Calcium negatively modulates calmodulin interaction with IQGAP1.
IQGAP1 regulates cytoskeletal dynamics through interactions with the Rho family GTPases Rac1 and Cdc42, F-actin, and beta-catenin. Calmodulin interaction with IQ motifs of IQGAP1 negatively influences these IQGAP1 interactions. Although, calmodulin interacts with IQGAP1 in the absence of Ca(2+) and was suggested to exhibit reduced binding when Ca(2+) bound, recent reports show substantially greater binding when Ca(2+) is present. Binding evaluations have primarily relied on IQGAP1 interaction with calmodulin conjugated to Sepharose 4B. In this study we evaluated the Ca(2+)-dependence of calmodulin interaction with native IQGAP1 using a series of independent biochemical approaches. We found the apparent binding of calmodulin to IQGAP1 was Ca(2+)-independent, being between 5- and 20-fold greater in the absence than in the presence of Ca(2+). In addition, calmodulin interaction with IQGAP1 was negatively regulated by buffer [Ca(2+)] (IC(50)=3.4x10(-7)M). Regulation was specific to Ca(2+), as Ba(2+) was approximately 400-fold less effective than Ca(2+) at modulating the interaction. Moreover, testing of calmodulin mutants demonstrated that apocalmodulin tightly binds IQGAP1 and that the N- and C-terminal pair of EF hands are important for Ca(2+) sensitivity. These data indicate that calmodulin may disassemble from IQGAP1 to facilitate IQGAP1 interaction with effectors of cytoskeletal reorganization during conditions of cell activation that promote increased cytosolic [Ca(2+)].
Li
Quanwen
Q
Department of Molecular and Integrative Physiology, 7807 Medical Sciences II Bldg., The University of Michigan, Ann Arbor, MI 48109-0622, USA.
Stuenkel
Edward L
EL
D000818:Animals; D002118:Calcium / Q000502:physiology*; D002147:Calmodulin / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D011233:Precipitin Tests; D011485:Protein Binding; D020703:ras GTPase-Activating Proteins
D002147:Calmodulin; D002352:Carrier Proteins; C101203:IQ motif containing GTPase activating protein 1; D020703:ras GTPase-Activating Proteins; D002118:Calcium