Expression of catalytic mutants of the mtDNA helicase Twinkle and polymerase POLG causes distinct replication stalling phenotypes

Sjoerd Wanrooij, Steffi Goffart, Jaakko L.O. Pohjoismäki, Takehiro Yasukawa, Johannes N. Spelbrink

研究成果: Contribution to journalArticle査読

108 被引用数 (Scopus)

抄録

The mechanism of mitochondrial DNA replication is a subject of intense debate. One model proposes a strand-asynchronous replication in which both strands of the circular genome are replicated semi-independently while the other model proposes both a bidirectional coupled leading- and lagging-strand synthesis mode and a unidirectional mode in which the lagging-strand is initially laid-down as RNA by an unknown mechanism (RITOLS mode). Both the strand-asynchronous and RITOLS model have in common a delayed synthesis of the DNA-lagging strand. Mitochondrial DNA is replicated by a limited set of proteins including DNA polymerase gamma (POLG) and the helicase Twinkle. Here, we report the effects of expression of various catalytically deficient mutants of POLG1 and Twinkle in human cell culture. Both groups of mutants reduced mitochondrial DNA copy number by severe replication stalling. However, the analysis showed that while induction of POLG1 mutants still displayed delayed lagging-strand synthesis, Twinkle-induced stalling resulted in maturated, essentially fully double-stranded DNA intermediates. In the latter case, limited inhibition of POLG with dideoxycytidine restored the delay between leading- and lagging-strand synthesis. The observed cause-effect relationship suggests that Twinkle-induced stalling increases lagging-strand initiation events and/ or maturation mimicking conventional strand-coupled replication.

本文言語英語
ページ(範囲)3238-3251
ページ数14
ジャーナルNucleic acids research
35
10
DOI
出版ステータス出版済み - 5 2007
外部発表はい

All Science Journal Classification (ASJC) codes

  • 遺伝学

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