Gonçalves Lisa, Filipe Mário, Marques Sara, Salgueiro Ana-Marisa, Becker Jorg D, Belo José António
Regenerative Medicine Program, Departamento de Ciências Biomédicas e Medicina, Universidade do Algarve, Campus de Gambelas, Faro, Portugal.
Int J Dev Biol. 2011;55(3):281-95. doi: 10.1387/ijdb.103273lg.
During early vertebrate development, the correct establishment of the body axes is critical. The anterior pole of the mouse embryo is established when Distal Visceral Endoderm (DVE) cells migrate to form the Anterior Visceral Endoderm (AVE). Symmetrical expression of Lefty1, Cer1 and Dkk1 determines the direction of DVE migration and the future anterior side. In addition to the establishment of the Anterior-Posterior axis, the AVE has also been implicated in anterior neural specification. To better understand the role of the AVE in these processes, we have performed a differential screening using Affymetrix GeneChip technology with AVE cells isolated from cer1P-EGFP transgenic mouse embryos. We found 175 genes which were upregulated in the AVE and 36 genes in the Proximal-posterior sample. Using DAVID software, we characterized the AVE cell population regarding cellular component, molecular function and biological processes. Among the genes that were found to be upregulated in the AVE, several novel genes were identified. Four of these transcripts displaying high-fold change in the AVE were further characterized by in situ hybridization in early stages of development in order to validate the screening. From those four selected genes, one, denominated Adtk1, was chosen to be functionally characterized by targeted inactivation in ES cells. Adtk1 encodes for a serine/threonine kinase. Adtk1 null mutants are smaller and present short limbs due to decreased mineralization, suggesting a potential role in chondrogenesis during limb development. Taken together, these data point to the importance of reporting novel genes present in the AVE.
在脊椎动物早期发育过程中，身体轴线的正确确立至关重要。当远端内脏内胚层（DVE）细胞迁移形成前内脏内胚层（AVE）时，小鼠胚胎的前极得以确立。Lefty1、Cer1和Dkk1的对称表达决定了DVE迁移的方向以及未来的前侧。除了前后轴的确立，AVE还与前神经特化有关。为了更好地理解AVE在这些过程中的作用，我们使用Affymetrix基因芯片技术，对从cer1P-EGFP转基因小鼠胚胎中分离出的AVE细胞进行了差异筛选。我们发现175个基因在AVE中上调，36个基因在近端后样本中上调。使用DAVID软件，我们从细胞成分、分子功能和生物学过程方面对AVE细胞群体进行了表征。在发现的在AVE中上调的基因中，鉴定出了几个新基因。其中四个在AVE中显示出高倍数变化的转录本，通过在发育早期阶段的原位杂交进一步表征，以验证筛选结果。从这四个选定的基因中，选择了一个名为Adtk1的基因，通过在胚胎干细胞中的靶向失活来进行功能表征。Adtk1编码一种丝氨酸/苏氨酸激酶。Adtk1基因敲除突变体较小，由于矿化减少而出现四肢短小，这表明其在肢体发育过程中的软骨形成中可能发挥作用。综上所述，这些数据表明报告AVE中存在的新基因的重要性。
