Wong Chung Fai, Barnes Liam M, Dahler Alison L, Smith Louise, Popa Claudia, Serewko-Auret Magdalena M, Saunders Nicholas A
Epithelial Pathobiology Group, Cancer Biology Programme, Centre for Immunology and Cancer Research, University of Queensland, Princess Alexandra Hospital, Woolloongabba, Queensland 4102, Australia.
Oncogene. 2005 May 12;24(21):3525-34. doi: 10.1038/sj.onc.1208372.
Recently, E2F function has expanded to include the regulation of differentiation in human epidermal keratinocytes (HEKs). We extend these findings to report that in HEKs, Sp1 is a differentiation-specific activator and a downstream target of E2F-mediated suppression of the differentiation-specific marker, transglutaminase type 1 (TG-1). Deletion of elements between -0.084 to -0.034 kb of the TG-1 promoter disabled E2F1-induced suppression of promoter activity. Electrophoretic mobility shift assays (EMSAs) demonstrated that Sp1 and Sp3 bound this region. Protein expression analysis suggested that squamous differentiation was accompanied by increased Sp1/Sp3 ratio. Cotransfection of proliferating HEKs or the squamous cell carcinoma (SCC) cell line, KJD-1/SV40, with an E2F inhibitor (E2Fd/n) and Sp1 expression plasmid was sufficient to activate the TG-1 promoter. The suppression of Sp1 activity by E2F in differentiated cells appeared to be indirect since we found no evidence of an Sp1/E2F coassociation on the TG-1 promoter fragment. Moreover, E2F inhibition in the presence of a differentiation stimulus induced Sp1 protein. These data demonstrate that (i) Sp1 can act as a differentiation stimulus, (ii) E2F-mediated suppression of differentiation-specific markers is indirect via Sp1 inhibition and (iii) a combination of E2F inhibition and Sp1 activation could form the basis of a differentiation therapy for SCCs.
最近，E2F的功能已扩展至包括对人表皮角质形成细胞（HEK）分化的调控。我们拓展了这些发现，报告在HEK中，Sp1是一种分化特异性激活因子，并且是E2F介导的对分化特异性标志物转谷氨酰胺酶1（TG-1）抑制作用的下游靶点。缺失TG-1启动子-0.084至-0.034 kb之间的元件会使E2F1诱导的启动子活性抑制失效。电泳迁移率变动分析（EMSA）表明Sp1和Sp3结合该区域。蛋白质表达分析提示鳞状分化伴随着Sp1/Sp3比值增加。用E2F抑制剂（E2Fd/n）和Sp1表达质粒共转染增殖性HEK或鳞状细胞癌（SCC）细胞系KJD-1/SV40，足以激活TG-1启动子。在分化细胞中E2F对Sp1活性的抑制似乎是间接的，因为我们未发现TG-1启动子片段上存在Sp1/E2F共缔合的证据。此外，在存在分化刺激的情况下抑制E2F可诱导Sp1蛋白。这些数据表明：（i）Sp1可作为分化刺激因子；（ii）E2F介导的对分化特异性标志物的抑制是通过抑制Sp1间接实现的；（iii）抑制E2F与激活Sp1相结合可能构成SCC分化治疗的基础。
