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SETD2 is highly expressed during cortical development and required for neuronal migration. a qPCR showed that the relative level of SETD2 mRNA was high in embryonic and early post-natal mouse cerebral cortex. All data were normalized to the P60 cerebral cortex and represented as the mean ± s.e.m. (n = 3 biological replicates). b Immunostaining showed a distribution of SETD2 in CP, IZ and VZ/SVZ of mouse somatosensory cortex at E14 and E16. Brain slices were stained for SETD2 (red), Tuj1 (green) and DAPI (blue) (n = 3 biological replicates). Scale bar: 25 μm. c SETD2 was localized in the NPCs of VZ/SVZ and the neurons of CP at E14 cerebral cortex. Dashed line indicated the lineament of cell and arrowheads indicated the SETD2 puncta in the cytoplasm. Brain slices were stained for SETD2 (red), Tuj1 (green) and DAPI (blue) (n = 3 biological replicates). Scale bar: 2.5 μm. d Representative images of coronal brain sections in somatosensory cortex at E16 showed the distribution of GFP + cells (green) electroporated with GFP reporter as well as control shRNA (shNC), mSETD2 shRNA1 or mSETD2 shRNA2 at E14. Sections were stained for DAPI (blue). Scale bar: 25 μm. e Quantitative analysis of d showed that the percentage of GFP + cells in IZ was significantly increased after SETD2 knockdown. All data were shown as the mean ± s.e.m. (n = 17, 10, 11, respectively). Data were analyzed using two-way ANOVA with Bonferroni's post-hoc test. ***P < 0.001 versus shNC. f Representative images of shNC and shRNA2-expressing neurons (green) at E16 in the IZ. Brain slices were stained for DAPI (blue). Scale bar: 10 μm. g Quantitative analysis of f showed that the percentage of GFP + neurons at the multipolar stage in IZ was significantly increased after SETD2 knockdown. All data were shown as the mean ± s. e.m. (n = 12, 16, respectively). Data were analyzed using two-way ANOVA with Bonferroni's post-hoc test. ***P < 0.001 versus shNC. Source data are provided as a Source Data file.
Source publication
Tri-methylation on lysine 40 of α-tubulin (α-TubK40me3) is a recently identified post-translational modification involved in mitosis and cytokinesis. However, knowledge about α-TubK40me3 in microtubule function and post-mitotic cells remains largely incomplete. Here, we report that α-TubK40me3 is required for neuronal polarization and migration by...
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... for the brain slices and 5 μm for the upper-right high magnification. Source data are provided as a Source Data file. SETD2 was continuously increased from E12 and reached a peak at E16, which was corresponding to the high level of α-TubK40me3 at E14 and E16. The SETD2 mRNA remained at a high level until P3 and afterwards was gradually decreased (Fig. 2a). Immunostaining showed that SETD2 was expressed both in the VZ/SVZ, IZ and CP of somatosensory cortex at E14 and E16 (Fig. 2b), consistent with the distribution pattern of α-TubK40me3 at the same stages (Fig. 1c). In detail, SETD2 puncta were not only localized in the nucleus but also in the cytoplasm of cells in VZ/SVZ and CP (Fig. ...
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... continuously increased from E12 and reached a peak at E16, which was corresponding to the high level of α-TubK40me3 at E14 and E16. The SETD2 mRNA remained at a high level until P3 and afterwards was gradually decreased (Fig. 2a). Immunostaining showed that SETD2 was expressed both in the VZ/SVZ, IZ and CP of somatosensory cortex at E14 and E16 (Fig. 2b), consistent with the distribution pattern of α-TubK40me3 at the same stages (Fig. 1c). In detail, SETD2 puncta were not only localized in the nucleus but also in the cytoplasm of cells in VZ/SVZ and CP (Fig. 2c), supporting its role in catalyzing α-TubK40me3 and fulfilling the non-chromatin function in these ...
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... decreased (Fig. 2a). Immunostaining showed that SETD2 was expressed both in the VZ/SVZ, IZ and CP of somatosensory cortex at E14 and E16 (Fig. 2b), consistent with the distribution pattern of α-TubK40me3 at the same stages (Fig. 1c). In detail, SETD2 puncta were not only localized in the nucleus but also in the cytoplasm of cells in VZ/SVZ and CP (Fig. 2c), supporting its role in catalyzing α-TubK40me3 and fulfilling the non-chromatin function in these ...
