ABSTRACT Primary Sjögren's syndrome (pSS), occurring in 1.4 million people in the United States, is the most female- predominant autoimmune disease with a female-to-male ratio of 14:1. While the sex chromosome has been proposed to impact the female bias in pSS, the exact molecular mechanism of X chromosome regulation in pSS- relevant tissues remains elusive. To close this knowledge gap, this project focuses on the regulatory mechanisms controlling the expression of escapees, genes that escape X-chromosome inactivation (XCI), in minor salivary gland-derived mesenchymal stromal cells (MSCs). The intriguing finding that escapees exhibit marked skewing in pSS but not control MSCs underscores the importance of maintaining allelic balance in preventing pSS pathogenesis. Therefore, the objective of this project is to elucidate the molecular checkpoints ensuring allelic balance of escapee expression. The project will test the central hypothesis that dysregulation of miR6891-5p, a HLA-encoded noncoding RNA, leads to skewed escape from XCI in Sjögren's syndrome: Aim 1. Elucidate the functional consequence of restoring miR6891-5p expression in reversing X skewing and inflammatory differentiation of pSS MSCs. Aim 2. Establish the molecular mechanism by which miR6891-5p regulates escape from XCI. Aim 3. Determine the genetic and epigenetic interplay of miR6891-5p-regulated XCI escape. This project will allow us to gain insights into the molecular underpinnings of pSS-associated X chromosomal abnormalities as well as their contribution to the female bias in pSS. By establishing miR6891-5p as a critical gatekeeper against X skewing, this work will provide a novel clinical target for the treatment of pSS and additional autoimmune diseases.

PROJECT NARRATIVE Women are more likely to develop Sjögren's syndrome than men, and the cause of this female bias in disease susceptibility is unknown. This project is designed to delineate the role of miR6891-5p, an RNA molecule, in maintaining accurate gene expression from the two X chromosomes in females and to elucidate how alterations in miR6891-5p may predispose females to Sjögren's syndrome. It has the potential to advance our understanding in the molecular causes for sex differences in autoimmune pathogenesis, and provide novel, pharmacologically targetable pathways for the clinical management of Sjögren's syndrome and additional autoimmune diseases.