AUTOCRINE ACTIONS OF THYROID HORMONE (T3) REGULATE EXPRESSION OF GENES INVOLVED ON POST-TRANSCRIPTIONAL AND POST-TRANSLACIONAL EVENTS IN THYROID FOLLICULAR CELLS ..

Thyroid hormone plays an important role in developmental, growth and metabolic processes. It acts through specific nuclear receptors bound to thyroid response elements on target genes activating/repressing their transcription or through receptors on the plasma membrane triggering intracellular signaling pathways, by non-genomic mechanism. Thyroid dysfunctions are the second major cause of metabolic disorders around the world. So, many studies have been carried out intending to clarify all aspects of thyroid hormone actions on different tissues. However, little is known about the role of TH on the gland in which it is produced. In this sense, the aim of this study was to evaluate, through New Generation Sequencing methodology (RNAseq) the gene expression profile of thyroid follicular cells in the presence or absence of TH. Pccl3 cells were maintained in TH-free medium (HYPO) for 24h. Cells were divided into two groups: one remained in the HYPO medium and the other was treated with 3,3',5 triiodothyronine (T3) at concentration of 10-7M for 24h. Afterwards, cells were lysed for total RNA extraction and submitted to ribosomal RNA depletion assay (RibominusTM). cDNA libraries were prepared using rRNA-free samples and then sequenced by SOLiD® commercial sequencing platform. The raw data obtained in the sequencing were statistically analyzed to build the list of genes differentially expressed, of which 85% were upregulated, while 15% were down-regulated by T3. Among genes that were upregulated two were selected for validation: gene Pfdn1 (LogFoldChange = 1.68, p = 9.2E-05) which is involved in protein folding and gene Fam103a1 (LogFoldChange = 1.94, p = 1.26E-05) which is responsible for adding CAP (7Rmethylguanosine) into mRNA during transcription. After validation of the sequencing data by RT-qPCR, we also evaluated the expression of both genes under T3 treatment at 10-9 M for 1h. In 10-7M of T3 the Pfdn1 mRNA expression increased 2 and 8-fold (vs. HIPO) after 1 and 24h, respectively. In the 10-9 M dose, it was observed an increase only after 24h. The Fam103a1 mRNA expression increased approximately 6-fold (vs. HYPO) only at the 10-7 M; no effect was observed at the 10-9 M. Data suggest that T3 plays an important role on the mRNAs mcappingc, which is essential for their stabilization and translation rate, as well on the protein folding, which point out to important actions played by TH on post-transcriptional and pos-translational control of gene expression.