在四膜虫提取物中鉴定出一种特定的端粒末端转移酶活性。

Identification of a specific telomere terminal transferase activity in Tetrahymena extracts.

作者信息

Greider C W, Blackburn E H

出版信息

Cell. 1985 Dec;43(2 Pt 1):405-13. doi: 10.1016/0092-8674(85)90170-9.

Abstract

We have found a novel activity in Tetrahymena cell free extracts that adds tandem TTGGGG repeats onto synthetic telomere primers. The single-stranded DNA oligonucleotides (TTGGGG)4 and TGTGTGGGTGTGTGGGTGTGTGGG, consisting of the Tetrahymena and yeast telomeric sequences respectively, each functioned as primers for elongation, while (CCCCAA)4 and two nontelomeric sequence DNA oligomers did not. Efficient synthesis of the TTGGGG repeats depended only on addition of micromolar concentrations of oligomer primer, dGTP, and dTTP to the extract. The activity was sensitive to heat and proteinase K treatment. The repeat addition was independent of both endogenous Tetrahymena DNA and the endogenous alpha-type DNA polymerase; and a greater elongation activity was present during macronuclear development, when a large number of telomeres are formed and replicated, than during vegetative cell growth. We propose that the novel telomere terminal transferase is involved in the addition of telomeric repeats necessary for the replication of chromosome ends in eukaryotes.

摘要

我们在四膜虫无细胞提取物中发现了一种新活性，它能在合成端粒引物上添加串联的TTGGGG重复序列。分别由四膜虫和酵母端粒序列组成的单链DNA寡核苷酸（TTGGGG）4和TGTGTGGGTGTGTGGGTGTGTGGG，均可作为延伸引物发挥作用，而（CCCCAA）4和两种非端粒序列DNA寡聚物则不能。TTGGGG重复序列的高效合成仅取决于向提取物中添加微摩尔浓度的寡聚物引物、dGTP和dTTP。该活性对热和蛋白酶K处理敏感。重复序列的添加既不依赖于内源性四膜虫DNA，也不依赖于内源性α型DNA聚合酶；并且在大核发育过程中，当形成并复制大量端粒时，比在营养细胞生长过程中具有更高的延伸活性。我们认为，这种新的端粒末端转移酶参与了真核生物染色体末端复制所需的端粒重复序列的添加。