Ui A, Satoh Y, Onoda F, Miyajima A, Seki M, Enomoto T
Molecular Cell Biology Laboratory, Graduate School of Pharmaceutical Sciences, Tohoku University, Sendai, Miyagi, Japan.
Mol Genet Genomics. 2001 Jul;265(5):837-50. doi: 10.1007/s004380100479.
The SGS1 gene of Saccharomyces (cerevisiae is a homologue of the genes affected in Bloom's syndrome, Werner's syndrome, and Rothmund-Thomson's syndrome. Disruption of the SGS1 gene is associated with high sensitivity to methyl methanesulfonate (MMS) and hydroxyurea (HU), and with hyper-recombination phenotypes, including interchromosomal recombination between heteroalleles. SGS1 encodes a protein which has a helicase domain similar to that of Escherichia coli RecQ. A comparison of amino acid sequences among helicases of the RecQ family reveals that Sgs1,WRN, and BLM share a conserved region adjacent to the C-terminal part of the helicase domain (C-terminal conserved region). In addition, Sgs1 contains two highly charged acidic regions in its N-terminal region and the HRDC (helicase and RNaseD C-terminal) domain at its C-terminal end. These regions were also found in BLM and WRN, and in Rqh1 from Schizosaccharomyces pombe. In this study, we demonstrate that the C-terminal conserved region, as well as the helicase motifs, of Sgs1 are essential for complementation of MMS sensitivity and suppression of hyper-recombination in sgs1 mutants. In contrast, the highly charged acidic regions, the HRDC domain, and the C-terminal 252 amino acids were dispensable for the complementation of these phenotypes. Surprisingly, the N-terminal 45 amino acids of Sgs1 were absolutely required for the suppression of the above phenotypes. Introduction of missense mutations into the region encoding amino acids 4-13 abolished the ability of Sgsl to complement MMS sensitivity and suppress hyper-recombination in sgs1 mutants, and also prevented its interaction with Top3, indicating that interaction with Top3 via the N-terminal region of Sgs1 is involved in the complementation of MMS sensitivity and the suppression of hyper-recombination.
酿酒酵母的SGS1基因是在布卢姆综合征、沃纳综合征和罗思蒙德 - 汤姆森综合征中受影响基因的同源物。SGS1基因的破坏与对甲磺酸甲酯（MMS）和羟基脲（HU）的高敏感性以及高重组表型有关，包括异等位基因之间的染色体间重组。SGS1编码一种蛋白质，该蛋白质具有与大肠杆菌RecQ相似的解旋酶结构域。RecQ家族解旋酶之间的氨基酸序列比较表明，Sgs1、WRN和BLM在解旋酶结构域的C末端部分附近共享一个保守区域（C末端保守区域）。此外，Sgs1在其N末端区域包含两个高度带电荷的酸性区域，并在其C末端含有HRDC（解旋酶和RNaseD C末端）结构域。这些区域也存在于BLM和WRN以及粟酒裂殖酵母的Rqh1中。在本研究中，我们证明Sgs1的C末端保守区域以及解旋酶基序对于sgs1突变体中MMS敏感性的互补和高重组的抑制是必不可少的。相比之下，高度带电荷的酸性区域、HRDC结构域和C末端的252个氨基酸对于这些表型的互补是可有可无的。令人惊讶的是，Sgs1的N末端45个氨基酸对于上述表型的抑制是绝对必需的。在编码氨基酸4 - 13的区域引入错义突变消除了Sgsl互补MMS敏感性和抑制sgs1突变体中高重组的能力，并且还阻止了它与Top3的相互作用，表明通过Sgs1的N末端区域与Top3的相互作用参与了MMS敏感性的互补和高重组的抑制。
