Gierlikowski Wojciech, Grzędzicka Jowita, Konieczek Katarzyna, Kotlarek-Łysakowska Marta
Department of Internal Medicine and Endocrinology, Medical University of Warsaw, Banacha 1a, 02-097 Warsaw, Poland.
Department of Genomic Medicine, Medical University of Warsaw, Nielubowicza 5, 02-097 Warsaw, Poland.
Int J Mol Sci. 2025 Aug 15;26(16):7889. doi: 10.3390/ijms26167889.
SLC5A8 is a protein coded by the 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 expression in PTC is reduced by microRNAs and can be modulated by their inhibition. We used real-time PCR to analyze the expression of 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 transcript. Luciferase assays were performed to confirm the direct binding of synthetic microRNAs to the 3'UTR of . 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 . Finally, we investigated if modulation of SLC5A8 induces changes in transcriptomes. We confirmed the downregulation of in PTC. In silico analysis revealed microRNAs potentially targeting . Luciferase assay confirmed direct binding between the 3'UTR of 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 mRNA, while silencing of miR-181a-5p, miR-182-5p, miR-494-3p, and all three microRNAs simultaneously increased expression; however, only simultaneous inhibition was able to induce changes visible for SLC5A8 protein. Changes in expression did not alter the whole transcriptome significantly. This study shows microRNA-dependent regulation of expression and underlines the potential effectiveness of simultaneous inhibition of a few microRNAs to derepress their common target.
SLC5A8是一种由该基因编码的蛋白质，被认为是一种肿瘤抑制因子和碘转运体。其在甲状腺乳头状癌（PTC）中的表达降低，但其潜在机制仍不清楚。我们推测PTC中该基因的表达受微小RNA调控，且可通过抑制微小RNA来调节。我们使用实时PCR分析了49对PTC/正常组织样本中该基因及相关微小RNA的表达。我们采用计算机分析方法来识别PTC中上调且可能与该基因转录本结合的微小RNA。进行荧光素酶检测以证实合成的微小RNA与该基因3'UTR的直接结合。随后，我们使用表达微小RNA的质粒和微小RNA海绵，包括一种设计用于同时抑制三种选定微小RNA的微小RNA海绵，来检测微小RNA调节对内源性该基因表达的影响。最后，我们研究了SLC5A8的调节是否会引起转录组的变化。我们证实了PTC中该基因表达下调。计算机分析揭示了可能靶向该基因的微小RNA。荧光素酶检测证实了该基因3'UTR与miR - 181a - 5p、miR - 182 - 5p和miR - 494 - 3p之间的直接结合。miR - 181a - 5p和miR - 182 - 5p在PTC中上调。在HEK293细胞系中，转染表达miR - 181a和miR - 182的质粒会降低内源性该基因的mRNA，而沉默miR - 181a - 5p、miR - 182 - 5p、miR - 494 - 3p以及同时沉默所有这三种微小RNA均会增加该基因的表达；然而，只有同时抑制能够使SLC5A8蛋白表达出现可见变化。该基因表达的变化并未显著改变整个转录组。这项研究显示了微小RNA对该基因表达的依赖性调节，并强调了同时抑制几种微小RNA以解除对其共同靶标的抑制作用的潜在有效性。