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... we addressed whether the high level of α-TubK40me3 in this period was involved in the regulation of neuronal morphology and migration. The levels of SETD2 and α-TubK40me3 were dramatically decreased in Neuro-2a cells expressing short-hairpin-interfering (sh) RNAs for mouse SETD2 (mSETD2) (Extended Data Fig. 2a, b), suggesting that acute knockdown of SETD2 by shRNA is a practical strategy to study α-TubK40me3-related functions. ...
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... utero electroporation (IUE), mixed plasmids containing shRNA and GFP reporter were co-transfected to radial glia progenitors in the cerebral cortex of E14 mice. Immunostaining showed that the levels of SETD2 and α-TubK40me3 were significantly decreased in GFP-positive (GFP + ) SETD2-deficient neurons of somatosensory cortex at E16 (Extended Data Fig. 2c-f). The level of H3K36me3 was also reduced in GFP + SETD2-deficient neurons though the extent of reduction was smaller than α-TubK40me3 (Extended Data Fig. 2e, f). The distribution of GFP + cells in somatosensory cortex, which was divided into CP, IZ, and VZ/SVZ, was analyzed at E16 (Fig. 2d). Approximate 27% of GFP + control cells ...
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... showed that the levels of SETD2 and α-TubK40me3 were significantly decreased in GFP-positive (GFP + ) SETD2-deficient neurons of somatosensory cortex at E16 (Extended Data Fig. 2c-f). The level of H3K36me3 was also reduced in GFP + SETD2-deficient neurons though the extent of reduction was smaller than α-TubK40me3 (Extended Data Fig. 2e, f). The distribution of GFP + cells in somatosensory cortex, which was divided into CP, IZ, and VZ/SVZ, was analyzed at E16 (Fig. 2d). ...
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... neurons of somatosensory cortex at E16 (Extended Data Fig. 2c-f). The level of H3K36me3 was also reduced in GFP + SETD2-deficient neurons though the extent of reduction was smaller than α-TubK40me3 (Extended Data Fig. 2e, f). The distribution of GFP + cells in somatosensory cortex, which was divided into CP, IZ, and VZ/SVZ, was analyzed at E16 (Fig. 2d). Approximate 27% of GFP + control cells expressing control shRNA (shNC) migrated into CP, but less than 10% of GFP + SETD2-deficient cells expressing mSETD2 shRNA1 or shRNA2 were able to migrate into CP and most of them were arrested in IZ (Fig. 2d, e). Most SETD2-deficient cells in IZ exhibited highly branched leading processes and ...
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... + cells in somatosensory cortex, which was divided into CP, IZ, and VZ/SVZ, was analyzed at E16 (Fig. 2d). Approximate 27% of GFP + control cells expressing control shRNA (shNC) migrated into CP, but less than 10% of GFP + SETD2-deficient cells expressing mSETD2 shRNA1 or shRNA2 were able to migrate into CP and most of them were arrested in IZ (Fig. 2d, e). Most SETD2-deficient cells in IZ exhibited highly branched leading processes and multipolar morphology, whereas most control cells in IZ were bipolar with one unbranched leading process (Fig. 2f, g). The percentage of GFP + cells in VZ/ SVZ showed no difference between control and SETD2 knockdown (Fig. 2e), neither the percentage of ...
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... less than 10% of GFP + SETD2-deficient cells expressing mSETD2 shRNA1 or shRNA2 were able to migrate into CP and most of them were arrested in IZ (Fig. 2d, e). Most SETD2-deficient cells in IZ exhibited highly branched leading processes and multipolar morphology, whereas most control cells in IZ were bipolar with one unbranched leading process (Fig. 2f, g). The percentage of GFP + cells in VZ/ SVZ showed no difference between control and SETD2 knockdown (Fig. 2e), neither the percentage of cells labeled by Ki67, a marker for the proliferative cells, PH3, a marker for the mitotic cells, or cleaved caspase 3, a marker for the apoptotic cells (Extended Data Fig. 3a-d), indicating that ...
