多药耐药性蛋白MRP5和MRP9在雄性生殖和线粒体功能中起协同作用

多药耐药性蛋白是一种转运蛋白，在癌症中起着关键作用，尽管这些转运蛋白的生理基础尚不清楚。在秀丽隐桿线虫中，MRP5/ABCC5是一种重要的血红素输出者，因为 MRP5突变体不能从肠道向肠外组织输出血红素，所以无法存活。补充血红素可恢复这些突变体的生存能力，但不能恢复男性生殖缺陷。相应地，细胞生物学研究表明，MRP5调节哺乳动物分泌通路的血红素水平，即使 MRP5基因敲除(KO)小鼠没有表现出生殖表型。MRP5最接近的同源基因是 MRP9/ABCC12，在秀丽隐杆线虫中不存在这种基因，提高了 MRP9可能在遗传上弥补 MRP5的可能性。本研究，我们表明 MRP5和 MRP9双 KO 小鼠是可行的，但显示了显着的雄性生殖缺陷。虽然 MRP9在精子中高度表达，但 MRP9 KO 小鼠只有在 MRP5缺失时才表现出生殖表型。这两种 ABCC 转运蛋白都定位于线粒体相关膜(MAMs)上，这是一种将线粒体和内质网结合在一起的动态支架。因此，DKO 小鼠发现了异常的精子线粒体，线粒体膜电位和受精率都降低了。

代谢组学研究显示，DKO 睾丸的代谢物谱和 rna seq 的代谢物谱有显著差异，其中与线粒体功能和维甲酸代谢有关的基因发生了显著变化。靶向功能代谢组学显示，DKO 睾丸的维甲酸水平较低，而线粒体的甘油三酯水平较高。这些发现建立了 MRP5和 MRP9在调节男性生殖功能和线粒体充足性方面发挥协同作用的模型。

Multidrug Resistance Proteins (MRPs) are transporters that play critical roles in cancer even though the physiological substrates of these enigmatic transporters are poorly elucidated. In Caenorhabditis elegans, MRP5/ABCC5 is an essential heme exporter as mrp-5 mutants are unviable due to their inability to export heme from the intestine to extra-intestinal tissues. Heme supplementation restores viability of these mutants but fails to restore male reproductive deficits. Correspondingly, cell biological studies show that MRP5 regulates heme levels in the mammalian secretory pathway even though MRP5 knockout (KO) mice do not show reproductive phenotypes. The closest homolog of MRP5 is MRP9/ABCC12, which is absent in C. elegans raising the possibility that MRP9 may genetically compensate for MRP5. Here, we show that MRP5 and MRP9 double KO mice are viable but reveal significant male reproductive deficits. Although MRP9 is highly expressed in sperm, MRP9 KO mice show reproductive phenotypes only when MRP5 is absent. Both ABCC transporters localize to mitochondrial-associated membranes (MAMs), dynamic scaffolds that associate the mitochondria and endoplasmic reticulum. Consequently, DKO mice reveal abnormal sperm mitochondria with reduced mitochondrial membrane potential and fertilization rates. Metabolomics show striking differences in metabolite profiles in the DKO testes and RNA-seq show significant alterations in genes related to mitochondrial function and retinoic acid metabolism. Targeted functional metabolomics reveal lower retinoic acid levels in the DKO testes and higher levels of triglycerides in the mitochondria. These findings establish a model in which MRP5 and MRP9 play a concerted role in regulating male reproductive functions and mitochondrial sufficiency.

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