Pancreas-specific primary regulatory targets of Nkx2.2

Nkx2.2 胰腺特异性主要调控靶标

• 批准号: 6781793
• 负责人: DERVLA M MELLERICK-DRESSLER
• 金额: $ 15.3万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-08-01 至 2006-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6781793
• 关键词: Drosophilidae; animal tissue; arthropod genetics; cell differentiation; developmental genetics; functional /structural genomics; gene expression; genetic models; genetically modified animals; high throughput technology; homeobox genes; in situ hybridization; messenger RNA; microarray technology; pancreas; pancreatic islets; polymerase chain reaction; transcription factor

DESCRIPTION (provided by applicant): Homeobox genes of the Nkx2.2/vnd family are required for pancreatic specification and ventral neural tube patterning. This gene family plays conserved roles in lineage specification in divergent animal species, including flies and man. Although invertebrates lack a pancreas, endocrine cells in the nervous and digestive systems secrete insulin to co-ordinate cellular metabolism and nutritional conditions. Seven insulin-like genes have recently been described in Drosophila. The pancreatic and neural lineages are ancestrally related, and share common regulatory factors. In mice, the insulin enhancer drives expression in the ventral neural tube and the pancreas, while fly neurons produce insulin. Moreover, ES cells, exposed to factors that generate neurons, develop into both motor neurons and islets. Nkx2.2/Vnd-type transcription factors are critical for aspects of pancreatic and neural specification, yet how this family functions is not understood, primarily because target genes remain largely unidentified. We previously described, ind, a dorsal-ventral patterning gene that is directly repressed by Drosophila Vnd. This is to date the only well-characterized target of the Nkx2.2/Vnd family. The simplicity, yet sophistication of the fruit fly, Drosophila, as a genetic model has provided novel insights into aspects of development ranging from insulin responsiveness to aging. Here we propose identifying targets of the Nkx2.2/vnd gene family using microarray analyses on mRNAs isolated from Drosophila embryos that transiently over-express vnd at times when only primary (direct) target genes should be activated. To assess the feasibility of this pilot project, initially we will focus upon those candidate vnd target genes that have previously been studied by other labs. Quantitative RT-PCR and in situ hybridization on RNAs from wild-type, vnd minus, and vnd over-expression mutant embryos will further establish whether the candidate vnd target genes are real targets. To address whether vertebrate homologues of the target fly genes we identify are critical for pancreatic development, expression of conserved vertebrate homologues will be assessed in the developing pancreas of wild-type and NKx2.2 mutants. This pilot study is designed to exploit the versatile Drosophila model to gain pioneering insights into NKx2.2 target genes. This knowledge may be highly instructive in defining those regulatory networks required for beta islet specification.

描述（由申请方提供）：Nkx2.2/vnd家族的同源框基因是胰腺特化和腹侧神经管模式形成所需的。这个基因家族在不同的动物物种（包括苍蝇和人类）的谱系特化中起着保守的作用。虽然无脊椎动物没有胰腺，但神经和消化系统中的内分泌细胞分泌胰岛素来协调细胞代谢和营养条件。最近在果蝇中描述了七种胰岛素样基因。胰腺和神经谱系是祖先相关的，并且共享共同的调节因子。在小鼠中，胰岛素增强子驱动腹侧神经管和胰腺中的表达，而苍蝇神经元产生胰岛素。此外，ES细胞暴露于产生神经元的因子，发育成运动神经元和胰岛。Nkx2.2/Vnd型转录因子对于胰腺和神经特化方面至关重要，但该家族如何发挥功能尚不清楚，主要是因为靶基因在很大程度上仍未被识别。我们以前描述过，ind，一个背腹图案的基因，是直接抑制果蝇VND。这是迄今为止Nkx2.2/Vnd家族中唯一一个得到充分表征的靶标。果蝇作为遗传模型的简单性和复杂性为从胰岛素反应到衰老的发育方面提供了新的见解。在这里，我们提出了确定目标的Nkx2.2/vnd基因家族使用微阵列分析的mRNA从果蝇胚胎，瞬时过度表达vnd的时候，只有主要的（直接）靶基因应该被激活。为了评估这个试点项目的可行性，我们首先将重点放在那些候选vnd靶基因，以前已被其他实验室研究。对来自野生型、vnd减和vnd过表达突变体胚胎的RNA进行定量RT-PCR和原位杂交将进一步确定候选vnd靶基因是否是真实的靶。为了解决我们鉴定的靶蝇基因的脊椎动物同源物是否对胰腺发育至关重要，将在野生型和NKx2.2突变体的发育胰腺中评估保守脊椎动物同源物的表达。这项初步研究旨在利用多功能果蝇模型来获得对NKx2.2靶基因的开创性见解。这方面的知识可能是非常有益的定义β胰岛规格所需的那些监管网络。