Interferon-Activated Genetic Programs and a Novel Short Isoform of the SARS-CoV-2 Receptor ACE2 in the Kidney

Background: Severe COVID-19 causes cytokine storm, worsening patient prognosis and contributing to acute kidney injury (AKI) development. Genetic programs activated in the renal epithelium by cytokines like interferon, as well as those ablated by JAK inhibitors, like ruxolitinib, were previously not investigated in detail. Additionally, a short isoform of ACE2, deltaACE2 (dACE2), of unknown function was recently identified as an interferon-stimulated gene, and it's presence, inducibility and regulation in the kidney was not explored. Methods: We treated Human Primary Proximal Tubule (HPPT) renal epithelial cells with IFNa, IFNβ, IFNg, IL-1β and ruxolitinib and used RNA-seq to explore gene expression patterns. We performed GSEA analysis and compared this data to available AKI and renal COVID-19 datasets, as well as to other human interferon-treated tissues. We also measured mRNA expression of both ACE2 isoforms by RT-qPCR before and after cytokine stimulation and identified changes in gene regulatory elements of the ACE2 locus using ChIP-seq. Results: RNA-seq analysis identified genes significantly induced by IFNα (746), IFNβ (1169), IFNγ (1280) and IL-1β (2142), mostly immunity related. We saw an overlap of 162 genes between IFNβ treatment and the post-AKI dataset and of only 35 with severe COVID-19. Comparison of kidney, lung and liver cells treated with IFNβ revealed a shared set of 153 genes and unique 685 renal genes. Using RT-qPCR we show 300-and 600-fold upregulation of dACE2 mRNA by IFNα and IFNβ, respectively, while full length ACE2 expression is almost unchanged. RNA-seq data revealed abundant fragment mapping to exons corresponding to dACE2 compared to rest of the transcript. ChIP-seq analysis showed additional putative regulatory elements in ACE2 locus, including intragenic enhancers and a dACE2 promoter. JAK inhibitor ruxolitinib successfully ablated 79.5% (929) of genes induced by IFNβ in HPPT cells, including dACE2. Conclusions: We generated and made available novel RNA-seq and ChIP-seq datasets for human renal proximal tubule cells stimulated with cytokines. We observed that type I interferons significantly upregulated only the short isoform of SARS-CoV-2 receptor ACE2 and we linked it to JAK/STAT pathway, which may be an important factor in COVID-19 therapies using JAK inhibitors.