Department of Immunology, University of Washington School of Medicine, Seattle, Washington, United States of America.
PLoS One. 2010 Jan 26;5(1):e8908. doi: 10.1371/journal.pone.0008908.
Exonuclease 1 (EXO1) and Flap endonuclease 1 (FEN1) are members of the RAD2 family of structure-specific nucleases. Genetic analysis has identified roles for EXO1 and FEN1 in replication, recombination, DNA repair and maintenance of telomeres. Telomeres are composed of G-rich repeats that readily form G4 DNA. We recently showed that human EXO1 and FEN1 exhibit distinct activities on G4 DNA substrates representative of intermediates in immunoglobulin class switch recombination.
METHODOLOGY/PRINCIPAL FINDINGS: We have now compared activities of these enzymes on telomeric substrates bearing G4 DNA, identifying non-overlapping functions that provide mechanistic insight into the distinct telomeric phenotypes caused by their deficiencies. We show that hFEN1 but not hEXO1 cleaves substrates bearing telomeric G4 DNA 5'-flaps, consistent with the requirement for FEN1 in telomeric lagging strand replication. Both hEXO1 and hFEN1 are active on substrates bearing telomeric G4 DNA tails, resembling uncapped telomeres. Notably, hEXO1 but not hFEN1 is active on transcribed telomeric G-loops.
CONCLUSION/SIGNIFICANCE: Our results suggest that EXO1 may act at transcription-induced telomeric structures to promote telomere recombination while FEN1 has a dominant role in lagging strand replication at telomeres. Both enzymes can create ssDNA at uncapped telomere ends thereby contributing to recombination.
核酸外切酶 1（EXO1）和核酸内切酶 1（FEN1）是 RAD2 家族结构特异性核酸酶的成员。遗传分析已经确定了 EXO1 和 FEN1 在复制、重组、DNA 修复和端粒维持中的作用。端粒由富含 G 的重复序列组成，这些序列很容易形成 G4 DNA。我们最近表明，人类 EXO1 和 FEN1 在代表免疫球蛋白类别转换重组中间产物的 G4 DNA 底物上表现出不同的活性。
方法/主要发现：我们现在比较了这些酶在具有 G4 DNA 的端粒底物上的活性，确定了它们在功能上没有重叠，这为它们的缺陷导致的不同端粒表型提供了机制上的见解。我们表明，hFEN1 而不是 hEXO1 切割具有端粒 G4 DNA 5'-flap 的底物，这与 FEN1 在端粒滞后链复制中的作用一致。hEXO1 和 hFEN1 都能在具有端粒 G4 DNA 尾巴的底物上发挥作用，类似于无帽端粒。值得注意的是，hEXO1 而不是 hFEN1 能在转录的端粒 G-环上发挥作用。
结论/意义：我们的结果表明，EXO1 可能在转录诱导的端粒结构中发挥作用，以促进端粒重组，而 FEN1 在端粒的滞后链复制中具有主导作用。两种酶都可以在无帽端粒末端产生单链 DNA，从而促进重组。
