Molecular mechanisms of negative regulation on gene expression.

谷胱甘肽转移酶-P(GST-P)基因在正常肝组织中很少表达，而在大鼠肝癌发生过程中高度诱导表达。我们鉴定了GST-P启动子中的沉默区域，并分离了沉默结合蛋白。1)我们克隆了核因子-1(NF1)作为一种沉默结合蛋白，NF1-A具有抑制活性。2)克隆了NF1-A的基因组克隆，并确定了所有的外显子-内含子边界；3)分离了NF1-B的剪接异构体，发现这些蛋白具有正负转录因子的功能；4)研究了NF1-A、-B、-C和-XDNA结合的特异性和亲和力。5)鉴定了NF1-A的核定位信号。6)C/eBPδ基因是通过下游的C/eBP结合位点通过自身调节机制调节的。7)酵母单杂交系统分离了新的沉默结合蛋白

The expression of glutathione transferase-P (GST-P) gene is rarely detected in normal liver, whereas it is highly induced during hepatocarcinogenesis of rat. We identified the silencer region in the GST-P promoter, and isolated the silencer binding proteins. In this project, we characterized the silencer binding proteins and mapped the functional (repressive) domains.1) We cloned nuclear factor-1 (NF1) as a silencer binding protein, and NF1-A had a repressive activity. The minimal repression domain was also identified.2) We cloned the genomic clone of NF1-A and identified the all exon-intron boundaries.3) The splicing isoforms of NF1-B was isolated, and these proteins were found to function as both positive and negative transcription factors.4) We characterized the DNA-binding specificities and affinities of NF1-A, -B, -C and -X.5) The nuclear localization signals in the NF1-A were identified.6) C/EBPδgene is regulated through C/EBP binding sites at the downstream region by autoregulation mechanism.7) The novel silencer binding proteins were isolated by yeast one-hybrid system