Jules Stein Eye Institute, David Geffen School of Medicine at UCLA, Brain Research Institute, University of California, Los Angeles, CA 90095, USA.
Cell Stress Chaperones. 2013 May;18(3):377-87. doi: 10.1007/s12192-012-0386-7. Epub 2012 Dec 23.
Epithelial cells and fibroblasts both express heat shock transcription factors, HSF1 and HSF4, yet they respond to heat shock differentially. For example, while HSP70 is induced in both cell types, the small heat shock protein, αB-crystallin gene (CRYAB) that contains a canonical heat shock promoter, is only induced in fibroblasts. A canonical heat shock promoter contains three or more inverted repeats of the pentanucleotide 5'-nGAAn-3' that make the heat shock element. It is known that, in vitro, promoter architecture (the order and spacing of these repeats) impacts the interaction of various heat shock transcription factors (HSFs) with the heat shock promoter, but in vivo relevance of these binding preferences so far as the expression is concerned is poorly understood. In this report, we first establish cell-type-dependent differential expression of CRYAB in four established cell lines and then working with adult human retinal pigment epithelial cells and NIH3T3 fibroblasts and employing chromatin immunoprecipitation, attempt to relate expression to promoter occupancy by HSF1 and HSF4. We show that HSF4 occupies only CRYAB and not HSP70 promoter in epithelial cells, while HSF1 occupies only HSP70 promoter in both cell types, and cryab promoter, only in heat shocked fibroblasts; HSF4, on the other hand, is never seen on these two promoters in NIH3T3 fibroblasts. This comparative analysis with CRYAB and HSP70 demonstrates that differential heat shock response is controlled by cell-type-dependent access of HSFs (HSF1 and HSF4) to specific promoters, independent of the promoter architecture.
上皮细胞和成纤维细胞都表达热休克转录因子 HSF1 和 HSF4,但它们对热休克的反应不同。例如,虽然 HSP70 在这两种细胞类型中都被诱导,但含有典型热休克启动子的小分子热休克蛋白 αB-晶状体蛋白基因 (CRYAB) 仅在成纤维细胞中被诱导。典型的热休克启动子包含三个或更多的 5'-nGAAn-3'反向重复,构成热休克元件。已知,在体外,启动子结构(这些重复的顺序和间隔)影响各种热休克转录因子(HSFs)与热休克启动子的相互作用,但就表达而言,这些结合偏好的体内相关性知之甚少。在本报告中,我们首先在四种已建立的细胞系中确定 CRYAB 的细胞类型依赖性差异表达,然后与成人视网膜色素上皮细胞和 NIH3T3 成纤维细胞一起工作,并采用染色质免疫沉淀,试图将表达与 HSF1 和 HSF4 对启动子的占据相关联。我们表明,HSF4 仅在上皮细胞中占据 CRYAB 而不占据 HSP70 启动子,而 HSF1 仅在这两种细胞类型中占据 HSP70 启动子,仅在热休克的成纤维细胞中占据 cryab 启动子;另一方面,HSF4 从未在 NIH3T3 成纤维细胞中的这两个启动子上被看到。这种与 CRYAB 和 HSP70 的比较分析表明,差异热休克反应是由 HSFs(HSF1 和 HSF4)对特定启动子的细胞类型依赖性进入控制的,与启动子结构无关。