The Traf2- and Nck-interacting kinase as a putative effector of Rap2 to regulate actin cytoskeleton.
Rap2 belongs to the Ras family of small GTP-binding proteins, but its specific roles in cell signaling remain unknown. In the present study, we have affinity-purified from rat brain a Rap2-interacting protein of approximately 155 kDa, p155. By liquid chromatography tandem mass spectrometry, we have identified p155 as Traf2- and Nck-interacting kinase (TNIK). TNIK possesses an N-terminal kinase domain homologous to STE20, the Saccharomyces cerevisiae mitogen-activated protein kinase kinase kinase kinase, and a C-terminal regulatory domain termed the citron homology (CNH) domain. TNIK induces disruption of F-actin structure, thereby inhibiting cell spreading. In addition, TNIK specifically activates the c-Jun N-terminal kinase (JNK) pathway. Among our observations, TNIK interacted with Rap2 through its CNH domain but did not interact with Rap1 or Ras. TNIK interaction with Rap2 was dependent on the intact effector region and GTP-bound configuration of Rap2. When co-expressed in cultured cells, TNIK colocalized with Rap2, while a mutant TNIK lacking the CNH domain did not. Rap2 potently enhanced the inhibitory function of TNIK against cell spreading, but this was not observed for the mutant TNIK lacking the CNH domain. Rap2 did not significantly enhance TNIK-induced JNK activation, but promoted autophosphorylation and translocation of TNIK to the detergent-insoluble cytoskeletal fraction. These results suggest that TNIK is a specific effector of Rap2 to regulate actin cytoskeleton.
Taira
Kiyohito
K
Division of Cell Biology, Graduate School of Medicine, University of the Ryukyus, 207 Uehara, Nishihara-cho, Okinawa 903-0215, Japan.
Umikawa
Masato
M
Takei
Kimiko
K
Myagmar
Bat-Erdene
BE
Shinzato
Manabu
M
Machida
Noriko
N
Uezato
Hiroshi
H
Nonaka
Shigeo
S
Kariya
Ken-ichi
K
D000199:Actins / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000378:metabolism; D002460:Cell Line; D002853:Chromatography, Liquid; D003599:Cytoskeleton / Q000378:metabolism*; D018076:DNA, Complementary / Q000378:metabolism; D003902:Detergents / Q000494:pharmacology; D017353:Gene Deletion; D000081410:Germinal Center Kinases; D005982:Glutathione Transferase / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000737:chemistry; D006801:Humans; D007313:Insecta; D047908:Intracellular Signaling Peptides and Proteins; D020930:MAP Kinase Kinase Kinases; D051379:Mice; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009154:Mutation; D041681:NIH 3T3 Cells; D028861:Nuclear Pore Complex Proteins / Q000737:chemistry; D010766:Phosphorylation; D011485:Protein Binding; D011487:Protein Conformation; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000502:physiology*; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D051381:Rats; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry; D020798:Two-Hybrid System Techniques; D020668:rap GTP-Binding Proteins / Q000378:metabolism*
D000199:Actins; D018076:DNA, Complementary; D003902:Detergents; D000081410:Germinal Center Kinases; D047908:Intracellular Signaling Peptides and Proteins; C080427:NUP155 protein, human; D028861:Nuclear Pore Complex Proteins; D029701:Saccharomyces cerevisiae Proteins; D006160:Guanosine Triphosphate; D005982:Glutathione Transferase; D017346:Protein Serine-Threonine Kinases; C000631454:TNIK protein, human; D020930:MAP Kinase Kinase Kinases; C078754:STE20 protein, S cerevisiae; C116756:RAP2A protein, human; D020668:rap GTP-Binding Proteins