Biology subjects

Egoldt, C.

Publications and source records attributed to Egoldt, C..

4 recordsLinked to original sources

bioRxiv2026

A biochemical probe for microtubule lattice integrity uncovers motor-caused lattice damage

Microtubules experience mechanical and enzymatic stresses that can compromise lattice integrity, yet where and how lattice damage forms remains poorly understood due to the lack of tools that directly visualize damage as it occurs. Existing approaches infer damage indirectly through repair events or rely on static ultrastructural snapshots, precluding dynamic analysis. Here, we introduce MT-DS (Microtubule Damage Sensor), a damage-selective fluorescent probe that directly labels microtubule lattice openings. MT-DS combines taxane-based microtubule binding with a multivalent protein scaffold to restrict intraluminal diffusion and selectively retain the probe at sites of lattice openings. Using MT-DS, we visualize intrinsic lattice defects in stabilized microtubules, uncover a strong enrichment of damage at annealing sites, and demonstrate that kinesin-1{Delta}6 actively generates de novo lattice damage during motility. By enabling direct, time-resolved detection of microtubule damage, MT-DS establishes lattice integrity as an experimentally accessible parameter and provides a chemical tool to investigate how mechanical stress reshapes the microtubule cytoskeleton.

biochemistry↗

bioRxiv2025

Microtubule lattice conformation and integrity regulate α-tubulin acetylation

Microtubule acetylation of lysine 40 of -tubulin is a hallmark of stable microtubules. This luminal modification is catalyzed by -tubulin acetyltransferase 1 (TAT1) and reversed by histone deacetylase 6 (HDAC6). However, acetylation regulation within the microtubule lumen and the influence of lattice architecture on enzymatic activity remain poorly understood. Here, we reconstitute microtubule acetylation in vitro using purified TAT1 and HDAC6 on microtubules assembled with defined lattice conformations. We show that TAT1 overwrites HDAC6 enzymatic activity, but its acetylation efficiency decreases upon microtubule damage. Importantly, TAT1 efficiently acetylates microtubules in expanded lattices and twisted tubulin states, while compacted lattices impede its activity. Our findings reveal that both microtubule integrity and lattice conformation are critical regulators for TAT1 enzymatic activity, suggesting that dynamic lattice transitions modulate the acetylation pattern of microtubules in cells.

cell biology↗

bioRxiv2024

SNAP-tag2: faster and brighter protein labeling

SNAP-tag is a powerful tool for labeling proteins with synthetic fluorophores in bioimaging. However, its utility in live-cell applications can be constrained by its relatively slow labeling kinetics and the limited cell permeability of its substrates. Here we introduce new labeling substrates and an engineered SNAP-tag for faster labeling in vitro and in live cells. SNAP-tag2 presents a second-order rate constant with rhodamine substrates that approaches 107 s-1 M-1, a 100-fold improvement over the corresponding SNAP-tag-substrate pairs. When labeled with highly fluorogenic dyes, SNAP-tag2 also shows a 5-fold increase in fluorescence brightness relative to currently used SNAP-tag. The increased labeling kinetics and brightness of SNAP-tag2 translates into a greatly improved performance in various live-cell (super-resolution) imaging applications.

biochemistry↗

bioRxiv2022

Running Kinesin-1 shapes the microtubule acetylation gradient

The properties of single microtubules within the microtubule network can be modulated through posttranslational modifications (PTMs), including acetylation within the lumen of microtubules. To access the lumen, the enzymes could either enter through the microtubule ends or at damage sites along the microtubule shaft. Here we show that the acetylation profile depends on damage sites, which can be caused by the motor protein kinesin-1. Indeed, the entry of the deacetylase HDAC6 into the microtubule lumen depends on kinesin-1-induced damage sites. In contrast, activity of the microtubule acetylase TAT1 is independent of kinesin-1 and shaft damage. On a cellular level, our results show that microtubule acetylation distributes in an exponential gradient. This gradient results from tight regulation of microtubule (de-)acetylation and scales with the size of the cells. The control of shaft damage represents a novel mechanism to regulate PTM inside the microtubule by giving access to the lumen.

cell biology↗