Host Genes Associated with BK Polyomavirus Entry and Intracellular Trafficking.

Abstract

BK polyomavirus (BKPyV) is a small DNA icosahedral virus measuring about 45nm in diameter, and it was first isolated in 1971. BKPyV infection is ubiquitous and usually asymptomatic; however, BKPyV reactivates in immunosuppressed transplant patients and causes two diseases, polyomavirus associated nephropathy (PVAN) and hemorrhagic cystitis (HC). Due to a lack of specific antiviral drugs, the first-line treatment for BKPyV reactivation is to reduce immunosuppression in PVAN patient or to target host DNA synthesis machinery in HC patient. None of the current treatments is optimal; therefore, elucidating details of the BKPyV life cycle will potentially benefit therapy development and uncover interesting viral and cell biology. Despite being isolated more than 40 year ago, details of the BKPyV life cycle require further elucidation. BKPyV has been considered to infect host cells via a caveolin-mediated pathway. In order to study viral entry in greater detail, caveolin 1, caveolin 2, and clathrin heavy chain were silenced with siRNA in renal proximal tubule epithelial (RPTE) cells. Our experiments showed that caveolin 1, caveolin 2, and clathrin heavy chain knockdown did not block BKPyV infection. However, knocking down UDP-glucose ceramide glucosyltransferase (UGCG), an enzyme required for ganglioside synthesis, decreased BKPyV infection. This suggests that there is a caveolin- and clathrin-independent pathway during BKPyV entry in RPTE cells, and BKPyV does require gangliosides for efficient infection. To further identify host factors associated with BKPyV entry and intracellular trafficking, a whole genome siRNA screen was performed on BKPyV in RPTE cells. The DnaJ heat shock protein family, which has previously been implicated in BKPyV entry, was our top hit. The other hits we identified have not been previously reported; however, many of them are involved in vesicular transport. After validating our top interesting hits, a protein complex was identified to be essential for BKPyV infection. Considering that all of these proteins localize to the ER membrane and participate in the Golgi to ER trafficking, the Golgi to ER trafficking pathway may play an important role in BKPyV infection.