Tau is an abundant microtubule-associated protein in neurons. Tau aggregation into insoluble fibrils is a hallmark of Alzheimer's disease and other dementias 1 , yet the physiological state of tau molecules within cells remains unclear. Using single molecule imaging, we directly observe that the microtubule lattice regulates reversible tau self-association, leading to localized, dynamic condensation of tau molecules on the microtubule surface. Tau condensates form selectively permissible barriers, spatially regulating the activity of microtubule severing enzymes and the movement of molecular motors through their boundaries. We propose that reversible selfassociation of tau molecules, gated by the microtubule lattice, is an important mechanism of tau's biological functions, and that oligomerization of tau is a common property shared between the physiological and disease forms of the molecule.

Tau is an abundant microtubule-associated protein in neurons. Tau aggregation into insoluble fibrils is a hallmark of Alzheimer's disease and other dementias, yet the physiological state of tau molecules within cells remains unclear. Using single molecule imaging, we directly observe that the microtubule lattice regulates reversible tau selfassociation, leading to dynamic condensation of tau molecules on the microtubule surface. Tau condensates form selectively permissible barriers, spatially regulating the activity of MT severing enzymes and the movement of molecular motors through their boundaries. We propose that reversible self-association of tau molecules, controlled by the microtubule, is an important mechanism of tau's biological functions, and that oligomerization of tau is a common property shared between the physiological and disease forms of the molecule.

KIF1A is a critical cargo transport motor within neurons. More than 100 known mutations result in KIF1A-associated neurological disorder (KAND), a degenerative condition for which there is no cure. A missense mutation, P305L, was identified in children diagnosed with KAND, but the molecular basis for the disease is unknown. We find that this conserved residue is part of an unusual 310 helix immediately adjacent to the family-specific K-loop, which facilitates a high microtubule-association rate. We find that the mutation negatively affects several biophysical parameters of the motor. However, the microtubule-association rate of the motor is most markedly affected, revealing that the presence of an intact K-loop is not sufficient for its function. We hypothesize that the 310 helix facilitates a specific K-loop conformation that is critical for its function. We find that the function of this proline is conserved in kinesin-1, revealing a fundamental principle of the kinesin motor mechanism.

KIF1A, a kinesin-3 family member, plays critical roles as a long-distance cargo-transporter within neurons. Over 100 known KIF1A mutations in humans result in KIF1A Associated Neurological Disease (KAND), developmental and degenerative neurological conditions for which there is no cure. A de novo missense mutation, P305L, was recently identified in several children diagnosed with KAND, but the underlying molecular basis for the disease phenotype is unknown. Interestingly, this residue is highly conserved in kinesin-family proteins, and together with adjacent conserved residues also implicated in KAND, forms an unusual 310-helical element immediately C-terminal to loop-12 (L12, also known as the K-loop in KIF1A) in the conserved kinesin motor core. In KIF1A, the disordered K-loop contains a highly charged insertion of lysines that is thought to endow the motor with a high microtubule-association rate. Here, we characterize the molecular defects of the P305L mutation in KIF1A using genetic, biochemical, and single-molecule approaches. We find the mutation negatively impacts the velocity, run-length, and force generation of the motor. However, a much more dramatic effect is observed on the microtubule-association rate of the motor, revealing that the presence of an intact K-loop is not sufficient for its function. We hypothesize that an elusive K-loop conformation, mediated by formation of a highly conserved adjacent 310-helix that is modulated via P305, is critically important for the kinesin-microtubule interaction. Importantly, we find that the function of this proline is conserved in the canonical kinesin, KIF5, revealing a fundamental principle of the kinesin motor mechanism.

ABD) preferentially engages actin in the presence of mechanical load across actin filaments (''mechanoaccumulation''), while vinculin's ABD does not. Simultaneous optical trapping and confocal microscopy experiments demonstrate that a load of 1pN across actin activates a-catenin ABD binding. Atomic-resolution cryo-EM structures of the metavinculin ABD-actin (2.9Å ) and a-catenin ABD-actin (3.2Å ) complexes demonstrate both ABDs undergo major conformational changes upon actin engagement, prominently at their N-and C-termini, and their C-terminal regions differentially refold to bind distinct sites on the filament surface. A C-terminal truncation of a-catenin's ABD constitutively binds actin regardless of force, and a chimeric protein of vinculin's ABD featuring a-catenin's flexible termini gains mechanoaccumulation activity, suggesting the a-catenin C-terminus-actin interaction is necessary and sufficient for mechanically regulated binding. This work, for the first time, establishes a force-regulated actin-binding mechanism in structural detail, and lays the groundwork for the rational design of therapeutics targeting cytoskeletal mechanotransduction pathways.