RRC ID 39577
著者 Etoh K, Fukuda M.
タイトル Structure-function analyses of the small GTPase Rab35 and its effector protein centaurin-β2/ACAP2 during neurite outgrowth of PC12 cells.
ジャーナル J Biol Chem
Abstract The small GTPase Rab35 is a molecular switch for membrane trafficking that regulates a variety of cellular events. We previously showed that Rab35 promotes neurite outgrowth of nerve growth factor-stimulated PC12 cells through interaction with centaurin-β2 (also called ACAP2). Centaurin-β2 is the only Rab35-binding protein reported thus far that exclusively recognizes Rab35 and does not recognize any of the other 59 Rabs identified in mammals, but the molecular basis for the exclusive specificity of centaurin-β2 for Rab35 has remained completely unknown. In this study, we performed deletion and mutation analyses and succeeded in identifying the residues of Rab35 and centaurin-β2 that are crucial for formation of a Rab35·centaurin-β2 complex. We found that two threonine residues (Thr-76 and Thr-81) in the switch II region of Rab35 are responsible for binding centaurin-β2 and that the same residues are dispensable for Rab35 recognition by other Rab35-binding proteins. We also determined the minimal Rab35-binding site of centaurin-β2 and identified two asparagine residues (Asn-610 and Asn-691) in the Rab35-binding site as key residues for its specific Rab35 recognition. We further showed by knockdown-rescue approaches that neither a centaurin-β2 binding-deficient Rab35(T76S/T81A) mutant nor a Rab35 binding-deficient centaurin-β2(N610A/N691A) mutant supported neurite outgrowth of PC12 cells, thereby demonstrating the functional significance of the Rab35/centaurin-β2 interaction during neurite outgrowth of PC12 cells.
巻・号 290(14)
ページ 9064-74
公開日 2015-4-3
DOI 10.1074/jbc.M114.611301
PII S0021-9258(19)81996-8
PMID 25694427
PMC PMC4423693
MeSH Amino Acid Sequence Animals Base Sequence DNA Primers GTPase-Activating Proteins / genetics GTPase-Activating Proteins / physiology* Molecular Sequence Data Mutagenesis, Site-Directed Neurites* PC12 Cells Polymerase Chain Reaction Rats Sequence Homology, Amino Acid Structure-Activity Relationship rab GTP-Binding Proteins / chemistry rab GTP-Binding Proteins / genetics rab GTP-Binding Proteins / physiology*
IF 4.238
引用数 10
WOS 分野 BIOCHEMISTRY & MOLECULAR BIOLOGY
リソース情報
遺伝子材料 pEGFP-C1-mouse Rab35 (RDB14197) pEGFP-C1-mouse Rab35(TS/TA) (RDB14198) pEGFP-C1-mouse Rab35(S5A) (RDB14199).