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

Abstract

Tyrosine phosphorylation of secretion machinery proteins is a crucial regulatory mechanism for exocytosis. However, the participation of protein tyrosine phosphatases (PTPs) in different exocytosis stages has not been defined. Here we demonstrate that PTP-MEG2 controls multiple steps of catecholamine secretion. Biochemical and crystallographic analyses reveal key residues that govern the interaction between PTP-MEG2 and its substrate, a peptide containing the phosphorylated NSF-pY(83) site, specify PTP-MEG2 substrate selectivity, and modulate the fusion of catecholamine-containing vesicles. Unexpectedly, delineation of PTP-MEG2 mutants along with the NSF binding interface reveals that PTP-MEG2 controls the fusion pore opening through NSF independent mechanisms. Utilizing bioinformatics search and biochemical and electrochemical screening approaches, we uncover that PTP-MEG2 regulates the opening and extension of the fusion pore by dephosphorylating the DYNAMIN2-pY(125) and MUNC18-1-pY(145) sites. Further structural and biochemical analyses confirmed the interaction of PTP-MEG2 with MUNC18-1-pY(145) or DYNAMIN2-pY(125) through a distinct structural basis compared with that of the NSF-pY(83) site. Our studies thus provide mechanistic insights in complex exocytosis processes.