The Mon1-Ccz1 Complex Is the GEF of the Late Endosomal Rab7 Homolog Ypt7

Rab GTPases coordinate membrane fusion reactions [1]. Rab-GDP requires a guanine nucleotide exchange factor (GEF) for its conversion to the active GTP form. It then binds to effectors such as multimeric tethering complexes and supports fusion [2]. GTPase-activating proteins (GAPs) promote GTP hydrolysis to inactivate the Rab. GEFs are thus critical activators of fusion reactions [3, 4]. The Rab GEF family is diverse, ranging from multimeric complexes [5] to monomeric GEFs [6–9]. At the late endosome, Rab7 activation is critical for endosomal maturation. The yeast Rab7 homolog Ypt7 binds to the homotypic fusion and protein sorting (HOPS) complex [10, 11]. Its subunit Vps39/Vam6 has been proposed as a GEF for Ypt7 [12] and the Rag GTPase Gtr1 [13], but other genetic evidence has implicated the endosomal protein Ccz1 as a GEF for Ypt7 [14]. Ccz1 and its binding partner Mon1 have been linked to endosomal transport and maturation [15–20]. We now provide evidence that the dimeric Mon1-Ccz1 complex is the Rab7/Ypt7 GEF. The Mon1-Ccz1 complex, but neither protein alone, counteracts GAP function in vivo, rescues in vitro fusion of vacuoles carrying Ypt7-GDP, and promotes nucleotide exchange on Ypt7 independently of Vps39/HOPS. Our data indicate that the Mon1-Ccz1 complex triggers endosomal maturation by activating Ypt7 on late endosomes. ► Mon1 and Ccz1 interact via their longin domains ► Both proteins work together to counteract GAP activity in vivo ► The Mon1-Ccz1 complex can rescue fusion of GAP-treated vacuoles in vitro ► The Mon1-Ccz1 complex promotes nucleotide exchange on Ypt7 in vitro