MUNC18-1 regulates the submembrane F-actin network, independently of syntaxin1 targeting, via hydrophobicity in β-sheet 10

MUNC18-1 is an essential protein for docking and fusion of secretory vesicles. Mouse chromaffin cells (MCCs) lacking MUNC18-1 show impaired secretory vesicle docking, but also (i) mistargeting of SNARE protein syntaxin1 and (ii) an abnormally dense submembrane F-actin network. Here, we tested the contribution of both phenomena to docking and secretion defects in MUNC18-1 deficient MCCs. We show that abnormal F-actin network and syntaxin1 targeting are not observed in other secretion-deficient cells, Snap25 or synaptotagmin1 knock-out MCCs. We identified a MUNC18-1 mutant (V263T in β-sheet 10) that fully restores syntaxin1 targeting but not F-actin abnormalities in Munc18-1 KO cells. MUNC18-2 or -3, which lack the hydrophobic residue at position 263, did not restore a normal F-actin network. However, these proteins did when a hydrophobic residue was introduced at the corresponding position. Munc18-1 knock-out MCCs expressing MUNC18-1-V263T showed normal vesicle docking and exocytosis. These results demonstrate that MUNC18-1 regulates F-actin network, independent of syntaxin1 targeting, via hydrophobicity in β-sheet 10. The abnormally dense F-actin network in Munc18-1 deficient cells is not a rate-limiting barrier in secretory vesicle docking or fusion.