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... and most of them were arrested in IZ (Fig. 2d, e). Most SETD2-deficient cells in IZ exhibited highly branched leading processes and multipolar morphology, whereas most control cells in IZ were bipolar with one unbranched leading process (Fig. 2f, g). The percentage of GFP + cells in VZ/ SVZ showed no difference between control and SETD2 knockdown (Fig. 2e), neither the percentage of cells labeled by Ki67, a marker for the proliferative cells, PH3, a marker for the mitotic cells, or cleaved caspase 3, a marker for the apoptotic cells (Extended Data Fig. 3a-d), indicating that proliferation and apoptosis of NPCs were not affected by acute SETD2 knockdown at E14. Furthermore, we did ...
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... cerebral cortex throughout embryonic stages (Fig. 2a), the high level of α-TubK40me3 only occurred during E14-E16 and rapidly decreased at E18 (Fig. 1a). qPCR showed that the mRNA level of MEC-17 and α-tubulin deacetylase HDAC6 was gradually increased from E14 to E16 and reached the peak at E18 and P0, respectively (Extended Data Fig. 8a), indicating a multifactorial regulation of ...
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... Quantitative analysis of e showed that the percentage of GFP + neurons at the multipolar and branched uni/bipolar stage was largely rescued by expressing either NES-SETD2(1469-1724) or α-tubulin K40F . All data were shown as the mean ± s.e.m. (n = 12, 14, 8, 8, respectively). Data were analyzed using two-way ANOVA with Bonferroni's post-hoc test. ...
Citations
... Less is known about the mechanism by which H3G34R/V mutations alter the global epigenetic profile; however, it is believed that the change to G34 impacts trimethylation of the nearby K36 residue by inhibiting the catalytic activity of the histone methylase SETD2 [47,48]. Loss of SETD2-mediated trimethylation has been associated with inhibition of neuronal activity and proper differentiation in other diseases and would therefore be consistent with the altered differentiation state seen in DHG [49,50]. ...
Pediatric high-grade gliomas (pHGG) are malignant and usually fatal central nervous system (CNS) WHO Grade 4 tumors. The majority of pHGG consist of diffuse midline gliomas (DMG), H3.3 or H3.1 K27 altered, or diffuse hemispheric gliomas (DHG) (H3.3 G34-mutant). Due to diffuse tumor infiltration of eloquent brain areas, especially for DMG, surgery has often been limited and chemotherapy has not been effective, leaving fractionated radiation to the involved field as the current standard of care. pHGG has only been classified as molecularly distinct from adult HGG since 2012 through Next-Generation sequencing approaches, which have shown pHGG to be epigenetically regulated and specific tumor sub-types to be representative of dysregulated differentiating cells. To translate discovery research into novel therapies, improved pre-clinical models that more adequately represent the tumor biology of pHGG are required. This review will summarize the molecular characteristics of different pHGG sub-types, with a specific focus on histone K27M mutations and the dysregulated gene expression profiles arising from these mutations. Current and emerging pre-clinical models for pHGG will be discussed, including commonly used patient-derived cell lines and in vivo modeling techniques, encompassing patient-derived xenograft murine models and genetically engineered mouse models (GEMMs). Lastly, emerging techniques to model CNS tumors within a human brain environment using brain organoids through co-culture will be explored. As models that more reliably represent pHGG continue to be developed, targetable biological and genetic vulnerabilities in the disease will be more rapidly identified, leading to better treatments and improved clinical outcomes.
... Interestingly, the K40 residue of α-tubulin is also targeted for tri-methylation (K40me3) by SETD2, a dual-function histone and microtubule methyltransferase [113]. K40me3 is associated with polymerized MTs in cells and promotes MT stabilization in vitro [113,114]. Furthermore, it has been observed that α-tubulin K40me3 is increased in the absence of K40 acetylation, and that this increase rescues the defects in radial migration and morphological transition of cortical neurons caused by ATAT1 depletion [114]. These results suggest that the two modifications may have overlapping effects, but it is still to be determined whether the two enzymes compete to access the K40 residue and their functional interactions in specific cellular events. ...
... K40me3 is associated with polymerized MTs in cells and promotes MT stabilization in vitro [113,114]. Furthermore, it has been observed that α-tubulin K40me3 is increased in the absence of K40 acetylation, and that this increase rescues the defects in radial migration and morphological transition of cortical neurons caused by ATAT1 depletion [114]. These results suggest that the two modifications may have overlapping effects, but it is still to be determined whether the two enzymes compete to access the K40 residue and their functional interactions in specific cellular events. ...
