Role of the Hippo pathway in scleroderma pathogenesis

ABSTRACT Systemic Sclerosis (SSc) is a multi-system fibrotic disease characterized by hard, thickened (fibrotic) skin and involvement of multiple internal organs, significantly impacting morbidity, mortality, and quality of life. The pathogenesis of SSc is complex, with contributions from genetic predisposition, striking female predominance, associated vascular alterations (vasculopathy), immune system dysregulation, and aberrant tissue fibrosis. Using single-cell sequencing, we have observed that both myofibroblasts and endothelial cells undergoing endothelial-mesenchymal transition (endo-MT) are the primary source of type I collagen in SSc skin and primarily interact with a subset of myeloid cells (LAMP3+ dendritic cells. We further identified Hippo signaling, an evolutionarily conserved signaling pathway implicated in female-biased autoimmunity through the vestigial like 3 (VGLL3) transcriptional co-regulator, through both direct inhibition and siRNA targeting, as an upstream regulator of both fibroblast-to-myofibroblast and endothelial-to-endo-MT transition in SSc skin. This forms the basis of our central hypotheses that LAMP3+ DC – stromal cell interactions drive myofibroblast and endo- MT transition underlying fibrosis in SSc skin through modulation of Hippo signaling. We will address this hypothesis through three specific aims, with aim 1 to mechanistically dissect the role of the Hippo-signaling pathway in both endo-MTs and myofibroblasts, aim 2 to assess the role of the sex-biased transcriptional co- factor VGLL3, which regulates the Hippo pathway at the nuclear level, and aim 3 to address LAMP3+ DCs, which secrete pro-inflammatory mediators and is the primary immune cell subsets interacting with Endo-MTs and myofibroblast in SSc skin. This proposal will address a novel pathway in SSc pathogenesis, assess cross-talk with pro-inflammatory and pro-fibrotic immune function, and assess the contribution of the sex-biased regulator VGLL3 in the process. Thus, this work may add to our understanding of SSc pathogenesis and the biological processes that drive the marked female bias observed in SSc.