Crenshaw E B, Kalla K, Simmons D M, Swanson L W, Rosenfeld M G
Department of Biology, School of Medicine, University of California, San Diego, La Jolla 92093.
Genes Dev. 1989 Jul;3(7):959-72. doi: 10.1101/gad.3.7.959.
Prolactin gene expression is restricted to the lactotrophic and somatomammotrophic cells of the anterior pituitary. In transgenic mice, a fusion gene consisting of 3 kb of prolactin 5'-flanking region fused to a firefly luciferase or human growth hormone (hGH) reporter gene is expressed at high levels with the strict tissue and cell-type specificity that is characteristic of the endogenous prolactin gene. High levels of expression require two cis-acting regions: a distal enhancer (-1.8 to -1.5 kb) and a proximal region (-422 to +33 bp). Each of these regions alone can direct low levels of fusion gene expression to prolactin-producing cell types in transgenic mice, but a synergistic interaction between these regions is necessary for high levels of expression. The ontogeny of the prolactin transgene expression closely follows the appearance of high levels of a POU homeo-domain transcription factor, Pit-1, that has been shown previously to bind structurally related sequences in both the distal enhancer and proximal regions and to activate the expression of the prolactin gene in vitro. Unexpectedly, transgenes containing the distal enhancer removed from its normal context are expressed in both the prolactin-producing lactotrophs and the thyroid-stimulating hormone (TSH)-producing thyrotrophs, thereby suggesting that sequences flanking this enhancer are necessary to restrict expression to the correct cell type within the pituitary. These data indicate that distinct processes of gene activation and restriction are necessary for the fidelity of cell-type-specific expression within an organ.
催乳素基因表达仅限于垂体前叶的催乳素分泌细胞和促生长催乳素细胞。在转基因小鼠中,由3 kb催乳素5′侧翼区与萤火虫荧光素酶或人生长激素(hGH)报告基因融合而成的融合基因,以与内源性催乳素基因相同的严格组织和细胞类型特异性高水平表达。高水平表达需要两个顺式作用区域:一个远端增强子(-1.8至-1.5 kb)和一个近端区域(-422至+33 bp)。单独的每个区域都能将融合基因低水平表达导向转基因小鼠中产生催乳素的细胞类型,但这些区域之间的协同相互作用对于高水平表达是必需的。催乳素转基因表达的个体发生密切跟随一种POU同源结构域转录因子Pit-1高水平的出现,此前已证明该转录因子能在远端增强子和近端区域结合结构相关序列,并在体外激活催乳素基因的表达。出乎意料的是,去除其正常背景的含远端增强子的转基因在产生催乳素的催乳素分泌细胞和产生促甲状腺激素(TSH)的促甲状腺细胞中均有表达,从而表明该增强子侧翼序列对于将表达限制在垂体内正确的细胞类型是必需的。这些数据表明,基因激活和限制的不同过程对于器官内细胞类型特异性表达的保真度是必要的。