Calcipressins are a family of calcineurin binding proteins conserved from fungi to yeast to humans. They have shown to be negative feedback regulators of the calcium/calmodulin phosphatase calcineurin, inihibit cardiac hypertrophy in mammals, and also may play a role in Down Syndrome in humans. We identified
rcn-1, a calcipressin homologue, in C. elegans on chromosome III in cosmid F54E7 and cloned the gene from a cDNA library. GFP expression of promoter regions of
rcn-1 was seen mainly in pharyngeal muscle, excretory cells, vulval epithelial cells, ventral and dorsal nerve cords and commissures, neurons, hypodermal cells and intestine. Whole-mount immunostaining patterns with DS-24 polyclonal antibody showed similar expression patterns. DS-24 antibody was raised against a 24 bp oligonucleotide of the most conserved region of DSCR-1, a human calcipressin. This expression not only confirms our previous GFP expression results, but also shows the conservation of calcipressins from humans to C. elegans . Preliminary data of calcineurin GFP and antibody expression patterns from our laboratory has shown much similarity with
rcn-1 suggesting a relationship between the two proteins. This relationship was further confirmed by GST in vitro binding assay. GST-fused
rcn-1 bound calcineurin A in a calcium-dependent manner suggesting that the activity of calcineurin may be important for the binding of the two proteins. Furthermore, we are interested in testing the effect of
rcn-1 on calcineurin activity by phosphatase assay, and are also currently raising antibodies against
rcn-1 for further protein analysis. Northern blot analysis has confirmed a low-level of expression of a 1.0 kb mRNA transcript. In addition, we are currently studying the effect of calcineurin on the transcriptional expression of
rcn-1 through GFP analysis with calcineurin mutants. We are planning to conduct RNAi of
rcn-1 and will attempt to obtain deletion mutants by UV-TMP mutagenesis to observe loss-of-function phenotypes.