Altered Metabolism in Heterozygous Mice With a Mutation in the Motor Domain of Cytoplasmic Dynein, DYNC1H1 (AAA4;...

DYNC1H1 encodes the cytoplasmic dynein heavy chain, a key motor protein involved in intracellular transport and neural development. While mutations in DYNC1H1 are known to cause a range of neurodevelopmental and motor disorders, the molecular mechanisms linking genotype to phenotype remain poorly defined. Here, we investigated how a patient-derived missense mutation in the motor domain of DYNC1H1 (c.9052C>T; P3018S) affects brain metabolism and behavior using a heterozygous knock-in mouse model. Behavioral phenotyping revealed increased locomotor activity without significant changes in sociability or novelty-seeking behavior. To uncover potential molecular correlates of this hyperactive phenotype, we performed label-free quantitative proteomics on cerebrum and cerebellum tissue from male and female mice. Over 80 mitochondrial proteins exhibited differential abundance in HET mice relative to WT controls, particularly in pathways related to oxidative phosphorylation and carbohydrate metabolism. These proteomic signatures were more pronounced in the cerebrum and showed sex-specific patterns. Our findings support the hypothesis that dysregulated mitochondrial metabolism contributes to the behavioral phenotype observed in DYNC1H1 HET mice, and they provide a molecular framework for understanding how cytoplasmic dynein mutations may lead to neurodevelopmental disorders. SUMMARY: Cytoplasmic dynein is a multi-subunit motor protein complex essential for intracellular cargo transport and proper neural function. Mutations in DYNC1H1 are increasingly recognized as a cause of pediatric motor and cognitive disorders, yet the molecular underpinnings of these phenotypes are not fully understood. The P3018S mutation in DYNC1H1 disrupts the motor domain and has been associated with abnormal neuronal migration, cortical malformations, and developmental delay. In this study, we used a knock-in mouse model carrying the P3018S mutation to explore how dynein dysfunction affects brain metabolism and behavior. Our proteomic analysis of cerebrum and cerebellum samples revealed widespread differential abundance of mitochondrial proteins, particularly those involved in oxidative phosphorylation and carbohydrate metabolism. These findings align with prior behavioral and metabolic observations of increased activity and energy expenditure in heterozygous mice. By stratifying data by sex and brain region, we identified molecular patterns that may underlie the distinct neurobehavioral profiles observed in DYNC1H1-related disorders. This study provides a valuable resource linking genotype, proteomic phenotype, and behavior, and lays the groundwork for future therapeutic strategies aimed at modulating mitochondrial metabolism in affected individuals.