REGULATION BY CALCITONIN AND GLUCOCORTICOIDS OF CALCITONIN RECEPTOR GENE-EXPRESSION IN MOUSE OSTEOCLASTS

We previously studied regulation of the calcitonin (CT) receptor (CTR) by glucocorticoid (GC) and CT in cultures of mature mouse osteoclast- like cells (OCLs). The present studies were designed to examine the in teraction of CT and GC in regulation of the CTR in osteoclasts and the molecular mechanisms involved. Treatment of OCLs with 10(-7) M dexame thasone (Dex) increased the CTR number in a time-dependent manner, whe reas treatment with 10(-9) M salmon CT (sCT) reduced CTR number; neith er treatment changed receptor affinity. Dex pretreatment somewhat anta gonized the CT-induced reduction in [I-125]sCT specific binding. Dex i ncreased, and sCT pretreatment decreased, the sCT-responsive adenylate cyclase activity in parallel with the change in receptor binding. Dex treatment resulted in an increase in CTR messenger RNA (mRNA) levels, as assessed by reverse transcription-PCR, indicating that the increas ed CTR number was mediated by de novo CTR synthesis. This effect was s pecific to GCs and was not reproduced by mineralocorticoids or sex ste roids. Treatment with sCT resulted in a rapid and profound reduction i n CTR mRNA expression, and this reductions was somewhat delayed by Dex pretreatment. OCLs were treated with 5,6-dichloro-1 beta-D-ribofurano syl benzimidazole to enable estimation of the mRNA decay rates in the absence of ongoing transcription. The stability of CTR mRNA was simila r to the control value in Dex-treated OCLs, suggesting that the effect of Dex may be due to changes in transcriptional activity. Interesting ly, transcriptional inhibition by 5,6-dichloro-1 beta-D-ribofuranosyl benzimidazole abolished the ability of CT to reduce CTR mRNA levels, s uggesting that CT may act by increasing the rate of CTR mRNA decay, an d that this effect requires ongoing transcription. The 3'-untranslated region of the mouse CTR mRNA contains four copies of the AUUUA motif, as well as other A/U-rich sequences, which have been shown to determi ne the stability of other mRNA transcripts. The stability results were consistent with the results of the nuclear transcript run-on assay, w hich indicated that treatment with Dex enhanced the rate of transcript ion, whereas CT had no effect. These results Show that GC and CT influ ence CTR expression by distinct mechanisms and provide the basis for i dentification of the cellular factors involved.