Quantitative Comparison of Monomeric StayGold Variants Using Protein Nanocages in Living Cells DOI Creative Commons

Giulia Viola,

Kyle A. Jacobs, Joël Lemière

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Sept. 17, 2024

Abstract To standardize comparison of fluorescent proteins and independently determine which monomeric StayGold variant is best for live microscopy, we analyzed protein tagged I3-01 peptides that self-assemble into stable sixty subunit dodecahedrons inside cells. We find mStayGold 3-fold brighter more photostable compared with EGFP superior to other variants in mammalian cytoplasm. In addition, analysis intracellular nanocage diffusion confirms the nature mStayGold.

Language: Английский

Doublecortin restricts neuronal branching by regulating tubulin polyglutamylation DOI Creative Commons
Muriel Sébastien, Alexandra Paquette, Emily N. P. Prowse

et al.

Nature Communications, Journal Year: 2025, Volume and Issue: 16(1)

Published: Feb. 18, 2025

Doublecortin is a neuronal microtubule-associated protein that regulates microtubule structure in neurons. Mutations cause lissencephaly and subcortical band heterotopia by impairing migration. We use CRISPR/Cas9 to knock-out the gene induced pluripotent stem cells differentiate into cortical DCX-KO neurons show reduced velocities of nuclear movements an increased number neurites early development, consistent with previous findings. Neurite branching regulated host proteins, as well polymerization dynamics. However, EB comet dynamics are unchanged Rather, we observe significant reduction α-tubulin polyglutamylation Polyglutamylation levels rescued expression or TTLL11, glutamylase. Using U2OS orthogonal model system, DCX TTLL11 act synergistically promote polyglutamylation. propose acts positive regulator restricts neurite branching. Our results indicate unexpected role for homeostasis tubulin code. Lissencephaly severe neurodevelopmental disease often caused mutations Dcx gene. human cellular lissencephaly, authors report activating

Language: Английский

Citations

1
Diverse microtubule-binding repeats regulate TPX2 activities at distinct locations within the spindle DOI
Zhuobi Liang, Junjie Huang, Yong Wang

et al.

The Journal of Cell Biology, Journal Year: 2025, Volume and Issue: 224(3)

Published: Jan. 16, 2025

TPX2 is an elongated molecule containing multiple α-helical repeats. It stabilizes microtubules (MTs), promotes MT nucleation, and essential for spindle assembly. However, the molecular basis of how performs these functions remains elusive. Here, we systematically characterized MT-binding activities all modules individually in combinations investigated their respective contributions both vitro cells. We show that contains repeats with opposite preferences “extended” “compacted” tubulin dimer spacing, distinct produce divergent outcomes, making activity highly robust yet tunable. Importantly, a repeat group at C terminus, R8-9, key determinant function. MTs by promoting rescues critical propose model where are spatially regulated via its diverse to accommodate varied locations within spindle. Furthermore, reveal synergy between HURP stabilizing MTs.

Language: Английский

Citations

0
Rethinking tubulin acetylation: From regulation to cellular adaptation DOI Creative Commons
Lisa Donker, Susana A. Godinho

Current Opinion in Cell Biology, Journal Year: 2025, Volume and Issue: 94, P. 102512 - 102512

Published: April 14, 2025

Language: Английский

Citations

0
Systematic Identification of Microtubule Inner Proteins Reveals JPT2 as a Key Regulator of Lumen Microenvironment and Drug Sensitivity DOI Open Access
Jinhui Shao,

Rui Zhang,

Peiyuan Wu

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2025, Volume and Issue: unknown

Published: Jan. 13, 2025

Abstract Microtubules are primarily studied for the interactions of proteins that bind to their outer surfaces and ends, while regulatory mechanisms within microtubule lumen, particularly in singlet microtubules critical essential cellular processes, remain largely unexplored. Our study provides first systematic identification key single lumen. Using proximity-dependent biotin (Bio-ID) coupled with mass spectrometry, we identified candidate inner (MIPs), including Jupiter microtubule-associated homolog 2 (JPT2). JPT2 binds directly specifically localizes where it modulates luminal environment by inhibiting acetylase MEC17 independently affects binding efficacy Paclitaxel. Furthermore, our screening additional MIPs influence sensitivity Paclitaxel, indicating a link between regulation drug responsiveness. These discoveries reveal JPT2’s role suggest new therapeutic targets enhancing cancer sensitivity.

Language: Английский

Citations

0
How does the tubulin code facilitate directed cell migration? DOI Creative Commons

Miguel Marques Simoes-da-Silva,

Marin Barišić

Biochemical Society Transactions, Journal Year: 2025, Volume and Issue: 53(01)

Published: Feb. 21, 2025

Besides being a component of the cytoskeleton that provides structural integrity to cell, microtubules also serve as tracks for intracellular transport. As building units mitotic spindle, distribute chromosomes during cell division. By distributing organelles, vesicles, and proteins, they play pivotal role in diverse cellular processes, including migration, which reorganize facilitate polarization. Structurally, are built up α/β-tubulin dimers, consist various tubulin isotypes undergo multiple post-translational modifications (PTMs). These PTMs allow differentiate into functional subsets, influencing associated processes. This text explores current understanding roles particularly detyrosination acetylation, their implications human diseases.

