Regulation of proteostasis by sleep through autophagy in Drosophila models of Alzheimer's disease.

Sleep and circadian rhythm dysfunctions are common clinical features of Alzheimer's disease (AD). Increasing evidence suggests that in addition to being a symptom, sleep disturbances can also drive the progression of neurodegeneration. Protein aggregation is a pathological hallmark of AD; however, the molecular pathways behind how sleep affects protein homeostasis remain elusive. Here we demonstrate that sleep modulation influences proteostasis and the progression of neurodegeneration in Drosophila models of tauopathy. We show that sleep deprivation enhanced Tau aggregational toxicity resulting in exacerbated synaptic degeneration. In contrast, sleep induction using gaboxadol led to reduced toxic Tau accumulation in neurons as a result of modulated autophagic flux and enhanced clearance of ubiquitinated Tau, suggesting altered protein processing and clearance that resulted in improved synaptic integrity and function. These findings highlight the complex relationship between sleep and regulation of protein homeostasis and the neuroprotective potential of sleep-enhancing therapeutics to slow the progression or delay the onset of neurodegeneration.

Loss of Fic causes progressive neurodegeneration in a Drosophila model of hereditary spastic paraplegia.

Hereditary Spastic Paraplegia (HSP) is a group of rare inherited disorders characterized by progressive weakness and spasticity of the legs. Recent newly discovered biallelic variants in the gene FICD were found in patients with a highly similar phenotype to early onset HSP. FICD encodes filamentation induced by cAMP domain protein. FICD is involved in the AMPylation and deAMPylation protein modifications of the endoplasmic reticulum (ER) chaperone BIP, a major constituent of the ER that regulates the unfolded protein response. Although several biochemical properties of FICD have been characterized, the neurological function of FICD and the pathological mechanism underlying HSP are unknown. We established a Drosophila model to gain mechanistic understanding of the function of FICD in HSP pathogenesis, and specifically the role of BIP in neuromuscular physiology. Our studies on Drosophila Fic null mutants uncovered that loss of Fic resulted in locomotor impairment and reduced levels of BIP in the motor neuron circuitry, as well as increased reactive oxygen species (ROS) in the ventral nerve cord of Fic null mutants. Finally, feeding Drosophila Fic null mutants with chemical chaperones PBA or TUDCA, or treatment of patient fibroblasts with PBA, reduced the ROS accumulation. The neuronal phenotypes of Fic null mutants recapitulate several clinical features of HSP patients and further reveal cellular patho-mechanisms. By modeling FICD in Drosophila, we provide potential targets for intervention for HSP, and advance fundamental biology that is important for understanding related rare and common neuromuscular diseases.

High-resolution, high-throughput analysis of Drosophila geotactic behavior.

Drosophila innate response to gravity, geotaxis, has been previously used to assess the impact of aging and disease on motor performance. Despite its rich history, fly geotaxis continues to be largely measured manually and assessed through simplistic metrics. The manual nature of this assay introduces substantial experimental variability while simplistic metrics provide limited analytic insights into the behavior. To address these shortcomings, we have constructed a fully automated, programable apparatus, and developed a multi-object tracking software capable of following sub-second movements of individual flies, thus allowing reproducible, detailed, and quantitative analysis of geotactic behavior. The apparatus triggers and monitors geotaxis of 10 fly cohorts simultaneously, with each cohort consisting of up to 7 flies. The tracking program isolates cohorts and records individual fly coordinate outputs allowing for simultaneous multi-group, multi-fly tracks per experiment, greatly improving throughput and resolution. The algorithm tracks individual flies during the entire run with ~97% accuracy, yielding detailed climbing curve, speed, and movement direction with 1/30 second resolution. Our tracking also allows the construction of multi-variable metrics and the detection of transitory movement phenotypes, such as slips and falls, which have thus far been neglected in geotaxis studies due to limited spatio-temporal resolution. Through a combination of automation and robust tracking, the platform is therefore poised to advance Drosophila geotaxis assay into a comprehensive assessment of locomotor behavior.

Reduction of spermine synthase enhances autophagy to suppress Tau accumulation.

