The Protein Phosphatase Calcineurin Determines Basal Parathyroid Hormone Gene Expression

Abstract

Calcium and phosphate regulate PTH mRNA stability through differences in binding of parathyroid (PT) proteins to a minimal 63-nucleotide (nt) cis-acting instability element in its 3'-untranslated region. One of these proteins is adenosine-uridine-rich binding factor (AUF1), whose levels are not regulated in PT extracts from rats fed the different diets. However, two-dimensional gels showed posttranslational modification of AUF1 that included phosphorylation. There is no PT cell line, but in HEK 293 cells the 63-nt element is recognized as an instability element, and RNA interference for AUF1 decreased human PTH secretion in cotransfection experiments. Stably transfected cells with a chimeric GH gene containing the PTH 63-nt cis-acting element were used to study the signal transduction pathway that regulates AUF1 modification and chimeric gene mRNA stability. Cyclosporine A, the calcineurin inhibitor, regulated AUF1 posttranslationally, and this correlated with an increase in the stability of GH-PTH 63-nt mRNA but not of the control GH mRNA. Mice with genetic deletion of the calcineurin Abeta gene had markedly increased PTH mRNA levels that were still regulated by low calcium and phosphorus diets. Therefore, calcineurin regulates AUF1 posttranslationally in vitro and PTH gene expression in vivo but still allows its physiological regulation by calcium and phosphate.

Increased Parathyroid Hormone Gene Expression in Secondary Hyperparathyroidism of Experimental Uremia Is Reversed by Calcimimetics

“…In the adenine high-phosphate model, the increased creatinine levels already at 3 days may have contributed to the differences in protein-RNA interactions and AUF1 modifications shown in our study. Similar changes in protein-RNA binding and posttranslational modifications of AUF1 were shown by dietary-induced hypocalcemia with attendant hyperphosphatemia in rats with normal kidney function (14). Moreover, recombinant AUF1 stabilized a PTH transcript in an in vitro degradation assay with parathyroid cytosolic proteins (13).…”

“…S-100 protein extracts from PT glands were run on one-(1-D) and two-dimension (2-D) SDS gels, and Western blots were performed with a mAb for AUF1 that was provided by G. Dreyfuss (University of Pennsylvania, Philadelphia) as described previously (14). The mAb for ␣-tubulin was purchased from Sigma (St. Louis, MO).…”

Destabilization of parathyroid hormone mRNA by extracellular Ca2+ and the calcimimetic R-568 in parathyroid cells: role of cytosolic Ca and requirement for gene transcription

“…Formation of the Ca:CaM complex activates CaM-dependent protein kinases and the protein phosphatase calcineurin. Calcineurin has been implicated in the regulation of basal levels of PTH gene expression (Bell et al 2005b). PTH mRNA levels were increased, though still regulated by low-Ca and phosphorus diets, in mice with genetic deletion of the calcineurin Ab gene.…”

mRNA stability control: a clandestine force in normal and malignant hematopoiesis

“…113 A decrease in AUF1 phosphorylation by cyclosporine A has been observed, suggesting that calcineurin can modulate AUF1 activity through phosphorylation. 127 AUF1 has also been shown to be phosphorylated on tyrosines by the NPM-ALK oncoprotein, as discussed below. The physiologic significance of basal and regulated phosphorylation of AUF1 has yet to be fully explored.…”

“…Transcript stability is controlled by kinases and other mediators characteristic of cytokine signal transduction pathways, including PKC, [146][147][148][149][150] PKB, 151 MAPK/Erk, 45 PI-3-kinase, 128,152 Src-family kinases, 153 p38 MAPK (reviewed in Dean et al 154 and Kotlyarov and Gaestel 155 ), mixed lineage kinase, 130 JNK kinases, 156,157 and phosphatases such as calcineurin 127 and PP2A. 158 Several of these signaling intermediaries, particularly p38 and ERK have been shown to be important in stabilizing inflammatory transcripts downstream of LPS or IL-1.…”