CrkL plays a role in SDF-1-induced activation of the Raf-1/MEK/Erk pathway through Ras and Rac to mediate chemotactic signaling in hematopoietic cells.
Intracellular signaling mechanisms regulating SDF-1-induced chemotaxis of hematopoietic cells have remained elusive. Here we demonstrate that overexpression of the adaptor molecule CrkL enhances SDF-1-induced chemotaxis of hematopoietic BaF3 and 32Dcl3 cells. Overexpression of CrkL also enhanced SDF-1-induced activation of the Raf-1/MEK/Erk signaling pathway as well as that of the small GTPases Ras, Rap1, and Rac, while a dominant negative mutant of Ras or Rac suppressed CrkL-enhanced Erk activation. SDF-1 stimulation induced tyrosine phosphorylation of CrkL, which was inhibited by the Src family kinase inhibitor PP1 or by dominant negative mutants of Lyn, thus indicating that Lyn mediated SDF-1-induced phosphorylation of CrkL. However, inhibition of the Lyn kinase activity failed to affect SDF-1-induced activation of the small GTPases and Erk. On the other hand, SDF-1-induced activation of the Erk signaling pathway as well as chemotaxis was inhibited by overexpression of a CrkL mutant lacking the N-terminal SH3 domain, which mediates interaction with various signaling molecules including guanine nucleotide exchange factors for the Ras and Rho family GTPases. SDF-1-induced chemotaxis was also inhibited by the dominant negative Ras or Rac mutant as well as by the MEK inhibitor PD98059. These results indicate that CrkL mediates SDF-1-induced activation of the Raf-1/MEK/Erk signaling pathway through Ras as well as Rac in hematopoietic cells and, thereby, plays important roles in the induction of chemotactic response.
Arai
Ayako
A
Department of Hematology, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8519, Japan.
Aoki
Mae
M
Weihua
Yan
Y
Jin
Aishun
A
Miura
Osamu
O
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000502:physiology*; D000818:Animals; D002460:Cell Line; D054377:Chemokine CXCL12; D019743:Chemokines, CXC / Q000494:pharmacology*; D002633:Chemotaxis / Q000187:drug effects*; D004318:Doxycycline / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D051379:Mice; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D008954:Models, Biological; D009154:Mutation / Q000235:genetics; D009687:Nuclear Proteins / Q000235:genetics / Q000502:physiology*; D010766:Phosphorylation / Q000187:drug effects; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D014162:Transfection; D014443:Tyrosine / Q000378:metabolism; D020744:rac GTP-Binding Proteins / Q000378:metabolism*; D018631:ras Proteins / Q000378:metabolism*; D018909:src Homology Domains / Q000235:genetics / Q000502:physiology; D019061:src-Family Kinases / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; C082869:CRKL protein; C515255:CXCL12 protein, human; D054377:Chemokine CXCL12; D019743:Chemokines, CXC; C515256:Cxcl12 protein, mouse; D009687:Nuclear Proteins; D014443:Tyrosine; C060902:lyn protein-tyrosine kinase; D019061:src-Family Kinases; D019908:Proto-Oncogene Proteins c-raf; D048049:Extracellular Signal-Regulated MAP Kinases; D020929:Mitogen-Activated Protein Kinase Kinases; D020744:rac GTP-Binding Proteins; D018631:ras Proteins; D004318:Doxycycline