Regulation of the NLRP1 inflammasome by HSV-1

Project Summary. The goals of this project are to identify how herpes simplex virus 1 (HSV-1) activates and inhibits the NLRP1 inflammasome pathway and to understand the contribution of NLRP1 to antiviral defense in human skin. NLRP1 is a guard protein expressed in human keratinocytes that detects pathogen-encoded virulence factor activities. Yet, the mechanisms regulating NLRP1 activity during viral infections are poorly understood. In preliminary studies, we find that HSV-1 infection of primary human keratinocytes promotes phosphorylation of stress activated protein kinases (SAPKs) and the addition of post-translational modifications to NLRP1 consistent with activation of this guard protein, but that downstream inflammasome formation is not observed in HSV-1 infected cells. We demonstrate that HSV-1 infection blocks the proteasomal degradation of an inhibitory component that restrains NLRP1 inflammasome formation, suggesting a mechanism of inhibition, and that expression of the immediate-early protein ICP0 was necessary and sufficient for this inhibition. Further, ICP0 expression in HSV-1 infected keratinocytes was also necessary for virus induced-SAPK phosphorylation, suggesting that ICP0 activities during infection may trigger the NRLP1 inflammasome. However, the mechanisms by which ICP0 engages and inhibits the NLRP1 inflammasome pathway are undefined. In this application, we will leverage our expertise in HSV-1 biology, inflammasome signaling, and skin immunology to understand these opposing functions of HSV-1 ICP0 in regulating NLRP1 activity and investigate the contribution of this immune sensor to cutaneous antiviral defense in human keratinocytes and an organotypic model of human skin. To this end, we will determine how ICP0 activates SAPKs and its consequences for NLRP1 inflammasome activation (Aim 1), define the mechanism by which HSV-1 ICP0 inhibits the NLRP1 inflammasome (Aim 2), and determine the functional consequences of NLRP1 activation for antiviral defense in human skin (Aim 3). Completion of the proposed studies will provide fundamental mechanistic insight into antiviral defense strategies that are active in human skin during the earliest stages of HSV-1 infection and define how this human pathogen successfully evades these defenses to help establish a life-long infection within its human host.