Murine Cytomegalovirus Encodes Proteins that Regulate Viral Late Transcription

Abstract

OF THE DISSERTATION Murine Cytomegalovirus Encodes Proteins that Regulate Viral Late Transcription by Travis James Chapa Doctor of Philosophy in Biology and Biomedical Sciences Molecular Microbiology and Microbial Pathogenesis Washington University in St. Louis, 2014 Professor Anthony French, Chair Professor Dong Yu, Co-Chair Human Cytomegalovirus (HCMV) is a Beta-herpesvirus that causes severe disease in immunocompromised individuals (including AIDS patients), and is the leading viral cause of congenital birth defects. Murine Cytomegalovirus (MCMV) is the primary surrogate model for HCMV, and resembles the human virus with respect to virion structure, genome organization, gene expression, tissue tropism, and clinical manifestations. In my graduate studies, I discovered the function of two novel MCMV proteins during in vitro infection. I demonstrated that ORF M79 encoded protein pM79, and ORF M92 encoded protein pM92. Both pM79 and pM92 have homologs in HCMV, and I showed that they regulate late viral gene transcription. During infection, a mutant virus for either M79 or M92 accumulated representative viral immediate early gene products, early gene products, and viral DNA sufficiently but had severe reduction in the accumulation of late gene products, thus unable to produce infectious progeny. Analysis of the viral transcriptome via tiled array and quantitative PCR analysis revealed that many late transcripts sensitive to a viral DNA synthesis inhibitor (phosphonoacetic acid) were markedly reduced by pM79 or pM92 mutation. Co-immunoprecipitation and mass spectrometry analysis revealed the interactions between pM92 and pM79, and suggests that they are part of a larger virus transcription complex. A colleague demonstrated that this function was conserved in the HCMV homolog pUL79, and I showed that pM92 expression in trans could complement the growth defect of pUL92 deficient HCMV. My work was the first evidence of this regulatory complex in MCMV. This complex represents a potential new