Concerted mechanism of DNA replication initiation and transcription in mammalian cells

哺乳动物细胞中 DNA 复制起始和转录的协同机制

基本信息

批准号：
03454489
负责人：
ARIGA Hiroyoshi
金额：
$ 4.1万
依托单位：
Hokkaido University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1991
资助国家：
日本
起止时间：
1991 至 1992
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-03454489/
关键词：
DNA replication transcriptional regulation c-myc N-myc hsp70 immunoglobulin SV40 Alu family Aluファミリー myc遺伝子 HSP70 免疫グロブリン Alu配列転写因子複製開始

项目摘要

Although the concerted mechanism of DNA replivation and transcription has been studied by using anima virus system, that in cellular system is stil not clarified. We have used the systems including c-myc, N-myc, immunogloburin heavy chain (IgH), heat shock protein 70 (hsp70), P53, repeated sequence Alu, and SV40 to dissolve such mechanisms.We have determined the origin of DNA replication (ori) functioning in living cells. The origins identified were located as following; 15-20 kbs upstream of the start site of transcription in c-myc gene, promoter in hsp70 gene, enhance in IgH gene, 10 kb downstream of the 11exon in p53 gene. Clones containing above regions could replicate autonomously (ARS) in mammalian cells and their regions overlapped the transcriptional regulatory regions. Thus these results clearly indicate the concerted mechanism of DNA replication initiation and transcription in mammalian cells.We then identified the protein factors recognizing the regions described above. MSSP recognized the ori/enhancer in c-myc gene and HSBP did the promoter/ori in hsp70 gene, both of which made complexes with c-myc protein. Two cDNAs of MSSP were cloned. Oct-1 also bound to the enhancer/ori in IgH gene. In N-myc gene, 45k and 110 K proteins bound to the enhancer region acting ARS. We have also identified the protein, SOAP, which bound AT rich sequence placed in ori core of SV40, and SOAP and MSSP-1 are most probably identical each other. Alu family DNA modulated both DNA replication and transcription, and was recognized by 37 K protein. All the proteins described here bind sequenece-specifically to both double and single stranded DNA and may be involved in DNA replication and transcription.

尽管已经通过使用Anima病毒系统研究了DNA重新启动和转录的协同机制，但在细胞系统中尚未阐明该机制。我们已经使用了包括C-MYC，N-MYC，免疫白素重链（IGH），热激蛋白70（HSP70），p53，重复序列ALU和SV40的系统来溶解此类机制。我们确定了活细胞中DNA复制（ORI）的起源。确定的起源如下； 15-20 kbs在C-MYC基因的开始位点的上游，Hsp70基因中的启动子，增强IGH基因中的IGH基因，p53基因的11 exon下游10 kb。包含上述区域的克隆可以在哺乳动物细胞及其区域中自主复制（ARS）重叠。因此，这些结果清楚地表明了哺乳动物细胞中DNA复制起始和转录的协同机制。然后，我们确定了识别上述区域的蛋白质因子。 MSSP识别C-MYC基因和汇丰（HSBP）中的ORI/增强子在HSP70基因中进行了启动子/ORI，这两者都与C-MYC蛋白制成了复合物。克隆两个MSSP的cDNA。 OCT-1也与IGH基因中的增强子/ORI结合。在N-MYC基因中，与增强子区域作用ARS结合的45K和110 K蛋白。我们还鉴定了蛋白质，肥皂，该蛋白质结合在SV40 ORI核心的丰富序列上，而SOAP和MSSP-1很可能相同。 Alu家族DNA调节了DNA复制和转录，并被37 K蛋白识别。此处描述的所有蛋白质特定于双重和单链DNA，并可能参与DNA复制和转录。