Biology subjects

Cappelletti, G.

Publications and source records attributed to Cappelletti, G..

5 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

Effective Tubulin Degradation by Rationally Designed Proteolysis Targeting Chimeras

Proteolysis targeting chimeras (PROTACs) are heterobifunctional molecules that induce the degradation of proteins of interest (POIs) via the ubiquitin-proteasome pathway by recruiting E3 ligases to form a ternary complex with the POI. In this study, we rationally designed and synthesized PROTACs targeting the {beta}-tubulin heterodimer, the building block of microtubules (MTs) that are essential for numerous cellular functions and represent important therapeutic targets in cancer and neurodegenerative diseases. Maytansinol, a known MT-destabilising agent, was selected as the POI ligand, functionalised and conjugated to linkers bearing cereblon or Von Hippel-Lindau ligands as E3 ligase recruiters. Four compounds were synthesized and characterized through structural, biophysical and cell biology studies to evaluate their ability to form degradation-prone tubulin-PROTAC-E3 ligase ternary complexes. We confirmed that the PROTACs effectively bind tubulin and recruit E3 ligases. Remarkably, two PROTACs exhibited cellular degradation activity, representing an important advancement in chemically inducing tubulin-E3-ligase interactions. This work integrates rational design, biophysical and structural validation, and cell-based studies to establish a robust framework for developing tubulin-targeting PROTACs, offering significant implications for basic research and therapeutic developments.

biochemistry↗

bioRxiv2025

PARylation in Parkinson's disease: a bridge between Lewy body formation and neuronal cell death

Poly-ADP-ribosylation (PARylation), catalyzed by the enzyme PARP1, involves the addition of poly-ADP-ribose polymers (PAR) and has been associated with -synuclein aggregation in Parkinsons disease (PD) models. This study aimed to unravel the role of PARylation in -synuclein aggregation and neuronal cell death in the complex environment of post-mortem human PD brains. Using high-resolution imaging and 3D reconstruction analysis, we observed that PAR accumulate in the cytoplasm in regions affected by PD pathology, preceding the formation of -synuclein oligomers. Additionally, we found that PAR and stress granules contribute to the formation of Lewy bodies. Increased colocalization of PAR with mitochondria in the substantia nigra of PD patients, along with the presence of PAR-positive condensed DNA, further suggests a role in neuronal cell death. Collectively, our findings reveal a critical involvement of PARylation in the pathological mechanisms underlying neurodegeneration in PD and position PARylation as a potential therapeutic target. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=160 SRC="FIGDIR/small/642849v1_ufig1.gif" ALT="Figure 1"> View larger version (73K): org.highwire.dtl.DTLVardef@64858aorg.highwire.dtl.DTLVardef@17b4706org.highwire.dtl.DTLVardef@73f865org.highwire.dtl.DTLVardef@1b8d619_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2022

Linking acetylated alpha-Tubulin redistribution to alpha-Synuclein pathology in brain of Parkinson's disease patients

Highly specialized microtubules in neurons are crucial to the health and disease of the nervous system, and their properties are strictly regulated by different post-translational modifications, including -Tubulin acetylation. An imbalance in the levels of acetylated -Tubulin has been reported in experimental models of Parkinsons disease (PD) whereas pharmacological or genetic modulation that leads to increased acetylated -Tubulin successfully rescues axonal transport defects and inhibits -Synuclein aggregation. However, the role of acetylation of -Tubulin in the human nervous system is largely unknown as most studies are based on in vitro evidence. To capture the complexity of the pathological processes in vivo, we analysed post-mortem human brain of PD patients and control subjects. In the brain of PD patients at Braak stage 6, we found a redistribution of acetylated -Tubulin, which accumulates in the neuronal cell bodies in subcortical structures but not in the cerebral cortex, and decreases in the axonal compartment, both in the central and peripheral nervous system. High-resolution and 3D reconstruction analysis linked acetylated -Tubulin redistribution to -Synuclein oligomerization, leading us to propose a model for Lewy body (LB) morphogenesis. Finally, for the first time in post-mortem human brain, we observed threadlike structures, resembling tunnelling nanotubes that contain -Synuclein oligomers and are associated with acetylated -Tubulin enriched neurons. In conclusion, we disclose a novel aspect of LB morphogenesis, indicating the role of acetylated -Tubulin in PD, that may provide clues to design novel therapeutic interventions.

neuroscience↗

bioRxiv2022

SARS-CoV-2 ORF3c impairs mitochondrial respiratory metabolism, oxidative stress and autophagic flow

Coronaviruses encode a variable number of accessory proteins that play a role in host-virus interactions, in the suppression of immune responses, or in immune evasion. Accessory proteins in SARS-CoV-2 consist of at least twelve viral proteins whose roles during infection have been extensively studied. Nevertheless, the role of the ORF3c accessory protein, an alternative open reading frame of ORF3a, has remained elusive. Herein, we characterized ORF3c in terms of cellular localization, hosts antiviral response modulation, and effects on mitochondrial metabolism. We show that ORF3c has a mitochondrial localization and alters mitochondrial metabolism, resulting in increased ROS production, block of the autophagic flux, and accumulation of autophagosomes/autolysosomes. Notably, we also found that ORF3c induces a shift from glucose to fatty acids oxidation and enhanced oxidative phosphorylation. This is similar to the condition observed in the chronic degenerative phase of COVID-19. Altogether these data suggest that ORF3c could be a key protein for SARS-CoV-2 pathogenesis and that it may play a role in disease progression.

microbiology↗