This thesis examines three key phenomena: mitochondrial branching, transport, and pearling, to illustrate how membrane mechanics, cytoskeletal forces, and genome organization together shape mitochondrial behavior. By comparing tip-to-side fusion with motor-driven pulling, we show that mitochondria branch through physically distinct pathways that yield branches of differing persistence and geometries. We identify cristae voids as favorable sites for membrane pulling initiation as these regions present a lower elastic resistance to pulling forces. Moreover, we report the importance of microtubules and molecular motors in the frequency and stability of pulled branches. Using a photoconversion-based strategy, we then reveal how perinuclear mitochondria mix more extensively with the cellular network than their peripheral counterparts, implicating adaptor proteins such as TRAK2 in modulating
this spatial heterogeneity. Finally, experiments in yeast demonstrate that transient spontaneous pearling events maintain a regular inter-nucleoid spacing, a feature that could not be explained by the combination of fission and fusion. Across these studies, a unifying theme emerges: mitochondrial function depends on a delicate balance between structural flexibility, cytoskeletal interactions, inner structure, and the organization of mtDNA. Collectively, these
findings underscore the fine-tuned interplay between mitochondrial membrane mechanics, cytoskeletal forces, molecular motors, and metabolic cues in shaping mitochondrial architecture. These observations deepen our understanding of the ways in which mitochondria maintain their dynamic organization and underscore the importance of multiple regulatory layers in preserving mitochondrial integrity.