Mitochondria house evolutionarily conserved pathways of carbon and nitrogen metabolism that drive cellular energy production. Mitochondrial bioenergetics is regulated by calcium uptake through the mitochondrial calcium uniporter (MCU), a multi-protein complex whose assembly in the inner mitochondrial membrane is facilitated by the scaffold factor MCUR1. Intriguingly, many fungi that lack MCU contain MCUR1 homologs, suggesting alternate functions. Herein, we characterize Saccharomyces cerevisiae homologs Put6 and Put7 of MCUR1 as regulators of mitochondrial proline metabolism. Put6 and Put7 are tethered to the inner mitochondrial membrane in a large hetero-oligomeric complex, whose abundance is regulated by proline. Loss of this complex perturbs mitochondrial proline homeostasis and cellular redox balance. Yeast cells lacking either Put6 or Put7 exhibit a pronounced defect in proline utilization, which can be corrected by the heterologous expression of human MCUR1. Our work uncovers an unexpected role of MCUR1 homologs in mitochondrial proline metabolism.

线粒体拥有进化上保守的碳和氮代谢途径，驱动细胞能量产生。线粒体生物能通过线粒体钙单向转运蛋白 (MCU) 的钙摄取进行调节，MCU 是一种多蛋白复合物，支架因子 MCUR1 促进其在线粒体内膜中的组装。有趣的是，许多缺乏 MCU 的真菌含有 MCUR1 同源物，这表明它们具有替代功能。在此，我们将 MCUR1 的酿酒酵母同源物 Put6 和 Put7 描述为线粒体脯氨酸代谢的调节剂。 Put6 和 Put7 在一个大型异源寡聚复合物中与线粒体内膜相连，其丰度受脯氨酸调节。这种复合物的丢失会扰乱线粒体脯氨酸稳态和细胞氧化还原平衡。缺乏 Put6 或 Put7 的酵母细胞在脯氨酸利用方面表现出明显的缺陷，这可以通过人 MCUR1 的异源表达来纠正。我们的工作揭示了 MCUR1 同源物在线粒体脯氨酸代谢中的意想不到的作用。