Age related obesity-induced shortening of GLUT4 mRNA poly(A) tail length in rat gastrocnemius skeletal muscle

Obese insulin resistant animals and humans have shown reduced GLUT4 gene expression. Yet, in skeletal muscle, discrepancy between mRNA and protein regulation has been frequently observed, suggesting a post-transcriptional modulation. We investigated the GLUT4 expression in adipose tissue and muscle of obese 12-month-old (12-mo) rats, comparing with lean 2-month-old (2-mo) animals. Obesity was accompanied by insulin resistance, and 65% reduction (P < 0.01) in GLUT4 mRNA and protein in adipose tissue. However, in muscle, despite increased (P < 0.05) mRNA content, GLUT4 protein was unchanged. RNase H and poly(A) test assays showed a reduction (P < 0.01) of ∼80 adenines in the GLUT4 mRNA poly(A) tail of muscle from 12-mo rats, recognizing that the poly(A) tail length correlates with translation efficiency. Concluding, age related obesity of 12-mo rats involves suppression of GLUT4 expression in adipose tissue; however, in muscle, GLUT4 mRNA content increases, but with a shorter poly(A) tail, thus unchanging the protein content. © 2007 Elsevier B.V. All rights reserved.

Servicios públicos y regulación: consecuencias legales de las fallas de mercado

Incluye Bibliografía

Busca de parceiros físicos por duplo-híbrido das proteínas ADAM23 e MX1, codificadas por genes metilados em câncer de cabeça e pescoço

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Servicios públicos y regulación

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Epigenetic and expression analysis of TRAIL-R2 and BCL2: on the TRAIL to knowledge of apoptosis in ovarian tumors

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Worldwide genetic variation at the 3 ' untranslated region of the HLA-G gene: balancing selection influencing genetic diversity

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Influência de diferentes períodos de chuva após a aplicação de Clethodim, Quizalofope - P- Metilico e Haloxifope em pós emergência no controle de plantas daninhas

Pós-graduação em Agronomia (Proteção de Plantas) - FCA

Evolution and genomic organization of muscle microRNAs in fish genomes

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Luteinizing Hormone Receptor (LHR): basic concepts in cattle and other mammals. a review

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Glyceraldehyde 3-phosphate dehydrogenase-telomere association correlates with redox status in trypanosoma cruzi

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Reduction in cytoplasmic lipid content in bovine embryos cultured in vitro with linoleic acid in semi-defined medium is correlated with increases in cryotolerance

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Expressão diferencial de microRNAs no câncer de próstata

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

T-3 Rapidly Increases SLC2A4 Gene Expression and GLUT4 Trafficking to the Plasma Membrane in Skeletal Muscle of Rat and Improves Glucose Homeostasis

Background: 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. Methods: Rats were surgically thyroidectomized and treated with T-3 (0.3 to 100 mu g/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). Results: 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. Conclusions: 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.

Potential Contribution of Translational Factors to Triiodo-L-Thyronine-Induced Insulin Synthesis by Pancreatic Beta Cells

Background: Thyroid hormones (THs) are known to regulate protein synthesis by acting at the transcriptional level and inducing the expression of many genes. However, little is known about their role in protein expression at the post-transcriptional level, even though studies have shown enhancement of protein synthesis associated with mTOR/p70S6K activation after triiodo-l-thyronine (T3) administration. On the other hand, the effects of TH on translation initiation and polypeptidic chain elongation factors, being essential for activating protein synthesis, have been poorly explored. Therefore, considering that preliminary studies from our laboratory have demonstrated an increase in insulin content in INS-1E cells in response to T3 treatment, the aim of the present study was to investigate if proteins of translational nature might be involved in this effect. Methods: INS-1E cells were maintained in the presence or absence of T3 (10(-6) or 10(-8) M) for 12 hours. Thereafter, insulin concentration in the culture medium was determined by radioimmunoassay, and the cells were processed for Western blot detection of insulin, eukaryotic initiation factor 2 (eIF2), p-eIF2, eIF5A, EF1A, eIF4E binding protein (4E-BP), p-4E-BP, p70S6K, and p-p70S6K. Results: It was found that, in parallel with increased insulin generation, T3 induced p70S6K phosphorylation and the expression of the translational factors eIF2, eIF5A, and eukaryotic elongation factor 1 alpha (eEF1A). In contrast, total and phosphorylated 4E-BP, as well as total p70S6K and p-eIF2 content, remained unchanged after T3 treatment. Conclusions: Considering that (i) p70S6K induces S6 phosphorylation of the 40S ribosomal subunit, an essential condition for protein synthesis; (ii) eIF2 is essential for the initiation of messenger RNA translation process; and (iii) eIF5A and eEF1A play a central role in the elongation of the polypeptidic chain during the transcripts decoding, the data presented here lead us to suppose that a part of T3-induced insulin expression in INS-1E cells depends on the protein synthesis activation at the post-transcriptional level, as these proteins of the translational machinery were shown to be regulated by T3.

MicroRNA miR-146b-5p regulates signal transduction of TGF-beta by repressing SMAD4 in thyroid cancer

MicroRNAs (miRNA) are small non-coding RNAs involved in post-transcriptional gene regulation that have crucial roles in several types of tumors, including papillary thyroid carcinoma (PTC). miR-146b-5p is overexpressed in PTCs and is regarded as a relevant diagnostic marker for this type of cancer. A computational search revealed that miR-146b-5p putatively binds to the 3' untranslated region (UTR) of SMAD4, an important member of the transforming growth factor beta (TGF-beta) signaling pathway. The TGF-beta pathway is a negative regulator of thyroid follicular cell growth, and the mechanism by which thyroid cancer cells evade its inhibitory signal remains unclear. We questioned whether the modulation of the TGF-beta pathway by miR-146b-5p can contribute to thyroid tumorigenesis. Luciferase reporter assay confirmed the direct binding of miR-146b-5p on the SMAD4 3'UTR. Specific inhibition of miR-146b-5p with a locked nucleic acid-modified anti-miR-146b oligonucleotide significantly increased SMAD4 levels in the human papillary carcinoma cell lines, TPC-1 and BCPAP. Moreover, suppression of miR-146b-5p increased the cellular response to the TGF-beta anti-proliferative signal, significantly decreasing the proliferation rate. The overexpression of miR-146b-5p in normal rat follicular PCCL3 cells decreased SMAD4 levels and disrupted TGF-beta signal transduction. MiR-146b-5p overexpression in PCCL3 cells also significantly increased cell proliferation in the absence of thyroid-stimulating hormone and conferred resistance to TGF-beta-mediated cell-cycle arrest. Additionally, the activation of thyroid most common oncogenes RET/PTC3 and BRAF in PCCL3 cells upregulated miR-146b-5p expression. Our results confirm the oncogenic role of miR-146b-5p in thyroid follicular cells and contribute to knowledge regarding the modulation of TGF-beta signal transduction by miRNAs in PTCs. Oncogene (2012) 31, 1910-1922; doi:10.1038/onc.2011.381; published online 29 August 2011