The small GTPase Rap1 is required for Mn(2+)- and antibody-induced LFA-1- and VLA-4-mediated cell adhesion.
In T-lymphocytes the Ras-like small GTPase Rap1 plays an essential role in stimulus-induced inside-out activation of integrin LFA-1 (alpha(L)beta(2)) and VLA-4 (alpha(4)beta(1)). Here we show that Rap1 is also involved in the direct activation of these integrins by divalent cations or activating antibodies. Inhibition of Rap1 either by Rap GTPase-activating protein (RapGAP) or the Rap1 binding domain of RalGDS abolished both Mn(2+)- and KIM185 (anti-LFA-1)-induced LFA-1-mediated cell adhesion to intercellular adhesion molecule 1. Mn(2+)- and TS2/16 (anti-VLA-4)-induced VLA-4-mediated adhesion were inhibited as well. Interestingly, both Mn(2+), KIM185 and TS2/16 failed to induce elevated levels of Rap1GTP. These findings indicate that available levels of GTP-bound Rap1 are required for the direct activation of LFA-1 and VLA-4. Pharmacological inhibition studies demonstrated that both Mn(2+)- and KIM185-induced adhesion as well as Rap1-induced adhesion require intracellular calcium but not signaling activity of the MEK-ERK pathway. Moreover, functional calmodulin signaling was shown to be a prerequisite for Rap1-induced adhesion. From these results we conclude that in addition to stimulus-induced inside-out activation of integrins, active Rap1 is required for cell adhesion induced by direct activation of integrins LFA-1 and VLA-4. We suggest that Rap1 determines the functional availability of integrins for productive binding to integrin ligands.
de Bruyn
Kim M T
KM
Department of Physiological Chemistry and Centre for Biomedical Genetics, University Medical Centre Utrecht, Utrecht University, Universiteitsweg 100, P. O. Box 85060, 3508 AB Utrecht, The Netherlands.
Rangarajan
Savithri
S
Reedquist
Kris A
KA
Figdor
Carl G
CG
Bos
Johannes L
JL
D000906:Antibodies / Q000502:physiology*; D018821:CD18 Antigens / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism; D002413:Cations, Divalent; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D039041:Integrin alpha4beta1; D019012:Integrin beta1 / Q000378:metabolism; D016023:Integrins / Q000502:physiology*; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism; D019169:Jurkat Cells; D020014:K562 Cells; D016169:Lymphocyte Function-Associated Antigen-1 / Q000502:physiology*; D008345:Manganese / Q000378:metabolism*; D016201:Receptors, Lymphocyte Homing / Q000502:physiology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D019010:Vascular Cell Adhesion Molecule-1 / Q000378:metabolism; D020827:rap1 GTP-Binding Proteins / Q000378:metabolism*
D000906:Antibodies; D018821:CD18 Antigens; D002147:Calmodulin; D002413:Cations, Divalent; D039041:Integrin alpha4beta1; D019012:Integrin beta1; D016023:Integrins; D016169:Lymphocyte Function-Associated Antigen-1; D016201:Receptors, Lymphocyte Homing; D019010:Vascular Cell Adhesion Molecule-1; D018799:Intercellular Adhesion Molecule-1; D008345:Manganese; D020827:rap1 GTP-Binding Proteins; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium