BIOCHEMISTRY OF SAMHD1-MEDIATED INNATE IMMUNITY RESPONSES

SAMHD1, a mammalian member of the HD-domain hydrolase family of enzymes, catalyzes hydrolysis ofdeoxynucleotides triphosphates (dNTPs) to triphosphate and unphosphorylated nucleosides, which is thoughtto be the main pathway for controlled depletion of cellular dNTPs. Discoveries that SAMHD1 is an immunefactor that restricts retroviral replication in non-cycling immune cells and regulates interferon signaling revealedthat dNTP depletion may act as a defense mechanism of innate antiviral immunity. Existence of suchmechanism implies that the enzymatic activity of SAMHD1 must be controlled by pathways of innate immunesensing and response, and that cellular regulation of SAMHD1 is key to understanding the functionalrelationship between antiviral immunity and dNTP metabolism. In the studies described here we will useunique experimental tools developed by my laboratory to elucidate how biochemical regulation of SAMHD1determines its immune function. This project will explore two novel regulatory mechanisms that have emergedfrom our preliminary work and establish their contribution to the SAMHD1-mediated anti-retroviral state in non-cycling immune cells. The studies will shed light on how and possibly why different molecular clues and cellularsignaling pathways alter susceptibility of myeloid and resting T cells to HIV infection, and thus elucidate thebiological significance of SAMHD1 function at the interface of dNTP metabolism and antiviral defense. In acontinued collaboration with the laboratory of Dr. Diaz-Griffero we will pursue two major specific aims. In Aim 1we will explore the role of nucleic acid binding in the immune function of SAMHD1, elucidate structural andbiochemical determinants of high-affinity interaction of SAMHD1 with oligonucleotides and determine whatnucleic acid species regulate SAMHD1 activity and why. Our preliminary data suggest that phosphorothioatelinkages in nucleic acids may act as a danger-associated molecular pattern or a second messenger in antiviralimmunity. In Aim 2 we will elucidate the mechanism linking redox transformations of SAMHD1 to the enzymaticactivity and the immune function of the protein. Our preliminary studies suggest that redox regulation ofSAMHD1 may offer insight into the emerging role of reactive oxygen species (ROS) in modulating innateantiviral immunity. We will determine what redox states are sampled by the redox-active cysteines of SAMHD1,how these transformations alter the biochemical properties of the protein and explore whether SAMHD1activity is controlled by specific sources of ROS and signaling pathways in the cell.