Ting Helen, Kouzminova Elena A, Kuzminov Andrei
Department of Microbiology, University of Illinois at Urbana-Champaign, Urbana, Illinois, USA.
J Bacteriol. 2008 Sep;190(17):5841-54. doi: 10.1128/JB.00711-08. Epub 2008 Jun 27.
Synthetic lethality is inviability of a double-mutant combination of two fully viable single mutants, commonly interpreted as redundancy at an essential metabolic step. The dut-1 defect in Escherichia coli inactivates dUTPase, causing increased uracil incorporation in DNA and known synthetic lethalities [SL(dut) mutations]. According to the redundancy logic, most of these SL(dut) mutations should affect nucleotide metabolism. After a systematic search for SL(dut) mutants, we did identify a single defect in the DNA precursor metabolism, inactivating thymidine kinase (tdk), that confirmed the redundancy explanation of synthetic lethality. However, we found that the bulk of mutations interacting genetically with dut are in DNA repair, revealing layers of damage of increasing complexity that uracil-DNA incorporation sends through the chromosomal metabolism. Thus, we isolated mutants in functions involved in (i) uracil-DNA excision (ung, polA, and xthA); (ii) double-strand DNA break repair (recA, recBC, and ruvABC); and (iii) chromosomal-dimer resolution (xerC, xerD, and ftsK). These mutants in various DNA repair transactions cannot be redundant with dUTPase and instead reveal "defect-damage-repair" cycles linking unrelated metabolic pathways. In addition, two SL(dut) inserts (phoU and degP) identify functions that could act to support the weakened activity of the Dut-1 mutant enzyme, suggesting the "compensation" explanation for this synthetic lethality. We conclude that genetic interactions with dut can be explained by redundancy, by defect-damage-repair cycles, or as compensation.
合成致死性是指两个完全可存活的单突变体的双突变组合表现出不可存活性，通常被解释为在一个关键代谢步骤上的冗余性。大肠杆菌中的 dut-1 缺陷会使 dUTP 酶失活，导致尿嘧啶在 DNA 中的掺入增加，并产生已知的合成致死性[SL(dut)突变]。根据冗余性逻辑，这些 SL(dut)突变中的大多数应该影响核苷酸代谢。在对 SL(dut)突变体进行系统搜索后，我们确实发现了 DNA 前体代谢中的一个单一缺陷，即胸苷激酶（tdk）失活，这证实了合成致死性的冗余性解释。然而，我们发现与 dut 发生遗传相互作用的大部分突变都与 DNA 修复有关，揭示了尿嘧啶-DNA 掺入在染色体代谢中引发的越来越复杂的损伤层次。因此，我们分离出了参与以下功能的突变体：（i）尿嘧啶-DNA 切除（ung、polA 和 xthA）；（ii）双链 DNA 断裂修复（recA、recBC 和 ruvABC）；以及（iii）染色体二聚体分辨率（xerC、xerD 和 ftsK）。这些在各种 DNA 修复过程中的突变体与 dUTP 酶不可能是冗余的，反而揭示了连接不相关代谢途径的“缺陷-损伤-修复”循环。此外，两个 SL(dut)插入突变（phoU 和 degP）确定了可能起到支持 Dut-1 突变酶减弱活性作用 的功能，这为这种合成致死性提供了“补偿”解释。我们得出结论，与 dut 的遗传相互作用可以通过冗余性、缺陷-损伤-修复循环或补偿来解释。
