Direct interaction between a myosin V motor and the Rab GTPases Ypt31/32 is required for polarized secretion.
Rab GTPases recruit myosin motors to endocytic compartments, which in turn are required for their motility. However, no Ypt/Rab GTPase has been shown to regulate the motility of exocytic compartments. In yeast, the Ypt31/32 functional pair is required for the formation of trans-Golgi vesicles. The myosin V motor Myo2 attaches to these vesicles through its globular-tail domain (GTD) and mediates their polarized delivery to sites of cell growth. Here, we identify Myo2 as an effector of Ypt31/32 and show that the Ypt31/32-Myo2 interaction is required for polarized secretion. Using the yeast-two hybrid system and coprecipitation of recombinant proteins, we show that Ypt31/32 in their guanosine triphosphate (GTP)-bound form interact directly with Myo2-GTD. The physiological relevance of this interaction is shown by colocalization of the proteins, genetic interactions between their genes, and rescue of the lethality caused by a mutation in the Ypt31/32-binding site of Myo2-GTD through fusion with Ypt32. Furthermore, microscopic analyses show a defective Myo2 intracellular localization in ypt31Delta/32ts and in Ypt31/32-interaction-deficient myo2 mutant cells, as well as accumulation of unpolarized secretory vesicles in the latter mutant cells. Together, these results indicate that Ypt31/32 play roles in both the formation of trans-Golgi vesicles and their subsequent Myo2-dependent motility.
Lipatova
Zhanna
Z
Laboratory for Molecular Biology, Department of Biological Sciences, University of Illinois at Chicago, Chicago, IL 60607, USA.
Tokarev
Andrei A
AA
Jin
Yui
Y
Mulholland
Jon
J
Weisman
Lois S
LS
Segev
Nava
N
D049109:Cell Proliferation; D005089:Exocytosis; D015966:Gene Expression Regulation, Fungal; D006056:Golgi Apparatus / Q000378:metabolism; D008968:Molecular Conformation; D009154:Mutation; D018995:Myosin Heavy Chains / Q000737:chemistry* / Q000378:metabolism; D024701:Myosin Type V / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism; D020798:Two-Hybrid System Techniques; D020691:rab GTP-Binding Proteins / Q000737:chemistry* / Q000378:metabolism
C077512:MYO2 protein, S cerevisiae; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; C087905:YPT31 protein, S cerevisiae; D024701:Myosin Type V; C121633:YPT32 protein, S cerevisiae; D018995:Myosin Heavy Chains; D020691:rab GTP-Binding Proteins