Expression of Cytosolic Double-Stranded DNA Sensors in Renal Tubular Epithelial Cells.

Abstract

Viral infections can hamper allograft survival and for kidney transplantation specifically, the double-stranded DNA (dsDNA) BK polyoma virus is a huge threat. In kidney transplant recipients with a BK virus infection, the viral particles can be found in the urine and renal tubular epithelium. Microarray analysis of BK virus infection of tubular epithelial cells showed a lack of transcriptional regulation of immune mediators (Abend et al Virology 2010;397). We set out to establish which dsDNA sensors are expressed in resting and stimulated tubular epithelial cells. By that, we aim to establish how BK virus can evade detection and elimination by these mechanisms. As an in vitro model, we used human primary tubular epithelial cells, isolated from the cortex of healthy tissue surrounding Grawitz tumors or from donor kidneys intended for transplantation that were declined for technical reasons. Cells were stimulated with synthetic pattern recognition receptor (PRR) ligands, poly(I:C) as positive dsRNA control and genomic dsDNA from E.coli (dsDNA-EC), both with Lyovec as a transfection reagent. Optimal concentration and incubation time were established. mRNA was isolated and via PCR, RT-qPCR and gel electrophoreses expression was determined. In resting primary tubular epithelial cells, expression of DAI and AIM2 could not be detected while these were clearly up-regulated upon stimulation with both dsDNA-EC and poly(I:C).[figure 1] DDX41, MRE11, LLRFIP1, and Ku70 were present but no altered regulation took place after stimulation. cGAS was only up-regulated upon poly(I:C) simulation. After stimulation with dsDNA-EC or poly(I:C), up-regulation of IFI16, ABCF1, and STING took place. We conclude that primary tubular epithelial cells do express several cytosolic dsDNA receptors, which can be triggered by synthetic PRR ligands. How BK virus can evade these functioning receptors remains to be elucidated.