Biology subjects

Bartolini, F.

Publications and source records attributed to Bartolini, F..

15 recordsLinked to original sources

bioRxiv2026

α-Synuclein and γ-Tubulin Cooperatively Regulate Activity-Evoked Presynaptic Microtubule Nucleation to Gate Dopamine Release

-Synuclein has long been implicated in the regulation of synaptic activity, but the molecular basis that underlies this function has been elusive. Here, we identify a microtubule (MT)-dependent mechanism through which -synuclein regulates synaptic dopamine release. Using live imaging of cultured dopaminergic neurons, we visualize dynamic MTs at individual presynaptic boutons and show that neuronal activity triggers local {gamma}-tubulin-dependent MT nucleation. We find that this nucleation is essential for interbouton synaptic vesicle (SV) transport and for sustained dopamine release during high activity. We further discover that -synuclein acts as a positive regulator of presynaptic MT nucleation by binding directly to {gamma}-tubulin and the /{beta}-tubulin heterodimer. Activity-evoked phosphorylation of -synuclein at serine 129, a modification that accumulates in synucleinopathies and a molecular switch for -synuclein binding to synaptic proteins, occurs in the region of /m tubulin binding and is both necessary and sufficient for MT initiation. Our findings reveal a previously unrecognized, activity-dependent role for -synuclein in the nucleation of axonal MTs that enables on-demand SV interbouton redistribution and dopamine release. This mechanism provides a novel molecular link between -synuclein phosphorylation and MT-dependent modulation of dopamine release, offering insight into how its dysregulation may contribute to dopaminergic synaptic dysfunction, a central feature of synucleinopathies.

cell biology↗

bioRxiv2025

Tubulin autoregulation factors SCAPER and TTC5 recruit γ-tubulin to non-centrosomal MTOCs for neuronal microtubule nucleation and axon regeneration

Neuronal function and survival depend on highly stereotyped non-centrosomal microtubule (MT) arrays. How these arrays form remains poorly understood. Here we identified a role for SCAPER and TTC5, two factors previously implicated in tubulin mRNA autoregulation, in controlling neuronal MT content through {gamma}-tubulin-dependent nucleation. In C. elegans neurons, loss of scpr-1, ttc-5, or both reduced MT numbers to a similar degree as depletion of {gamma}-tubulin, the main MT nucleator. Using conditional single-cell degradation alleles and endogenous tagging, we find that {gamma}-tubulin nucleates MTs in the neuronal cell body from endosomal puncta, and that scpr-1 and ttc-5 are required to recruit {gamma}-tubulin to these structures. SCPR-1 is also instructive, as its overexpression drastically increases {gamma}-tubulin levels and enhances MT density. We propose that these mechanisms are conserved since human SCAPER rescues C. elegans mutants, and SCAPER knockdown in rat hippocampal neurons reduces both {gamma}-tubulin clustering at presynaptic sites and activity-dependent synaptic MT nucleation. Finally, while scpr-1, ttc-5, and{gamma} -tubulin are not required for developmental axon elongation, they are essential for regeneration, where SCPR-1 directs {gamma}-tubulin to the growth cone to facilitate regrowth following injury. These findings reveal mechanisms governing neuronal cytoskeleton assembly and function, and suggest potential crosstalk between tubulin autoregulation and microtubule nucleation.

cell biology↗

bioRxiv2025

Preventing neuropathy and improving anti-cancer chemotherapy with a carbazole-based compound

Advances in cancer treatment have led to a steady increase in the rate of disease remission. However, while many treatment-related adverse effects gradually resolve after therapy, chemotherapy-induced peripheral neuropathy (CIPN) often persists, with no means of prevention or direct treatment available. Herein, we present Carba1, a novel bi-functional carbazole that mitigates neuropathy through two distinct mechanisms. First, by interacting with tubulin, Carba1 reduces the required dose of taxanes, widely used chemotherapy drugs notorious for their toxic side effects, including CIPN. Second, Carba1 activates nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme in the NAD salvage pathway, triggering a metabolic rewiring that enhances the resilience of neurons and Schwann cells against chemotherapy-induced toxicity. We demonstrate the neuroprotective efficacy of Carba1 both in vitro, against neurotoxicity induced by paclitaxel (PTX), cisplatin, and bortezomib, and in vivo in a rat model of PTX-induced neuropathy. Importantly, we establish that Carba1 does not compromise the therapeutic efficacy of PTX nor promotes tumor growth. Comparative analyses of Carba1 derivatives further suggest the potential of designing compounds with either dual synergistic and neuroprotective activity or exclusive neuroprotective properties. Altogether, our findings position Carba1 as a promising therapeutic candidate for preventing CIPN, with the potential, if successfully translated to clinical settings, to improve both the quality of life and treatment outcome for cancer patients.

