Fernández-Ayala Daniel J M, Jiménez-Gancedo Sandra, Guerra Ignacio, Hernández-Camacho Juan D, Neto Marta, Scialo Filippo, Astillero-López Verónica, Cortés-Rodríguez Ana Belén, Santos-Ocaña Carlos, Rodríguez-Aguilera Juan Carlos, Casares Fernando, Sanz Alberto, López-Lluch Guillermo, Navas Plácido
Centro Andaluz de Biología del Desarrollo, Universidad Pablo de Olavide-CSIC-JA, Sevilla, Spain; CIBERER, U729, Instituto de Salud Carlos III, Madrid, Spain.
Centro Andaluz de Biología del Desarrollo, Universidad Pablo de Olavide-CSIC-JA, Sevilla, Spain.
Free Radic Biol Med. 2025 Mar 16;230:95-111. doi: 10.1016/j.freeradbiomed.2024.12.056. Epub 2025 Jan 27.
The interference of the expression of each of the genes involved in the synthesis of coenzyme Q (CoQ) in Drosophila melanogaster can help to understand the pathophysiology of CoQ-dependent mitochondrial diseases in humans. We have knocked-down all genes involved in the CoQ biosynthesis pathway at different temperatures to induce depletion of CoQ at different levels throughout the body and in a tissue-specific manner. The efficiency of the knockdowns was quantified by Q-RTPCR and determination of CoQ levels by HPLC-UV + ECD. We performed mitochondria purification and quantified respiratory chain activity, both mitochondrial hydrogen peroxide and superoxide production, resistance to mechanical stress and determination of life expectancy. Finally, we evaluated the effect of CoQ10 supplementation as phenotype rescue therapy. D. melanogaster presents 3 isoforms of CoQ: CoQ8, CoQ9 and CoQ10. The level of depletion depended on the efficiency of the RNAi used and is specific for each gene. The interference of some genes interrupted fly development in embryogenesis (pdss2) or during metamorphosis (pdss1, coq3, coq5, coq8 and coq10), while in other cases viable adults can be obtained (coq2, coq6 and coq7). The knockdown of coq7 accumulated intermediates of the CoQ biosynthesis pathway at all stages of development, altered electron transfer with poor assembly of mitochondrial complexes, and deregulated mitochondrial hydrogen peroxide and superoxide production. Coq7 mutant flies showed partial lethality in metamorphosis, bang sensitivity and reduced life span of surviving animals. CoQ10 supplementation rescued the coq7-mutant phenotypes. Knock-down in the imaginal disc generated gene-specific eye deformities that can be mitigated by CoQ10 supplementation. Our results indicate that interference of the CoQ biosynthesis pathway in D. melanogaster shows a great diversity of phenotypes depending on the target gene, mirroring the heterogeneity of CoQ deficiency syndrome in humans and point to why mutations in certain genes are rarely found in patients.
干扰果蝇中参与辅酶Q（CoQ）合成的每个基因的表达，有助于了解人类中CoQ依赖性线粒体疾病的病理生理学。我们在不同温度下敲低了CoQ生物合成途径中的所有基因，以在全身和组织特异性方式下诱导不同水平的CoQ消耗。通过Q-RTPCR定量敲低效率，并通过HPLC-UV + ECD测定CoQ水平。我们进行了线粒体纯化，并定量了呼吸链活性、线粒体过氧化氢和超氧化物的产生、对机械应激的抗性以及预期寿命的测定。最后，我们评估了补充CoQ10作为表型挽救疗法的效果。果蝇呈现3种CoQ同工型：CoQ8、CoQ9和CoQ10。消耗水平取决于所用RNAi的效率，并且对每个基因都是特异性的。一些基因的干扰在胚胎发生（pdss2）或变态期间（pdss1、coq3、coq5、coq8和coq10）中断了果蝇发育，而在其他情况下可以获得存活的成虫（coq2、coq6和coq7）。coq7的敲低在发育的所有阶段积累了CoQ生物合成途径的中间体，改变了电子传递，线粒体复合物组装不良，并使线粒体过氧化氢和超氧化物的产生失调。coq7突变果蝇在变态中表现出部分致死性、对巨响敏感以及存活动物的寿命缩短。补充CoQ10挽救了coq7突变体表型。在成虫盘上敲低产生了基因特异性的眼睛畸形，补充CoQ10可以减轻这种畸形。我们的结果表明，果蝇中CoQ生物合成途径的干扰根据靶基因显示出多种表型，反映了人类中CoQ缺乏综合征的异质性，并指出了为什么某些基因的突变在患者中很少发现。
