title: Distinct biochemical and functional properties of two Rab5 homologs from the rice blast fungus Magnaporthe oryzae.
abstract: Rab5 is a key regulator of early endocytosis by promoting early endosomal fusion and motility. In this study, we have unexpectedly found distinct properties of the two Rab5 homologs (MoRab5A and MoRab5B) from Magnaporthe oryzae, a pathogenic fungus in plants whose infection causes rice blast disease. Like mammalian Rab5, MoRab5A and MoRab5B can bind to several Rab5 effectors in a GTP-dependent manner, including EEA1, Rabenosyn-5, and Rabaptin-5. However, MoRab5A shows distinct binding characteristics in the sense that both the wild-type and the GTP hydrolysis-defective constitutively active mutant bind the effectors equally well in GST pull-down assays, suggesting that MoRab5A is defective in GTP hydrolysis and mostly in the GTP-bound conformation in the cell. Indeed, GTP hydrolysis assays indicate that MoRab5A GTPase activity is dramatically lower than MoRab5B and human Rab5 and is insensitive to RabGAP5 stimulation. We have further identified a Pro residue in the switch I region largely responsible for the distinct MoRab5A properties by characterization of MoRab5A and MoRab5B chimeras and mutagenesis. The differences between MoRab5A and MoRab5B extend to their functions in the cell. Although they both target to early endosomes, only MoRab5B closely resembles human Rab5 in promoting early endosome fusion and stimulating fluid phase endocytosis. In contrast, MoRab5A correlates with another related early endosomal Rab, Rab22, in terms of the presence of the switch I Pro residue and the blocked GTPase activity. Our data thus identify MoRab5B as the Rab5 ortholog and suggest that MoRab5A specializes to perform a non-redundant function in endosomal sorting.
authors.affiliation: From the Key Laboratory of Biopesticides and Chemical Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, Fujian, 350002, China, and Departments of Biochemistry and Molecular Biology.
authors.forename: Yaoyao
authors.initials: Y
authors.lastname: Qi
authors.affiliation: Departments of Biochemistry and Molecular Biology.
authors.forename: M Caleb
authors.initials: MC
authors.lastname: Marlin
authors.affiliation: Departments of Biochemistry and Molecular Biology.
authors.forename: Zhimin
authors.initials: Z
authors.lastname: Liang
authors.affiliation: Cell Biology, and.
authors.forename: William L
authors.initials: WL
authors.lastname: Berry
authors.affiliation: Cell Biology, and Peggy and Charles Stephenson Cancer Center, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104.
authors.forename: Ralf
authors.initials: R
authors.lastname: Janknecht
authors.affiliation: From the Key Laboratory of Biopesticides and Chemical Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, Fujian, 350002, China, and.
authors.forename: Jie
authors.initials: J
authors.lastname: Zhou
authors.affiliation: From the Key Laboratory of Biopesticides and Chemical Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, Fujian, 350002, China, and.
authors.forename: Zonghua
authors.initials: Z
authors.lastname: Wang
authors.affiliation: From the Key Laboratory of Biopesticides and Chemical Biology, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, Fujian, 350002, China, and gdlufafu@163.com.
authors.forename: Guodong
authors.initials: G
authors.lastname: Lu
authors.affiliation: Departments of Biochemistry and Molecular Biology, Peggy and Charles Stephenson Cancer Center, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma 73104 guangpu-li@ouhsc.edu.
authors.forename: Guangpu
authors.initials: G
authors.lastname: Li
mesh_terms: D000595:Amino Acid Sequence; D004705:Endocytosis; D011992:Endosomes / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal; D006160:Guanosine Triphosphate / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D020082:Magnaporthe / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D050506:Mutant Chimeric Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012275:Oryza / Q000382:microbiology; D010935:Plant Diseases / Q000382:microbiology; D011485:Protein Binding; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D033921:Vesicular Transport Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020696:rab5 GTP-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D005656:Fungal Proteins; D050506:Mutant Chimeric Proteins; D020033:Protein Isoforms; D033921:Vesicular Transport Proteins; D006160:Guanosine Triphosphate; D020696:rab5 GTP-Binding Proteins
keywords: Endocytosis; Endosome; Fungi; GTP Hydrolysis; GTPase; Magnaporthe; Rab