Axonal transport is an essential mechanism that supports the proper physiological functioning of neurons by efficiently channeling the intracellular cargo trafficking (Huang et al., 2022). Axonal transport acts as a central network in neurons by facilitating essential cargoes to different parts of neurons, by aiding in vital processes like synaptic plasticity and neurotransmission. In neuron axonal transport, the core component, microtubules serve as a primary channel for intracellular cargo trafficking, where the structural organization of microtubules controls the efficiency of axonal transport. Microtubules are a well-organized array bundle and spatially aligned scaffold proteins that efficiently enable axonal transport’s pace and functioning (Stephan et al., 2015). Furthermore, other properties of microtubules, like their arrangement, polarity and stability, regulate the axonal transport efficacy and direction, resulting in adequate neuronal functioning and integrity. Disruptions in microtubule physiology affect the axonal transport mechanism and eventually cause neurodegenerative disease (Yogev et al., 2016; Costa and Sousa, 2022). Microtubules exhibit a polarized orientation, with the plus-end leading toward the axon terminal and the minus-end toward the cell body. This polarized orientation helps guide the motor protein directions, which facilitates kinesin and dynein with anterograde and retrograde transport directions, respectively. PTMs on microtubules like acetylation and tyrosination influence the efficiency of these motor proteins movements by regulating their affinity and sorting (Tas et al., 2017). Microtubule stability in axonal transport determines the motor protein recruitment and movement efficiency for cargo transport. By stabilizing microtubules, tubulin acetylation increases the ability to bind and retain the motor proteins, specifically for dynein in retrograde transport. Motor proteins like kinesin and dynein bind with the microtubule and other end of the motor proteins binds to the cargoes via adaptors, this interaction between the motor protein and cargo is facilitated by palmitoylation of adaptor proteins illustrated in Figure 1.

Motor proteins like kinesin and dynein transport along the microtubule network effectively carrying the cargo movement are essential for axonal transport. Kinesin and dynein support the anterograde and retrograde transportation respectively, where the motor proteins that transports the cargo from the cell body to the axonal terminal at the synaptic terminals are called anterograde transportation, and the reverse is called retrograde transportation. These movements make it easier for kinesin and dynein to effectively transport necessary cargo like organelles, proteins, vesicles and cell signaling responses for processes like synaptic functions, recycling cell organelles, neuronal development and maintenance. Kinesin and dynein work coordinately in both directions, which upon disruption of one motor protein’s function tends to affect the other, indicating their connective interdependence (Encalada et al., 2011; Lee et al., 2011). Neurofilament transport is the paramount example of the concerted action of kinesin and dynein in axonal transport, where kinesin follows the anterograde movement, and dynein follows the retrograde movement. Their actions are tightly coupled to regulate efficient bi-directional transport. Kinesin-1 drives the transport of mitochondria to high energy demand regions like synaptic terminals and dynein drives the degradation and recycling of damaged or aged mitochondria toward the cell body (Pilling et al., 2006; Uchida et al., 2009; Twelvetrees et al., 2016). When impaired, the mutually dependent bi-directional movements of kinesin and dynein can lead to mitochondrial buildup, energy deficit, and neuronal deterioration, which are hallmarks of many neurodegenerative diseases (Wilson et al., 2023).

Additionally to motor proteins, MAPs are also essential to maintain the microtubules stability, axonal structure and transport efficiency by promoting microtubule bundling (Tortosa et al., 2013; Tas et al., 2017). MAPs are critically important in regulating microtubule stability and neuronal integrity, which influences axonal transport’s direction, organization and speed by interacting with microtubules and motor proteins. MAPs dysregulations can cause disruption in the transport mechanisms leading to synaptic dysfunction and neuron degeneration. The classic example of such a case is tau protein hyperphosphorylation followed by aggregation, an important pathological feature in neurodegenerative diseases. Tau is an essential MAP member that promotes axon elongation, microtubule stability and intracellular transport support. Tau, in a normal state, binds to the microtubules to stabilize them and acts as a scaffold for motor proteins to transport cargo. However, hyperphosphorylated and aggregated conditions of tauopathies, such as AD and frontotemporal dementia (FTD), impair the transport mechanism and destabilize microtubules (Iqbal et al., 2010; Alonso et al., 2018). The tangles formed by aggregated hyperphosphorylated proteins called neurofibrillary tangles (NFT), are a hallmark of AD. These tangles disrupt the anterograde and retrograde transport movement of motor proteins, causing an imbalance in the homeostasis of neuron and synaptic functions. This imbalance in the transport mechanism of tau proves to have differential effects on kinesin and dynein. It is observed that tau potentially affects anterograde transport more than retrograde transport by inhibiting Kinesin-1. This results in increased retrograde transport and accumulation of cargo in the axon, increasing the synaptic function impairment (Dixit et al., 2008; Chaudhary et al., 2018). Dynein-mediated transport is also affected by tau modifications. Although dynein is less susceptible to tau inhibition compared to kinesin, high tau levels can still impair dynein-dependent transport, particularly in early phagosomes, by lowering the interacted dynein motors numbers (Dixit et al., 2008; Vershinin et al., 2008). The characteristic features of tauopathies are synaptic dysfunction and neuronal degeneration; transport imbalance can result in both disturbances leading to tauopathies.

