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.

尽管已经利用阿尼玛病毒系统研究了DNA复制和转录的协同机制，但在细胞系统中的协同机制仍不清楚。我们利用c-myc、N-myc、免疫球蛋白重链（IgH）、热休克蛋白70（hsp70）、P53、重复序列Alu、SV40等系统来溶解这些机制。我们已经确定了DNA复制的起点。 (ori) 在活细胞中发挥作用。已确定的起源如下： c-myc 基因转录起始位点上游 15-20 kb，hsp70 基因启动子，IgH 基因增强，p53 基因 11 外显子下游 10 kb。含有上述区域的克隆可以在哺乳动物细胞中自主复制（ARS），并且它们的区域与转录调控区域重叠。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，我们鉴定了识别上述区域的蛋白质因子。 MSSP识别c-myc基因中的ori/增强子，HSBP识别hsp70基因中的启动子/ori，两者均与c-myc蛋白形成复合物。克隆了 MSSP 的两个 cDNA。 Oct-1 还与 IgH 基因中的增强子/ori 结合。在 N-myc 基因中，45k 和 110 K 蛋白与发挥 ARS 作用的增强子区域结合。我们还鉴定了蛋白质 SOAP，它与 SV40 的 ori 核心中富含 AT 的序列结合，并且 SOAP 和 MSSP-1 很可能彼此相同。 Alu 家族 DNA 调节 DNA 复制和转录，并被 37 K 蛋白识别。这里描述的所有蛋白质都与双链和单链 DNA 序列特异性结合，并且可能参与 DNA 复制和转录。