Precise polyamine metabolism regulation is vital for cells and organisms. Mutations in spermine synthase (SMS) cause Snyder-Robinson intellectual disability syndrome (SRS), characterized by significant spermidine accumulation and autophagy blockage in the nervous system. Emerging evidence connects polyamine metabolism with other autophagy-related diseases, such as Tauopathy, however, the functional intersection between polyamine metabolism and autophagy in the context of these diseases remains unclear. Here, we altered SMS expression level to investigate the regulation of autophagy by modulated polyamine metabolism in Tauopathy in Drosophila and human cellular models. Interestingly, while complete loss of Drosophila spermine synthase (dSms) impairs lysosomal function and blocks autophagic flux recapitulating SRS disease phenotype, partial loss of dSms enhanced autophagic flux, reduced Tau protein accumulation, and led to extended lifespan and improved climbing performance in Tauopathy flies. Measurement of polyamine levels detected a mild elevation of spermidine in flies with partial loss of dSms. Similarly, in human neuronal or glial cells, partial loss of SMS by siRNA-mediated knockdown upregulated autophagic flux and reduced Tau protein accumulation. Importantly, proteomics analysis of postmortem brain tissue from Alzheimer's disease (AD) patients showed a significant albeit modest elevation of SMS level. Taken together, our study uncovers a functional correlation between polyamine metabolism and autophagy in AD: SMS reduction upregulates autophagy, suppresses Tau accumulation, and ameliorates neurodegeneration and cell death. These findings provide a new potential therapeutic target for AD.

Copper enhances aggregational toxicity of mutant huntingtin in a Drosophila model of Huntington's Disease.

Huntington's disease (HD) is a progressive neurodegenerative disorder with clinical presentations of moderate to severe cognitive, motor, and psychiatric disturbances. HD is caused by the trinucleotide repeat expansion of CAG of the huntingtin (HTT) gene. The mutant HTT protein containing pathological polyglutamine (polyQ) extension is prone to misfolding and aggregation in the brain. It has previously been observed that copper and iron concentrations are increased in the striata of post-mortem human HD brains. Although it has been shown that the accumulation of mutant HTT protein can interact with copper, the underlying HD progressive phenotypes due to copper overload remains elusive. Here, in a Drosophila model of HD, we showed that copper induces dose-dependent aggregational toxicity and enhancement of Htt-induced neurodegeneration. Specifically, we found that copper increases mutant Htt aggregation, enhances the accumulation of Thioflavin S positive ß-amyloid structures within Htt aggregates, and consequently alters autophagy in the brain. Administration of copper chelator D-penicillamine (DPA) through feeding significantly decreases ß-amyloid aggregates in the HD pathological model. These findings reveal a direct role of copper in potentiating mutant Htt protein-induced aggregational toxicity, and further indicate the potential impact of environmental copper exposure in the disease onset and progression of HD.

Difluoromethylornithine rebalances aberrant polyamine ratios in Snyder-Robinson syndrome.

Snyder-Robinson syndrome (SRS) results from mutations in spermine synthase (SMS), which converts the polyamine spermidine into spermine. Affecting primarily males, common manifestations of SRS include intellectual disability, osteoporosis, hypotonia, and seizures. Symptom management is the only treatment. Reduced SMS activity causes spermidine accumulation while spermine levels are reduced. The resulting exaggerated spermidine:spermine ratio is a biochemical hallmark of SRS that tends to correlate with symptom severity. Our studies aim to pharmacologically manipulate polyamine metabolism to correct this imbalance as a therapeutic strategy for SRS. Here we report the repurposing of 2-difluoromethylornithine (DFMO), an FDA-approved inhibitor of polyamine biosynthesis, in rebalancing spermidine:spermine ratios in SRS patient cells. Mechanistic in vitro studies demonstrate that, while reducing spermidine biosynthesis, DFMO also stimulates the conversion of spermidine into spermine in hypomorphic SMS cells and induces uptake of exogenous spermine, altogether reducing the aberrant ratios. In a Drosophila SRS model characterized by reduced lifespan, DFMO improves longevity. As nearly all SRS patient mutations are hypomorphic, these studies form a strong foundation for translational studies with significant therapeutic potential.

The Architecture of the Presynaptic Release Site.

The architecture of the presynaptic release site is exquisitely designed to facilitate and regulate synaptic vesicle exocytosis. With the identification of some of the building blocks of the active zone and the advent of super resolution imaging techniques, we are beginning to understand the morphological and functional properties of synapses in great detail. Presynaptic release sites consist of the plasma membrane, the cytomatrix, and dense projections. These three components are morphologically distinct but intimately connected with each other and with postsynaptic specializations, ensuring the fidelity of synaptic vesicle tethering, docking, and fusion, as well as signal detection. Although the morphology and molecular compositions of active zones may vary among species, tissues, and cells, global architectural design of the release sites is highly conserved.

Presynaptic Cytomatrix Proteins.

The Cytomatrix Assembled at the active Zone (CAZ) of a presynaptic terminal displays electron-dense appearance and defines the center of the synaptic vesicle release. The protein constituents of CAZ are multiple-domain scaffolds that interact extensively with each other and also with an ensemble of synaptic vesicle proteins to ensure docking, fusion, and recycling. Reflecting the central roles of the active zone in synaptic transmission, CAZ proteins are highly conserved throughout evolution. As the nervous system increases complexity and diversity in types of neurons and synapses, CAZ proteins expand in the number of gene and protein isoforms and interacting partners. This chapter summarizes the discovery of the core CAZ proteins and current knowledge of their functions.

