CLONING OF SENESCENT CELL-DERIVED INHIBITORS OF DNA-SYNTHESIS USING AN EXPRESSION SCREEN
CLONING OF SENESCENT CELL-DERIVED INHIBITORS OF DNA-SYNTHESIS USING AN EXPRESSION SCREEN
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DOI：
10.1006/excr.1994.1063
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发表时间：
1994-03-01
影响因子：
3.7
通讯作者：
SMITH, JR
中科院分区：
文献类型：
--
作者：
NODA, A;NING, Y;SMITH, JR
我们在这里描述了一种快速而简单的表达筛选方法，该方法使我们能够从衰老的人二倍体成纤维细胞中分离编码DNA合成抑制物的cDNA。该检测包括在增殖细胞中瞬时转录过表达由cDNA编码的基因产物，假设这足以在使用氚胸苷放射自显影的短期检测中阻止DNA合成。三个被称为衰老细胞衍生抑制物(SDI)的cDNA被成功地鉴定出来，当它们被引入年轻的周期细胞中时，它们表现出DNA合成抑制活性。其中一个cDNAs sdi1的表达在衰老的细胞中比年轻的细胞增加10-20倍，并且RNA的增加与衰老表型的开始和细胞增殖的丧失密切相关。在被剥夺生长因子或接触抑制而不分裂(静止)的年轻细胞中，SDI1的表达也增加。血清刺激后，静止细胞中SDI1RNA水平开始升高，但在细胞进入S时相之前下降到较低水平。相反，衰老细胞中sdi1的RNA水平没有下降，这表明该基因在维持衰老表型方面发挥了作用。Sdi1基因已被定位在6号染色体的p臂上。(C)1994年学术出版社，Inc.
We here describe a rapid and simple expression screen method that has allowed us to isolate cDNAs coding for inhibitors of DNA synthesis from senescent human diploid fibroblasts. The assay involved transient transcriptional overexpression of a gene product encoded by a cDNA in a proliferating cell, on the assumption that this would be sufficient to block DNA synthesis in a short-term assay using tritiated thymidine autoradiography. Three cDNAs, referred to as senescent cell-derived inhibitors (sdi), that exhibit DNA synthesis-inhibitory activity when introduced into young cycling cells, were successfully identified. Expression of one of the cDNAs, sdi1, increased 10- to 20fold in senescent compared with young cells and the increase in RNA closely paralleled the onset of the senescent phenotype and loss of cell proliferation. sdi1 expression was also increased in young cells made nondividing (quiescent) by deprivation of growth factors or contact inhibition. Following serum stimulation, RNA levels of sdi1 in quiescent cells were initially increased, but then declined to low levels just prior to the entry of the cells into S phase. In contrast, RNA levels of sdi1 in senescent cells failed to decline, suggesting a role for this gene in maintaining the senescent phenotype. The sdi1 gene has been mapped to the p arm of chromosome 6. (C) 1994 Academic Press, Inc.