Transcriptional regulation of the human cystathionine beta-synthase -1b basal promoter: synergistic transactivation by transcription factors NF-Y and Sp1/Sp3

• Published in: Biochemical journal 2001-07, Vol.357 (Pt 1), p.97-105
• Main Authors: Ge, Y, Konrad, M A, Matherly, L H, Taub, J W
• Format: Article
• Language: English
• Subjects: Base Sequence; Binding Sites; CBS gene; CCAAT-Binding Factor - metabolism; Cystathionine ^b-synthase; Cystathionine beta-Synthase - genetics; Drosophila melanogaster; Exons; Gene Expression Regulation, Enzymologic; Humans; Introns; Molecular Sequence Data; NF-Y gene; Oligodeoxyribonucleotides - chemistry; Promoter Regions, Genetic; Recombinant Proteins - biosynthesis; Regulatory Sequences, Nucleic Acid; Reverse Transcriptase Polymerase Chain Reaction; Sp1 gene; Sp1 Transcription Factor - metabolism; Transcription, Genetic; Transcriptional Activation; Transfection; Tumor Cells, Cultured
• ISSN: 0264-6021; 1470-8728
• DOI: 10.1042/0264-6021:3570097

Cystathionine beta-synthase (CBS) catalyses the condensation of serine and homocysteine to form cystathionine, an intermediate step in the synthesis of cysteine. Human CBS encodes five distinct 5' non-coding exons, the most frequent termed CBS -1a and CBS -1b, each transcribed from its own unique GC-rich TATA-less promoter. The minimal transcriptional region (-3792 to -3667) of the CBS -1b promoter was defined by 5'- and 3'-deletions, and transient transfections of reporter gene constructs in HepG2 cells, characterized by CBS transcription exclusively from the -1b promoter. Included in this 125 bp region are 3 GC-boxes (termed GC-a, GC-b and GC-c), an inverted CAAT-box and an E-box. By gel-shift and supershift assays, binding of specificity protein (Sp)1 and Sp3 to the GC-box elements, upstream stimulatory factor 1 (USF-1) to the E-box, and both nuclear factor (NF)-Y and an NF-1-like factor to the CAAT box could be demonstrated. By transient trans fections and reporter gene assays in HepG2 and Drosophila SL2 cells, a functional interplay was indicated between NF-Y binding to the CAAT-box, or between USF-1 binding to the E-box, and Sp1/Sp3 binding to the GC-box elements. In SL2 cells, NF-Y and Sp1/Sp3 were synergistic. Furthermore, both Sp1 and the long Sp3 isoform transactivated the CBS -1b minimal promoter; however, the short Sp3 isoforms were potent repressors. These results may explain the cell- or tissue-specific regulation of CBS transcription, and clarify the bases for alterations in CBS gene expression in human disease and Down's syndrome.