Specific localization of scallop gill epithelial calmodulin in cilia

Abstract

Calmodulin has been isolated and characterized from the gill of the bay scallop Aequipecten irradians. Quantitative electrophoretic analysis of epithelial cell fractions shows most of the calmodulin to be localized in the cilia, specifically in the detergent-solubilized membrane-matrix fraction . Calmodulin represents 2.2 ± 0.3% of the membrane-matrix protein or 0.41 ± 0.05% of the total ciliary protein. Its concentration is at least 10 -4 M if distributed uniformly within the matrix . Extraction in the presence of calcium suggests that the calmodulin is not bound to the axoneme proper . The ciliary protein is identified as a calmodulin on the basis of its calcium-dependent binding to a fIuphenazine-Sepharose affinity column and its comigration with bovine brain calmodulin on alkaline-urea and SDS polyacrylamide gels in both the presence and absence of calcium . Scallop ciliary calmodulin activates bovine brain phosphodiesterase to the same extent as bovine brain and chicken gizzard calmodulins. Containing trimethyllysine and lacking cysteine and tryptophan, the amino acid composition of gill calmodulin is typical of known calmodulins, except that it is relatively high in serine and low in methionine . Its composition is less acidic than other calmodulins, in agreement with an observed isoelectric point~0 .2 units higher than that of bovine brain. Comparative tryptic peptide mapping of scallop gill ciliary and bovine brain calmodulins indicates coincidence of over 75% of the major peptides, but at least two major peptides in each show no nearequivalency. Preliminary results using ATP-reactivated gill cell models show no effect of calcium at micromolar levels on ciliary beat or directionality of the lateral cilia, the cilia which constitute the vast majority of those isolated . However, ciliary arrest will occur at calcium levels >150 uM . Because calmodulin usually functions in the micromolar range, its role in this system is unclear. Scallop gill ciliary calmodulin may be involved in the direct regulation of dyneintubule sliding, or it may serve some coupled calcium transport function . At the concentration in which it is found, it must also at least act as a calcium buffer .There is substantial evidence for the existence of calmodulin in cilia and flagella . However, its specific localization and function in these organelles is uncertain. Satir and co-workers (20; see also reference 17) find Tetrahymena calmodulin to be concentrated considerably more in cilia than in the cell body, using immunofluorescent techniques, whereas Walter and Schultz (27) find a greater concentration of Paramecium calmodulin in cell bodies than in cilia, using SDS-polyacrylamide gel analysis . Gitelman and Witman (10) report that calmodulin from Chlamydomonas occurs in both the cell body and in isolated flagella, with the latter appearing to contain a much greater concentration of calmodulin . About half of the flagellar calmodulin remains tightly bound to the axoneme after deter-622 E. W. STOMMEL, R. E. STEPHENS, H . R. MAS...

Molecular characteristics and expression of calmodulin cDNA from the freshwater pearl mussel, Hyriopsis schlegelii

“…In bivalves, the gill is the primary organ for calcium uptake from the water (Elijah, 1984;Rousseau et al, 2003). Particularly in the scallop, the gill is thought to play a regulatory role in the membrane Ca 2+ -ATPase system or to act as a "calcium sink" (Stommel et al, 1982). The high expression level of hsCaM mRNA in the gill supports the theory that CaM plays an important regulatory role in calcium uptake and accumulation in the gill of H. schlegelii.…”

Drosophilapericentrin requires interaction with calmodulin for its function at centrosomes and neuronal basal bodies but not at sperm basal bodies

“…In addition, CaM is a well-known axonemal component (Gitelman and Witman, 1980; Stommel et al ., 1982) and is important for ciliary function (AbouAlaiwi et al ., 2009; DiPetrillo and Smith, 2010; Plotnikova et al ., 2012). For these reasons, we investigated the importance of the PLP-CaM interaction for cilia function.…”

A calcium regenerative potential controlling ciliary reversal is propagated along the length of ctenophore comb plates.

“…The length constant for a cilium of "infinite"length has been calculated to be as high as one to several millimeters (4, 31), indicating that the short ,um) cilia of protozoa are essentially isopotential along their length and with the cell body (7,32 (12)(13)(14)(15). Fluorescence and immunoelectron microscopy have shown that calmodulin is localized along the entire length of the axoneme (16,17 Therefore, our finding that Ca2+ influx occurs over most of the length of comb-plate cilia implies that the Ca2+-sensitive machinery controlling beat direction is also distributed along the length of the axoneme.…”

“…Calmodulin, a putative ciliary Ca2+ sensor (12)(13)(14)(15), has been detected along the entire length of cilia (16,17). In contrast, local application of Ca2+ to reactivated cilia and flagella indicate that the Ca2+-sensitive machinery controlling various motor responses is localized to a specific region of the axoneme (18)(19)(20).…”