The zDHHC family comprises multiple isoforms of PATs, each with distinct substrate specificities and tissue distributions. In neurons, specific zDHHC enzymes have been associated with palmitoylation of key synaptic proteins, thereby modulating the neurotransmission and synaptic plasticity. The zDHHC enzymes localization and activity are tightly regulated, ensuring precise spatial and temporal control over protein palmitoylation states within neuronal circuits. In neurons, the palmitoylation landscape is orchestrated by a repertoire of zDHHC palmitoyl acyltransferases, each exhibiting unique substrate specificities and regulatory mechanisms. PATs are the crucial factor in palmitoylation for regulating protein trafficking, targeting and function. They are established PATs with specificity to involve in certain processes which are discussed below. The following are the list of a few important PATs involved in several neuronal functioning and pathways.

In AD, interaction between tau protein and Aβ peptides is one of the major players in triggering synaptic dysfunction and neuronal degeneration particularly by disrupting the neuronal transport mechanisms. Tau is one of the largely involved MAPs that helps in facilitating microtubule stability and axonal transport. While direct palmitoylation of tau has not been established, emerging evidence suggests that palmitoylation-related pathways indirectly influence tau phosphorylation, aggregation, and its interactions with key proteins involved in axonal transport and synaptic toxicity. The imbalance between tau 3R and 4R isoform populations can impair the transport of APP, where the 3R isoform dominance favors anterograde movement and 4R isoform dominance favors the retrograde movement in axonal transport. This imbalance between anterograde and retrograde transport leads to disruption of APP dynamics, causing neurodegeneration (Lacovich et al., 2017). More importantly, tau is essential for Aβ-induced neurotoxicity. Suppose tau-expressing neurons degenerate in the presence of Aβ while tau-depleted neurons exhibit resistance. In that case, it is evident that tau plays a critical role in the disruption and neurodegeneration of Aβ-mediated transport (Vossel et al., 2015). Tau hyperphosphorylation driven by Aβ oligomers can disrupt microtubules, causing degeneration of neurons. Due to which, tau’s function in regulating axonal transport can be directly impacted by Aβ, leading to synaptic dysfunction and neurodegeneration (Decker et al., 2010). Furthermore, tau pathology is primarily caused by the ratio of Aβ42/40. Elevated aggregation and tau deposition are linked with an increased Aβ42/40 ratio, which further impairs neuronal transport and builds up to the pathophysiology of AD (Salvadores et al., 2020). These studies collectively indicate that tau and Aβ play a crucial role in disrupting transport mechanisms in AD by the involvement of Aβ oligomer interactions and imbalance in tau isoform impairing axonal transport and synaptic function mechanisms.

Palmitoylation at the cysteine residue is a crucial PTM controlling synaptic protein localization, stability and functionality. Dysregulated palmitoylation causes neurodegeneration due to mutations in zDHHC enzymes and appears to have significant effects on synaptic proteins. For example, flotillin-2, an important synaptic protein that maintains synaptic integrity, is not palmitoylated due to the zDHHC5 enzyme mutation disrupting neuron development and synaptic plasticity (Li et al., 2012). zDHHC2 enzyme mutations disrupt the palmitoylation of AKAP-79/150, important for maintaining synaptic potential and AMPA receptor trafficking. Studies imply that palmitoylation of AKAP-79/150 is important for maintaining synaptic plasticity, where knockdown of zDHHC2 enzymes resulted in impairing AMPA receptor exocytosis, dendritic spine expansion and synaptic strengthening (Woolfrey et al., 2015). GABAergic transmissions are affected by the disrupted palmitoylation of the GABA_A receptor due to a mutation in the zDHHC3 enzyme, also called golgi-specific DHHC zinc finger protein (GODZ). GABAergic transmissions are affected by GODZ enzyme mutation, which impacts inhibitory synapse formation and function (Kilpatrick et al., 2016). δ-catenin palmitoylation is essential for enhancing spine stability, receptor recruitment and synaptic cadherin interactions. Disruption in this process impacts the synaptic plasticity and adhesion with cadherins, severely disturbing memory formation (Brigidi et al., 2014). The interaction between protein interacting with C kinase 1 (PICK1) and zDHHC8 palmitoylates is necessary for cerebellar long-term synaptic depression (LTD), which is disrupted due to loss of zDHHC8 results in impaired LTD and synaptic functions (Thomas et al., 2013). The overall stability and function of the DATs are effectively maintained by zDHHC enzymes such as zDHHC2, zDHHC3, zDHHC8, zDHHC15, and zDHHC17. Defects in these zDHHC enzymes can induce disruptions in the dopaminergic system, causing neurodegenerative disorders (Rastedt et al., 2014). The above finding exhibits that mutations in zDHHC enzymes can affect the role of essential synaptic protein palmitoylation, which consequently impairs synaptic plasticity and neurotransmission and increases the chances of neurodegenerative diseases. Studying the effects of zDHHC mutations on the palmitoylation of synaptic proteins could provide essential knowledge on the molecular mechanisms behind neuronal degeneration and identify potential opportunities for intervention.

