Chen Zhenhua, Lu Junpeng, Tang Qinghai, Yang Zhili
Systems Biology Laboratory, School of Marine Science and Technology, Zhejiang Ocean University, Zhoushan, Zhejiang, China.
Systems Biology Laboratory, School of Marine Science and Technology, Zhejiang Ocean University, Zhoushan, Zhejiang, China.
J Biol Chem. 2025 Apr 26;301(6):108553. doi: 10.1016/j.jbc.2025.108553.
FabZ, a β-hydroxyacyl-acyl carrier protein dehydratase in the type II fatty acid synthesis pathway, is essential for the viability of Pseudomonas aeruginosa by ensuring proper fatty acid elongation and membrane stability. However, the precise genetic interactions between fabZ and lipid A biosynthesis genes, such as lpxA and lpxC, as well as the potential existence of other suppressor genes of fabZ in P. aeruginosa, remain unclear. To explore these genetic interactions and identify potential suppressor genes, we constructed a conditional fabZ mutant, ΔfabZ(p_ts-fabZ), by deleting the chromosomal fabZ gene and complementing it with a temperature-sensitive plasmid-borne copy. The ΔfabZ(p_ts-fabZ) mutant exhibited lethality and cell morphology defects at a restrictive temperature, confirming its essentiality. Genetic interaction analyses revealed that deletion of lpxA or lpxC failed to rescue ΔfabZ(p_ts-fabZ) lethality at restrictive temperature. Through suppressor screening, we isolated a mutant strain capable of rescuing ΔfabZ lethality and identified lpxH as the suppressor gene using genome resequencing. Further analysis revealed that the fabZ and lpxH double mutant (ΔfabZΔlpxH) produced odd-chain fatty acids, identified as pentadecanoic acid (C15:0) and heptadecanoic acid (C17:0) through fatty acid methyl ester analysis coupled with GC-MS, and supplementation with these fatty acids restored the growth and morphology of ΔfabZ(p_ts-fabZ) and ΔlpxH(p_ts-lpxH) mutants at restrictive temperature, suggesting their critical role in membrane stability. These results indicate that deletion of lpxH serves as a genetic suppressor of ΔfabZ lethality, highlighting a previously unrecognized compensatory mechanism involving odd-chain fatty acid synthesis essential for membrane stability in P. aeruginosa.
FabZ是II型脂肪酸合成途径中的一种β-羟基酰基-酰基载体蛋白脱水酶，通过确保适当的脂肪酸延长和膜稳定性，对铜绿假单胞菌的生存能力至关重要。然而，fabZ与脂质A生物合成基因（如lpxA和lpxC）之间的确切遗传相互作用，以及铜绿假单胞菌中fabZ其他潜在抑制基因的存在，仍不清楚。为了探索这些遗传相互作用并鉴定潜在的抑制基因，我们构建了一个条件性fabZ突变体ΔfabZ(p_ts-fabZ)，方法是删除染色体上的fabZ基因并用温度敏感型质粒携带的拷贝进行互补。ΔfabZ(p_ts-fabZ)突变体在限制温度下表现出致死性和细胞形态缺陷，证实了其必要性。遗传相互作用分析表明，删除lpxA或lpxC未能挽救ΔfabZ(p_ts-fabZ)在限制温度下的致死性。通过抑制子筛选，我们分离出一个能够挽救ΔfabZ致死性的突变菌株，并通过基因组重测序鉴定lpxH为抑制基因。进一步分析表明，fabZ和lpxH双突变体(ΔfabZΔlpxH)产生奇数链脂肪酸，通过脂肪酸甲酯分析结合GC-MS鉴定为十五烷酸(C15:0)和十七烷酸(C17:0)，补充这些脂肪酸可恢复ΔfabZ(p_ts-fabZ)和ΔlpxH(p_ts-lpxH)突变体在限制温度下的生长和形态，表明它们在膜稳定性中起关键作用。这些结果表明，删除lpxH可作为ΔfabZ致死性的遗传抑制子，并突出了一种以前未被认识的补偿机制，该机制涉及奇数链脂肪酸合成，对铜绿假单胞菌的膜稳定性至关重要。
