Involvement of the adapter protein CRKL in integrin-mediated adhesion

CRKL, an SH2-SH3-SH3 adapter protein, is one of the major tyrosine phosphop roteins detected in primary leukemic neutrophils from patients with CML. CR KL binds directly to BCR/ABL through its N-terminal SH3 domain, suggesting it may be involved in BCR/ABL signal transduction. However, the biological function of CRKL in either normal or leukemic cells is still largely unknow n. In this study, we have examined the effects of overexpressing full lengt h or deletion mutants of CRKL in hematopoietic cell Lines. Full length, SH2 - and SH3(N)-domain deletion mutants of CRKL were transfected into an inter leukin-3-dependent hematopoietic cell line, Ba/F3, and 3-5 individual subli nes which stably overexpressed each transgene were obtained [Bal F-CRKL, Ba /F-CRKL Delta SH2, and Ba/F-CRKL Delta SH3(N)]. The growth properties of th ese transfected cells in the presence or absence of IL-3 were not different from mock transfected or untransfected Ba/F3 cells. However, Ba/F3 cells o verexpressing full length CRKL, but not deletion mutants of CRKL, were foun d to have an increase in their ability to bind to fibronectin-coated surfac es. Further, expression of full length, but not Delta SH2- or Delta SH3-CRK L deletion mutants, was found to alter cell morphology on fibronectin-coate d plates, an effect which was further enhanced by certain kinds of stress s timuli, such as ionizing radiation. Similar results were obtained when CRKL was transiently overexpressed in Ba/F3 cells, and were also obtained in a second IL-3 dependent hematopoietic cell line, 32Dcl3. Adhesion to fibronec tin was blocked by anti-beta 1 integrin monoclonal antibody, but overexpres sion of CRKL did not affect surface expression of beta 1 integrins, nor did it spontaneously induce expression of the beta 1 integrin 'activation' epi tope recognized by the 9EG7 monoclonal antibody. These data suggest a role for CRKL in signaling pathways which regulate adhesion to fibronectin.