Ligand-mediated decrease of thyroid hormone receptor-alpha1 in cardiomyocytes by proteosome-dependent degradation and altered mRNA stability.
Tri-iodo-L-thyronine (T3) is essential for maintaining normal cardiac contractile function by regulating transcription of numerous T3-responsive genes. Both hormone availability and relative amounts of nuclear thyroid hormone receptor isoforms (TRalpha1, TRbeta1) determine T3 effectiveness. Cultured neonatal rat ventricular myocytes grown in T3-depleted medium expressed predominantly TRalpha1 protein, but within 4 h of T3 treatment, TRbeta1 protein increased significantly, whereas TRalpha1 was decreased by 46 +/- 5%. Using replication-defective adenoviruses to overexpress TRalpha1 in cardiomyocytes, we studied the mechanisms by which T3 mediated the decrease in TRalpha1 protein. Inhibitors of the proteosome pathway resulted in an accumulation of ubiquitylated TRalpha1 in the nucleus and prevented T3-induced degradation of ubiquitylated TRalpha1, suggesting that T3 induced proteosome-mediated degradation of TRalpha1; however, TR ubiquitylation was T3 independent. TRalpha1 transcriptional activity, measured using transient transfection of a thyroid hormone-responsive element (TRE) reporter plasmid, was T3 dose dependent and inversely proportional to nuclear TRalpha1 content, with 10 nM T3 having maximum effect. Quantitative RT-PCR showed that both endogenous and adenovirus-expressed TRalpha1 mRNAs were significantly decreased to 54 +/- 11 and 25 +/- 5%, respectively, within 4 h of T3 treatment. Measurements of TRalpha1 mRNA half-life in actinomycin D-treated cardiomyocytes showed that T3 treatment significantly decreased TRalpha1 mRNA half-life from 4 h to less than 2 h, whereas it had no effect of TRbeta1 mRNA half-life. These data support a role for both the proteosome degradation pathway and altered mRNA stability in T3-induced decrease of nuclear TRalpha1 in the cardiomyocyte and provide novel cellular targets for therapeutic development.
Kenessey
Agnes
A
North Shore-Long Island Jewish Research Institute, Manhasset, New York 11030, USA. kojamaa@nshs.edu
Ojamaa
Kaie
K
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D000831:Animals, Newborn; D002478:Cells, Cultured; D015870:Gene Expression / Q000187:drug effects / Q000235:genetics; D008024:Ligands; D032383:Myocytes, Cardiac / Q000166:cytology / Q000502:physiology*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D020871:RNA Stability / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D037021:Thyroid Hormone Receptors alpha / Q000235:genetics* / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects / Q000502:physiology; D014284:Triiodothyronine / Q000494:pharmacology
D008024:Ligands; D012333:RNA, Messenger; D037021:Thyroid Hormone Receptors alpha; D014284:Triiodothyronine; D046988:Proteasome Endopeptidase Complex