Department of Integrated Biosciences, Graduate School of Frontier Sciences, The University of Tokyo, Kashiwa, Japan.
PLoS Genet. 2010 Dec 16;6(12):e1001245. doi: 10.1371/journal.pgen.1001245.
A fundamental step in the evolution of the visual system is the gene duplication of visual opsins and differentiation between the duplicates in absorption spectra and expression pattern in the retina. However, our understanding of the mechanism of expression differentiation is far behind that of spectral tuning of opsins. Zebrafish (Danio rerio) have two red-sensitive cone opsin genes, LWS-1 and LWS-2. These genes are arrayed in a tail-to-head manner, in this order, and are both expressed in the long member of double cones (LDCs) in the retina. Expression of the longer-wave sensitive LWS-1 occurs later in development and is thus confined to the peripheral, especially ventral-nasal region of the adult retina, whereas expression of LWS-2 occurs earlier and is confined to the central region of the adult retina, shifted slightly to the dorsal-temporal region. In this study, we employed a transgenic reporter assay using fluorescent proteins and P1-artificial chromosome (PAC) clones encompassing the two genes and identified a 0.6-kb "LWS-activating region" (LAR) upstream of LWS-1, which regulates expression of both genes. Under the 2.6-kb flanking upstream region containing the LAR, the expression pattern of LWS-1 was recapitulated by the fluorescent reporter. On the other hand, when LAR was directly conjugated to the LWS-2 upstream region, the reporter was expressed in the LDCs but also across the entire outer nuclear layer. Deletion of LAR from the PAC clones drastically lowered the reporter expression of the two genes. These results suggest that LAR regulates both LWS-1 and LWS-2 by enhancing their expression and that interaction of LAR with the promoters is competitive between the two genes in a developmentally restricted manner. Sharing a regulatory region between duplicated genes could be a general way to facilitate the expression differentiation in duplicated visual opsins.
视觉系统进化的一个基本步骤是视觉视蛋白的基因复制，以及在视网膜中的吸收光谱和表达模式上的差异。然而，我们对表达分化机制的理解远远落后于视蛋白光谱调谐的理解。斑马鱼（Danio rerio）有两个红色敏感的视锥蛋白基因，LWS-1 和 LWS-2。这些基因以头尾的方式排列，并且都在视网膜中的长双圆锥体（LDC）中表达。较长波长敏感的 LWS-1 的表达在发育过程中较晚发生，因此局限于成年视网膜的周边，特别是鼻侧区域，而 LWS-2 的表达较早，局限于成年视网膜的中央区域，略微向背颞区域移动。在这项研究中，我们使用荧光蛋白和包含这两个基因的 P1 人工染色体（PAC）克隆进行了转基因报告基因分析，确定了 LWS-1 上游的一个 0.6kb 的“LWS 激活区”（LAR），它调节两个基因的表达。在包含 LAR 的 2.6kb 侧翼上游区域的调控下，荧光报告基因重现了 LWS-1 的表达模式。另一方面，当 LAR 直接与 LWS-2 的上游区域连接时，报告基因在 LDC 中表达，但也在整个外核层中表达。从 PAC 克隆中删除 LAR 大大降低了两个基因的报告基因表达。这些结果表明，LAR 通过增强它们的表达来调节 LWS-1 和 LWS-2，并且 LAR 与启动子的相互作用在发育上受到限制，在两个基因之间是竞争的。在复制的视觉视蛋白之间共享一个调控区域可能是促进表达分化的一种普遍方式。
