Determinants of Archetype BK Polyomavirus Replication.

Abstract

BK polyomavirus (BKPyV) is a widespread, small double-stranded DNA virus that is an emerging pathogen in immunocompromised individuals. Following an initial asymptomatic infection, it establishes persistence in the urinary tract of its human host. BKPyV can reactivate and cause diseases such as polyomavirus-associated nephropathy and hemorrhagic cystitis under conditions of immune suppression in renal and bone marrow transplant recipients, respectively.

The focus of this dissertation is to better understand viral mechanisms of regulation that control replication, specifically how the virus is able to establish and maintain a state of persistence following initial infection. Therefore we set out to characterize the transmissible form of BKPyV, archetype virus, which is thought to establish persistence in the host. This virus is defined as having a linear O-P-Q-R-S arrangement of sequence blocks in the non-coding control region (NCCR), of the genomic DNA, as opposed to rearranged variants which have deletions and duplications in their blocks. Rearranged variants can arise during reactivation and are found almost exclusively in patients with BKPyV-associated diseases. Our laboratory developed a natural cell culture model of rearranged variant replication in primary renal proximal tubule epithelial (RPTE) cells. However, archetype virus does not produce progeny in RPTE cells. We determined that of the three major genetic regions that make up the BKPyV genome, the NCCR is responsible for the replication phenotype that differentiates archetype and rearranged virus in RPTE cells. However, we can artificially induce archetype virus propagation in 293TT cells, which overexpress large T antigen, a protein that is required for DNA replication. Furthermore, archetype virus uniquely displays differential promoter activity, driving high levels of a regulatory miRNA early, prior to DNA replication. This miRNA targets early mRNA, thereby limiting virus replication in RPTE cells. This means of repressing viral replication likely contributes to the mechanism of archetype virus persistence in the host. Since there is currently no model of persistence for any human polyomavirus, we have begun to directly examine archetype BKPyV persistence in a cell culture model of urothelial cells. Overall, our results advance our current understanding of archetype polyomavirus replication and persistence.