Luminal Localization of α-tubulin K40 Acetylation by Cryo-EM Analysis of Fab-Labeled Microtubules

Soppina, Virupakshi; Herbstman, Jeffrey F.; Skiniotis, Georgios; Verhey, Kristen J.

doi:10.1371/journal.pone.0048204

Cited by 98 publications

(91 citation statements)

References 47 publications

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“…40 Acetylation of the Lys40 residue in a-tubulin is a well-noted MT modification, and acetylated a-tubulin (Ac-tubulin) is enriched at interpolar and kinetochore MTs in mitotic cells. 24 In our research, we showed that depletion of EWSR1 reduced the level of MT acetylation but did not affect the levels of MT polyglutamylation and detyrosination in mitotic cells, indicating that EWSR1 has a specific role in the regulation of MT acetylation. Moreover, EWSR1 is distributed in the nucleus during interphase but is localized to spindle MTs during mitosis.…”

Section: Discussionmentioning

confidence: 52%

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EWSR1 regulates mitosis by dynamically influencing microtubule acetylation

et al. 2016

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EWSR1, participating in transcription and splicing, has been identified as a translocation partner for various transcription factors, resulting in translocation, which in turn plays crucial roles in tumorigenesis. Recent studies have investigated the role of EWSR1 in mitosis. However, the effect of EWSR1 on mitosis is poorly understood. Here, we observed that depletion of EWSR1 resulted in cell cycle arrest in the mitotic phase, mainly due to an increase in the time from nuclear envelope breakdown to metaphase, resulting in a high percentage of unaligned chromosomes and multipolar spindles. We also demonstrated that EWSR1 is a spindle-associated protein that interacts with a-tubulin during mitosis. EWSR1 depletion increased the cold-sensitivity of spindle microtubules, and decreased the rate of spindle assembly. EWSR1 regulated the level of microtubule acetylation in the mitotic spindle; microtubule acetylation was rescued in EWSR1-depleted mitotic cells following suppression of HDAC6 activity by its specific inhibitor or siRNA treatment. In summary, these results suggest that EWSR1 regulates the acetylation of microtubules in a cell cycledependent manner through its dynamic location on spindle MTs, and may be a novel regulator for mitosis progress independent of its translocation.

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Section: Discussionmentioning

confidence: 52%

“…24 Thus, we examined the effects of EWSR1 on a-tubulin acetylation. As shown in Figure 5A, cells transfected with control vector or Myc-EWSR1 were synchronized in mitosis with thymidine-MG132 treatment.…”

Section: Knockdown Of Ewsr1 Decreases A-tubulin Acetylationmentioning

confidence: 99%

EWSR1 regulates mitosis by dynamically influencing microtubule acetylation

et al. 2016

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“…Acetylated microtubules are generally thought to be more stable than unmodified microtubules, but no direct evidence has shown that acetylation modification of microtubules directly influences tubulin polymerization or depolymerization kinetics in vitro (18). In addition, no clear differences in microtubule structure or tubulin conformation have been found between electron cryomicroscopy reconstructions of maximally deacetylated or acetylated microtubules (19), and the levels at which kinesin-1 binds to acetylated microtubules and deacetylated microtubules are similar (20,21). These findings require further investigation into the mechanism by which acetylation of ␣-tubulin enhances the fusion of IBs of HPIV3 via interaction with the N-P complex.…”

Section: Discussionmentioning

confidence: 99%

Inclusion Body Fusion of Human Parainfluenza Virus Type 3 Regulated by Acetylated α-Tubulin Enhances Viral Replication

