with a novel predicted structure.