Kenessey Agnes, Sullivan Elizabeth Ann, Ojamaa Kaie
Institute for Medical Research, North Shore-LIJ Health System, 350 Community Dr., Manhasset, NY 11030, USA.
Am J Physiol Heart Circ Physiol. 2006 Jan;290(1):H381-9. doi: 10.1152/ajpheart.00576.2005. Epub 2005 Sep 9.
Maladaptive cardiac hypertrophy results in phenotypic changes in several genes that are thyroid hormone responsive, suggesting that thyroid hormone receptor (TR) function may be altered by cellular kinases, including protein kinase C (PKC) isozymes that are activated in pathological hypertrophy. To investigate the role of PKC signaling in regulating TR function, cultured neonatal rat ventricular myocytes were transduced with adenovirus (Ad) expressing wild-type (wt) or kinase-inactive (dn) PKC alpha or constitutively active (ca) PKC delta and PKC epsilon. Overexpression of wtPKC alpha, but not caPKC delta or caPKC epsilon, induced a 28-fold increase (P < 0.001) in TR alpha1 protein in the nuclear compartment and a smaller increase in the cytosol. Furthermore, TR alpha1 mRNA was increased 55-fold (P < 0.001). This effect of PKC alpha was dependent on its kinase activity because dnPKC alpha was without effect. Phorbol 12-myristate 13-acetate (PMA) induced nuclear translocation of endogenous PKC alpha and Ad-wtPKC alpha concomitantly with an increase in nuclear TR alpha1 protein. In contrast, PMA-induced nuclear translocation of dnPKC alpha resulted in a decrease of TR alpha1. The increase in TR alpha1 protein in Ad-wtPKC alpha-transduced cardiomyocytes was not the result of a reduced rate of protein degradation, nor was the half-life of TR alpha1 mRNA prolonged, suggesting a PKC alpha-mediated effect on TR alpha transcription. Although phosphorylation of ERK1/2 was increased in Ad-wtPKC alpha-transduced cells, inhibition of phospho-ERK did not change TR alpha1 expression. PKC alpha overexpression in cardiomyocytes caused marked repression of triiodothyronine (T3)-responsive genes, alpha-myosin heavy chain, and the sarcoplasmic reticulum calcium-activated adenosinetriphosphatase SERCA2. Treatment with T3 for 4 h resulted in significant reductions of PKC alpha in nuclear and cytosolic compartments, and decreased TR alpha1 mRNA and protein, with normalization of phenotype. These results implicate PKC alpha as a regulator of TR function and suggest that nuclear localization of PKC alpha may control transcription of the TR alpha gene, and consequently, affect cardiac phenotype.
适应性不良的心脏肥大导致多个甲状腺激素反应性基因发生表型变化，这表明甲状腺激素受体（TR）的功能可能会被细胞激酶改变，包括在病理性肥大中被激活的蛋白激酶C（PKC）同工酶。为了研究PKC信号通路在调节TR功能中的作用，用表达野生型（wt）或激酶失活型（dn）PKCα或组成型激活型（ca）PKCδ和PKCε的腺病毒（Ad）转导培养的新生大鼠心室肌细胞。wtPKCα的过表达而非caPKCδ或caPKCε的过表达导致核内TRα1蛋白增加28倍（P<0.001），胞质中增加较少。此外，TRα1 mRNA增加了55倍（P<0.001）。PKCα的这种作用依赖于其激酶活性，因为dnPKCα没有作用。佛波酯12-肉豆蔻酸酯13-乙酸酯（PMA）诱导内源性PKCα和Ad-wtPKCα核转位，同时核内TRα1蛋白增加。相反，PMA诱导的dnPKCα核转位导致TRα1减少。Ad-wtPKCα转导的心肌细胞中TRα1蛋白的增加不是蛋白质降解速率降低的结果，TRα1 mRNA的半衰期也没有延长，提示PKCα对TRα转录有介导作用。虽然Ad-wtPKCα转导的细胞中ERK1/2的磷酸化增加，但抑制磷酸化的ERK并没有改变TRα1的表达。心肌细胞中PKCα过表达导致三碘甲状腺原氨酸（T3）反应性基因、α-肌球蛋白重链和肌浆网钙激活腺苷三磷酸酶SERCA2明显受到抑制。用T3处理4小时导致核和胞质部分的PKCα显著减少，TRα1 mRNA和蛋白减少，表型恢复正常。这些结果表明PKCα是TR功能的调节因子，并提示PKCα的核定位可能控制TRα基因的转录，从而影响心脏表型。
