Synergistic regulation of human cystathionine-β-synthase-1b promoter by transcription factors NF-YA isoforms and Sp1

Abstract

Cystathionine-β-synthase (CBS) catalyzes the condensation of serine and homocysteine to form cystathionine, an intermediate step in the synthesis of cysteine. We previously described essential transactivating roles for specificity protein 1 (Sp1), Sp3, nuclear factor Y (NF-Y), and USF-1 in the regulation of the CBS-1b promoter. Differential binding of Sp1/Sp3 to the CBS-1b promoter due to differences in Sp1/Sp3 phosphorylation, and Sp1/Sp3 synergism with NF-Y might, in part, explain cell-specific patterns of CBS expression. In this report, the roles of various NF-YA isoforms in influencing cell-specific differences in CBS gene expression were determined in HT1080 and HepG2 cells. Seven unique NF-YA isoforms were detected in HT1080 by reverse transcriptase-PCR (RT-PCR) and DNA sequencing, characterized by deletions in the glutamine-rich and/or serine/threonine-rich domains. Only four of the seven NF-YA isoforms were found in HepG2 cells. The six alternatively spliced NF-YA isoforms all showed significantly less synergistic transactivation of the CBS-1b promoter with Sp1 than wild-type NF-YA, as determined by cotransfections in Drosophila SL2 cells with NF-YB and NF-YC. Further, all six alternatively spliced NF-YA isoforms inhibited the synergistic transactivation of the CBS-1b promoter by wild-type NF-Y and Sp1. Thus, the cellular distributions of these alternatively spliced NF-YA isoforms could impart an important cell-specific component to CBS transcriptional regulation, by virtue of their abilities to directly synergize with Sp1/Sp3 and interfere with transactivation of the CBS-1b promoter by wild-type NF-Y. Characterization of CBS promoter structure and function should clarify the molecular bases for variations in CBS gene expression in genetic diseases and the relationship between CBS and Down's syndrome (DS).