MicroRNA-Mediated Obstruction of Stem-loop Alternative Splicing (MIMOSAS): a global mechanism for the regulation of alternative splicing.

While RNA secondary structures are critical to regulate alternative splicing of long-range pre-mRNA, the factors that modulate RNA structure and interfere with the recognition of the splice sites are largely unknown. Previously, we identified a small, non-coding microRNA that sufficiently affects stable stem structure formation of Nmnat pre-mRNA to regulate the outcomes of alternative splicing. However, the fundamental question remains whether such microRNA-mediated interference with RNA secondary structures is a global molecular mechanism for regulating mRNA splicing. We designed and refined a bioinformatic pipeline to predict candidate microRNAs that potentially interfere with pre-mRNA stem-loop structures, and experimentally verified splicing predictions of three different long-range pre-mRNAs in the Drosophila model system. Specifically, we observed that microRNAs can either disrupt or stabilize stem-loop structures to influence splicing outcomes. Our study suggests that MicroRNA-Mediated Obstruction of Stem-loop Alternative Splicing (MIMOSAS) is a novel regulatory mechanism for the transcriptome-wide regulation of alternative splicing, increases the repertoire of microRNA function and further indicates cellular complexity of post-transcriptional regulation. One-Sentence Summary: MicroRNA-Mediated Obstruction of Stem-loop Alternative Splicing (MIMOSAS) is a novel regulatory mechanism for the transcriptome-wide regulation of alternative splicing.

Sorbitol reduction via govorestat ameliorates synaptic dysfunction and neurodegeneration in sorbitol dehydrogenase deficiency.

Sorbitol dehydrogenase (SORD) deficiency has been identified as the most frequent autosomal recessive form of hereditary neuropathy. Loss of SORD causes high sorbitol levels in tissues due to the inability to convert sorbitol to fructose in the 2-step polyol pathway, leading to degenerative neuropathy. The underlying mechanisms of sorbitol-induced degeneration have not been fully elucidated, and no current FDA-approved therapeutic options are available to reduce sorbitol levels in the nervous system. Here, in a Drosophila model of SORD deficiency, we showed synaptic degeneration in the brain, neurotransmission defect, locomotor impairment, and structural abnormalities in the neuromuscular junctions. In addition, we found reduced ATP production in the brain and ROS accumulation in the CNS and muscle, indicating mitochondrial dysfunction. Applied Therapeutics has developed a CNS-penetrant next-generation aldose reductase inhibitor (ARI), AT-007 (govorestat), which inhibits the conversion of glucose to sorbitol. AT-007 significantly reduced sorbitol levels in patient-derived fibroblasts, induced pluripotent stem cell-derived (iPSC-derived) motor neurons, and Drosophila brains. AT-007 feeding in Sord-deficient Drosophila mitigated synaptic degeneration and significantly improved synaptic transduction, locomotor activity, and mitochondrial function. Moreover, AT-007 treatment significantly reduced ROS accumulation in Drosophila CNS, muscle, and patient-derived fibroblasts. These findings uncover the molecular and cellular pathophysiology of SORD neuropathy and provide a potential treatment strategy for patients with SORD deficiency.

Reduction of Spermine Synthase Suppresses Tau Accumulation Through Autophagy Modulation in Tauopathy.

Tauopathy, including Alzheimer Disease (AD), is characterized by Tau protein accumulation and autophagy dysregulation. Emerging evidence connects polyamine metabolism with the autophagy pathway, however the role of polyamines in Tauopathy remains unclear. In the present study we investigated the role of spermine synthase (SMS) in autophagy regulation and tau protein processing in Drosophila and human cellular models of Tauopathy. Our previous study showed that Drosophila spermine synthase (dSms) deficiency impairs lysosomal function and blocks autophagy flux. Interestingly, partial loss-of-function of SMS in heterozygous dSms flies extends lifespan and improves the climbing performance of flies with human Tau (hTau) overexpression. Mechanistic analysis showed that heterozygous loss-of-function mutation of dSms reduces hTau protein accumulation through enhancing autophagic flux. Measurement of polyamine levels detected a mild elevation of spermidine in flies with heterozygous loss of dSms. SMS knock-down in human neuronal or glial cells also upregulates autophagic flux and reduces Tau protein accumulation. Proteomics analysis of postmortem brain tissue from AD patients showed a significant albeit modest elevation of SMS protein level in AD-relevant brain regions compared to that of control brains consistently across several datasets. Taken together, our study uncovers a correlation between SMS protein level and AD pathogenesis and reveals that SMS reduction upregulates autophagy, promotes Tau clearance, and reduces Tau protein accumulation. These findings provide a new potential therapeutic target of Tauopathy.

