Developmentally Regulated GTP-binding Protein Drg1 defines a translational decision point that protects mitochondrial integrity

Abstract

Coordination of the functionalities among key organelles is essential for maintaining the complexity and adaptability of the eukaryotic cell. Functionally versatile proteins, such as GTPases, often assume important roles in this process. A ribosome-binding GTPase called Developmentally-regulated GTP-binding (Drg) protein is a conserved family of GTPases recently implicated in promoting protein synthesis and regulating cytoskeletal dynamics, suggesting their importance for cell growth and proliferation. Here, we show that Drg1, one of the two Drg paralogues in eukaryotic cells, is critical for maintaining mitochondria morphology, dynamics, and function. Results from APEX2-proximity labeling show that Drg1 is in proximity to proteins associated with the nucleus, mitochondria, endoplasmic reticulum membrane, cytoplasmic ribosomal subunits, and cytoskeleton components. We demonstrate that Drg1 associates with the outer mitochondrial membrane and that loss of Drg1 leads to reduced mitochondrial membrane potential, decreased protein import, and ATP production. Data from our mammalian in vitro translation assay show that the presence of Drg1/Dfrp1 increases puromycin incorporation in the stalled ribosome, indicating a molecular mechanism of enhanced translation. Data from candidate approaches further reveal that loss of Drg1 leads to changes in mRNA abundance and translation of proteins critical for mitochondrial ATP production, fusion, and fission. Thus, at the molecular level, loss of cellular homeostasis that is caused by compromised translation of cytoplasmically synthesized proteins functionally related to mitochondria underpins the observed phenomenon. Given the evolutionary conservation of Drg proteins, our findings suggest that this mechanism is broadly shared across other living organisms. Scientific SignificanceMitochondria are essential organelles that not only generate ATP but also coordinate numerous cellular processes vital to cell survival and adaptation. Most mitochondrial proteins are synthesized in the cytoplasm and imported into the organelle. Our study reveals that the conserved ribosome-binding GTPase Drg1 facilitates translation of mitochondrial proteins at the outer mitochondrial membrane. By coupling cytoplasmic protein synthesis with mitochondrial function, Drg1 plays a critical role in maintaining mitochondrial morphology, dynamics, and functions. These findings provide new insights into the coordination between cytoplasmic translation, protein homeostasis, and organelle function, which informs future studies on diseases involving mitochondrial dysfunction and defective proteostasis.