Regulation of avian calbindin-D_28K gene expression in primary chick kidney cells: Importance of posttranscriptional mechanisms and calcium ion concentration

Vitamin D-dependent calcium-binding protein (calbindin-D_28K), is regulated by 1,25-dihydroxyvitamin D₃ [1,25-(OH)₂D₃], and several other factors in a tissue-specific manner, but the controlling mechanisms are still poorly understood. In this study we examined the relative contributions of transcriptional and posttranscriptional mechanisms in the 1,25-(OH)₂D₃ control of calbindin-D_28K mRNA expression in primary chick kidney cells and investigated the effect of extracellular Ca²⁺ on calbindin-D_28K gene expression in the presence and absence of hormone. 1,25-(OH)₂D₃ treatment (10^-8 M) of cells grown in serum-free medium resulted in a marked 20- to 30-fold increase in calbindin-D_28K mRNA peaking at 12-18 h, which then rapidly declined to basal levels by 24 h. The abrupt decline in mRNA appeared to be associated with a reduction in size of the calbindin-D_28K transcripts. Nuclear run-off assays showed only a slight (1.5-fold) increase in calbindin-_D28K gene transcription 2 h after 1,25-(OH)₂D₃, whereas parallel assays clearly demonstrated a marked (7-fold) induction in the rate of metallothionein gene transcription 2 h after treatment of chick kidney cells with 10 μM zinc. The induction of calbindin-D mRNA by 1,25-(OH)₂D₃ required ongoing protein synthesis, since it was blocked by cycloheximide. Calbindin-D_28K mRNA was stable for 12 h in the presence of actinomycin-D in both vitamin D-deficient and 1,25-(OH)₂D₃-treated cells. Both basal and 1,25-(OH)₂D₃-induced calbindin-D_2sK mRNA were modulated by the extracellular Ca²⁺, with maximum expression occurring at 1-2 mM. We conclude that 1,25-(OH)₂D₃ induces kidney calbindin-D_28K mRNA by producing a small increase in its transcriptional rate, which is accompanied by pronounced posttranscriptional effects(s). The striking modulation of calbindin-D_28K expression by extracellular Ca²⁺ is consistent with a putative role for this protein in the regulation of this ion in the kidney cell.