The Role of Type VII Collagen in Mediating Extracellular Matrix Protein Secretion in Recessive Dystrophic Epidermolysis Bullosa

Recessive Dystrophic Epidermolysis Bullosa (RDEB) is a devastating skin blistering disease caused by mutations in the gene encoding type VII collagen (C7). The major phenotypes of RDEB include epidermal fragility, trauma-induced blistering and hard-to-heal wounds. Fibrosis develops rapidly in RDEB skin and contributes to chronic wounds and squamous cell carcinoma (SCC). RDEB patients with chronic fibrosis carry a high risk of metastatic SCC with more than 80% mortality by age 50, making RDEB a deadly disease. A number of studies have shown that RDEB pathogenesis is driven by radical change in extracellular matrix (ECM) composition and increased transforming growth factor-beta (TGFb) signaling that is a direct result of C7 loss of function in dermal fibroblasts. However, the exact mechanism of how C7 loss results in extensive fibrosis is unclear, particularly how TGFb signaling is activated and then sustained through complex networks. Thrombospondin-1 (TSP1) is known as a major activator stimulating TGFb signaling pathways. Previous work has revealed that extracellular TSP1 can activate TGFb signaling by releasing the TGFb ligand from the TGFb complex, particularly in RDEB fibroblasts. In the meantime, significant impairment of TSP1 secretion is also observed when C7 is absent, implying that more TSP1 is accumulated intracellularly in pathological conditions. Therefore, this suggests that accumulated intracellular TSP1 caused by aberrant secretion also promotes TGFb activation. In this thesis, my aims were: 1. identify how loss of C7 impairs TSP1 and other ECM protein secretion, and 2. how the accumulated TSP1 caused by impaired TSP1 secretion contributes to TGFb activation independent from extracellular TSP1 and increased TGFb ligand levels. I demonstrate that C7 directs a specialized secretory pathway by which other large ECM proteins (e.g., TSP1) are also secreted. It is known that C7 secretion is facilitated by Transport ANd Golgi Organization protein 1 (TANGO1), a transmembrane protein promoting large protein secretion at the ER exit sites. Lack of C7 impairs the TANGO1-dependent secretory pathway, and secretion of other large ECM proteins is also impeded. Meanwhile, TSP1 is still observed in the RDEB ECM, implying that TSP1 may also be secreted via TANGO1-independent pathways. By silencing Type I Collagen (C1) secretion previously shown to be independent from TANGO1-mediated secretion, we observe that TSP1 secretion is reduced and that TSP1 is also secreted with C1. The accumulated TSP1 in pathological conditions induces cellular stress and elevates TGFb signaling. Collectively, these data demonstrate a role for C7 in loading TANGO1 vesicle cargo and provide a mechanism for disrupted proteostasis, elevated cellular stress and increased TGF signaling in patients with RDEB. This also implies that the disabled TANGO1-mediated pathway leads to pathology previously associated with two major complications in RDEB: cancer and fibrosis