Transcription of human cathepsin B is mediated by Sp1 and Ets family factors in glioma

Cathepsin B expression is increased at both the mRNA and protein levels in a wide variety of tumors. The mechanisms responsible for this regulation ar e not well elucidated. We have isolated a 2.2-kb cathepsin B genomic fragme nt that contains the 5'-flanking region of the cathepsin B gene. Using repo rter gene analysis in human glioblastoma U87MG cells, we have mapped a 228- bp fragment (-172 to +56) having high promoter activity. This promoter regi on has a high G+C content; contains potential Spl, Ets, and USF binding mot ifs; and lacks canonical TATA and CAAT boxes immediately upstream of the ma jor transcriptional initiation site. Cotransfection experiments demonstrate d that Sp1 and Ets1 could trans-activate cathepsin B transcription,whereas Ets2 could not. Electrophoretic mobility shift assays and supershift assays revealed that three of the four putative Sp1 sites in this promoter region form a specific complex containing the Spl transcription factor. Mutating all four of the Sp1 binding sites individually markedly reduced the promote r activity of transfected reporter genes in U87 cells. Cotransfection of th is cathepsin B promoter construct with Sp1 family expression vectors in Sch neider's Drosophila line 2 (SL2) cells demonstrated that Sp1 and Sp3, but n ot Sp3, activated cathepsin B transcription. Taken together, these results suggest that Sp1, Sp3, and Ets1 are important factors in cathepsin B transc ription. The regulation of cathepsin B transcription by Sp1- and Sp1-relate d factors is mediated through multiple GC boxes.