cancer biology↗

bioRxiv2023

Rho GTPase signaling and mDia facilitate endocytosis via presynaptic actin

Neurotransmission at synapses is mediated by the fusion and subsequent endocytosis of synaptic vesicle membranes. Actin has been suggested to be required for presynaptic endocytosis but the mechanisms that control actin polymerization and its mode of action within presynaptic nerve terminals remain poorly understood. We combine optical recordings of presynaptic membrane dynamics and ultrastructural analysis with genetic and pharmacological manipulations to demonstrate that presynaptic endocytosis is controlled by actin regulatory diaphanous-related formins mDia1/3 and Rho family GTPase signaling. We show that impaired presynaptic actin assembly in the near absence of mDia1/3 and reduced RhoA activity is partly compensated by hyperactivation of Rac1. Inhibition of Rac1 signaling further aggravates impaired presynaptic endocytosis elicited by loss of mDia1/3. Our data suggest that interdependent mDia1/3-Rho and Rac1 signaling pathways cooperatively act to facilitate synaptic vesicle endocytosis by controlling presynaptic F-actin.

neuroscience↗

bioRxiv2023

Dual spatio-temporal regulation of axon growth and microtubule dynamics by RhoA signaling pathways

RhoA plays a crucial role in neuronal polarization, where its action restraining axon outgrowth has been thoroughly studied. We now report that RhoA has not only inhibitory but also a stimulatory effect on axon development depending on when and where exerts its action and the downstream effectors involved. In cultured hippocampal neurons, FRET imaging revealed that RhoA activity selectively localizes in growth cones of undifferentiated neurites, while in developing axons it displays a biphasic pattern, being low in nascent axons and high in elongating ones. RhoA-Rho kinase (ROCK) signaling prevents axon initiation but has no effect on elongation, while formin inhibition reduces axon extension without significantly altering initial outgrowth. Besides, RhoA-mDia promotes axon elongation by stimulating growth cone microtubule stability and assembly, as opposed to RhoA-ROCK that restrains growth cone microtubule assembly and protrusion. Finally, we show that similar mechanisms might operate during axonal regeneration, with RhoA-ROCK slowing axon regrowth after axotomy and RhoA-mDia favoring extension of regenerated axons.

neuroscience↗

bioRxiv2023

MFN2-dependent recruitment of ATAT1 coordinates mitochondria motility with alpha-tubulin acetylation and is disrupted in CMT2A

Acetylated microtubules play key roles in the regulation of mitochondria dynamics. It has however remained unknown if the machinery controlling mitochondria dynamics functionally interacts with the -tubulin acetylation cycle. Mitofusin-2 (MFN2), a large GTPase residing in the mitochondrial outer membrane and mutated in Charcot-Marie-Tooth type 2 disease (CMT2A), is a regulator of mitochondrial fusion, transport and tethering with the endoplasmic reticulum. The role of MFN2 in regulating mitochondrial transport has however remained elusive. Here we show that mitochondrial contacts with microtubules are sites of -tubulin acetylation, which occurs through the MFN2-mediated recruitment of -tubulin acetyltransferase 1 (ATAT1). We discover that this activity is critical for MFN2-dependent regulation of mitochondria transport, and that axonal degeneration caused by CMT2A MFN2 associated mutations, R94W and T105M, may depend on the inability to release ATAT1 at sites of mitochondrial contacts with microtubules. Our findings reveal a function for mitochondria in regulating acetylated -tubulin and suggest that disruption of the tubulin acetylation cycle play a pathogenic role in the onset of MFN2-dependent CMT2A. O_FIG O_LINKSMALLFIG WIDTH=193 HEIGHT=200 SRC="FIGDIR/small/532838v1_ufig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@3db8eorg.highwire.dtl.DTLVardef@143821corg.highwire.dtl.DTLVardef@1132b13org.highwire.dtl.DTLVardef@1710383_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIMitochondria contacts with MTs are hotspots of -tubulin acetylation through the recruitment of ATAT1 by MFN2 C_LIO_LIMutations in MFN2 associated with CMT2A disease lose this activity by sequestering ATAT1 C_LIO_LIDistal axonal degeneration caused by loss of MFN2 depends on acetylated tubulin-mediated mitochondria transport C_LI eTOCRecruitment of ATAT1 to mitochondria by MFN2 is critical for axonal viability through the regulation of mitochondria transport, and is disrupted in CMT2A

