Sumoy L, Wang C K, Lichtler A C, Pierro L J, Kosher R A, Upholt W B
Department of BioStructure and Function, School of Dental Medicine, University of Connecticut Health Center, Farmington 06030, USA.
Dev Biol. 1995 Jul;170(1):230-42. doi: 10.1006/dbio.1995.1210.
Msx-2 is a member of the Msx family of homeobox-containing genes expressed in a variety of embryonic tissues involved in epithelial-mesenchymal interactions and pattern formation. In the developing chick limb bud, Msx-2 is expressed in the apical ectodermal ridge, which plays a crucial role in directing the growth and patterning of limb mesoderm. In addition, Msx-2 is expressed in the anterior nonskeletal-forming mesoderm of the limb bud, in the posterior necrotic zone, and in the interdigital mesenchyme. Studies of the altered expression patterns of Msx-2 in amelic and polydactylous mutant chick limbs have suggested that the apical ectodermal ridge and mesodermal domains of Msx-2 expression are independently regulated and that there might be separate cis-regulatory elements in the Msx-2 gene controlling its spatially distinct domains of expression. To test this hypothesis, we have isolated the chicken Msx-2 gene and have tested the ability of various regions of the gene to target expression of LacZ reporter gene to specific regions of the limbs of transgenic mice. A variety of these constructs are consistently expressed only in the apical ectodermal ridge and the ectoderm of the genital tubercle and are not expressed in the mesoderm of the limb bud or in other regions of the embryo where the endogenous Msx-2 gene is expressed. These results suggest the presence of spatially specific cis-regulatory elements in the Msx-2 gene. We identified a 348-bp region in the 5' flanking region of the Msx-2 gene which can act as an apical ectodermal ridge enhancer element when placed in reverse orientation in front of the reporter gene with transcription initiation directed by the minimal hsp68 promoter.
Msx - 2是含同源盒基因的Msx家族成员，在参与上皮 - 间充质相互作用和模式形成的多种胚胎组织中表达。在发育中的鸡胚肢芽中，Msx - 2在顶端外胚层嵴中表达，顶端外胚层嵴在引导肢中胚层的生长和模式形成中起关键作用。此外，Msx - 2在肢芽的前部非骨骼形成中胚层、后部坏死区和指间间充质中表达。对无肢和多指突变鸡胚肢中Msx - 2表达模式改变的研究表明，Msx - 2表达的顶端外胚层嵴和中胚层结构域是独立调控的，并且Msx - 2基因中可能存在单独的顺式调控元件来控制其在空间上不同的表达结构域。为了验证这一假设，我们分离了鸡Msx - 2基因，并测试了该基因不同区域将LacZ报告基因靶向表达至转基因小鼠肢体特定区域的能力。多种这些构建体仅在顶端外胚层嵴和生殖结节的外胚层中持续表达，而不在肢芽的中胚层或胚胎中内源性Msx - 2基因表达的其他区域中表达。这些结果表明Msx - 2基因中存在空间特异性顺式调控元件。我们在Msx - 2基因的5'侧翼区域鉴定出一个348 bp的区域，当以反向方向置于由最小hsp68启动子指导转录起始的报告基因之前时，该区域可作为顶端外胚层嵴增强子元件。
