Role of zebrafish POU-domain gene, ZFPOU2, in early development

pou结构域蛋白是一个转录调控蛋白大家庭，其中许多与早期发育过程中基因表达的控制有关。我们在这里描述了斑马鱼pou2的克隆和表达，一个新的与小鼠生殖系特异性转录因子Oct-3相关的pou2结构域基因。斑马鱼pou2在母体中表达，从单细胞期到原肠期都存在转录本。原位杂交分析表明，该转录本在囊胚中期之前的所有卵裂球中都存在，而在原肠胚形成期间，该转录本仅在外胚层中表达。我们发现选择性剪接转录物t-pou2 RNA也在胚胎中表达。与Pou2产物相比，t-Pou2产物由于其不完整的Pou2结构域结构而缺乏dna结合活性。为了研究Pou2和t-Pou2产物的作用，我们在胚胎中通过向受精卵中微量注射合成的Pou2和t-Pou2来增加它们在胚胎中的表达。大多数由注射了pou2 RNA的卵子发育的胚胎没有表现出任何明显的发育缺陷。相反，过表达t-Pou2产物对胚胎发育的影响很大：由于原肠胚发育不完全，出现强烈的发育迟缓或发育停滞。在受影响的胚胎中，斑马鱼T基因表达减少，下胚形成受到干扰。这些产物的时空表达模式和过表达对发育的影响与Pou2和t-Pou2蛋白参与斑马鱼胚胎早期发育的观点一致。它们可能参与囊胚期未确定状态的卵裂球增殖和/或原胚期的早期细胞承诺事件。此外，我们的研究结果表明，由于同一基因的选择性剪接而产生的不同产物具有不同的功能能力。少

POU-domain proteins are a large family of transcriptional regulatory proteins, many of which are implicated in the control of gene expression during early development. We describe here the cloning and expression of zebrafish pou2, a novel POU-domain gene related to the mouse germ line-specific transcription factor Oct-3. Zebrafich pou2 is maternally expressed and the transcripts are present from the one-cell stage to the gastruls stage. In situ hybridization analyzes revealed that the transcripts were present in all blastomeres until the midblastula stage and that the expression was restricted to the epiblast during gastrulation. We found that alternatively spliced transcripts, t-pou2 RNA,were also expressed in the embryos. In contrast to the Pou2 product, t-Pou2 product lacks DNA-binding activity because of its incomplete POU-domain structure. To examine the roles of the Pou2 and t-Pou2 products, we increased their expression in the embryo by microinjection of synthetic pou2 and t-pou … More 2 RNAs into the fertilized eggs at the one-cell stage. Most embryos that developed from the eggs injected with pou2 RNA did not show any obvious developmental defects. In contrast, overexpression of the t-Pou2 product greatly affected the embryonic development : there was strong developmental retardation or arrest due to the incomplete gastrulation. In the affected embryos, expression of zebrafish T gene was reduced and the hypoblast formation was disturbed. Temporal and spatial expression patterns and the effects of overxpression of these products on development are consistent with the idea that the Pou2 and t-Pou2 proteins are involved in early development of zebrafish embryos. They may be involved in the proliferation of blastomeres in undetermined state at the blastula stage and/or the early cell commitment events at the gastrula stage. Also, our results indicate that different products generated as a result of alternative splicing from the same gene possess distinct functional capacities. Less