Cholecalcin (a 9‐kDa cholecalciferol‐induced calcium‐binding protein) messenger RNA

Abstract

In view of the possible physiological importance of the 9-kDa cholecalcin (a 9000-Mr cholecalciferol-induced calcium-binding protein) in the intestinal transport of calcium in mammals, the gene expression of this protein has been analysed. Its regulation in the digestive tract of the growing rat by calcitriol (1,25-dihydroxycholecalciferol) was studied using a specific cloned [32P]cDNA to 9-kDa cholecalcin. Northern hybridisation studies show that the cDNA sequence hybridises to a single 500 -600-nucleotide species throughout the digestive tract and therefore demonstrate identical 9-kDa-cholecalcin mRNA processing in the whole of the intestine and caecum. The highest concentrations of cholecalcin mRNA occur in the duodenum, proximal jejunum and caecum. The observed differences in 9-kDa-cholecalcin mRNA levels correlate well with both the in vivo variations in cholecalcin itself and with the known intestinal sites of calcium absorption. The whole intestine is able to respond to exogenous calcitriol but the response of the distal intestine and caecum, as measured by the increase in cholecalcin mRNA and corresponding protein, was proportionally higher than in the duodenum. The rapid production of fully functional cholecalcin mRNA, which was detectable as early as 1 h after a single dose of calcitriol to vitamin-D-deficient rats, provides convincing evidence that calcitriol increases 9-kDa cholecalcin production by increasing cholecalcin gene expression at the transcriptional level.Cholecalcin, a cholecalciferol-induced calcium-binding protein which was first isolated from the chick intestine by Wasserman and Taylor [l], belongs to the large family of intracellular proteins which bind calcium with high affinity While the chick produces a single 28000-M, protein (28-kDa cholecalcin), the rat possesses both the 28-kDa cholecalcin, found mainly in the kidney and cerebellum, and a smaller, 9000-M, protein (9-kDa cholecalcin) which is characteristic of the duodenum [3]. The rat intestinal 9-kDa cholecalcin is concentrated in the cytoplasm of the absorptive cells of the duodenum [4] where it accounts for 2% of the soluble protein [3]. Although its exact molecular function remains unclear, mammalian 9-kDa cholecalcin is probably involved in the complex process of intestinal calcium absorption [5], which is under the control of (calcitriol) (1,25-dihydroxycholecalciferol), a calciotropic hormone required for the optimal absorption of calcium by the intestine. Its mechanism of action is apparently steroid-hormone-like. Evidence for this includes, in the rat, the localisation of calcitriol within the nuclei of the intestinal absorptive cells [6], the identification of a specific high-affinity receptor protein [7] and the calcitriol-dependent level of intestinal 9-kDa cholecalcin [3, 81. The synthesis of 9-kDa cholecalcin in a cellfree system from rat duodenal poly(A)-rich RNA has recently been described [9] and a complementary DNA has been synthesised and cloned in pBR322 [lo]. The resulting cloned cDNA (pC 109) hybridis...

Estradiol-dependent uterine leiomyomas in transgenic mice.

“…Total RNAs were electrophoresed in 1.5% (wt/vol) agarose-formaldehyde gel and blotted on filters (21). Mouse CaBP9K mRNA was detected with a rat CaBP9K cDNA clone (15).Tag mRNA was detected with the StuI-BamHI fragment containing the coding region of the large T antigen.…”

“…CaBP9K is found in several rat tissues, such as enterocytes, myometrium, and epithelial alveolar cells of the lung (14)(15)(16)(17).The uterine CaBP9K gene is under the positive control of estradiol; the gene is silent in ovariectomized rodents (18,19) and strongly activated after estradiol treatment (20,21). CaBP9K is also expressed in the kidney of the mouse (22,23).…”

Identification of a 1,25-dihydroxyvitamin D3-response element in the 5'-flanking region of the rat calbindin D-9k gene.

“…In the intestine, vitamin D appears to be the primary regulator for the calbindin D-9k gene at the transcriptional level (22)(23)(24)(25). However, no regulation by vitamin D is exerted on the gene in the uterus, where it seems to be primarily under estrogen regulation (7,26).…”

Regulation of metallothionein gene expression by 1 alpha,25-dihydroxyvitamin D3 in cultured cells and in mice.

“…Calcium binding protein was shown to be induced in rat intestine by la,25(0H)2D3 (18). DeLuca and his colleagues (19) reported that four distinct mRNAs were significantly increased in rat intestine when la,25(0H)2D3 was applied to vitamin D-deficient animals: these were vitamin D-dependent calcium-binding protein, mitochondrial ATP synthetase, and cytochrome oxidase subunits I and III.…”