Zhang

Jiang

Cheng

et al. 2017

J Virol

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Viral inclusion bodies (IBs), or replication factories, are unique structures generated by viral proteins together with some cellular proteins as a platform for efficient viral replication, but little is known about the mechanism underlying IB formation and fusion. Our previous study demonstrated that the interaction between the nucleoprotein (N) and phosphoprotein (P) of human parainfluenza virus type 3 (HPIV3), an enveloped virus with great medical impact, can form IBs. In this study, we found that small IBs can fuse with each other to form large IBs that enhance viral replication. Furthermore, we found that acetylated ␣-tubulin interacts with the N-P complex and colocalizes with IBs of HPIV3 but does not interact with the N-P complex of human respiratory syncytial virus or vesicular stomatitis virus and does not colocalize with IBs of human respiratory syncytial virus. Most importantly, enhancement of ␣-tubulin acetylation using the pharmacological inhibitor trichostatin A (TSA), RNA interference (RNAi) knockdown of the deacetylase enzymes histone deacetylase 6 (HDAC6) and sirtuin 2 (SIRT2), or expression of ␣-tubulin acetyltransferase 1 (␣-TAT1) resulted in the fusion of small IBs into large IBs and effective viral replication. In contrast, suppression of acetylation of ␣-tubulin by overexpressing HDAC6 and SIRT2 profoundly inhibited the fusion of small IBs and viral replication. Our findings offer previously unidentified mechanistic insights into the regulation of viral IB fusion by acetylated ␣-tubulin, which is critical for viral replication.IMPORTANCE Inclusion bodies (IBs) are unique structures generated by viral proteins and some cellular proteins as a platform for efficient viral replication. Human parainfluenza virus type 3 (HPIV3) is a nonsegmented single-stranded RNA virus that mainly causes lower respiratory tract disease in infants and young children. However, no vaccines or antiviral drugs for HPIV3 are available. Therefore, understanding virus-host interactions and developing new antiviral strategies are increasingly important. Acetylation on lysine (K) 40 of ␣-tubulin is an evolutionarily conserved modification and plays an important role in many cellular processes, but its role in viral IB dynamics has not been fully explored. To our knowledge, our findings are the first to show that acetylated ␣-tubulin enhances viral replication by regulating HPIV3 IB fusion.KEYWORDS inclusion bodies, human parainfluenza virus type 3, acetylated ␣-tubulin H uman parainfluenza virus type 3 (HPIV3) is a nonsegmented single-stranded RNA virus (NSV) that belongs to the Paramyxoviridae family and mainly causes lower respiratory tract disease in infants and young children (1). However, no vaccines are available for HPIV3, and therefore, a deeper understanding of the replication of HPIV3

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“…A consecutive change in MT conformation might therefore enhance motor recruitment to MTs (Dompierre et al, 2007;Hammond et al, 2009;Reed et al, 2006). Conversely, the role of tubulin acetylation upon molecular motor recruitment and function has been questioned in motility assays using tubulin acetylated in vitro (Soppina et al, 2012;Walter et al, 2012). These studies proposed that the recruitment of kinesin-1 to acetylated MTs and its velocity were not affected, but other tubulin modifications (de-tyrosination and polyglutamylation) could have masked the effect of acetylation.…”

Section: Autophagy and Microtubules 1075mentioning

confidence: 99%

Autophagy and microtubules – new story, old players

Mackeh

Perdiz

Lorin

et al. 2013

Journal of Cell Science

192

181

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SummaryBoth at a basal level and after induction (especially in response to nutrient starvation), the function of autophagy is to allow cells to degrade and recycle damaged organelles, proteins and other biological constituents. Here, we focus on the role microtubules have in autophagosome formation, autophagosome transport across the cytoplasm and in the formation of autolysosomes. Recent insights into the exact relationship between autophagy and microtubules now point to the importance of microtubule dynamics, tubulin posttranslational modifications and microtubule motors in the autophagy process. Such factors regulate signaling pathways that converge to stimulate autophagosome formation. They also orchestrate the movements of pre-autophagosomal structures and autophagosomes or more globally organize and localize immature and mature autophagosomes and lysosomes. Most of the factors that now appear to link microtubules to autophagosome formation or to autophagosome dynamics and fate were identified initially without the notion that sequestration, recruitment and/or interaction with microtubules contribute to their function. Spatial and temporal coordination of many stages in the life of autophagosomes thus underlines the integrative role of microtubules and progressively reveals hidden parts of the autophagy machinery.

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Luminal Localization of α-tubulin K40 Acetylation by Cryo-EM Analysis of Fab-Labeled Microtubules

Cited by 98 publications

References 47 publications

EWSR1 regulates mitosis by dynamically influencing microtubule acetylation

EWSR1 regulates mitosis by dynamically influencing microtubule acetylation

Inclusion Body Fusion of Human Parainfluenza Virus Type 3 Regulated by Acetylated α-Tubulin Enhances Viral Replication

Autophagy and microtubules – new story, old players

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