Isolation, characterization and expression of calmodulin genes from carrot and Arabidopsis

Abstract

Calmodulin is a highly conserved calcium-binding protein universally distributed among eukaryotes. Although the role of calmodulin in regulatory pathways in animals is well characterized, information on similar pathways in plants is limited. To better understand the role of calmodulin in plant systems, the structure and expression of calmodulin genes from carrot suspension cells and Arabidopsis was studied.A full-length calmodulin cDNA clone was isolated from a $\lambda$gt10 library and this cDNA was used as a probe to measure steady state calmodulin mRNA levels during the growth cycle of carrot cells. During the exponential phase of culture growth, when mitotic activity and oxidative respiration rates of the culture were maximal, calmodulin mRNA levels were four to five-fold higher than they were during the later stages of culture growth. Net calmodulin polypeptide synthesis paralleled the changes in steady state calmodulin mRNA levels during the growth cycle. Calmodulin polypeptide levels, in contrast, remained constant throughout the growth cycle. The data suggest that the calmodulin polypeptide is turned over more rapidly during periods of high mitotic activity and respiration.A 2.3-kb genomic sequence and its corresponding cDNA of 755 bp, encoding Arabidopsis calmodulin were isolated. These sequences represent a third Arabidopsis calmodulin gene (Acam 3), distinct from the two previously isolated cDNAs Acam 1 and Acam 2 (Ling, V., Perera, I., and Zielinski, R. E., 1991. Plant Physiol. 96, 1196-1202). Genomic Southern blots confirmed the presence of a calmodulin multi-gene family in Arabidopsis. The three cDNAs share 83-87% nucleotide sequence identity within their coding regions. Acam 2 and 3 encode identical calmodulin polypeptides. The relative levels of expression of the three genes varied in different plant organs. Both Acam 1 and 3 mRNAs were two to three-fold more abundant than Acam 2 mRNA in Arabidopsis leaves, flowers and developing siliques. Acam 2 and 3 mRNAs were not expressed at detectable levels in Arabidopsis roots. There were detectable differences in the relative levels of transcription and the kinetics of touch induction of Acam 1, 2, and 3, suggesting that the three genes may serve different functions in the plant.U of I OnlyETDs are only available to UIUC Users without author permissio