In PD, α-synuclein, a presynaptic neuronal protein, impairs mitochondrial transport and function when pathologically aggregated. These aggregations cause morphological disruptions in mitochondria by fragmentation and swelling, which, in process, damages the membrane potential of mitochondria and results in neuronal apoptosis. It is due to the subtle interaction of α-synuclein with mitochondrial proteins involved in the mitochondrial permeability transition pore, which disturbs the energy production and homeostasis in mitochondria (Lurette et al., 2023). Additionally, the dynamics of mitochondria are adversely affected by α-synuclein aggregation, where the balance between the fission and fusion process is impaired, causing mitochondrial fragmentation. This disruption changes the transport of mitochondria in the axonal transport mechanism, triggering oxidative stress and energy deficits within neurons (Pozo Devoto and Falzone, 2017). Additionally, translocase of the outer membrane 40 (TOM40), a mitochondrial protein involved in import mechanisms, was observed to be reduced by aggregated α-synuclein. Downregulation of TOM40 impairs the import of necessary proteins across mitochondria, increasing oxidative stress, decreasing energy levels, and compromising neuron viability (Vasquez et al., 2024). Aggregation of α-synuclein also impairs the microtubule-dependent transport mechanisms, which mislocalizes the mitochondria in neurons, causing severe energy depletion. Energy depletion due to mislocalized mitochondria occurs precisely at the synaptic terminal, contributing to neurodegeneration in PD (Zaltieri et al., 2015). Palmitoylation plays an important role in vesicle trafficking and synaptic activity regulation and increases the hydrophobicity of the proteins, thus accelerating their association with cellular membranes and affecting their localization and function (Ji and Skup, 2021).

Palmitoylation, in synaptic vesicle trafficking, regulates the synaptic protein involved in neurotransmitter release, such as the Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex. For example, Synaptosomal-associated protein of 25 kDa (SNAP25), a component from the SNARE complex, is required for proper localization of the SNARE complex toward the plasma membrane. Disruption in palmitoylation of SNAP25 impacts the synaptic vesicle exocytosis and impairs synaptic transmission and neurotransmitter release (Peng et al., 2024). Furthermore, palmitoylation impacts synaptic strength and plasticity by modulating the clustering and function of the postsynaptic receptor. For instance, the trafficking and surface expression of AMPA-type glutamate receptors is affected by palmitoylation, which is vital for the synaptic plasticity activities that represent learning and memory (Sohn and Park, 2019). Interestingly, changes in palmitoylation have been linked with PD pathogenesis. Focusing on the palmitoylation pathways may provide the potential for therapy in addressing α-synuclein-related neurodegeneration (Zarêba-Kozioł et al., 2021). The aggregation of α-synuclein in PD disrupts mitochondrial transport and function through multiple mechanisms through several processes, such as mitochondrial impairment, interfering with mitochondrial dynamics, reduction in protein import in mitochondria and disrupting microtubule-based transport. This proves that palmitoylation serves as a significant regulatory mechanism in vesicle trafficking and synaptic function, which, when dysregulated, causes synaptic deficits in PD.

The clinical application of palmitoylation-targeted therapies is obstructed by several challenges, despite many promising directions. Many palmitoylating inhibitors like 2-bromopalmitate (2-BP) targets several zDHHC enzymes, which may have neurotoxic effects by interacting with off-target zDHHC enzymes (Chaturvedi et al., 2024). A study conducted by Bai et al. (2024) studied the use of proteolysis-targeting chimeras (PROTACs) to target the membrane bound zDHHC and PAT interactions, and succeeded in degradation of the zDHHC5 and zDHHC20 enzymes and along with reduction in palmitoylation of their substrate. Additionally, the palmitoylation process on the substrate are observed to substituted by other non-targeted zDHHCs. This study in overall demonstrated the feasibility of targeting the zDHHC and PATs interactions, but with the limited enzyme redundancy where other non-targeted zDHHCs are observed to be compensating the loss of targeted zDHHCs. This functional redundancy among zDHHC enzymes complicates targeted therapeutic strategies, as inhibition of one isoform may be compensated by others, limiting long-term efficacy unless combinatorial or context-specific approaches are developed. Many zDHHC enzymes are complicated to target due to insufficient knowledge on the substrate specificity and regulatory mechanisms.

