General transcription factor IIB (GTF2B) was predicted by several bioinformatic tools to regulate AIP expression transcriptionally. Regulation by GTF2B was evaluated using chromatin immunoprecipitation (ChIP), reverse transcription PCR, luciferase reporter, and western blot experiments in SH-SY5Y cells. Furthermore, the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay, transwell invasive assay, ELISA, western blot, immunohistochemical staining, and terminal deoxynucleotidyl transferase dUTP nick end labeling were performed to investigate the effects of GTF2B and AIP on tumor cell proliferation, apoptosis, growth hormone secretion, and invasiveness in GH3 cells and mouse xenograft models. Moreover, correlations between GTF2B and AIP expression were explored in GHPA cases.
ChIP and luciferase reporter studies demonstrated that the regulation of AIP expression by GTF2B was dependent on the intergenic-5\' untranslated region element of AIP and the initial residual S65 of GTF2B. In vitro and in vivo experiments indicated that GTF2B regulated AIP expression to impact the GHPA phenotype; this was confirmed by data from 33 GHPA cases.
We determined the regulation by GTF2B of AIP transcription in GHPA and its impact on tumor phenotype. Our findings suggest that GTF2B may be a potential therapeutic target for GHPA with low AIP expression.
通过几种生物信息学工具预测了通用转录因子IIB(GTF2B)在转录上调节AIP表达。使用染色质免疫沉淀(ChIP)评估GTF2B的调节,逆转录PCR,荧光素酶报告基因,和SH-SY5Y细胞的Western印迹实验。此外,3-(4,5-二甲基噻唑-2-基)-2,5-二苯基-2H-四唑溴化物(MTT)测定,Transwell侵入性分析,ELISA,westernblot,免疫组织化学染色,和末端脱氧核苷酸转移酶dUTP缺口末端标记,以研究GTF2B和AIP对肿瘤细胞增殖的影响,凋亡,生长激素分泌,以及在GH3细胞和小鼠异种移植模型中的侵袭性。此外,在GHPA病例中探讨了GTF2B和AIP表达之间的相关性。
ChIP和荧光素酶报告基因研究表明,GTF2B对AIP表达的调节取决于AIP的基因间5'非翻译区元件和GTF2B的初始残留S65。体外和体内实验表明GTF2B调节AIP表达以影响GHPA表型;来自33个GHPA病例的数据证实了这一点。
我们确定了GTF2B对GHPA中AIP转录的调节及其对肿瘤表型的影响。我们的发现表明GTF2B可能是低AIP表达的GHPA的潜在治疗靶标。