Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins of the syntaxin, SNAP-25, and VAMP families mediate intracellular membrane fusion through the formation of helical bundles that span opposing membranes. Soluble SNARE domains that lack their integral membrane anchors inhibit membrane fusion by forming nonfunctional complexes with endogenous SNARE proteins. In this study we investigate the dependence of membrane fusion on the concentration of a soluble SNARE coil domain derived from VAMP2. The increase in the inhibition of fusion observed with increasing concentration of inhibitor is best fit to a function that suggests three SNARE complexes cooperate to mediate fusion of a single vesicle. These three complexes likely contribute part of a protein and lipidic fusion pore.

BACKGROUND Endometriosis is frequently associated with and thought of having propensity to develop into ovarian clear cell carcinoma (OCCC), although the molecular transformation mechanism is not completely understood. METHODS We employed immunohistochemical (IHC) staining for marker expression along the potential progression continuum. Expression profiling of microdissected endometriotic and OCCC cells from patient-matched formalin-fixed, paraffin-embedded samples was performed to explore the carcinogenic pathways. Function of novel biomarkers was confirmed by knockdown experiments. RESULTS PTEN was significantly lost in both endometriosis and invasive tumor tissues, while estrogen receptor (ER) expression was lost in OCCC relative to endometriosis. XRCC5, PTCH2, eEF1A2, and PPP1R14B were significantly overexpressed in OCCC and associated endometriosis, but not in benign endometriosis (p≤0.004). Knockdown experiments with XRCC5 and PTCH2 in a clear cell cancer cell line resulted in significant growth inhibition. There was also significant silencing of a panel of target genes with histone H3 lysine 27 trimethylation, a signature of polycomb chromatin-remodeling complex in OCCC. IHC confirmed the loss of expression of one such polycomb target gene, the serous ovarian cancer lineage marker WT1 in OCCC, while endometriotic tissues showed significant co-expression of WT1 and ER. CONCLUSIONS Loss of PTEN expression is proposed as an early and permissive event in endometriosis development, while the loss of ER and polycomb-mediated transcriptional reprogramming for pluripotency may play an important role in the ultimate transformation process. Our study provides new evidence to redefine the pathogenic program for lineage-specific transformation of endometriosis to OCCC.