ORIGINAL PAPER AtCML8, a calmodulin-like protein, differentially activating CaM-dependent enzymes in Arabidopsis thaliana Hyeong Cheol Park • Chan Young Park • Sung Cheol Koo • Mi Sun Cheong • Kyung Eun Kim • Min Chul Kim • Chae Oh Lim • Sang Yeol Lee • Dae-Jin Yun • Woo Sik Chung Received: 10 May 2010 / Revised: 21 July 2010 / Accepted: 17 August 2010 / Published online: 4 September 2010 Ó Springer-Verlag 2010 Abstract Plants express many calmodulins (CaMs) and calmodulin-like (CML) proteins that sense and transduce different Ca 2? signals. Previously, we reported divergent soybean (Glycine max) CaM isoforms (GmCaM4/5) with differential abilities to activate CaM-dependent enzymes. To elucidate biological functions of divergent CaM pro- teins, we isolated a cDNA encoding a CML protein, AtCML8, from Arabidopsis. AtCML8 shows highest identity with GmCaM4 at the protein sequence level. Expression of AtCML8 was high in roots, leaves, and ﬂowers but low in stems. In addition, the expression of AtCML8 was induced by exposure to salicylic acid or NaCl. AtCML8 showed typical characteristics of CaM such as Ca 2? -dependent electrophoretic mobility shift and Ca 2? binding ability. In immunoblot analyses, AtCML8 was recognized only by antiserum against GmCaM4 but not by GmCaM1 anti- bodies. Interestingly, AtCML8 was able to activate phos- phodiesterase (PDE) but did not activate NAD kinase. These results suggest that AtCML8 acts as a CML protein in Arabidopsis with characteristics similar to soybean divergent GmCaM4 at the biochemical levels. Keywords Arabidopsis  Calcium  Calmodulin (CaM)  Isoform  Calmodulin-like protein (CML)  Phosphodiesterase (PDE)  NAD kinase Introduction In plants, intracellular Ca 2? concentration is one of most important second messengers in the regulation of cellular events such as developmental cues and environmental biotic and abiotic stimuli (Sanders et al. 2002). Calcium is decoded by many calcium-binding proteins in calcium mediated signaling (DeFalco et al. 2010). Calmodulins (CaMs), representative Ca 2? binding proteins, are highly conserved and ubiquitous proteins in plants (McCormack et al. 2005). CaM also mediates Ca 2? signals in depen- dence on developmental and environmental cues and transmits signals to various target enzymes and signaling proteins, including metabolic enzymes, transcription fac- tors, ion channels, protein kinases/phosphatases and structural proteins (Snedden and Fromm 2001; Bouche ´ et al. 2005; Kim et al. 2009). In addition to CaMs, plants also possess a large reper- toire of CaM-like proteins (CMLs) that encode potential calcium sensors and exhibit signiﬁcant structural diver- gence from the typical CaM (McCormack and Braam 2003; McCormack et al. 2005; Ranty et al. 2006; Boonburapong and Buaboocha 2007). Based on sequence analysis of CML proteins, amino acids have been substituted in the EF-hand loop motif, which binds a single Ca 2? ion. Recent data indicated that CaM and CML proteins differ in their Ca 2? afﬁnity and target-binding activities (Hua et al. 2003; Communicated by J. R. Liu. H. C. Park  C. Y. Park  S. C. Koo  M. S. Cheong  K. E. Kim  M. C. Kim  C. O. Lim  S. Y. Lee  D.-J. Yun  W. S. Chung Division of Applied Life Science (BK21 Program), Plant Molecular Biology and Biotechnology Research Center, Jinju, Korea H. C. Park (&)  M. C. Kim  C. O. Lim  S. Y. Lee  D.-J. Yun  W. S. Chung (&) Division of Applied Life Science (BK21 Program), and Environmental Biotechnology National Core Research Center, Gyeongsang National University, Jinju 660-701, Korea e-mail: hcpark@gnu.ac.kr W. S. Chung e-mail: chungws@gnu.ac.kr 123 Plant Cell Rep (2010) 29:1297–1304 DOI 10.1007/s00299-010-0916-7