Laboratorio de Biomedicina Molecular I, Escuela Nacional de Medicina y Homeopatía, Instituto Politécnico Nacional, Mexico City, Mexico.
Departamento de Infectómica y Patogénesis Molecular, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional, Mexico City, Mexico.
Front Cell Infect Microbiol. 2017 Nov 29;7:492. doi: 10.3389/fcimb.2017.00492. eCollection 2017.
Transcriptional regulation of the multidrug resistance gene in is induced by emetine stress. overexpression alters the chloride-dependent currents that cause trophozoite swelling, diminishing induced programmed cell death (PCD) susceptibility. In contrast, antisense inhibition of P-glycoprotein (PGP) expression produces synchronous death of trophozoites and the enhancement of the biochemical and morphological characteristics of PCD induced by G418. Transcriptional gene regulation analysis identified a 59 bp region at position -170 to -111 bp promoter as putative emetine response elements (EREs). However, insights into transcription factors controlling gene transcription are missing; to fill this knowledge gap, we used deletion studies and transient CAT activity assays. Our findings suggested an activating motif (-151 to -136 bp) that corresponds to a heat shock element (HSE). Gel-shift assays, UV-crosslinking, binding protein purification, and western blotting assays revealed proteins of 94, 66, 62, and 51 kDa binding to the HSE that could be heat shock-like transcription factors that regulate the transcriptional activation of the gene in the presence of emetine drug.
依托咪酯应激诱导 基因的转录调控。过表达改变了导致滋养体肿胀的氯离子依赖性电流，降低了诱导程序性细胞死亡（PCD）的易感性。相比之下，反义抑制 P-糖蛋白（PGP）的表达会导致滋养体同步死亡，并增强 G418 诱导的 PCD 的生化和形态特征。转录基因调控分析确定了一个位于 -170 到 -111bp 启动子的 59bp 区域为依托咪酯反应元件（EREs）。然而，控制 基因转录的转录因子的见解仍然缺失；为了填补这一知识空白，我们使用了缺失研究和瞬时 CAT 活性测定。我们的研究结果表明存在一个激活基序（-151 到 -136bp），它对应于热休克元件（HSE）。凝胶迁移分析、UV 交联、结合蛋白纯化和 Western 印迹分析显示，与 HSE 结合的蛋白质为 94、66、62 和 51kDa，这些可能是热休克样转录因子，在依托咪酯药物存在的情况下调节 基因的转录激活。
