The Calmodulin-related Calcium Sensor CML42 Plays a Role in Trichome Branching

Abstract

Calcium (Ca(2+)) is a key second messenger in eukaryotes where it regulates a diverse array of cellular processes in response to external stimuli. An important Ca(2+) sensor in both animals and plants is calmodulin (CaM). In addition to evolutionarily conserved CaM, plants possess a unique family of CaM-like (CML) proteins. The majority of these CMLs have not yet been studied, and investigation into their physical properties and cellular functions will provide insight into Ca(2+) signal transduction in plants. Here we describe the characterization of CML42, a 191-amino acid Ca(2+)-binding protein from Arabidopsis. Ca(2+) binding to recombinant CML42 was assessed by fluorescence spectroscopy, NMR spectroscopy, microcalorimetry, and CD spectroscopy. CML42 displays significant alpha-helical secondary structure, binds three molecules of Ca(2+) with affinities ranging from 30 to 430 nm, and undergoes a Ca(2+)-induced conformational change that results in the exposure of one or more hydrophobic regions. Gene expression analysis revealed CML42 transcripts at various stages of development and in many cell types, including the support cells, which surround trichomes (leaf hairs) on the leaf surface. Using yeast two-hybrid screening we identified a putative CML42 interactor; kinesin-interacting Ca(2+)-binding protein (KIC). Because KIC is a protein known to function in trichome development, we examined transgenic CML42 knockout plants and found that they possess aberrant trichomes with increased branching. Collectively, our data support a role for CML42 as a Ca(2+) sensor that functions during cell branching in trichomes.