Transcription profiling of mouse skins of embryos lacking Grainyhead-like epithelial transactivator (Get-1)

Defective permeability barrier is an important feature of many skin diseases and causes mortality in premature infants. To investigate the control of barrier formation, we characterized the epidermally expressed Grainyhead-like epithelial transactivator (Get-1)/Grhl3, a conserved mammalian homologue of Grainyhead, which plays important roles in cuticle development in Drosophila. Get-1 interacts with the LIM-only protein LMO4, which is co-expressed in the developing mammalian epidermis. The epidermis of Get-1(-/-) mice showed a severe barrier function defect associated with impaired differentiation of the epidermis, including defects of the stratum corneum, extracellular lipid composition and cell adhesion in the granular layer. The Get-1 mutation affects multiple genes linked to terminal differentiation and barrier function, including most genes of the epidermal differentiation complex. Get-1 therefore directly or indirectly regulates a broad array of epidermal differentiation genes encoding structural proteins, lipid metabolizing enzymes and cell adhesion molecules. Although deletion of the LMO4 gene had no overt consequences for epidermal development, the epidermal terminal differentiation defect in mice deleted for both Get-1 and LMO4 is much more severe than in Get-1(-/-) mice with striking impairment of stratum corneum formation. These findings indicate that the Get-1 and LMO4 genes interact functionally to regulate epidermal terminal differentiation. Experiment Overall Design: The same region of the mouse back skin was excised from three Get1 +/+ and three Get1 -/- mice at e18.5.皮肤屏障功能缺损是多种皮肤疾病的典型特征，亦是导致早产儿死亡的重要诱因。为探究皮肤屏障形成的调控机制，我们对表皮特异性表达的Grainyhead样上皮转录激活因子（Grainyhead-like epithelial transactivator, Get-1）/Grhl3进行了系统表征——该蛋白是果蝇Grainyhead的保守哺乳动物同源物，在果蝇表皮发育过程中发挥关键调控作用。Get-1可与仅含LIM结构域蛋白LMO4（LIM-only protein LMO4）相互作用，二者在发育中的哺乳动物表皮中共表达。Get-1基因纯合敲除（Get-1(-/-)）小鼠的表皮出现严重屏障功能缺损，伴随表皮分化受损，具体表现为角质层（stratum corneum）异常、细胞外脂质组成改变以及颗粒层细胞黏附功能障碍。Get-1突变可影响多个与表皮终末分化及屏障功能相关的基因，其中绝大多数属于表皮分化复合体（epidermal differentiation complex）。因此，Get-1可直接或间接调控一系列表皮分化相关基因，这些基因编码结构蛋白、脂质代谢酶及细胞黏附分子。尽管单独敲除LMO4基因并不会对表皮发育产生明显影响，但同时敲除Get-1与LMO4的小鼠，其表皮终末分化缺陷程度远高于单独敲除Get-1的小鼠，角质层形成受到显著破坏。上述研究结果表明，Get-1与LMO4基因可通过功能互作协同调控表皮终末分化过程。实验总体设计：在胚胎第18.5天（e18.5），分别从3只Get1野生型（Get1 +/+）与3只Get1纯合敲除（Get1 -/-）小鼠背部皮肤取材同一区域的组织。