cell biology↗

bioRxiv2022

Alterations to the broad-spectrum formin inhibitor SMIFH2 improve potency

SMIFH2 is a small molecule inhibitor of the formin family of cytoskeletal regulators that was originally identified in a screen for suppression of actin polymerization induced by the mouse formin Diaphanous 1 (mDia1). Despite widespread use of this compound, it is unknown whether SMIFH2 inhibits all human formins. Additionally, the nature of protein/inhibitor interactions remains elusive. We assayed SMIFH2 against human formins representing six of the seven mammalian classes and found inhibitory activity against all formins tested. We synthesized a panel of SMIFH2 derivatives and found that, while many alterations disrupt SMIFH2 activity, substitution of an electron-donating methoxy group in place of the bromine along with halogenation of the furan ring increases potency by approximately five-fold. Similar to SMIFH2, the active derivatives are also pan-inhibitors for the formins tested. This result suggests that while potency can be improved, the goal of distinguishing between highly conserved FH2 domains may not be achievable using the SMIFH2 scaffold.

biochemistry↗

bioRxiv2022

Reactive microglia and cytokine release entails microtubule remodeling from acentrosomal to centrosomal arrays

Microglia reactivity entails a large-scale remodeling of cellular geometry, but the role of the microtubule cytoskeleton during these changes remains unexplored. Here we show that reactive proinflammatory microglia provide a heretofore unique example of microtubule reorganization from a non-centrosomal array of parallel and stable microtubules to a radial array of more dynamic microtubules. While in the homeostatic state microglia nucleate microtubules at Golgi outposts, proinflammatory signaling induces recruitment of nucleating material nearby the centrosome and inhibition of centrosomal maturation enhances NLRP3 inflammasome activation and secretion of IL-1{beta}. Our results demonstrate that a hallmark of microglia reactivity is a striking remodeling of the microtubule cytoskeleton and suggest that pericentrosomal microtubule nucleation may serve as a distinct marker of microglia activation as well as a novel target to modulate cytokine-mediated inflammatory responses in chronic disease and tissue injury.

cell biology↗

bioRxiv2021

NSun2 deficiency promotes tau hyperphosphorylation and neurodegeneration through epitranscriptomic regulation of miR-125b

Overproduction or suppression of certain microRNAs (miRNAs) in Alzheimers disease (AD) brains promote alterations in tau proteostasis and neurodegeneration. However, the mechanisms governing how specific miRNAs are dysregulated in AD brains are still under investigation. Epitranscriptomic regulation adds a layer of post-transcriptional control to brain function during development and adulthood. NOP2/Sun RNA methyltransferase 2 (NSun2) is one of the few known brain-enriched methyltransferases able to modify mammalian non-coding RNAs and loss of function autosomal-recessive mutations in NSUN2 have been associated with neurological abnormalities in humans. Here, we provide evidence that NSun2 is expressed in adult human neurons in the hippocampal formation and prefrontal cortex. When we evaluated NSun2 protein expression in post-mortem brain tissue from AD patients we find is dysregulated which was also found in mice and human cellular AD models. To probe these observed alterations were unique to AD we further evaluated brain tissue from other tauopathies, observing NSun2 protein levels were similar between cases and controls. In a well-established Drosophila melanogaster model of tau-induced toxicity we investigated the pathological role of NSun2 observing that reduction of NSun2 protein levels exacerbated tau toxicity, while overexpression of NSun2 partially abrogated the toxic effects. We further show using human induced pluripotent stem cell (iPSC) derived neuronal cultures that NSun2 deficiency results in tau hyperphosphorylation and we found in primary hippocampal neuronal cultures NSun2 levels decrease in response to amyloid-beta oligomers (A{beta}O). Furthermore, in mice, we observed that NSun2 deficiency promotes aberrant levels of m6A methylated miR-125b and tau hyperphosphorylation. Altogether, our study supports that neuronal NSun2 deficiency in AD promotes neurodegeneration by altering tau phosphorylation and tau toxicity through an epitranscriptomic regulatory mechanism and highlights a novel avenue for therapeutic targeting.

