Regulation of Parathyroid Hormone mRNA Stability by Calcium and Phosphate

Calcium and phosphate regulate parathyroid hormone (PTH) secretion, gene expression and if prolonged also parathyroid proliferation. The regulation of PTH gene expression by Ca2+ and Pi is post-transcriptional, affecting mRNA stability. The regulation of PTH mRNA stability is mediated by the binding of protective trans acting factors to a cis acting instability element in the 3’UTR. In hypocalcemia there is increased binding that protects the PTH mRNA from degradation by cytosolic ribonudeases resulting in higher levels of PTH mRNA. In hypophosphatemia there is decreased binding, with a subsequent increased degradation of the PTH mRNA resulting in low levels of PTH mRNA. We have identified an AU rich protein binding sequence in the PTH mRNA 3’-UTR and determined its functional-ity. A 63 nt PTH mRNA 3’ UTR element was inserted into a growth hormone (GH) reporter transcript and led to destabilization of the GH transcript. Moreover, this element was sufficient to reproduce the regulation of the reporter transcript’s stability by calcium and phosphate in an in vitro degradation assay (IVDA). The PTH mRNA 63 nt element destabilized mRNA for the reporter genes GFP and growth hormone, in transient transfection experiments in HEK293 cells, similar to its effect in the IVDA. Therefore, the PTH RNA-protein binding region is a destabilizing element and is sufficient to confer responsiveness to calcium and phosphate of PTH mRNA. We have identified one of the PTH mRNA 3’-UTR trans acting proteins as AU rich binding factor 1 (AUF1). Recombinant AUF1 stabilized the PTH mRNA transcript in the IVDA with parathyroid proteins. In parathyroid extracts from rats fed the different diets there was no difference in AUF1 protein levels by Western blots despite differences in RNA -protein binding. However, 2D gels showed post-translational modification of AUF1 in these extracts, suggesting that calcium and phosphate alter the AUF1 protein and thus its ability to bind and stabilize the PTH mRNA.