Department of Physiology and Biophysics, Institute of Biomedical Sciences, University of São Paulo, São Paulo, Brazil.
Thyroid. 2012 Jan;22(1):70-9. doi: 10.1089/thy.2010.0409. Epub 2011 Dec 2.
Glucose transporter 4 (GLUT4) is highly expressed in muscle and fat tissue, where triiodothyronine (T(3)) induces solute carrier family 2 facilitated glucose transporter member 4 (SLC2A4) gene transcription. T(3) was also shown to rapidly increase glucose uptake in myocytes exposed to cycloheximide, indicating that it might act nongenomically to regulate GLUT4 availability. We tested this hypothesis by evaluating, in thyroidectomized rats (Tx rats), the acute and/or chronic T(3) effects on GLUT4 mRNA expression and polyadenylation, protein content, and trafficking to the plasma membrane (PM) in skeletal muscle, as well as on blood glucose disappearance rate (kITT) after insulin administration.
Rats were surgically thyroidectomized and treated with T(3) (0.3 to 100 μg/100 g body weight) from 10 minutes to 5 days, and killed thereafter. Sham-operated (SO) rats were used as controls. Total RNA was extracted from the skeletal muscles (soleus [SOL] and extensorum digitalis longus [EDL]) and subjected to Northern blotting analysis using rat GLUT4 cDNA probe. Total protein was extracted and subjected to specific centrifugations for subcellular fractionation, and PM as well as microsomal (M) fractions were subjected to Western blotting analysis, using anti-GLUT4 antiserum as a probe. GLUT4 mRNA polyadenylation was examined by a rapid amplification of cDNA ends-poly(A) test (RACE-PAT).
Thyroidectomy reduced skeletal muscle GLUT4 mRNA, mRNA poly(A) tail length, protein content, and trafficking to the PM, as well as the kITT. The acute T(3) treatment rapidly (30 minutes) increased all these parameters compared with Tx rats. The 5-day T(3) treatment increased GLUT4 mRNA and protein expression, and restored GLUT4 trafficking to the PM and kITT to SO values.
The results presented here show for the first time that, in parallel to its transcriptional action on the SLC2A4 gene, T(3) exerts a rapid post-transcriptional effect on GLUT4 mRNA polyadenylation, which might increase transcript stability and translation efficiency, leading to the increased GLUT4 content and availability to skeletal muscle, as well as on GLUT4 translocation to the PM, improving the insulin sensitivity, as shown by the kITT.
葡萄糖转运蛋白 4（GLUT4）在肌肉和脂肪组织中高度表达，三碘甲状腺原氨酸（T(3)）诱导溶质载体家族 2 促进葡萄糖转运体成员 4（SLC2A4）基因转录。T(3) 还被证明可以在细胞暴露于环己酰亚胺的情况下迅速增加葡萄糖摄取，表明它可能通过非基因组途径调节 GLUT4 的可用性。我们通过评估甲状腺切除（Tx）大鼠的急性和/或慢性 T(3) 对骨骼肌中 GLUT4 mRNA 表达和多聚腺苷酸化、蛋白含量以及向质膜（PM）的转运以及胰岛素给药后血糖消失率（kITT）的影响来检验这一假设。
大鼠接受手术甲状腺切除术，并接受 10 分钟至 5 天的 T(3)（0.3 至 100 μg/100 g 体重）治疗，然后处死。假手术（SO）大鼠作为对照。从骨骼肌（比目鱼肌 [SOL] 和伸趾长肌 [EDL]）中提取总 RNA，并使用大鼠 GLUT4 cDNA 探针进行 Northern 印迹分析。提取总蛋白并进行特定的离心分离进行亚细胞分级，PM 和微粒体（M）级分用抗 GLUT4 抗血清作为探针进行 Western 印迹分析。GLUT4 mRNA 多聚腺苷酸化通过 cDNA 末端快速扩增-多聚（A）试验（RACE-PAT）进行检查。
甲状腺切除术降低了骨骼肌 GLUT4 mRNA、mRNA 多聚（A）尾长、蛋白含量和向 PM 的转运以及 kITT。与 Tx 大鼠相比，急性 T(3) 处理在 30 分钟内迅速增加了所有这些参数。5 天 T(3) 处理增加了 GLUT4 mRNA 和蛋白表达，并将 GLUT4 向 PM 的转运和 kITT 恢复到 SO 值。
本研究首次表明，除了对 SLC2A4 基因的转录作用外，T(3) 还对 GLUT4 mRNA 多聚腺苷酸化产生快速的转录后作用，这可能增加转录本的稳定性和翻译效率，导致 GLUT4 含量增加和骨骼肌的可用性增加，以及 GLUT4 向 PM 的易位增加，从而提高胰岛素敏感性，如 kITT 所示。
