Interrogating the Grainyhead-like 2 (Grhl2) genomic locus identifies an enhancer element that regulates palatogenesis in mouse.
Interrogating the Grainyhead-like 2 (Grhl2) genomic locus identifies an enhancer element that regulates palatogenesis in mouse.
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DOI：
10.1016/j.ydbio.2019.11.015
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发表时间：
2020-03-15
影响因子：
2.7
通讯作者：
Dworkin S
中科院分区：
文献类型：
--
作者：
de Vries M;Carpinelli M;Rutland E;Hatzipantelis A;Partridge D;Auden A;Anderson PJ;De Groef B;Wu H;Osterwalder M;Visel A;Jane SM;Dworkin S
高度保守的颗粒头样转录因子(GRHL)是胚胎发生的关键调节因子，调节细胞的存活、增殖、迁移和上皮完整性，特别是在头面部骨骼的形成过程中。Grhl2家族成员在发育过程中表达于上皮组织，Grhl2功能的丧失会导致神经形成、腹壁关闭、面部形成和上颌/腭部融合等方面的严重缺陷。虽然已经确定了许多Grhl2下游的靶基因，但对这种关键的发育转录因子本身是如何调控的知之甚少。在这里，利用对小鼠的硅胶和宫内表达分析和功能缺失，我们已经鉴定出一种新的2.4kb增强子元件(Mm1286)，它以一种强烈概括颅面部原基内源性Grhl2的模式驱动报告基因的表达，调节这些组织中Grhl2的表达，并增强Grhl2介导的次级腭部的关闭。在Ghhl2的一个等位基因失活的情况下(通过产生双重杂合的Ghl2+/−；Mm1286+/−小鼠)，该基因组元件的缺失导致出生时极易发生腭裂。此外，我们发现MM1286的一个高度保守的325bp区域对于调节该区域的头面部特异性增强子活性是必要和充分的，并且该元件中非常保守的12bp序列(CTGTCAAACAGGT)基本上决定了完整的增强子功能。总之，这些数据为上游基因组调控环境提供了有价值的新见解，这些基因组调控环境负责在腭闭合过程中对Grhl2的转录控制。头部和面部的骨骼、神经、血管、肌肉和皮肤的正确形成和整合是一个异常复杂的过程。多个发育基因的协同作用对于正确的颅面发育是必不可少的。当这些严格控制的过程出错时--就像他们经常做的那样--由此产生的头面部缺陷会给受影响的个人带来严重的心理、社会和功能困难。通过了解这些发育过程是如何在基因水平上调节的，我们可以对如何解决这些衰弱的疾病形成更全面的理解。在这里，我们描述了一个已知对头部、面部和颌骨发育至关重要的基因(Grainyhead-like 2；Grhl2)是如何在转录水平上受到一个新的颅面部特异性增强子区域的调控。这种增强子与Grhl2合作，在发育中的面部和颌骨内正确驱动表达。重要的是，这种增强子的功能丧失是小鼠腭裂病因学中的一个重要的预处理因素，考虑到脊椎动物之间的大量序列保守，最终也可能被证明是人类颅面部疾病的一个因素。
The highly-conserved Grainyhead-like (Grhl) transcription factors are critical regulators of embryogenesis that regulate cellular survival, proliferation, migration and epithelial integrity, especially during the formation of the craniofacial skeleton. Family member Grhl2 is expressed throughout epithelial tissues during development, and loss of Grhl2 function leads to significant defects in neurulation, abdominal wall closure, formation of the face and fusion of the maxilla/palate. Whereas numerous downstream target genes of Grhl2 have been identified, very little is known about how this crucial developmental transcription factor itself is regulated. Here, using in silico and in utero expression analyses and functional deletion in mice, we have identified a novel 2.4kb enhancer element (mm1286) that drives reporter gene expression in a pattern that strongly recapitulates endogenous Grhl2 in the craniofacial primordia, modulates Grhl2 expression in these tissues, and augments Grhl2-mediated closure of the secondary palate. Deletion of this genomic element, in the context of inactivation of one allele of Grhl2 (through generation of double heterozygous Grhl2+/−;mm1286+/− mice), results in a significant predisposition to palatal clefting at birth. Moreover, we found that a highly conserved 325 bp region of mm1286 is both necessary and sufficient for mediating the craniofacial-specific enhancer activity of this region, and that an extremely well-conserved 12-bp sequence within this element (CTGTCAAACAGGT) substantially determines full enhancer function. Together, these data provide valuable new insights into the upstream genomic regulatory landscape responsible for transcriptional control of Grhl2 during palatal closure. The correct formation and integration of the bones, nerves, blood vessels, muscles and skin of the head and face is an exceptionally complex process. The synergy of multiple developmental genes is essential for correct craniofacial development. When these tightly-regulated processes go wrong – as they often do – the resultant craniofacial defects lead to substantial psychological, social and functional difficulties for affected individuals. By understanding how these developmental processes are regulated at the genetic level, we can form a more holistic understanding of how to tackle these debilitating disorders. Here we describe how a gene known to be critical for development of the head, face and jaws (Grainyhead-like 2; Grhl2) is regulated at the transcriptional level by a new craniofacial-specific enhancer region. This enhancer co-operates with Grhl2 to drive expression correctly within the developing face and jaws. Importantly, functional loss of this enhancer is a significant pre-disposing factor in the aetiology of palatal clefts in mice, and given the substantial sequence conservation across vertebrates, may also ultimately prove to be a factor in human craniofacial disorders.
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