neuroscience↗

bioRxiv2021

Impaired α-tubulin re-tyrosination leads to synaptic dysfunction and is a feature of Alzheimer's disease

In neurons, dynamic microtubules play regulatory roles in neurotransmission and synaptic plasticity. While stable microtubules contain detyrosinated tubulin, dynamic microtubules are composed of tyrosinated tubulin, suggesting that the tubulin tyrosination/detyrosination (Tyr/deTyr) cycle modulates microtubule dynamics and synaptic function. In the Tyr/deTyr cycle, the C-terminal tyrosine of -tubulin is re-added by tubulin-tyrosine-ligase (TTL). Here we show that TTL+/- mice exhibit decreased tyrosinated microtubules, synaptic plasticity and memory deficits, and that reduced TTL expression is a feature of sporadic and familial Alzheimers disease (AD), with human APPV717I neurons having less dynamic microtubules. We find that spines visited by dynamic microtubules are more resistant to Amyloid{beta}1-42 and that TTL, by promoting microtubule entry into spines, prevents A{beta}1-42-induced spine pruning. Our results demonstrate that the Tyr/deTyr cycle regulates synaptic plasticity, is protective against spine injury, and that tubulin re-tyrosination is lost in AD, providing evidence that a defective Tyr/deTyr cycle may contribute to neurodegeneration.

neuroscience↗

bioRxiv2021

Transthyretin promotes axon growth via regulation of microtubule dynamics and tubulin acetylation

Transthyretin (TTR), a plasma and cerebrospinal fluid protein, increases axon growth and organelle transport in sensory neurons. These TTR functions were suggested to underlie its activity in promoting nerve regeneration. While neurons extend their axons, the microtubule (MT) cytoskeleton is crucial for the segregation of functional compartments and axonal outgrowth. Herein, we investigated the hypothesis that TTR promotes axon elongation and regeneration by modulating MT dynamics. Indeed, we found that TTR KO mice have an intrinsic increase in dynamic MTs and reduced levels of acetylated -tubulin in uninjured peripheral axons, and fail to modulate microtubule dynamics in response to sciatic nerve injury. Importantly, restoring acetylated -tubulin levels of TTR KO DRG neurons using an HDAC6 inhibitor was sufficient to completely revert defective MT dynamics and neurite outgrowth. In summary, our results revealed a new role for TTR in the modulation of MT dynamics by regulating -tubulin acetylation and support that this activity underlies TTR neuritogenic function.

neuroscience↗

bioRxiv2019

Integrins protect nociceptive neurons in models of paclitaxel-mediated peripheral sensory neuropathy

Chemotherapy induced peripheral neuropathy (CIPN) is a major side effect from cancer treatment with no known method for prevention or cure in clinics. CIPN primarily affects unmyelinated nociceptive sensory terminals. Despite the high prevalence of CIPN, molecular and cellular mechanisms that lead to CIPN are still poorly understood. Here, we used a genetically tractable Drosophila model and primary sensory neurons isolated from adult mouse to examine the mechanisms underlying CIPN and identify protective pathways. We found that chronic treatment of Drosophila larvae with paclitaxel caused sensory neuron degeneration, altered the terminal branching pattern of nociceptive neurons, and reduced thermal nociceptive responses. We found that nociceptive neuron-specific overexpression of integrins, which are known to support neuronal maintenance in several systems, conferred protection from paclitaxel-mediated cellular and behavioral phenotypes. Live imaging and superresolution approaches provide evidence that paclitaxel treatment causes cellular changes that are consistent with alterations in endosome-mediated trafficking of integrins. We used primary dorsal root ganglia neuron cultures to test conservation of integrin-mediated protection. We show that overexpression of a human integrin {beta} subunit 1 (ITGB1) also prevented degeneration following paclitaxel treatment. Altogether, our study supports conserved mechanisms of paclitaxel-induced perturbation of integrin trafficking and a therapeutic potential of restoring integrin levels to antagonize paclitaxel-mediated toxicity in sensory neurons.

neuroscience↗

bioRxiv2019

Arfgef1 haploinsufficiency in mice alters neuronal endosome composition and decreases membrane surface postsynaptic GABAA receptors

