Tsubouchi H, Ogawa H
Department of Biology, Graduate School of Science, Osaka University, Toyonaka, Japan.
Mol Cell Biol. 1998 Jan;18(1):260-8. doi: 10.1128/MCB.18.1.260.
Using complementation tests and nucleotide sequencing, we showed that the rad58-4 mutation was an allele of the MRE11 gene and have renamed the mutation mre11-58. Two amino acid changes from the wild-type sequence were identified; one is located at a conserved site of a phosphodiesterase motif, and the other is a homologous amino acid change at a nonconserved site. Unlike mre11 null mutations, the mre11-58 mutation allowed meiosis-specific double-strand DNA breaks (DSBs) to form at recombination hot spots but failed to process those breaks. DSB ends of this mutant were resistant to lambda exonuclease treatment. These phenotypes are similar to those of rad50S mutants. In contrast to rad50S, however, mre11-58 was highly sensitive to methyl methanesulfonate treatment. DSB end processing induced by HO endonuclease was suppressed in both mre11-58 and the mre11 disruption mutant. We constructed a new mre11 mutant that contains only the phosphodiesterase motif mutation of the Mre11-58 protein and named it mre11-58S. This mutant showed the same phenotypes observed in mre11-58, suggesting that the phosphodiesterase consensus sequence is important for nucleolytic processing of DSB ends during both mitosis and meiosis.
通过互补试验和核苷酸测序，我们发现rad58 - 4突变是MRE11基因的一个等位基因，并将该突变重新命名为mre11 - 58。我们鉴定出了两个与野生型序列不同的氨基酸变化；一个位于磷酸二酯酶基序的保守位点，另一个是在非保守位点的同源氨基酸变化。与mre11基因敲除突变不同，mre11 - 58突变允许减数分裂特异性双链DNA断裂（DSB）在重组热点处形成，但无法处理这些断裂。该突变体的DSB末端对λ核酸外切酶处理具有抗性。这些表型与rad50S突变体相似。然而，与rad50S不同的是，mre11 - 58对甲基磺酸甲酯处理高度敏感。在mre11 - 58和mre11基因破坏突变体中，HO内切酶诱导的DSB末端加工均受到抑制。我们构建了一个新的mre11突变体，它仅包含Mre11 - 58蛋白的磷酸二酯酶基序突变，并将其命名为mre11 - 58S。该突变体表现出与mre11 - 58相同的表型，这表明磷酸二酯酶共有序列在有丝分裂和减数分裂过程中对DSB末端的核酸酶加工很重要。
