E2F suppression and Sp1 overexpression are sufficient to induce the differentiation-specific marker, transglutaminase type 1, in a squamous cell carcinoma cell line.
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.
Wong
Chung Fai
CF
Epithelial Pathobiology Group, Cancer Biology Programme, Centre for Immunology and Cancer Research, University of Queensland, Princess Alexandra Hospital, Woolloongabba, Queensland 4102, Australia.
Barnes
Liam M
LM
Dahler
Alison L
AL
Smith
Louise
L
Popa
Claudia
C
Serewko-Auret
Magdalena M
MM
Saunders
Nicholas A
NA
D014408:Biomarkers, Tumor / Q000032:analysis; D002294:Carcinoma, Squamous Cell / Q000235:genetics* / Q000473:pathology*; D018797:Cell Cycle Proteins / Q000096:biosynthesis* / Q000235:genetics*; D002454:Cell Differentiation; D004268:DNA-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics*; D015536:Down-Regulation; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; D024202:Electrophoretic Mobility Shift Assay; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D015603:Keratinocytes; D016329:Sp1 Transcription Factor / Q000096:biosynthesis* / Q000235:genetics*; D014157:Transcription Factors / Q000096:biosynthesis* / Q000235:genetics*; D014162:Transfection; D011503:Transglutaminases / Q000096:biosynthesis*; D014407:Tumor Cells, Cultured; D015854:Up-Regulation
D014408:Biomarkers, Tumor; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; C493753:E2F1 protein, human; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; D011503:Transglutaminases; C105804:transglutaminase 1