The acetylation of α-tubulin on lysine 40 is a well-studied post-translational modification which has been associated with the presence of long-lived stable microtubules that are more resistant to mechanical breakdown. The discovery of α-tubulin acetyltransferase 1 (ATAT1), the enzyme responsible for lysine 40 acetylation on α-tubulin in a wide range of species, including protists, nematodes, and mammals, dates to about a decade ago. However, the role of ATAT1 in different cellular activities and molecular pathways has been only recently disclosed. This review comprehensively summarizes the most recent knowledge on ATAT1 structure and substrate binding and analyses the involvement of ATAT1 in a variety of cellular processes such as cell motility, mitosis, cytoskeletal organization, and intracellular trafficking. Finally, the review highlights ATAT1 emerging roles in human diseases and discusses ATAT1 potential enzymatic and non-enzymatic roles and the current efforts in developing ATAT1 inhibitors.
... Statistical significance was determined using two-way ANOVA followed by Sidak's multiple comparisons test *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001. neuronal migration (Xie et al., 2021). Analyzing cPcdh-null dorsal forebrains would help differentiate the two possibilities. ...
Proper formation of the hippocampus is crucial for the brain to execute memory and learning functions. However, many questions remain regarding how pyramidal neurons (PNs) of the hippocampus mature and precisely position. Here we revealed that Setd2, the methyltransferase for histone 3 lysine 36 trimethylation (H3K36me3), is essential for the precise localization and maturation of PNs in the hippocampal CA1. The ablation of Setd2 in neural progenitors leads to irregular lamination of the CA1 and increased numbers of PNs in the stratum oriens. Setd2 deletion in postmitotic neurons causes mis-localization and immaturity of CA1 PNs. Transcriptome analyses revealed that SETD2 maintains the expressions of clustered protocadherin ( cPcdh) genes. Together, Setd2 is required for proper hippocampal lamination and maturation of CA1 PNs.
... Notably, western blot experiments show much higher SETD2 expression in the prenatal and neonatal mouse brain when compared to the adult brain (Koenning et al., 2021). This result has been corroborated with qPCR data from the developing mouse cortex, which shows that Setd2 mRNA expression is at its highest at around E14-E18 and then decreases postnatally (Xie et al., 2021). Despite this, immunohistochemistry has confirmed that SETD2 protein is still expressed across several brain regions in the adult mouse (Koenning et al., 2021). ...
... The role of α-TubK40me3 in microtubule formation has been explored further in another study (Xie et al., 2021). Setd2 knockdown in the developing cortex of mice led to neuronal migration deficits that were rescued with the expression of a truncated SETD2 construct containing only the AWS, SET and post-SET functional domains, and nuclear export signals for cytoplasmic localisation. ...
... The deficit in microtubule formation is proposed to impede the dendritic outgrowth required for neuronal polarisation. These unpolarised neurons lack the ability to efficiently migrate through the cortex, leading to lamination deficits and potentially abnormal cortical function (Xie et al., 2021). ...
The covalent modification of histones is critical for many biological functions in mammals, including gene regulation and chromatin structure. Posttranslational histone modifications are added and removed by specialised ‘writer’ and ‘eraser’ enzymes, respectively. One such writer protein implicated in a wide range of cellular processes is SET domain-containing 2 (SETD2), a histone methyltransferase that catalyses the trimethylation of lysine 36 on histone H3 (H3K36me3). Recently, SETD2 has also been found to modify proteins other than histones, including actin and tubulin. The emerging roles of SETD2 in the development and function of the mammalian central nervous system (CNS) are of particular interest as several SETD2 variants have been implicated in neurodevelopmental disorders, such as autism spectrum disorder and the overgrowth disorder Luscan–Lumish syndrome. Here, we summarise the numerous roles of SETD2 in mammalian cellular functions and development, with a focus on the CNS. We also provide an overview of the consequences of SETD2 variants in human disease and discuss future directions for understanding essential cellular functions of SETD2.
... In addition, these SETD2 knockout mice also displayed defects in social intera tion, motor learning, and spatial memory, resembling LLS patients [27]. Moreover, knoc out of SETD2 results in defects in neuronal morphology transition, and therefore, in rad migration transition [28]. ...
