Analysis of the gene network patterning the hindhtain in vertebrates.

1.在斑马鱼胚胎后脑形成过程中，hoxb 1b在神经板中表达，其前边界位于r3-r4边界，提供了位置信息。为了了解hoxb 1b转录的调控机制，我用GFP作为报告基因进行了基因转移启动子分析。结果，在上游DNA（-1.8和-4.6kb）中鉴定出在晚期（体节发生）驱动GFP表达的两个调控区，在下游DNA（+1.5和+4.7）中鉴定出在早期（原肠胚）驱动GFP表达的两个调控区。它也变得很清楚，下游两个区域介导的反应，视黄酸是已知的图案后脑在脊椎动物胚胎。2.gbx2和fgf 8都表达在原始前后脑的早期神经板和图案的中脑-后脑边界区（MHB）。我对hoxb 1b进行了启动子分析。对于gbx 2，-6.9 kb的上游区域显示足以在前脑和间脑中转录，而fgf 8的+10.6和+15.1 kb的两个下游区域可以驱动报告基因在前脑中以及耳泡和视柄中表达。在fgf 8的侧翼区域中也已经鉴定了另外的调控区域，其驱动了fgf 8在胚胎中表达的许多胚胎区域中的表达。3.为了鉴定无任何偏差地支配脑形成的调控基因，作为先导实验，筛选ENU处理的雄鱼的后代鱼的胚胎脑中的异常，导致建立了几个显示异常表型的突变株系，例如在中脑、背侧间脑和轴形成中。

1.During the formation of hindbrain in zebrafish embryos, hoxb1b is expressed in the neural plate with its anterior boundary at the r3-r4 border, providing the positional information. In order to know the regulatory mechanism of hoxb1b transcription, I conducted the promoter analysis using gene transfer with GFP as a reporter. As a result, two regulatory regions were identified in the upstream DNA (-1.8 and -4.6 kb) which drove the GFP expression at late stages (somitogenesis) and two regulatory regions in the downstream DNA (+1.5 and +4.7) that drove GFP at earlier stages (gastrula). It also became clear that the downstream two regions mediate the responsiveness to retinoic acid that is known to pattern the hindbrain in vertebrate embryos.2.gbx2 and fgf8 are both expressed in the primordial anterior hindbrain of the early neural plate and pattern the midbrain-hindbrain boundary region (MHB). I conducted the promoter analyses as for hoxb1b. For gbx2, the upstream region at -6.9 kb was shown to be sufficient for transcription in the anterior hindbrain and diencephalon, while two downstream regions of fgf8 at +10.6 and +15.1 kb could drive reporter expression in the anterior hindbrain in addition to the otic vesicle and optic stalk. Additional regulatory regions have also been identified in the flanking region of fgf8 that drove expression in a number of embryonic regions where fgf8 is expressed in embryos.3.In order to identify regulatory genes that govern the brain formation without any bias, progeny fish of ENU-treated male fish were screened for anomalies in the embryonic brain as a pilot experiment, leading to establishment of several mutant lines showing abnormal phenotypes such as in the midbrain, dorsal diencephalon, and axis formation.