ARFGEF1 encodes a guanine exchange factor involved in intracellular vesicle trafficking, and is a candidate gene for childhood genetic epilepsies. To model ARFGEF1 haploinsufficiency observed in a recent Lennox Gastaut Syndrome patient, we studied a frameshift mutation (Arfgef1fs) in mice. Arfgef1fs/+ pups exhibit signs of developmental delay, and Arfgef1fs/+ adults have a significantly decreased threshold to induced seizures but do not experience spontaneous seizures. Histologically, the Arfgef1fs/+ brain exhibits a disruption in the apical lining of the dentate gyrus and altered spine morphology of deep layer neurons. In primary hippocampal neuron culture, dendritic surface and synaptic but not total GABAA receptors (GABAAR) are reduced in Arfgef1fs/+ neurons with an accompanying decrease in GABAAR-containing recycling endosomes in cell body. Arfgef1fs/+ neurons also display differences in the relative ratio of Arf6+:Rab11+:TrfR+ recycling endosomes. Although the GABAAR-containing early endosomes in Arfgef1fs/+ neurons are comparable to wildtype, Arfgef1fs/+ neurons show an increase in GABAAR-containing lysosomes in dendrite and cell body. Together, the altered endosome composition and decreased neuronal surface GABAAR results suggests a mechanism whereby impaired neuronal inhibition leads to seizure susceptibility.\n\nHighlightsO_LIArfgef1fs/+ mice have lower seizure threshold but no spontaneous seizure.\nC_LIO_LIArfgef1fs/+ neurons show reduced dendritic surface GABAAR.\nC_LIO_LIArfgef1fs/+ neurons have decreased GABAAR-containing recycling endosome accompanied with an increase in GABAAR-containing lysosomes.\nC_LI

genetics↗

bioRxiv2019

Pathogenic Role of Delta 2 Tubulin in Bortezomib Induced Peripheral Neuropathy

The pathogenesis of chemotherapy induced peripheral neuropathy (CIPN) is still poorly understood. Herein, we found that the CIPN-causing drug, bortezomib (Bort), induces delta 2 tubulin (D2) while affecting MT stability and dynamics in sensory neurons, and that accumulation of D2 is a hallmark of Bort-induced peripheral neuropathy in humans. Furthermore, while induction of D2 was sufficient to cause axonopathy and inhibit mitochondria motility, reducing D2 alleviated both axonal degeneration and loss of mitochondria motility promoted by Bort. Altogether, our data demonstrate that Bort, structurally unrelated to tubulin poisons, can affect the tubulin cytoskeleton in sensory neurons in vitro, in vivo and in humans, indicating that the pathogenic mechanisms of seemingly unrelated CIPN drugs may converge on tubulin damage. They further reveal a previously unrecognized pathogenic role for D2 in bortezomib-causing CIPN through its regulation of mitochondria dynamics.

neuroscience↗

bioRxiv2019

Activity-dependent nucleation of dynamic microtubules at presynaptic boutons is required for neurotransmission

Control of microtubule (MT) dynamics is critical for neuronal function. Whether MT nucleation is regulated at presynaptic boutons and influences overall presynaptic activity remains unknown. By visualizing MT dynamics at individual excitatory en passant boutons in axons of hippocampal neurons we found that MTs preferentially grow from presynaptic boutons as a result of {gamma}-tubulin and augmin-dependent nucleation. MT nucleation at boutons is promoted by neuronal activity, functionally coupled to synaptic vesicle (SV) transport, and required for neurotransmission. Hence, en passant boutons act as hotspots for activity-dependent MT nucleation, which is required for neurotransmission by providing the tracks for a rate-limiting supply of SVs to sites of neurotransmitter release.\n\nHighlightsO_LIExcitatory boutons are hotspots for neuronal activity-induced {gamma}-tubulin dependent MT nucleation\nC_LIO_LIThe augmin complex is required for the correct polarity of presynaptic de novo nucleated MTs\nC_LIO_LIPresynaptic MT nucleation promotes SV motility and exocytosis at sites of release\nC_LI\n\nIn BriefOur results demonstrate that excitatory en passant boutons are hotspots for neuronal activity-induced {gamma}-tubulin- and augmin-dependent oriented MT nucleation, and that the resulting presynaptic de novo nucleated MTs promote inter-bouton SV motility which is rate-limiting for neurotransmitter release.

cell biology↗