ATAD3A Knockout Cell Line (HEK293)

The ATPase Family AAA Domain Containing 3A (ATAD3A), is a mitochondrial inner membrane protein conserved in metazoans. ATAD3A has been associated with several mitochondrial functions, including nucleoid organization, cholesterol metabolism, and mitochondrial translation. To address its primary role, we generated a neuronal-specific conditional knockout (Atad3 nKO) mouse model, which developed a severe encephalopathy by 5 months of age. Pre-symptomatic mice showed aberrant mitochondrial cristae morphogenesis in the cortex as early as 2 months. Using a multi-omics approach in the CNS of 2-to-3-month-old mice, we found early alterations in the organelle membrane structure. We also show that human ATAD3A associates with different components of the inner membrane, including OXPHOS complex I, Letm1, and prohibitin complexes. Stochastic Optical Reconstruction Microscopy (STORM) shows that ATAD3A is regularly distributed along the inner mitochondrial membrane, suggesting a critical structural role in inner mitochondrial membrane and its organization, most likely in an ATPase-dependent manner.

Presenilin 1 (PS1), a key pathogenic factor in familial Alzheimer's disease, is implicated in the regulation of mitochondrial functions, yet its precise sub-mitochondrial localization and underlying mechanisms remain poorly understood. In this study, we generate knockout cell lines to investigate the role of PS1 in mitochondrial structure and function. Our results indicate that PS1 is directly localized in the mitochondrial inner membrane. deficiency leads to reduced ATP production, impaired mitochondrial respiration capacity, decreased mitochondrial membrane potential, disrupted Ca homeostasis, and elevated ROS accumulation. Moreover, loss of leads to abnormal mitochondrial cristae structure. Further analysis reveals that PS1 interacts with mitochondrial inner membrane proteins. Its absence promotes ATAD3A oligomerization and disrupts its arrangement at mitochondrial cristae junctions, leading to expansion of the mitochondria-associated membrane and instability of mitochondrial DNA. Our findings demonstrate that PS1 acts as a central regulator of mitochondrial cristae morphogenesis by modulating protein interaction networks at cristae junctions, thereby illuminating fundamental molecular mechanisms contributing to mitochondrial dysfunction in Alzheimer's disease.