Abstract
In human embryonic lung fibroblasts, transforming growth factor-beta 1 (TGF-beta 1) induced a time-dependent down-regulation of M2 muscarinic receptor binding sites as measured with the nonselective hydrophilic ligand [3H]N-methylscopolamine (NMS). This down-regulation was slow, with 58% loss of all receptors after 24 hr of treatment. The affinity of [3H]NMS for the remaining sites was unaltered by TGF-beta 1. The loss in [3H]NMS binding was accompanied by reduced adenylyl cyclase activity and functional desensitization of M2 muscarinic receptors. Northern blot analyses showed a 72% decrease in the steady state levels of m2 muscarinic receptor mRNA after 24-hr TGF-beta 1 treatment. Recovery of m2 muscarinic receptor mRNA after TGF-beta 1 treatment was slow, with a half-life of approximately 8 hr. There was no effect of TGF-beta 1 on the m2 muscarinic receptor mRNA half-life measured in the presence of actinomycin D, but the rate of m2 muscarinic receptor gene transcription measured with nuclear run-on assay was reduced by 50%, indicating reduced gene transcription. Cycloheximide (10 micrograms/ml) pretreatment abolished the TGF-beta 1 effect, indicating that de novo protein synthesis was required for receptor downregulation. In summary, we have shown that TGF-beta 1 induced desensitization and down-regulation of M2 muscarinic receptor protein and gene that was mediated through reduction in the rate of m2 receptor gene transcription.
MolPharm articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|