Cells depend on a complex and precisely regulated subcellular organization, largely driven by the cytoskeleton and motor proteins that control intracellular transport. This review explores innovative strategies to manipulate cellular architecture using targeted protein design and engineering of cytoskeletal elements and molecular motors. We highlight advances in inducible dimerization techniques, which enable precise control over cytoskeletal dynamics through light- and small-molecule-sensitive domains. In addition, we discuss modifications to motor proteins that alter directionality, processivity, and cargo specificity, providing insights into their roles in cellular transport. Rapid advances in de novo protein design offer new tools to hijack natural cytoskeletal machinery and create synthetic elements for cellular architecture, including membraneless organelles and synthetic cytoskeletal tracks. This research promises to deepen our understanding of cellular organization, uncover regulatory mechanisms, and provide new proteins for therapeutic applications and synthetic cell development.