Miao Feng, Gonzalo Irene Gaw, Lanting Linda, Natarajan Rama
Gonda Diabetes Center, Beckman Research Institute of City of Hope, Duarte, California 91010, USA.
J Biol Chem. 2004 Apr 23;279(17):18091-7. doi: 10.1074/jbc.M311786200. Epub 2004 Feb 19.
The transcription factor NF-kappaB (NF-kappaB) plays a pivotal role in regulating inflammatory gene expression. Its effects are optimized by various coactivators including histone acetyltransferases (HATs) such as CBP/p300 and p/CAF. Evidence shows that high glucose (HG) conditions mimicking diabetes can activate the transcription of NF-kappaB-regulated inflammatory genes. However, the underlying in vivo transcription and nuclear chromatin remodeling events are unknown. We therefore carried out chromatin immunoprecipitation (ChIP) assays in monocytes to identify 1) chromatin factors bound to the promoters of tumor necrosis factor-alpha (TNF-alpha) and related NF-kappaB-regulated genes under HG or diabetic conditions, 2) specific lysine (Lys (K)) residues on histone H3 (HH3) and HH4 acetylated in this process. HG treatment of THP-1 monocytes increased the transcriptional activity of NF-kappaB p65, which was augmented by CBP/p300 and p/CAF. ChIP assays showed that HG increased the recruitment of NF-kappaB p65, CPB, and p/CAF to the TNF-alpha and COX-2 promoters. Interestingly, ChIP assays also demonstrated concomitant acetylation of HH3 at Lys(9) and Lys(14), and HH4 at Lys(5), Lys(8), and Lys(12) at the TNF-alpha and COX-2 promoters. Overexpression of histone deacetylase (HDAC) isoforms inhibited p65-mediated TNF-alpha transcription. In contrast, a HDAC inhibitor stimulated gene transcription and histone acetylation. Finally, we demonstrated increased HH3 acetylation at TNF-alpha and COX-2 promoters in human blood monocytes from type 1 and type 2 diabetic subjects relative to nondiabetic. These results show for the first time that diabetic conditions can increase in vivo recruitment of NF-kappaB and HATs, as well as histone acetylation at the promoters of inflammatory genes, leading to chromatin remodeling and transcription.
转录因子核因子-κB（NF-κB）在调节炎症基因表达中起关键作用。其作用通过包括组蛋白乙酰转移酶（HATs）如CBP/p300和p/CAF等多种共激活因子得以优化。有证据表明，模拟糖尿病的高糖（HG）条件可激活NF-κB调节的炎症基因转录。然而，潜在的体内转录和核染色质重塑事件尚不清楚。因此，我们在单核细胞中进行了染色质免疫沉淀（ChIP）分析，以确定：1）在HG或糖尿病条件下与肿瘤坏死因子-α（TNF-α）及相关NF-κB调节基因启动子结合的染色质因子；2）在此过程中组蛋白H3（HH3）和HH4上特定赖氨酸（Lys（K））残基的乙酰化情况。用HG处理THP-1单核细胞可增加NF-κB p65的转录活性，CBP/p300和p/CAF可增强这种活性。ChIP分析表明，HG增加了NF-κB p65、CPB和p/CAF与TNF-α和COX-2启动子的结合。有趣的是，ChIP分析还证明在TNF-α和COX-2启动子处，HH3的Lys(9)和Lys(14)以及HH4的Lys(5)、Lys(8)和Lys(12)同时发生了乙酰化。组蛋白去乙酰化酶（HDAC）亚型的过表达抑制了p65介导的TNF-α转录。相反，HDAC抑制剂可刺激基因转录和组蛋白乙酰化。最后，我们证明1型和2型糖尿病患者的人血单核细胞中，相对于非糖尿病患者，TNF-α和COX-2启动子处的HH3乙酰化增加。这些结果首次表明，糖尿病条件可增加体内NF-κB和HATs的募集，以及炎症基因启动子处的组蛋白乙酰化，从而导致染色质重塑和转录。
