Multiple evolutionary routes to cytoskeletal arborization revealed by the rhizarian amoeba Filoreta ramosa.

The eukaryotic cytoskeleton generates remarkable diversity in cellular architecture despite being built from conserved actin and tubulin polymers. Diversification of cytoskeletal regulators, motors, and filament-organizing proteins produces varied morphologies, yet certain architectures repeatedly emerge in distantly related lineages. One notable example is cytoskeletal arborization, occurring in metazoan neurons and amoeboid lineages throughout the eukaryotic tree. Whether these similar branched architectures arise through conserved cytoskeletal organization, shared molecular reuse, or convergence driven by common physical constraints remains unresolved. Here, we investigate the rhizarian amoeba Filoreta ramosa, which forms a multinucleate reticulated network through branching and anastomosis. Using live imaging, immunofluorescence, morphometric analyses, and cytoskeletal drugs, we define how actin and microtubule systems organize branch formation, intracellular transport, and large-scale network architecture. Actin-rich protrusions initiate exploratory branchlets that become selectively stabilized through microtubule incorporation. Longitudinal microtubule arrays reinforce mature branches and support rapid bidirectional organelle transport, while branch nodes function as distributed sites of microtubule nucleation. These cytoskeletal features parallel key mechanisms underlying neuronal arborization, yet unlike neurons, Filoreta develops a decentralized reticulated network through repeated anastomosis and distributed microtubule organization, thereby scaling transport capacity and structural support with network growth. Our findings suggest that shared cytoskeletal mechanisms repeatedly support branching architectures, while distinct topologies and modes of organization allow evolution to reach arborization through different routes as increasing cellular scale imposes common demands on transport and structural integrity. Filoreta therefore provides a tractable model for investigating how conserved cytoskeletal systems generate arborized architectures that scale with cellular size.