KIC, a novel Ca2+ binding protein with one EF-hand motif, interacts with a microtubule motor protein and regulates trichome morphogenesis.
Kinesin-like calmodulin binding protein (KCBP) is a microtubule motor protein involved in the regulation of cell division and trichome morphogenesis. Genetic studies have shown that KCBP is likely to interact with several other proteins. To identify KCBP-interacting proteins, we used the C-terminal region of KCBP in a yeast two-hybrid screen. This screening resulted in the isolation of a novel KCBP-interacting Ca2+ binding protein (KIC). KIC, with its single EF-hand motif, bound Ca2+ at a physiological concentration. Coprecipitation with bacterially expressed protein and native KCBP, gel-mobility shift studies, and ATPase assays with the KCBP motor confirmed that KIC interacts with KCBP in a Ca2+-dependent manner. Interestingly, although both Ca2+-KIC and Ca2+-calmodulin were able to interact with KCBP and inhibit its microtubule binding activity, the concentration of Ca2+ required to inhibit the microtubule-stimulated ATPase activity of KCBP by KIC was threefold less than that required for calmodulin. Two KIC-related Ca2+ binding proteins and a centrin from Arabidopsis, which contain one and four EF-hand motifs, respectively, bound Ca2+ but did not affect microtubule binding and microtubule-stimulated ATPase activities of KCBP, indicating the specificity of Ca2+ sensors in regulating their targets. Overexpression of KIC in Arabidopsis resulted in trichomes with reduced branch number resembling the zwichel/kcbp phenotype. These results suggest that KIC modulates the activity of KCBP in response to changes in cytosolic Ca2+ and regulates trichome morphogenesis.
Reddy
Vaka S
VS
Department of Biology and Program in Cell and Molecular Biology, Colorado State University, Fort Collins, Colorado 80523, USA.
Day
Irene S
IS
Thomas
Tyler
T
Reddy
Anireddy S N
AS
D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism; D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism; D002148:Calmodulin-Binding Proteins / Q000235:genetics / Q000378:metabolism; D022081:Cell Surface Extensions / Q000235:genetics* / Q000502:physiology; D020832:EF Hand Motifs / Q000235:genetics*; D018507:Gene Expression Regulation, Developmental; D018506:Gene Expression Regulation, Plant; D008869:Microtubule-Associated Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D008969:Molecular Sequence Data; D010802:Phylogeny; D025941:Protein Interaction Mapping; D017386:Sequence Homology, Amino Acid
D029681:Arabidopsis Proteins; D002135:Calcium-Binding Proteins; D002147:Calmodulin; D002148:Calmodulin-Binding Proteins; C099551:KCBP protein, Arabidopsis; C484060:KIC protein, Arabidopsis; D008869:Microtubule-Associated Proteins; D000251:Adenosine Triphosphatases; D002118:Calcium