Human cytomegalovirus (HCMV) is of clinical importance and is the leading viral cause of birth defects. HCMV infections are also problematic for immunocompromised individuals and organ transplant recipients. This virus infects a variety of cell types in vivo, and although laboratory studies have demonstrated that HCMV does not productively infect embryonic stem cells, the main symptoms in newborns constitute defects in early nervous system development. While the HCMV life cycle has been studied extensively in human fibroblasts, our understanding is limited in terms of how different cell types in the host contribute to the overall infection. The HCMV genome not only has the capacity to encode hundreds of protein products, but also itself encodes microRNAs (miRNAs), and these viral miRNAs reach high abundance during the infection. Here, I first describe comprehensive deep-sequencing characterization of both viral and human miRNAs during lytic HCMV infection of primary fibroblasts. In addition to the identification of two new HCMV-encoded miRNAs, my results also include the finding that the genomic cluster of human miRNAs 182/96/183 is highly upregulated by late stages of infection. For the second stage of my HCMV studies, I present analysis of the full RNA transcriptomes of three different primary cell types undergoing infection. This work was initiated to compare infected neural progenitors with other more permissive cell types, namely fibroblasts and endothelial cells. In my comparative transcriptome analysis I uncovered a wealth of new findings related to conserved effects of HCMV infection on host RNA processing patterns. Finally, I report preliminary results from high-throughput chromosome conformation studies, which have suggested the existence of virus-host DNA interactions during HCMV infection

Host and virus interactions occurring at the post-transcriptional level are critical for infection but remain poorly understood. Here, we performed comprehensive transcriptome-wide analyses revealing that human cytomegalovirus (HCMV) infection results in widespread alternative splicing (AS), shortening of 3' untranslated regions (3' UTRs) and lengthening of poly(A)-tails in host gene transcripts. We found that the host RNA-binding protein CPEB1 was highly induced after infection, and ectopic expression of CPEB1 in noninfected cells recapitulated infection-related post-transcriptional changes. CPEB1 was also required for poly(A)-tail lengthening of viral RNAs important for productive infection. Strikingly, depletion of CPEB1 reversed infection-related cytopathology and post-transcriptional changes, and decreased productive HCMV titers. Host RNA processing was also altered in herpes simplex virus-2 (HSV-2)-infected cells, thereby indicating that this phenomenon might be a common occurrence during herpesvirus infections. We anticipate that our work may serve as a starting point for therapeutic targeting of host RNA-binding proteins in herpesvirus infections.

Alternative splicing (AS) enables programmed diversity of gene expression across tissues and development. We show here that binding in distal intronic regions (>500 nucleotides (nt) from any exon) by Rbfox splicing factors important in development is extensive and is an active mode of splicing regulation. Similarly to exon-proximal sites, distal sites contain evolutionarily conserved GCATG sequences and are associated with AS activation and repression upon modulation of Rbfox abundance in human and mouse experimental systems. As a proof of principle, we validated the activity of two specific Rbfox enhancers in KIF21A and ENAH distal introns and showed that a conserved long-range RNA-RNA base-pairing interaction (an RNA bridge) is necessary for Rbfox-mediated exon inclusion in the ENAH gene. Thus we demonstrate a previously unknown RNA-mediated mechanism for AS control by distally bound RNA-binding proteins.