Bisig, G. | Search bioRxiv

bioRxiv2025

Kinesin drive meiotic chromosome dynamics via interaction with the KASH5-LINC complex

Telomere-led rapid prophase chromosome movements (RPMs) during meiotic prophase are critical for homologous chromosome pairing and proper meiotic progression. These movements are generated by the cytoskeleton and are transmitted to the telomeres via the LINC complex, yet the cytoplasmic components that generate these forces remain poorly defined. Among candidates of microtubule-associated motor proteins in mouse primary spermatocytes, we confirmed KIF5B as a specific interactor of the KASH5-LINC complex. Total internal reflection fluorescence microscopy and microtubule sedimentation assays performed with purified recombinant proteins suggest a direct interaction between KASH5 and KIF5B on microtubules, enhanced by MAP7, a known KIF5B-recruiting and activating cofactor. Mapping the KIF5B-binding surface of KASH5 revealed that KASH5 N-terminal EF-hand domains mediate the interaction. Further, in vivo KIF5B-KASH5 interaction and KIF5B role in RPMs are evidenced as (1) KIF5B is recruited by KASH5-SUN1 to the nuclear envelope in two different cultured somatic cell models, (2) KIF5B is telomere-associated and colocalizes with KASH5, and microtubules associated with the nuclear envelope in mouse spermatocytes, and (3) chemical inhibition of KIF5B reduces telomere-led chromosome motions. Altogether, our findings identify the KIF5B kinesin as a previously unrecognized component of the force-generating machinery that drives chromosome movement during meiotic prophase I, acting through KASH5 as a specific nuclear membrane adaptor.

cell biology↗

bioRxiv2024

L-Dopa incorporation into tubulin alters microtubule dynamics and reduces dendritic spine invasion and synapse maintenance

Previous studies have shown that L-Dopa, a tyrosine analog used in Parkinsons disease treatment, can be incorporated into -tubulin C-terminal tail via the tubulin tyrosine ligase (TTL) and polymerize into microtubules. In this work, we demonstrated that mature wild type hippocampal neurons treated with L-Dopa exhibited reduced dendritic spine density, primarily affecting mature dendritic spines. In these neurons, L-Dopa treatment significantly reduced tyrosinated -tubulin levels without altering detyrosinated or {Delta}2 -tubulin levels, suggesting the formation of a new tubulin pool, likely composed of L-Dopa--tubulin. In vitro analysis of the activity of the purified VASH1-SVBP complex, the most abundant tubulin carboxypeptidase in brain, revealed that L-Dopa incorporation into -tubulin modified the binding of the complex to microtubules and reduced its carboxypeptidase activity. These results suggest that L-Dopa incorporation into tubulin alters the properties of microtubules and affects their ability to interact with the enzyme. To confirm the implication of L-Dopa-microtubules in dendritic spine alterations observed in wild type neurons, we analyzed the effect of L-Dopa treatment in neurons lacking the enzymes of the -tubulin detyrosination/tyrosination cycle. In these cells, L-Dopa cannot be incorporated into -tubulin due to the absence of the ligase (in TTL KO neurons) or the reduction of detyrosinated -tubulin levels (in SVBP KO neurons). L-Dopa treatment did not modify dendritic spine density in TLL KO or SVBP KO neurons, clearly demonstrating that the alterations in dendritic spines seen in WT neurons are due to the incorporation of L-Dopa into tubulin. Further analysis revealed that L-Dopa treatment decreased the percentage of spines containing excitatory synapses in wild type neurons, but not in TTL KO or SVBP KO neurons, suggesting a cumulative synaptic defect due to L-Dopa incorporation into microtubules. Additionally, L-Dopa altered microtubule dynamics by increasing catastrophe frequency and reducing comet lifetime, which led to fewer microtubules entering dendritic spines and decreased spine resistance to pruning. Taken together, our results demonstrate that L-Dopa incorporation into -tubulin drastically affects synaptic homeostasis, reaffirming the importance of balanced detyrosination/tyrosination of tubulin within the synaptic compartment. The abnormal dynamics of L-Dopa-microtubules and the reduction of dendritic spines and excitatory synapses highlight a novel mechanism of L-Dopa-induced synaptotoxicity.

neuroscience↗