MicroRNAs and Their Inhibition in Modulating SLC5A8 Expression in the Context of Papillary Thyroid Carcinoma

SLC5A8 is a protein coded by the <i>SLC5A8</i> gene, and has been proposed as a tumor suppressor and iodide transporter. Its expression is reduced in papillary thyroid carcinoma (PTC), yet the mechanisms underlying this phenomenon are largely unknown. We hypothesized that <i>SLC5A8</i> expression in PTC is reduced by microRNAs and can be modulated by their inhibition. We used real-time PCR to analyze the expression of <i>SLC5A8</i> and the microRNAs of interest in a set of 49 PTC/normal tissue pairs. We used an in silico approach to identify microRNAs upregulated in PTC and putatively binding to the <i>SLC5A8</i> transcript. Luciferase assays were performed to confirm the direct binding of synthetic microRNAs to the 3'UTR of <i>SLC5A8</i>. Subsequently, using mir-expressing plasmids and microRNA sponges, including a microRNA sponge designed to simultaneously inhibit three selected microRNAs, we checked the impact of the modulation of microRNAs on endogenous <i>SLC5A8</i>. Finally, we investigated if modulation of SLC5A8 induces changes in transcriptomes. We confirmed the downregulation of <i>SLC5A8</i> in PTC. In silico analysis revealed microRNAs potentially targeting <i>SLC5A8</i>. Luciferase assay confirmed direct binding between the 3'UTR of <i>SLC5A8</i> and miR-181a-5p, miR-182-5p, and miR-494-3p. MiR-181a-5p and miR-182-5p were upregulated in PTC. In HEK293 cell lines, transfection with mir-181a- and mir-182-expressing plasmids decreased endogenous <i>SLC5A8</i> mRNA, while silencing of miR-181a-5p, miR-182-5p, miR-494-3p, and all three microRNAs simultaneously increased <i>SLC5A8</i> expression; however, only simultaneous inhibition was able to induce changes visible for SLC5A8 protein. Changes in <i>SLC5A8</i> expression did not alter the whole transcriptome significantly. This study shows microRNA-dependent regulation of <i>SLC5A8</i> expression and underlines the potential effectiveness of simultaneous inhibition of a few microRNAs to derepress their common target.