PTP-MEG2 regulates quantal size and fusion pore opening through two distinct structural bases and substrates

The tyrosine phosphorylation of secretion machinery proteins is a crucial regulatory mechanism for exocytosis. However, the participation of tyrosine phosphatases in different exocytosis stages has not been defined. Here, we demonstrated that PTP-MEG2 controls multiple steps of catecholamine secretion. Biochemical and crystallographic analysis revealed key residues that govern the PTP-MEG2 and NSF-pY83 site interactions, specify PTP-MEG2 substrate selectivity and modulate the fusion of catecholamine-containing vesicles. Unexpectedly, delineation of the PTP-MEG2 mutants along with the NSF interface revealed that PTP-MEG2 controls the fusion pore opening through another unknown substrate. Utilizing a bioinformatics search and biochemical and electrochemical screening, we uncovered that PTP-MEG2 regulates the opening and extension of the fusion pore by dephosphorylating the MUNC18-1 Y145 site, which is associated with epileptic encephalopathy. The crystal structure of the PTP-MEG2/phospho-MUNC18-1-pY145 segment confirmed the interaction of PTP-MEG2 with MUNC18-1 through a distinct structural basis. Our studies extended mechanism insights in complex exocytosis processes.