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Profilin 1 (PFN1) is an actin binding protein that is vital for the polymerization of monomeric actin into filaments. Here we screened knockout cells for novel functions of PFN1 and discovered that mitophagy, a type of selective autophagy that removes defective or damaged mitochondria from the cell, was significantly upregulated in the absence of PFN1. Despite successful autophagosome formation and fusion with the lysosome, and activation of additional mitochondrial quality control pathways, PFN1 knockout cells still accumulate damaged, dysfunctional mitochondria. Subsequent imaging and functional assays showed that loss of PFN1 significantly affects mitochondria morphology, dynamics, and respiration. Further experiments revealed that PFN1 is located to the mitochondria matrix and is likely regulating mitochondria function from within rather than through polymerizing actin at the mitochondria surface. Finally, PFN1 mutants associated with amyotrophic lateral sclerosis (ALS) fail to rescue PFN1 knockout mitochondrial phenotypes and form aggregates within mitochondria, further perturbing them. Together, these results suggest a novel function for PFN1 in regulating mitochondria and identify a potential pathogenic mechanism of ALS-linked PFN1 variants., Competing Interests: Declaration of Interests The authors declare no competing interests.

Microtubules, intermediate filaments, and actin are cytoskeletal polymer networks found within the cell. While each has unique functions, all the cytoskeletal elements must work together for cellular mechanics to be fully operative. This is achieved through crosstalk mechanisms whereby the different networks influence each other through signaling pathways and direct interactions. Because crosstalk can be complex, it is possible for perturbations in one cytoskeletal element to affect the others in ways that are difficult to predict. Here we investigated how long-term changes to the actin cytoskeleton affect microtubules and intermediate filaments. Reducing F-actin or actomyosin contractility increased acetylated microtubules and intermediate filament expression, with the effect being significantly more pronounced in neuronal processes. Changes to microtubules were completely reversible if F-actin and myosin activity is restored. Moreover, the altered microtubules in neuronal processes resulting from F-actin depletion caused significant changes to microtubule-based transport, mimicking phenotypes that are linked to neurodegenerative disease. Thus, defects in actin dynamics cause a compensatory response in other cytoskeleton components which profoundly alters cellular function., Competing Interests: Declaration of Interests The authors declare no competing interests.

Directed transport of cellular components is often dependent on the processive movements of cytoskeletal motors. Myosin 2 motors predominantly engage actin filaments of opposing orientation to drive contractile events, and are therefore not traditionally viewed as processive. However, recent in vitro experiments with purified non-muscle myosin 2 (NM2) demonstrated myosin 2 filaments could move processively. Here, we establish processivity as a cellular property of NM2. Processive runs in central nervous system-derived CAD cells are most apparent as processive movements on bundled actin in protrusions that terminate at the leading edge. We find that processive velocities in vivo are consistent with in vitro measurements. NM2 makes these processive runs in its filamentous form against lamellipodia retrograde flow, though anterograde movement can still occur in the absence of actin dynamics. Comparing the processivity of NM2 isoforms, we find that NM2A moves slightly faster than NM2B. Finally, we demonstrate that this is not a cell-specific property, as we observe processive-like movements of NM2 in the lamella and subnuclear stress fibers of fibroblasts. Collectively, these observations further broaden NM2 functionality and the biological processes in which the already ubiquitous motor can contribute.