Interaction of Rab31 and OCRL-1 in oligodendrocytes: its role in transport of mannose 6-phosphate receptors.
Rab31, a protein that we cloned from an oligodendrocyte cDNA library, is required for transport of mannose 6-phosphate receptors (MPRs) from the trans-Golgi network (TGN) to endosomes and for Golgi/TGN organization. Here we extend the knowledge of the mechanism of action of Rab31 by demonstrating its interaction with OCRL-1, a phosphatidylinositol 4,5-diphosphate 5-phosphatase (PI(4,5)P(2) 5-phosphatase) that regulates the levels of PI(4,5)P(2) and PI(4)P, molecules involved in transport and Golgi/TGN organization. We show that Rab31 interacts with OCRL-1 in a yeast two-hybrid system, GST-Rab31 pull-down experiments, and coimmunoprecipitation of OCRL-1 using oligodendrocyte culture lysates. Rab31 and OCRL-1 colocalize in the TGN, post-TGN carriers, and endosomes. Cation-dependent MPR (CD-MPR) is sorted to OCRL-1-containing carriers, but CD63 and vesicular stomatitis virus G (VSVG) are not. siRNA-mediated depletion of endogenous Rab31 causes collapse of the TGN apparatus and markedly decreases the levels of OCRL-1 in the TGN and endosomes. Our observations indicate that the role of Rab31 in the Golgi/TGN structure and transport of MPRs depends on its capability to recruit OCRL-1 to domains of the TGN where the formation of carriers occurs. The importance of our observations is highlighted by the fact that mutation of OCRL-1 causes demyelination in humans.
Rodriguez-Gabin
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AG
Department of Neurology, Albert Einstein College of Medicine, Yeshiva University, Bronx, New York 10461, USA.
Ortiz
E
E
Demoliner
K
K
Si
Q
Q
Almazan
G
G
Larocca
J N
JN
D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D001921:Brain / Q000378:metabolism; D002412:Cations / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D011992:Endosomes / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D009836:Oligodendroglia / Q000378:metabolism*; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics / Q000378:metabolism*; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism; D011485:Protein Binding / Q000235:genetics; D011487:Protein Conformation; D051381:Rats; D017527:Receptor, IGF Type 2 / Q000378:metabolism*; D060149:Tetraspanin 30; D020691:rab GTP-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D021601:trans-Golgi Network / Q000378:metabolism
D015703:Antigens, CD; C554586:CD63 protein, human; D002412:Cations; C554587:Cd63 protein, mouse; D010980:Platelet Membrane Glycoproteins; D017527:Receptor, IGF Type 2; D060149:Tetraspanin 30; D010744:Phosphoric Monoester Hydrolases; D020691:rab GTP-Binding Proteins