Bidirectional regulation between AP-1 and SUMO genes modulates inflammatory signaling during Salmonella Typhimurium infection

Abstract

Gram-negative bacterium Salmonella Typhimurium (STm) is the causative agent of gastroenteritis. Among the various gut pathogens, STm is still one of the most frequent culprits posing a significant health challenge. STm utilizes its effector proteins to highjack host cell processes. Alteration of SUMOylation, a post-translational modification mechanism, is one such change caused by STm. STm mediated simultaneous downregulation of SUMO-pathway genes, Ubc9 and PIAS1, is required for an efficient infection. In the present study, the regulation of SUMO pathway genes during STm infection was investigated. Promoters of both UBC9 and PIAS1, were seen to harbor binding motifs of AP-1, Activator protein-1 (c-Jun:c-Fos heterodimers or c-Jun:c-Jun homodimers). Using electrophoretic mobility shift assays, a direct binding of c-Fos to the identified motifs was observed. Perturbation of c-Fos led to changes in expression of Ubc9 and PIAS1, while its SUMO-modifications resulted in differential regulation of its target genes. In line with this, STm infection of fibroblasts with SUMOylation deficient c-Fos (c-FOS-KOSUMO-def-FOS) resulted in uncontrolled activation of target genes, as revealed by 3mRNA-Seq analysis and mathematical modelling, resulting in massive activation of inflammatory pathways. Infection of c-FOS-KOSUMO-def-FOS cells favored STm replication, indicating misdirected immune mechanisms in these cells. Finally, chromatin Immuno-precipitation assays confirmed a context dependent differential binding and release of AP-1 to/from target genes due to its Phosphorylation and SUMOylation respectively. Overall, our data point towards existence of a bidirectional cross-talk between c-Fos and the SUMO pathway and highlighting its importance in AP-1 function relevant to STm infections and beyond. Author summaryFood borne infections caused Salmonella Typhimurium pose a major health challenge in developing and developed world. Unfortunately, many aspects of Salmonella-host crosstalk still remain unknown. In the current work, using sophisticated computational tools along with cell culture experiments and mathematical modeling, we demonstrate how Salmonella controls SUMOylation, a post-translational modification (PTM) pathway of host. SUMOylation governs fundamental processes of the host cell, and its alteration is required for a successful Salmonella infection. We show that SUMO-pathway genes, Ubc9 and Pias1, are direct target genes of AP-1 transcription factor. C-Fos, a component of AP-1 transcriptionally regulates SUMO-genes by binding to their promoters. During Salmonella infection, a selective activation of target genes of c-Fos was observed. The selective regulation of target genes relied on c-fos PTMs. Experimental perturbation of c-Fos PTMs led to global transcriptional dysregulation including immune hyperactivation. Thus, we show existence of a complex interplay between the SUMO-pathway genes and AP-1 transcription factors which mediate selective gene regulation during Salmonella infection.