HDAC3 ensures stepwise epidermal stratification via NCoR/SMRT-reliant mechanisms independent of its histone deacetylase activity (KLF4 microarray)

Chromatin modifiers play critical roles in epidermal development, but the functions of histone deacetylases in this context are poorly understood. We find that the Class I HDAC, HDAC3, is expressed broadly in embryonic epidermis, and is required for its orderly stepwise stratification. Stability of HDAC3 protein in vivo is reliant on NCoR and SMRT, which function redundantly in epidermal development. However, point mutations in the NCoR and SMRT Deacetylase Activating Domains, which are required for HDAC3's enzymatic function, permit normal stratification, indicating that HDAC3's roles in this context are independent of its histone deacetylase activity. HDAC3 functions both in conjunction with, and independent of, KLF4 to repress premature expression of different sets of terminal differentiation genes and suppresses expression of inflammatory cytokines through a RelA-dependent mechanism. These data identify HDAC3 as a hub coordinating multiple aspects of epidermal barrier acquisition. We used microarrays to determine transcriptional changes in Klf4 deleted epidermis compared to control. Epidermis was collected from E18.5 from 6 control and 5 mutant embryos derived from two separate litters.染色质调控因子在表皮发育中发挥关键作用，但目前学界对组蛋白去乙酰化酶（histone deacetylases）在此过程中的功能仍知之甚少。本研究发现，I类组蛋白去乙酰化酶（Class I HDAC）HDAC3在胚胎表皮中广泛表达，且对胚胎表皮的有序逐步分层过程不可或缺。体内HDAC3蛋白的稳定性依赖于NCoR与SMRT，二者在表皮发育中功能冗余。然而，NCoR与SMRT的去乙酰化酶激活结构域（Deacetylase Activating Domains）的点突变——该结构域是HDAC3发挥酶催化活性所必需的——却可使表皮分层过程恢复正常，这表明HDAC3在此过程中的功能不依赖于其组蛋白去乙酰化酶活性。HDAC3可分别与Krüppel样因子4（KLF4）协同或独立发挥作用，抑制多组终末分化基因的过早表达，并通过依赖于RelA的机制抑制炎症细胞因子的表达。上述研究结果表明，HDAC3是协调表皮屏障建立多个环节的核心调控枢纽。本研究通过微阵列（microarray）技术，对比分析了Klf4敲除表皮与对照表皮的转录组变化。实验样本取自两窝独立繁育的胚胎，均采集其胚胎发育第18.5天（E18.5）的表皮组织，其中对照组6只、突变组5只。