Janes Sam M, Ofstad Tyler A, Campbell Douglas H, Watt Fiona M, Prowse David M
Keratinocyte Laboratory, Cancer Research UK, London Research Institute, 44 Lincoln's Inn Fields, London WC2A 3PX, UK.
J Cell Sci. 2004 Aug 15;117(Pt 18):4157-68. doi: 10.1242/jcs.01302.
The forkhead transcription factor FOXN1 is required for normal cutaneous and thymic epithelial development. Mutations in FOXN1 give rise to the nude phenotype in mice, rats and man. However, the genes that are regulated by FOXN1 are unknown. To investigate FOXN1 function we expressed an inducible form of the protein, FOXN1ER, that is activated by 4-hydroxytamoxifen in primary human epidermal keratinocytes. Transient activation of FOXN1 decreased the proportion of keratinocytes that formed actively growing clones attributable to stem cell founders and increased the number of abortive clones, without inducing apoptosis. Within 24 hours the majority of cells had initiated terminal differentiation, as assessed by involucrin expression. We performed a cDNA microarray experiment to analyse changes in the transcription of approximately 6,000 genes. Following FOXN1 activation we detected increases of two fold or greater in the RNA levels of over 30 genes. Genes promoting growth arrest, survival and differentiation featured prominently and markers of early events in keratinocyte differentiation were also detected. Since one of the induced genes was Akt we investigated whether Akt played a role in terminal differentiation. Activation of PI 3-kinase but not Akt was necessary for FOXN1-induced differentiation. In reconstituted epidermis FOXN1 promoted early stages of terminal differentiation whereas Akt activation was sufficient to induce late stages, including formation of the cornified layers. These results establish a role for FOXN1 in initiation of terminal differentiation and implicate Akt in subsequent events.
叉头转录因子FOXN1是正常皮肤和胸腺上皮发育所必需的。FOXN1的突变会导致小鼠、大鼠和人类出现裸鼠表型。然而，受FOXN1调控的基因尚不清楚。为了研究FOXN1的功能，我们在原代人表皮角质形成细胞中表达了一种可诱导形式的蛋白质FOXN1ER，它可被4-羟基他莫昔芬激活。FOXN1的瞬时激活降低了由干细胞祖细胞形成的活跃生长克隆的角质形成细胞比例，并增加了流产克隆的数量，且未诱导细胞凋亡。在24小时内，通过外皮蛋白表达评估，大多数细胞已开始终末分化。我们进行了一项cDNA微阵列实验，以分析约6000个基因转录的变化。FOXN1激活后，我们检测到30多个基因的RNA水平增加了两倍或更多。促进生长停滞、存活和分化的基因显著富集，同时也检测到了角质形成细胞分化早期事件的标志物。由于诱导基因之一是Akt，我们研究了Akt是否在终末分化中发挥作用。PI 3-激酶而非Akt的激活是FOXN1诱导分化所必需的。在重组表皮中，FOXN1促进终末分化的早期阶段，而Akt激活足以诱导晚期阶段，包括角质层的形成。这些结果确立了FOXN1在终末分化起始中的作用，并表明Akt参与随后的事件。