Language: Английский

Citations

0
Microtubule acetylation is a biomarker of cytoplasmic health during cellular senescence DOI Creative Commons
Yusheng Shen, Roilea Maxson, Richard J. McKenney

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2025, Volume and Issue: unknown

Published: April 1, 2025

Cellular senescence is marked by cytoskeletal dysfunction, yet the role of microtubule post-translational modifications (PTMs) remains unclear. We demonstrate that acetylation increases during drug-induced in human cells and natural aging Drosophila . Elevating via HDAC6 inhibition or α TAT1 overexpression BEAS-2B disrupts anterograde Rab6A vesicle transport, but spares retrograde transport Rab5 endosomes. Hyperacetylation results slowed polymerization decreased cytoplasmic fluidity, impeding diffusion micron-sized condensates. These effects are distinct from enhanced detyrosination, correlate with altered viscoelasticity resistance to osmotic stress. Modulating viscosity reciprocally perturbs dynamics, revealing bidirectional mechanical regulation. Senescent phenocopy hyperacetylated cells, exhibiting analogous on polymerization. Our findings establish as a biomarker for health potential driver age-related densification organelle decline, linking PTMs biomechanical feedback loops exacerbate senescence. This work highlights bridging changes broader hallmarks.

Language: Английский

Citations

0
Acetylation of α-tubulin restores endothelial cell injury and blood–brain barrier disruption after intracerebral hemorrhage in mice DOI Creative Commons

Xuejiao Lei,

Eryi Sun, Xufang Ru

et al.

Experimental & Molecular Medicine, Journal Year: 2025, Volume and Issue: unknown

Published: May 7, 2025

Abstract Damage to endothelial cells (ECs) is a key factor in blood–brain barrier (BBB) disruption after intracerebral hemorrhage (ICH). While microtubules are essential for EC structure, their role BBB injury remains unclear. Here we investigated the of acetylated α-tubulin (α-Ac-Tub) integration ICH. Using an autologous blood injection model striatum, showed that expression α-Ac-Tub and MEC17, acetyltransferase, significantly decreased along vessels around hematoma Conditional MEC17 knockout ECs further reduced levels exacerbated leakage, brain edema, expansion, inflammation motor dysfunction. Conversely, selective upregulation via intravenous delivery AAV-BI30-MEC17-GFP alleviated dysfunction improved recovery. Similarly, HDAC6 inhibitor tubastatin A enhanced levels, mitigating damage neurological deficits. Mechanistically, deficiency tight junction proteins (ZO-1 Claudin5) increased F-actin stress fibers through RhoA activation. Together, our findings highlighted as therapeutic target restoring function reducing

Language: Английский

Citations

0
Doublecortin restricts neuronal branching by regulating tubulin polyglutamylation DOI Creative Commons
Muriel Sébastien, Alexandra Paquette, Emily N. P. Prowse

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2023, Volume and Issue: unknown

Published: June 3, 2023

Doublecortin (DCX) is a neuronal microtubule-associated protein (MAP) that binds directly to microtubules via two (DC) domains. The DC domains sense the nucleotide state, longitudinal curvature, and protofilament number of microtubule lattice, indicating role in regulation structure neurons. Mutations DCX cause lissencephaly subcortical band heterotopia (also known as double-cortex syndrome) due impaired migration. To better understand migration, we developed model system based on induced pluripotent stem cells (iPSCs). We used CRISPR/Cas9 knock-out Dcx gene iPSCs differentiated into cortical Compared control neurons, DCX-KO neurons showed reduced velocities nuclear movements. coincided with an increase neurites early development, consistent migration phenotype previous findings mouse model. Neurite branching regulated by host MAPs other factors, well polymerization dynamics. However, EB comet dynamics were unchanged similar growth rates, lifetimes, numbers. Rather, observed significant reduction α-tubulin polyglutamylation Polyglutamylation levels rescued expression or TTLL11, glutamylase. Using U2OS orthogonal system, show TTLL11 act synergistically promote polyglutamylation. regulates numerous MAPs, severing enzymes, molecular motors. Consistently, observe lysosomes their processivity. propose acts positive regulator restricts neurite branching. Our results indicate unexpected for homeostasis tubulin code.

Language: Английский

Citations

4
Excess microtubule and F-Actin formation mediates primary cilia shortening and loss in response to hyperosmotic milieu DOI Creative Commons
H. Otani, Ryota Nakazato,

Kanae Koike

et al.

Journal of Cell Science, Journal Year: 2024, Volume and Issue: 137(16)

Published: Aug. 15, 2024

ABSTRACT The primary cilium is a small organelle protruding from the cell surface that receives signals extracellular milieu. Although dozens of studies have reported several genetic factors can impair structure cilia, evidence for environmental stimuli affecting cilia structures limited. Here, we investigated an stress affected morphology and its underlying mechanisms. Hyperosmotic shock induced reversible shortening disassembly murine intramedullary collecting duct cells. caused by hyperosmotic followed delocalization pericentriolar material (PCM). Excessive microtubule F-actin formation in cytoplasm coincided with shock-induced changes to PCM. Treatment microtubule-disrupting agent, nocodazole, partially prevented almost completely An actin polymerization inhibitor, latrunculin A, also We demonstrate induces morphological PCM manner dependent on excessive F-actin.

Language: Английский

Citations

1
Microtubule-associated protein, MAP1B, encodes functionally distinct polypeptides DOI Creative Commons

Tracy Tan,

Yusheng Shen,

Lily B Stine

et al.

Journal of Biological Chemistry, Journal Year: 2024, Volume and Issue: unknown, P. 107792 - 107792

Published: Sept. 1, 2024

Language: Английский

Citations

1