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中文摘要：
为探索人高亲和力钠离子依赖性二羧酸转运蛋白基因（high affinity sodium-dependent dicarboxylate transporter,SDCT2）3′端非翻译区是否在基因表达调控中起作用,首先通过生物信息学分析发现,在SDCT2βmRNA的3′端非翻译区内存在585nt的AU富含区（AU-rich region,AUR）,其中包括3个AU富含元件（AU-rich element,ARE）,然后将SDCT2β的AU富含区DNA片段插入报告基因GFP表达载体pcDNA-GFP的下游,构建pcDNA-GFP-AUR表达载体并转染HEK293、HKC和LLC-PK1细胞系,用Western blot和流式细胞仪检测细胞中GFP的表达水平.结果显示,SDCT2β的AU富含区序列可显著降低GFP的表达水平（P〈0.01）.利用放线菌素D阻断RNA转录后,每隔2h从稳定转染的HEK293细胞中提取总RNA,用RNA印迹分析GFP mRNA的稳定性.结果显示GFP-AURmRNA较GFP mRNA不稳定.这些结果提示,在SDCT2β3′非翻译区的AU富含区内存在基因表达负调控区,该区可降低mRNA的稳定性、促进mRNA的降解,从而在转录后水平调控基因的表达.
英文摘要：
To explore whether the 3′-untranslated region （3′-UTR） of human high-affinity sodium-dependent dicarboxylate transporter （hSDCT2） plays a role in the regulation of gene expression, sequence characteristics of the 3′-UTR was analyzed using bioinformatics. The results found that within the 3′-UTR of SDCT2β mRNA there is an AU-rich region （AUR） of 585 nt, which contains three AU-rich elements （ARE）. The AUR fragment was inserted into the 3′-UTR of GFP reporter gene within a expression vector pcDNA-GFP; and a pcDNA-GFP-AUR recombinant vector was constructed and transfected into HEK293, HKC and LLC-PK1 cell lines. The expression level of intracellular GFP was determined by Western blot and flow cytometry. The results showed that the AUR of SDCT2β could significantly reduce the expression level of GFP in the pcDNA-GFP-AUR- transfected cells （P 〈 0.01）. After blockade of RNA transcription with actinomycin D, total RNA was extracted from stably transfected HEK293 cells with an interval of 2 h, and the stability of GFP-AUR and GFP mRNAs was analyzed by Northern blot. The results indicated that the stability of GFP-AUR mRNA is less than that of GFP mRNA. These results suggest that within the AU-rich region of 3′-UTR of SDCT2β mRNA there is a negative regulation region which can decrease the stability of mRNA, accelerate the degradation of GFP mRNA and may play a negative regulation role for gene expression at post-transcriptional level.
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