Bompada Pradeep, Atac David, Luan Cheng, Andersson Robin, Omella Judit Domènech, Laakso Emilia Ottosson, Wright Jason, Groop Leif, De Marinis Yang
Diabetes and Endocrinology, Department of Clinical Sciences, Lund University, Malmö, SE-20502, Sweden.
Lund University Diabetes Centre, Department of Clinical Sciences, Lund University, Malmö, SE-20502, Sweden.
Int J Biochem Cell Biol. 2016 Dec;81(Pt A):82-91. doi: 10.1016/j.biocel.2016.10.022. Epub 2016 Oct 29.
Thioredoxin-interacting protein (TXNIP) has been shown to be associated with glucose-induced deterioration of pancreatic beta cell function in diabetes. However, whether epigenetic mechanisms contribute to the regulation of TXNIP gene expression by glucose is not clear. Here we studied how glucose exerts its effect on TXNIP gene expression via modulation of histone acetylation marks. To achieve this, we applied clustered regularly interspaced short palindromic repeats/Cas9 (CRISPR/Cas9) to knock out histone acetyltransferase (HAT) p300 in a rat pancreatic beta cell line INS1 832/13. We also treated the cells and human islets with chemical inhibitors of HAT p300 and histone deacetylase (HDAC). In human islets, diabetes and high glucose resulted in elevated TXNIP and EP300 expression, and glucose-induced TXNIP expression could be reversed by p300 inhibitor C646. In INS1 832/13 cells, Ep300 knock-out by CRISPR/Cas9 elevated glucose-induced insulin secretion and greatly reduced glucose-stimulated Txnip expression and cell apoptosis. This effect could be ascribed to decrease in histone marks H3K9ac and H4ac at the promoter and first coding region of the Txnip gene. Histone marks H3K9ac and H4ac in the Txnip gene in the wild-type cells was inhibited by HDAC inhibitor at high glucose, which most likely was due to enhanced acetylation levels of p300 after HDAC inhibition; and thereby reduced p300 binding to the Txnip gene promoter region. Such inhibition was absent in the Ep300 knock-out cells. Our study provides evidence that histone acetylation serves as a key regulator of glucose-induced increase in TXNIP gene expression and thereby glucotoxicity-induced apoptosis.
硫氧还蛋白相互作用蛋白（TXNIP）已被证明与糖尿病中葡萄糖诱导的胰腺β细胞功能恶化有关。然而，表观遗传机制是否参与葡萄糖对TXNIP基因表达的调控尚不清楚。在此，我们研究了葡萄糖如何通过调节组蛋白乙酰化标记来影响TXNIP基因表达。为此，我们应用成簇规律间隔短回文重复序列/Cas9（CRISPR/Cas9）敲除大鼠胰腺β细胞系INS1 832/13中的组蛋白乙酰转移酶（HAT）p300。我们还用HAT p300和组蛋白去乙酰化酶（HDAC）的化学抑制剂处理细胞和人胰岛。在人胰岛中，糖尿病和高糖导致TXNIP和EP300表达升高，p300抑制剂C646可逆转葡萄糖诱导的TXNIP表达。在INS1 832/13细胞中，CRISPR/Cas9敲除Ep300可提高葡萄糖诱导的胰岛素分泌，并大大降低葡萄糖刺激的Txnip表达和细胞凋亡。这种作用可归因于Txnip基因启动子和第一个编码区组蛋白标记H3K9ac和H4ac的减少。在野生型细胞中，高糖时HDAC抑制剂可抑制Txnip基因中的组蛋白标记H3K9ac和H4ac，这很可能是由于HDAC抑制后p300的乙酰化水平增强，从而减少了p300与Txnip基因启动子区域的结合。在Ep300敲除细胞中不存在这种抑制作用。我们的研究提供了证据，表明组蛋白乙酰化是葡萄糖诱导的TXNIP基因表达增加以及由此导致的糖毒性诱导凋亡的关键调节因子。
