IDENTIFICATION OF A NONPROCESSIVE TELOMERASE ACTIVITY FROM MOUSE CELLS
IDENTIFICATION OF A NONPROCESSIVE TELOMERASE ACTIVITY FROM MOUSE CELLS
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DOI：
10.1073/pnas.90.4.1493
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发表时间：
1993-02-15
影响因子：
11.1
通讯作者：
GREIDER, CW
中科院分区：
文献类型：
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作者：
PROWSE, KR;AVILION, AA;GREIDER, CW
在几种不同小鼠细胞系的提取物中鉴定了端粒酶活性。端粒TTAGGG重复序列的添加对端粒寡核苷酸引物具有特异性，并且对RNA酶A的预处理敏感。与人类和四膜虫端粒酶在体外合成的数百个重复序列相反，小鼠端粒酶在端粒引物上仅合成一个或两个TTAGGG重复序列。用环状排列的（TTAGGG）3序列延长引物后和用ddATP或ddTTP终止链后观察到的产物表明，在序列TTAGGG中添加第一个dG残基后，小鼠端粒酶暂停。由小鼠端粒酶合成的产物的短长度不是由于小鼠提取物中的可扩散抑制剂，因为当与小鼠组分混合时，人端粒酶继续合成长产物。引物激发实验表明，人的酶合成长TTAGGG重复序列在体外processive，而小鼠端粒酶似乎是少得多的进行性。在小鼠提取物中鉴定出短的端粒酶反应产物表明，来自其他生物体的提取物也可能仅产生短的产物。这些知识可能有助于在尚未检测到活性的生物体中鉴定端粒酶活性。
Telomerase activity was identified in extracts from several different mouse cell lines. Addition of telomeric TTAGGG repeats was specific to telomeric oligonucleotide primers and sensitive to pretreatment with RNase A. In contrast to the hundreds of repeats synthesized by the human and Tetrahymena telomerase enzymes in vitro, mouse telomerase synthesized only one or two TTAGGG repeats onto telomeric primers. The products observed after elongation of primers with circularly permuted (TTAGGG)3 sequences and after chain termination with ddATP or ddTTP indicated that mouse telomerase pauses after the addition of the first dG residue in the sequence TTAGGG. The short length of the products synthesized by mouse telomerase was not due to a diffusible inhibitor in the mouse extract, because the human telomerase continued to synthesize long products when mixed with mouse fractions. Primer challenge experiments showed that the human enzyme synthesized long TTAGGG repeats processively in vitro, whereas the mouse telomerase appeared to be much less processive. The identification of short telomerase reaction products in mouse extracts suggests that extracts from other organisms may also generate only short products. This knowledge may aid in the identification of telomerase activity in organisms where activity has not yet been detected.