... In addition, these SETD2 knockout mice also displayed defects in social interaction, motor learning, and spatial memory, resembling LLS patients [27]. Moreover, knockout of SETD2 results in defects in neuronal morphology transition, and therefore, in radial migration transition [28]. ...
SETD2 belongs to the family of histone methyltransferase proteins and has been associated with three nosologically distinct entities with different clinical and molecular features: Luscan–Lumish syndrome (LLS), intellectual developmental disorder, autosomal dominant 70 (MRD70), and Rabin–Pappas syndrome (RAPAS). LLS [MIM #616831] is an overgrowth disorder with multisystem involvement including intellectual disability, speech delay, autism spectrum disorder (ASD), macrocephaly, tall stature, and motor delay. RAPAS [MIM #6201551] is a recently reported multisystemic disorder characterized by severely impaired global and intellectual development, hypotonia, feeding difficulties with failure to thrive, microcephaly, and dysmorphic facial features. Other neurologic findings may include seizures, hearing loss, ophthalmologic defects, and brain imaging abnormalities. There is variable involvement of other organ systems, including skeletal, genitourinary, cardiac, and potentially endocrine. Three patients who carried the missense variant p.Arg1740Gln in SETD2 were reported with a moderately impaired intellectual disability, speech difficulties, and behavioral abnormalities. More variable findings included hypotonia and dysmorphic features. Due to the differences with the two previous phenotypes, this association was then named intellectual developmental disorder, autosomal dominant 70 [MIM 620157]. These three disorders seem to be allelic and are caused either by loss-of-function, gain-of-function, or missense variants in the SETD2 gene. Here we describe 18 new patients with variants in SETD2, most of them with the LLS phenotype, and reviewed 33 additional patients with variants in SETD2 that have been previously reported in the scientific literature. This article offers an expansion of the number of reported individuals with LLS and highlights the clinical features and the similarities and differences among the three phenotypes associated with SETD2.
... The stoichiometry of Lys40 methylation and acetylation within MTs is not known, but a recent study in mouse cortical neurons showed, as expected, an inverse relationship between the levels of MT Lys40 trimethylation (which decreases between embryonic day 17.5 and adulthood) and Lys40 acetylation (which increases during this period) [155]. Moreover, another study showed that α-tubulin Lys40 trimethylation is able to rescue the defects of radial migration and morphological transition of cortical neurons caused by α-tubulin Lys40 acetylation deficiency [156]. Another crosstalk may exist between tubulin tyrosination and α-tubulin Lys40 acetylation. ...
Microtubules (MTs), dynamic polymers of α/β-tubulin heterodimers found in all eukaryotes, are involved in cytoplasm spatial organization, intracellular transport, cell polarity, migration and division, and in cilia biology. MTs functional diversity depends on the differential expression of distinct tubulin isotypes and is amplified by a vast number of different post-translational modifications (PTMs). The addition/removal of PTMs to α- or β-tubulins is catalyzed by specific enzymes and allows combinatory patterns largely enriching the distinct biochemical and biophysical properties of MTs, creating a code read by distinct proteins, including microtubule-associated proteins (MAPs), which allow cellular responses. This review is focused on tubulin-acetylation, whose cellular roles continue to generate debate. We travel through the experimental data pointing to α-tubulin Lys40 acetylation role as being a MT stabilizer and a typical PTM of long lived MTs, to the most recent data, suggesting that Lys40 acetylation enhances MT flexibility and alters the mechanical properties of MTs, preventing MTs from mechanical aging characterized by structural damage. Additionally, we discuss the regulation of tubulin acetyltransferases/desacetylases and their impacts on cell physiology. Finally, we analyze how changes in MT acetylation levels have been found to be a general response to stress and how they are associated with several human pathologies.
... [61][62][63] Cilia dynamics, actin filament dynamics, and microtubule formation are essential for this movement. [64][65][66][67][68] Interestingly, the GO enrichment analysis showed that multiple microtubule-related and ciliarelated GO items were affected in miR-96 MT retinas, including microtubule-based processes (GO:0007017), microtubule cytoskeleton organization processes (GO:0000226), actin filament bundle assembly (GO:0051017), cilium organization (GO:0044782), and cilium movement (GO:0003341). ...
