Expression data from E18 mouse embryonic retina in wild-type mouse and Sox11 knockout mouse

The early retinal progenitor-expressed gene Sox11 regulates the timing of the differentiation of retinal cells. Sry-related HMG box (Sox) proteins play diverse and critical roles in a variety of morphogenetic processes during embryonic development. Sox11 and Sox4 are members of the SoxC subtype, and we found that Sox11 was strongly expressed in early retinal progenitor cells, and that when expression of Sox11 subsided around birth, Sox4 expression began. To analyze the role of Sox11 and Sox4 in retinal development, we perturbed their expression pattern by expressing them ectopically in retinal explant culture. Overexpression of Sox11 or Sox4 in retinal progenitors resulted in similar phenotypes, that is, increased cone cells and decreased Muller glia. Sox11-knockout retinas showed delayed onset and progress of differentiation of early-born retinal cells during the embryonic period. After birth, retinal differentiation took place relatively normally, probably because of the redundant activity of Sox4, which starts to differentiate around birth. Neither overexpression nor loss-of-function analysis gave any evidence that Sox11 and Sox4 directly regulate transcription of genes critical to early-born retinal cells. However, histone H3 acetylation status of the early neurogenic genes was lowered in knockout retinas, suggesting that Sox11 regulates the timing of differentiation in early-born retinas by creating an epigenetic state that helps to establish the competency to differentiate. We also found that the unique expression patterns of Sox11 and Sox4 may be achieved by the Notch signaling pathway and by epigenetic regulation. Taking our findings together, we propose that the precise regulation of Sox11 and Sox4 expression during retinogenesis by multiple mechanisms leads to the fine adjustment of retinal differentiation. To delineate the molecular mechanisms underlying the retinal action of Sox11, we performed microarray analysis of E18 retinas from wild-type and Sox11 knockout mice. Total RNA was obtained from each one retina of Sox11 knockout and wild-type littermate embryos at E18.早期视网膜祖细胞表达的Sox11基因可调控视网膜细胞的分化时序。Sry相关HMG盒（Sry-related HMG Box，Sox）蛋白在胚胎发育阶段的诸多形态发生过程中兼具多样且关键的调控功能。Sox11与Sox4同属于SoxC亚型家族成员，本研究发现Sox11在早期视网膜祖细胞中呈高表达；当Sox11的表达在围产期逐渐减弱时，Sox4的表达随即启动。为解析Sox11与Sox4在视网膜发育中的功能，我们通过在视网膜外植体培养体系中异位表达二者以扰乱其内源表达模式。在视网膜祖细胞中过表达Sox11或Sox4，会引发相似的表型改变：视锥细胞数量增多，而米勒胶质细胞（Muller glia）数量减少。Sox11基因敲除的视网膜在胚胎发育阶段，其早期产生的视网膜细胞的分化起始与进程均出现延迟。出生后，视网膜分化过程则相对正常，这可能得益于Sox4的功能冗余——Sox4的表达恰好于围产期开始激活。无论是过表达还是功能缺失实验，均未发现Sox11与Sox4可直接调控早期产生的视网膜细胞关键基因的转录。但在基因敲除视网膜中，早期神经发生相关基因的组蛋白H3乙酰化水平显著降低，这提示Sox11可通过构建有助于获得分化潜能的表观遗传状态，调控早期产生的视网膜细胞的分化时序。本研究同时发现，Sox11与Sox4的独特表达模式可能通过Notch信号通路（Notch signaling pathway）与表观遗传调控机制实现。综合本研究各项发现，我们提出：在视网膜发生过程中，多种机制对Sox11与Sox4的表达进行精准调控，从而实现视网膜分化过程的精细调节。为阐明Sox11在视网膜中发挥功能的分子机制，我们对野生型（wild-type）与Sox11基因敲除（knockout）小鼠的胚胎第18天（E18）视网膜开展了基因芯片分析（microarray analysis）。本研究从胚胎第18天的Sox11基因敲除与野生型同窝胚胎的单侧视网膜中提取了总RNA。