Complete Title of Thesis:
"Biogenesis and Biological Functions of the Gammaherpesvirus 68 Encoded MicroRNAs"
Prepared under the direction of: Linda F. van Dyk, Ph.D.
Recently it has been discovered that viruses, like their host cell counterparts, have the ability to encode biologically functional microRNAs (miRNAs). Current research dictates that an overwhelming majority of miRNA encoding viruses are found within the Herpesviridae family. This thesis focuses on furthering our understanding of the biogenesis and biological functions associated with the miRNAs of Gammaherpesvirus 68 (γHV68). γHV68 is a virus of wild murid rodents used as a small animal model to study gammaherpesvirus pathogenesis and encodes 10 miRNAs originally detected from a latently infected tumor cell line that reside adjacent to sequences in the γHV68 genome proposed to encode viral tRNA-like genes (vtRNAs). miRNAs are short, noncoding RNAs that have the ability to post-transcriptionally regulate the translation of target mRNAs which, in most cases, results in the decrease of protein expression from the target mRNA. Most cellular and viral miRNAs are transcribed by RNA polymerase II to generate a primary-miRNA transcript that becomes sequentially processed by RNaseIII family enzymes, Drosha and Dicer, into biologically functional RNAs 20-25 nucleotides in length. Due to the close proximity of the vtRNAs and miRNAs within the γHV68 genome and the known role of RNA polymerase III transcription of tRNA genes, it was predicted that the γHV68 miRNAs may be transcribed by RNA polymerase III rather than by RNA polymerase II. Additionally, due to the discovery of many herpesvirus miRNAs, including the γHV68 miRNAs, in latently infected cell lines; roles for herpesvirus miRNAs in latency have also been predicted.
Here, we demonstrate that the γHV68 miRNAs are biologically active during infection and that their biogenesis is linked to RNA polymerase III transcription and an alternative processing pathway that is a hybrid between the established tRNA and miRNA processing pathways. We also have generated multiple mRNA target prediction lists to define possible roles for herpesvirus miRNAs during latency. Through the use of this targeting list we have identified CD79B as a conserved target of regulation by gammaherpesvirus miRNAs encoded by EBV, KSHV, and γHV68.