Xu P X, Zhang X, Heaney S, Yoon A, Michelson A M, Maas R L
Division of Genetics, Department of Medicine and Howard Hughes Medical Institute, Brigham and Women's Hospital and Harvard Medical School, Boston MA 02115, USA.
Development. 1999 Jan;126(2):383-95. doi: 10.1242/dev.126.2.383.
Pax6 plays a key role in visual system development throughout the metazoa and the function of Pax6 is evolutionarily conserved. However, the regulation of Pax6 expression during eye development is largely unknown. We have identified two physically distinct promoters in mouse Pax6, P0 and P1, that direct differential Pax6 expression in the developing eye. P0-initiated transcripts predominate in lens placode and corneal and conjunctival epithelia, whereas P1-initiated transcripts are expressed in lens placode, optic vesicle and CNS, and only weakly in corneal and conjunctival epithelia. To further investigate their tissue-specific expression, a series of constructs for each promoter were examined in transgenic mice. We identified three different regulatory regions which direct distinct domains of Pax6 expression in the eye. A regulatory element upstream of the Pax6 P0 promoter is required for expression in a subpopulation of retinal progenitors and in the developing pancreas, while a second regulatory element upstream of the Pax6 P1 promoter is sufficient to direct expression in a subset of post-mitotic, non-terminally differentiated photoreceptors. A third element in Pax6 intron 4, when combined with either the P0 or P1 promoter, accurately directs expression in amacrine cells, ciliary body and iris. These results indicate that the complex expression pattern of Pax6 is differentially regulated by two promoters acting in combination with multiple cis-acting elements. We have also tested whether the regulatory mechanisms that direct Pax6 ocular expression are conserved between mice and flies. Remarkably, when inserted upstream of either the mouse Pax6 P1 or P0 promoter, an eye-enhancer region of the Drosophila eyeless gene, a Pax6 homolog, directs eye- and CNS-specific expression in transgenic mice that accurately reproduces features of endogenous Pax6 expression. These results suggest that in addition to conservation of Pax6 function, the upstream regulation of Pax6 has also been conserved during evolution.
在整个后生动物中，Pax6在视觉系统发育中起关键作用，且Pax6的功能在进化上是保守的。然而，眼睛发育过程中Pax6表达的调控在很大程度上尚不清楚。我们在小鼠Pax6中鉴定出两个物理上不同的启动子P0和P1，它们在发育中的眼睛中指导Pax6的差异表达。P0启动的转录本在晶状体基板以及角膜和结膜上皮中占主导，而P1启动的转录本在晶状体基板、视泡和中枢神经系统中表达，在角膜和结膜上皮中表达较弱。为了进一步研究它们的组织特异性表达，在转基因小鼠中检测了每个启动子的一系列构建体。我们鉴定出三个不同的调控区域，它们在眼睛中指导Pax6表达的不同结构域。Pax6 P0启动子上游的一个调控元件是视网膜祖细胞亚群和发育中的胰腺中表达所必需的，而Pax6 P1启动子上游的第二个调控元件足以指导有丝分裂后、未终末分化的光感受器亚群中的表达。Pax6内含子4中的第三个元件与P0或P1启动子结合时，可准确指导无长突细胞、睫状体和虹膜中的表达。这些结果表明，Pax6的复杂表达模式受两个启动子与多个顺式作用元件联合作用的差异调控。我们还测试了指导Pax6眼部表达的调控机制在小鼠和果蝇之间是否保守。值得注意的是，当果蝇无眼基因（一种Pax6同源物）的眼睛增强子区域插入小鼠Pax6 P1或P0启动子上游时，它在转基因小鼠中指导眼睛和中枢神经系统特异性表达，准确再现了内源性Pax6表达的特征。这些结果表明，除了Pax6功能的保守性外，Pax6的上游调控在进化过程中也得到了保守。
