ZNF750 Drives Terminal Epidermal Differentiation via Induction of Klf4

Disrupted differentiation is a hallmark of numerous diseases, which in epidermis alone impact >25% of the population. In a search for dominant mediators of differentiation, we defined a requirement for the ZNF750 nuclear protein in terminal epidermal differentiation. ZNF750 controlled genes mutated in numerous human skin diseases, including FLG, LOR, LCE3B, ALOXE3, and SPINK5. ZNF750 potently induced progenitor differentiation via an evolutionarily conserved C2H2 zinc finger motif. The epidermal master regulator, p63, bound the ZNF750 promoter and was necessary for its induction. ZNF750 restored differentiation to p63-deficient tissue, suggesting it acts downstream of p63. A search for functionally important ZNF750 targets via analysis of ZNF750-regulated genes identified KLF4, a transcription factor that activates late epidermal differentiation genes. ZNF750 binds the Klf4 promoter and controls its expression. ZNF750 thus provides a direct link between a tissue-specifying factor, p63, and an effector of terminal differentiation, Klf4, and represents a potential future target for disorders of this process. Gene expression analysis: To establish a differentiation signature for primary human keratinocytes, with ZNF750-depleted, and Klf4-depleted, total RNA was isolated in biologic duplicate from cells in different conditions and hybridized to Affymetrix HG-U133 2.0 Plus arrays.

分化受阻是众多疾病的标志性特征，仅在表皮组织中就影响超过25%的人群。为探寻分化过程中的关键调控因子，我们明确了核蛋白ZNF750在表皮终末分化（terminal epidermal differentiation）中的必需作用。ZNF750所调控的基因在多种人类皮肤疾病中发生突变，包括FLG、LOR、LCE3B、ALOXE3及SPINK5。ZNF750通过进化保守的C2H2型锌指结构域（C2H2 zinc finger motif），高效诱导表皮祖细胞分化。作为表皮核心调控因子的p63，可结合ZNF750的启动子区域，且对ZNF750的诱导表达不可或缺。ZNF750可恢复p63缺陷组织的分化能力，这提示其作用通路位于p63的下游。通过分析ZNF750的调控基因以筛选其功能关键靶标，我们发现了转录因子KLF4——该因子可激活表皮晚期分化相关基因。ZNF750可结合Klf4的启动子区域并调控其表达。由此，ZNF750搭建起了组织特异性调控因子p63与终末分化效应因子Klf4之间的直接调控链路，同时可为该分化过程异常相关疾病的未来干预提供潜在靶点。基因表达分析：为构建原代人角质形成细胞在ZNF750敲低、Klf4敲低及正常培养条件下的分化特征谱，我们从不同培养状态的细胞中提取了生物学重复的总RNA，并将其与Affymetrix HG-U133 2.0 Plus基因芯片进行杂交。