Despite the central role of chromosomal context in gene transcription, human noncoding DNA variants are generally studied outside of their endogenous genomic location. This poses major limitations to understand the true consequences of causal regulatory variants. We focused on a cis-regulatory mutation (c.-331C>G) in the INS gene promoter that is recurrently mutated in unrelated patients with recessive neonatal diabetes. We created mice in which a ~3.1 kb human INS upstream region carrying -331C or -331G alleles replaced the orthologous mouse Ins2 region. This human sequence drove cell-specific transcription in mice. It also recapitulated poised chromatin during pancreas development and active chromatin in differentiated β-cells. The c.-331C>G mutation, however, blocked active chromatin formation in differentiated β-cells. We further show that another neonatal diabetes gene product, GLIS3, had a singular pioneer-like ability to activate INS chromatin in non-pancreatic cells, which was hampered by the c.-331C>G mutation. This in vivo analysis of human regulatory defects, therefore, uncovered cis and trans components of a mechanism that is essential to activate the endogenous INS gene.

中文翻译：

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新生儿糖尿病突变破坏了激活人类胰岛素基因的染色质启动功能

尽管染色体背景在基因转录中起着中心作用，但人类非编码DNA变体通常在其内源基因组位置之外进行研究。这对理解因果调节变体的真实后果构成了主要限制。我们着重研究了INS基因启动子中的顺式调控突变（c.-331C> G），该突变在不相关的隐性新生儿糖尿病患者中反复突变。我们创建了约3.1 kb人INS的小鼠携带-331C或-331G等位基因的上游区域替换了直系同源小鼠Ins2区。该人类序列驱动小鼠中的细胞特异性转录。它也概括了胰腺发育过程中的平衡染色质和分化的β细胞中的活性染色质。然而，c.-331C> G突变阻止了分化的β细胞中活跃的染色质形成。我们进一步显示，另一种新生儿糖尿病基因产品GLIS3具有非胰腺细胞激活INS染色质的独特的先锋样能力，这受到c.-331C> G突变的阻碍。因此，对人体调节缺陷的体内分析发现了激活内源INS基因必不可少的机制的顺式和反式成分。