ABSTRACT Systemic Lupus Erythematosus (SLE) is an autoimmune disorder that is heavily biased 9:1 towards females. Sex bias in SLE reflects both hormonal and sex chromosome differences between men and women. This proposal focuses on identifying and functionally establishing genetic variants on the X chromosome that contribute to SLE sex bias. Current efforts to understand the genetic architecture of lupus using genome-wide association studies (GWAS) have identified variants at 13 X-linked genes that are associated with SLE. However, causative variants at these loci remain unknown. These GWAS loci may contribute to female bias in SLE if they influence how genes respond to X chromosome inactivation (XCI), the early developmental process that epigenetically silences most X-linked genes in females in order to maintain dosage equivalence with males. Although XCI represses most genes on one X in females, some genes escape XCI and are expressed from both active and inactive Xs. XCI states for >20% of human X genes are variable, and escape XCI from a subset of Xs, but are silenced in other females. Our recent data indicate that heritability at genes that variably escape XCI is enriched in sex-biased disorders including SLE. We hypothesize that causal variants on the X chromosome underlie aberrant inactive X expression in SLE. The objective of this proposal is to identify these causal variants and directly evaluate their response to XCI. To address this hypothesis, we propose in Aim 1 to fine map causal variants on the X chromosome and identify causal genes. Integration of results with eQTL and functional genome annotation will further refine loci and link to causal genes. Association effects will be compared between males and females to further differentiate those likely influenced by X inactivation. In Aim 2 we will functionally evaluate variants mapping to Xq28 for altered gene response to XCI and B cell expression. We will perform multiplex reporter gene assays as an initial screen for functional variants with regulatory potential. We will also establish a novel system to functionally evaluate X chromosome variants by editing stem cells that will be differentiated to B cells and undergo XCI. We expect that results from experiments will be the first to provide a critical functional link between associated variants and X chromosome inactivation, and as such will have a profound impact on understanding mechanisms of SLE and lupus biology.