UBL5 Modulates Expression of Crucial TLR Pathway Genes and Inflammasome Activation in Human Macrophage Cells

Abstract We conducted a genome-wide siRNA screen for regulators of LPS-driven TNF expression in macrophages. We used a splicing-independent reporter for the screen, and while we identified very few core spliceosome components among the strongest screen hits, we observed a clear enrichment for peripheral splicing regulators. Among these, a novel putative splicing regulator, UBL5, gave a strong screen phenotype, and a UBL5 shRNA-expressing cell line showed a selective defect in LPS-driven gene expression. While many genes were induced normally by LPS in the UBL5 deficient cells, a subset of genes, including core components of the LPS-activated TLR4 signaling pathway, were substantially diminished in UBL5-deficient cells. We conducted both Illumina and Pacbio long-read sequencing, and we observed marked defects in the LPS-induced splicing of numerous genes, however the primary impact on known components of the TLR4 pathway was observed at the level of overall gene expression rather than alternative splicing. Our data suggests that the putative splicing regulator UBL5 also has key roles in the maintenance and induction of gene expression for several core components of the TLR4 signaling pathway. Key role of UBL5 in the induction of TLR4 signaling pathway was further evidenced by the defective NLRP3 inflammasome activation and protection of OC43 coronavirus infection in UBL5-deficient cells. This work was supported by the Intramural Research Program of NIAID, NIH Intramural Research Program of NIAID, NIH