Biology subjects

Slivka, J.

Publications and source records attributed to Slivka, J..

3 recordsLinked to original sources

bioRxiv2026

Microtubule Lattice Spacing Governs MAP-Motor Regulation

Microtubules (MTs) serve as intracellular tracks that enable molecular motors to transport cargos to specific cellular destinations. It has been proposed that the signals directing motor-driven transport are encoded on MTs through different isotypes, lattice conformations, and post- translational modifications (PTMs) of tubulin, or MT-associated proteins (MAPs) that decorate the MT surface. However, molecular predictions of these models have not been rigorously tested in vitro. Using isotypically pure recombinant tubulin and biochemical reconstitution, we examined how tubulin PTMs and MT lattice spacing influence MAP binding and kinesin-1 motility. We found that kinesin-1 is largely insensitive to tubulin PTMs but is strongly regulated by MT lattice spacing. Likewise, the MAPs tau, MAP7, MAP4, DCX, and MAP9 exhibit little sensitivity to tubulin PTMs, whereas the MT-binding affinities of tau, DCX, and MAP7 depend on lattice spacing. In the presence of activating (MAP7) and inhibitory (tau) MAPs, lattice spacing determines MAP occupancy and thereby controls kinesin-1 motility. These findings support a two-layer transport code in which MT lattice spacing directs MAP binding, and MAPs determine which motors can move along individual MT tracks.

biochemistry↗

bioRxiv2024

Stepping dynamics of dynein characterized by MINFLUX

Cytoplasmic dynein is the principal motor for minus-end-directed motility and force generation functions along microtubules (MTs)1. Dynein converts the chemical energy of ATP hydrolysis into coordinated structural changes to step processively along MTs, but how dynein couples ATP hydrolysis to a minus-end-directed step remains controversial2, 3. The dynamics of dynein stepping have previously been characterized by tracking flexible regions of the motor with limited resolution4-6. Here, we site-specifically labeled yeast dynein at its MT-binding domain by developing a cysteine-light mutant and tracked its stepping at sub-millisecond and nanometer resolution at physiological ATP concentrations using MINFLUX7. We show that dynein hydrolyzes one ATP per step and takes multiples of 8 nm steps. Steps are preceded by a transient movement towards the plus end. These backward "dips" correspond to MT release upon ATP binding and subsequent diffusion of the stepping monomer around its MT-bound partner. Functional assays showed that dips terminate with a minus-end-directed movement upon ATP hydrolysis. These results provide critical insights into the order of mechanochemical events that result in a productive step of dynein.

biophysics↗

bioRxiv2019

Mechanism of processive telomerase catalysis revealed by high-resolution optical tweezers

Telomere maintenance by telomerase is essential for continuous proliferation of human cells and is vital for the survival of stem cells and 90% of cancer cells. To compensate for telomeric DNA lost during DNA replication, telomerase processively adds GGTTAG repeats to chromosome ends by copying the template region within its RNA subunit. Between repeat additions, the RNA template must be recycled. How telomerase remains associated with substrate DNA during this critical translocation step remains unknown. Using a newly developed single-molecule telomerase activity assay utilizing high-resolution optical tweezers, we demonstrate that stable substrate DNA binding at an anchor site within telomerase facilitates the processive synthesis of telomeric repeats. After release of multiple telomeric repeats from telomerase, we observed folding of product DNA into G-quadruplex structures. Our results provide detailed mechanistic insights into telomerase catalysis, a process of critical importance in aging and cancer.

biochemistry↗