Munc18-2, a functional partner of syntaxin 3, controls apical membrane trafficking in epithelial cells

The Sec1-related proteins bind to syntaxin family t-SNAREs with high affini ty, thus controlling the interaction of syntaxins with their cognate SNARE partners. Munc18-2 is a Sec1 homologue enriched in epithelial cells and for ms a complex with syntaxin 3, a t-SNARE localized to the apical plasma memb rane. We generated here a set of Munc18-2 point mutants with substitutions in conserved amino acid residues. The mutants displayed a spectrum of diffe rent syntaxin binding efficiencies. The in vitro and in vitro binding patte rns were highly similar, and the association of the Munc18-2 variants with syntaxin 3 correlated well with their ability to displace SNAP-23 from synt axin 3 complexes when overexpressed in Caco-2 cells. Even the Munc18-2 muta nts that do not detectably bind syntaxin 3 were membrane associated in Caco -2 cells, suggesting that the syntaxin interaction is not the sole determin ant of Sec1 protein membrane attachment, Overexpression of the wild-type Mu nc18-2 was shown to inhibit the apical delivery of influenza virus hemagglu tinin (HA), Interestingly, mutants unable to bind syntaxin 3 behaved differ ently in the HA transport assay. While one of the mutants tested had no eff ect, one inhibited and one enhanced the apical transport of RA This implies that Munc18-2 function in apical membrane trafficking involves aspects ind ependent of the syntaxin 3 interaction.