Purpose:
Abundant retinal microRNA-183 cluster (miR-183C) has been reported to be a key player in photoreceptor development and functionality in mice. However, whether there is a protagonist in this cluster remains unclear. Here, we used a mutant mouse model to study the role of miR-96, a member of miR-183C, in photoreceptor development and functionality.
Methods:
The mature miR-96 sequence was removed using the CRISPR/Cas9 genome-editing system. Electroretinogram (ERG) and optical coherence tomography (OCT) investigated the changes in structure and function in mouse retinas. Immunostaining determined the localization and morphology of the retinal cells. RNA sequencing was conducted to observe retinal transcription alterations.
Results:
The miR-96 mutant mice exhibited cone developmental delay, as occurs in miR-183/96 double knockout mice. Immunostaining of cone-specific marker genes revealed cone nucleus mislocalization and exiguous Opn1mw/Opn1sw in the mutant (MT) mouse outer segments at postnatal day 10. Interestingly, this phenomenon could be relieved in the adult stages. Transcriptome analysis revealed activation of microtubule-, actin filament-, and cilia-related pathways, further supporting the findings. Based on ERG and OCT results at different ages, the MT mice displayed developmental delay not only in cones but also in rods. In addition, a group of miR-96 potential direct and indirect target genes was summarized for interpretation and further studies of miR-96-related retinal developmental defects.
Conclusions:
Depletion of miR-96 delayed but did not arrest photoreceptor development in mice. This miRNA is indispensable for mouse photoreceptor maturation, especially for cones.
... However, at the later stages of development, i.e., adult neurons, MTs have minus-end-out orientation with hundreds of micrometers in length and act as major long-distance railways for organelle transport. During development, MT orientation according to a network of feedback loops is essential for maintaining proper neuronal shape and inducing neuronal polarization [77,78]. Upon neuronal polarization, posttranslational modifications of MTs in the nascent axon provide selective transport routes that increase the neurite length-dependent feedback and anterograde transport [79,80]. ...
Microtubules (MTs) are highly dynamic polymers essential for a wide range of cellular physiologies, such as acting as directional railways for intracellular transport and position, guiding chromosome segregation during cell division, and controlling cell polarity and morphogenesis. Evidence has established that maintaining microtubule (MT) stability in neurons is vital for fundamental cellular and developmental processes, such as neurodevelopment, degeneration, and regeneration. To fulfill these diverse functions, the nervous system employs an arsenal of microtubule-associated proteins (MAPs) to control MT organization and function. Subsequent studies have identified that the disruption of MT function in neurons is one of the most prevalent and important pathological features of traumatic nerve damage and neurodegenerative diseases and that this disruption manifests as a reduction in MT polymerization and concomitant deregulation of the MT cytoskeleton, as well as downregulation of microtubule-associated protein (MAP) expression. A variety of MT-targeting agents that reverse this pathological condition, which is regarded as a therapeutic opportunity to intervene the onset and development of these nervous system abnormalities, is currently under development. Here, we provide an overview of the MT-intrinsic organization process and how MAPs interact with the MT cytoskeleton to promote MT polymerization, stabilization, and bundling. We also highlight recent advances in MT-targeting therapeutic agents applied to various neurological disorders. Together, these findings increase our current understanding of the function and regulation of MT organization in nerve growth and regeneration.
Posttranslational modifications (PTMs) of tubulin, termed as the "tubulin code", play important roles in regulating microtubule functions within subcellular compartments for specialized cellular activities. While numerous tubulin PTMs have been identified, a comprehensive understanding of the complete repertoire is still underway. In this study, we report that α-tubulin lactylation catalyzed by HDAC6 by using lactate to increase microtubule dynamics in neurons. We identified lactylation on lysine 40 of α-tubulin in the soluble tubulin dimers. Notably, lactylated α-tubulin enhanced microtubule dynamics and facilitated neurite outgrowth and branching in cultured hippocampal neurons. Moreover, we discovered a novel function of HDAC6, acting as the primary “writer” for α-tubulin lactylation. HDAC6-catalyzed lactylation was a reversible process, dependent on lactate concentrations. Intracellular lactate concentration triggered HDAC6 to lactylate α-tubulin, a process dependent on its deacetylase activity. Additionally, the catalytic activity for lactylation was conserved in HDAC family proteins. Our study reveals the primary role of HDAC6 in regulating α-tubulin lactylation, establishing a link between cell metabolism and cytoskeleton functions.