Palmitoylation inhibitors, such as 2-BP, cerulenin, tunicamycin and cyano-myracrylamide (CMA) targets almost all palmitoylation modification, but they lack isoform specificity and broadly inhibit the entire zDHHC family of palmitoyl PATs as well as other unrelated proteins, leading to widespread off-target effects and toxicity (Jennings et al., 2009; Azizi et al., 2021; Yu et al., 2024). The catalytic domains of DHHC enzymes are highly conserved, making it technically challenging to design isoform or substrate-specific inhibitors (De and Sadhukhan, 2018). Natural flavonoids such as lutein, 5-hydroxyflavone, and 6-hydroxyflavone have been identified based on in silico screening as selective inhibitors of zDHHC20-mediated palmitoylation, represents a different therapeutic approach with potentially reduced adverse consequences (Chaturvedi et al., 2024). Several researchers are actively involved in developing palmitoylation inhibitors or activators as a neuroprotective strategy. For example, 2-BP and tunicamycin, a zDHHC inhibitor have been explored to check for their potential in mitigating the pathological palmitoylation in neurodegenerative diseases, but their lack of specificity and unintended cellular effects along with cytotoxic effects sets the limitations (Pei and Piao, 2024).

Lipid-based therapeutics provides additional avenue for modulating protein localization and function through palmitoylation. But, lipid-based molecules often exhibit poor bioavailability, necessitating the development of novel delivery systems to ensure effective therapeutic concentrations in neuronal tissues. A study on nitro-fatty acids (a lipid based therapy) by Hansen et al. (2018) observed that covalent modification of signaling protein stimulator of IFN genes (STING) via nitro-alkylation inhibits the palmitoylation and reduces the STING mediated signaling, a signaling pathway when triggered excessively leads to uncontrolled inflammation and tissue damage in inflammatory and autoimmune disease. Since the nitro-fatty acids utilized in Hansel et al. study is formed in response to viral infection, heir long-term safety, efficacy, and potential side effects remain unclear. Given the conditions, dysregulated palmitoylation modifications on specific zDHHC enzymes likes zDHHC9, zDHHC13, and zDHHC21 are targeted in AD, HD, schizophrenia, and glioma, suggesting as a therapeutic approach (Liao et al., 2024). Moreover, modulating palmitoyl-protein thioesterases (PPTs), which are responsible for depalmitoylation, could restore balance in protein palmitoylation, preventing the toxic accumulation of misfolded proteins characteristic of neurodegenerative disorders. There is a lack of robust clinical data demonstrating efficacy or safety in humans, and the blood-brain barrier remains a significant obstacle for CNS-targeted delivery (Wlodarczyk et al., 2024). The major challenge in developing lipid-based inhibitors is the poor solubility of small molecules. However, recent advances in the liposomal transport delivery of hydrophobic medications could offer a potential solution.

This review consolidates current mechanistic understanding of palmitoylation’s role in modulating microtubules, motor proteins, and microtubule-associated proteins, emphasizing their dysregulation in Alzheimer’s, Parkinson’s, and ALS. It critically appraises therapeutic opportunities to selectively target palmitoylation enzymes, acknowledges challenges such as inhibitor specificity and delivery, and underscores promising research directions including gene therapy, RNA-based modulation, and lipid nanoparticle delivery systems. Future research could explore the development of isoform- and substrate-specific modulators by targeting unique N- or C-terminal regions of DHHC enzymes or leveraging structural distinctions identified in recent crystallography studies. Additionally, investigating advanced delivery systems, such as nanoparticles or viral vector-based approaches may enhance brain bioavailability while minimizing peripheral toxicity and systemic exposure. Another promising avenue could involve identifying palmitoylation-specific biomarkers (e.g., through S-acylation profiling) to monitor target engagement in clinical trials. Advancements in acyl-biotin exchange assays and mass spectrometry-based S-palmitoylome profiling may enable the identification of palmitoylation-specific biomarkers, offering tools for both diagnosis and therapeutic monitoring (Zhao et al., 2010). Integrating palmitoylation modulation into neurodegeneration treatments requires multifaceted strategies, such as combining palmitoylation modulators with existing neuroprotective drugs could enhance synaptic stability and function in diseases like AD and PD. Personalized medicine approaches, involving the identification of patient-specific zDHHC mutations, could enable tailored treatments for various neurodegenerative disorders. Overall, while the therapeutic modulation of palmitoylation holds significant promise, realizing its clinical impact will require overcoming technical, biological, and regulatory barriers through collaborative, interdisciplinary research.