Overexpression of regucalcin enhances its nuclear localization and suppresses L-type Ca2+ channel and calcium-sensing receptor mRNA expressions in cloned normal rat kidney proximal tubular epithelial NRK52E cells.
The effect of regucalcin (RC), a regulatory protein in intracellular signaling pathway, on the gene expression of various mineral ion transport-related proteins was investigated using the cloned normal rat kidney proximal tubular epithelial NRK52E cells overexpressing RC. NRK52E cells (wild-type) and stable RC/pCXN2 transfectant were cultured for 72 h in medium containing 5% bovine serum (BS) to obtain subconfluent monolayers. After culture for 72 h, cells were further cultured 24-72 h in a medium containing either vehicle, aldosterone (10(-8) or 10(-7) M), or parathyroid hormone (PTH) (1-34) (10(-8) or 10(-7) M) without BS. RC was markedly localized in the nucleus of transfectants. Overexpression of RC caused a significant increase in rat outer medullary K(+) channel (ROMK) mRNA expression, while it caused a remarkable decrease in L-type Ca(2+) channel and calcium-sensing receptor (CaR) mRNA expressions. Overexpression of RC did not have an effect on epithelial sodium channel (ENaC), Na, K-ATPase (alpha-subunit), Type II Na-Pi cotransporter (NaPi-IIa), angiotensinogen, Na(+)-Ca(2+) exchanger, and glyceroaldehyde-3-phosphate dehydrogenase (G3PDH) mRNA expressions. Hormonal effect on gene expression, moreover, was examined. Culture with aldosterone (10(-8) or 10(-7) M) caused a significant increase in ENaC, Na, K-ATPase, and ROMK mRNA expressions in the wild-type cells. Those increases were weakened in the transfectants. Culture with PTH (10(-8) or 10(-7) M) significantly decreased NaPi-IIa mRNA expression in the wild-type cells. This effect was not altered in the transfectants. PTH significantly decreased angiotensinogen mRNA expression in the wild-type cells and the transfectants, while aldosterone had no effect. Culture with PTH (10(-8) or 10(-7) M) caused a significant decrease in L-type Ca(2+) channel and CaR mRNA expressions in the wild-type cells, while the hormone significantly increased Na(+)-Ca(2+) exchanger mRNA expression. The effects of PTH on L-type Ca(2+) channel, CaR, and Na(+)-Ca(2+) exchanger mRNA expressions were also seen in the transfectants. This study demonstrates that overexpression of RC caused a remarkable increase in its nuclear localization, and that it has suppressive effects on the gene expression of L-type Ca(2+) channel or CaR, which regulates intracellular Ca(2+) signaling, among various regulator proteins for mineral ions in NRK52E cells.
Nakagawa
Taeko
T
Laboratory of Endocrinology and Molecular Metabolism, Graduate School of Nutritional Sciences, 52-1 Yada, Suruga-ku, Shizuoka 422-8526, Japan.
Yamaguchi
Masayoshi
M
D000450:Aldosterone / Q000494:pharmacology; D000808:Angiotensinogen / Q000235:genetics; D000818:Animals; D002118:Calcium / Q000378:metabolism; D020746:Calcium Channels, L-Type / Q000235:genetics* / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D002265:Carboxylic Ester Hydrolases; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism*; D002999:Clone Cells / Q000187:drug effects / Q000378:metabolism; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D017136:Ion Transport / Q000187:drug effects; D007687:Kidney Tubules, Proximal / Q000166:cytology*; D010281:Parathyroid Hormone / Q000494:pharmacology; D021381:Protein Transport / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D044169:Receptors, Calcium-Sensing / Q000235:genetics* / Q000378:metabolism; D050613:Sodium-Phosphate Cotransporter Proteins, Type IIa / Q000235:genetics
D020746:Calcium Channels, L-Type; D002135:Calcium-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; D010281:Parathyroid Hormone; D012333:RNA, Messenger; D044169:Receptors, Calcium-Sensing; C493556:Slc34a1 protein, rat; D050613:Sodium-Phosphate Cotransporter Proteins, Type IIa; D000808:Angiotensinogen; D000450:Aldosterone; D002265:Carboxylic Ester Hydrolases; C513325:Rgn protein, rat; D002118:Calcium