Specific binding of Hsp27 and phosphorylated Tau mitigates abnormal Tau aggregation-induced pathology.

Amyloid aggregation of phosphorylated Tau (pTau) into neurofibrillary tangles is closely associated with Alzheimer's disease (AD). Several molecular chaperones have been reported to bind Tau and impede its pathological aggregation. Recent findings of elevated levels of Hsp27 in the brains of patients with AD suggested its important role in pTau pathology. However, the molecular mechanism of Hsp27 in pTau aggregation remains poorly understood. Here, we show that Hsp27 partially co-localizes with pTau tangles in the brains of patients with AD. Notably, phosphorylation of Tau by microtubule affinity regulating kinase 2 (MARK2), dramatically enhances the binding affinity of Hsp27 to Tau. Moreover, Hsp27 efficiently prevents pTau fibrillation in vitro and mitigates neuropathology of pTau aggregation in a Drosophila tauopathy model. Further mechanistic study reveals that Hsp27 employs its N-terminal domain to directly interact with multiple phosphorylation sites of pTau for specific binding. Our work provides the structural basis for the specific recognition of Hsp27 to pathogenic pTau, and highlights the important role of Hsp27 in preventing abnormal aggregation and pathology of pTau in AD.

Phenylbutyrate modulates polyamine acetylase and ameliorates Snyder-Robinson syndrome in a Drosophila model and patient cells.

Polyamine dysregulation plays key roles in a broad range of human diseases from cancer to neurodegeneration. Snyder-Robinson syndrome (SRS) is the first known genetic disorder of the polyamine pathway, caused by X-linked recessive loss-of-function mutations in spermine synthase. In the Drosophila SRS model, altered spermidine/spermine balance has been associated with increased generation of ROS and aldehydes, consistent with elevated spermidine catabolism. These toxic byproducts cause mitochondrial and lysosomal dysfunction, which are also observed in cells from SRS patients. No efficient therapy is available. We explored the biochemical mechanism and discovered acetyl-CoA reduction and altered protein acetylation as potentially novel pathomechanisms of SRS. We repurposed the FDA-approved drug phenylbutyrate (PBA) to treat SRS using an in vivo Drosophila model and patient fibroblast cell models. PBA treatment significantly restored the function of mitochondria and autolysosomes and extended life span in vivo in the Drosophila SRS model. Treating fibroblasts of patients with SRS with PBA ameliorated autolysosome dysfunction. We further explored the mechanism of drug action and found that PBA downregulates the first and rate-limiting spermidine catabolic enzyme spermidine/spermine N1-acetyltransferase 1 (SAT1), reduces the production of toxic metabolites, and inhibits the reduction of the substrate acetyl-CoA. Taken together, we revealed PBA as a potential modulator of SAT1 and acetyl-CoA levels and propose PBA as a therapy for SRS and potentially other polyamine dysregulation-related diseases.

Human Nmnat1 Promotes Autophagic Clearance of Amyloid Plaques in a Drosophila Model of Alzheimer's Disease.

Alzheimer's disease (AD) is a progressive neurodegenerative disease characterized by irreversible cognitive decline with limited therapeutic approaches. We characterized a Drosophila model of amyloid pathology that expresses human amyloid-beta precursor protein (APP695) and ß-site APP cleaving enzyme (BACE) in the nervous system. Our model recapitulates in vivo the age-dependent accumulation of BACE-derived C-terminal fragment (CTF) and amyloid plaques in the brain, one of the key pathological hallmarks of AD. Using this model, we assessed the effects on plaque formation of Nicotinamide mononucleotide adenylyltransferase (Nmnat), an evolutionarily conserved nicotinamide adenine dinucleotide (NAD+) synthase involved in cellular metabolism and neuroprotection. We compared the effects of overexpression of Drosophila Nmnat (dNmnat), human Nmnat1 (hNmnat1), human Nmnat2 (hNmnat2), and human Nmnat3 (hNmnat3), and observed that hNmnat1 has the highest efficacy in reducing amyloid aggregation and APP-CTF accumulation. Interestingly, we demonstrated that overexpression of hNmnat1 reduces amyloid plaques by promoting autophagic clearance. Our findings uncover a role of hNmnat1 in amyloid clearance and suggest an exciting neuroprotective potential of hNmnat1 in amyloid pathology.

NMNAT promotes glioma growth through regulating post-translational modifications of P53 to inhibit apoptosis.

Gliomas are highly malignant brain tumors with poor prognosis and short survival. NAD+ has been shown to impact multiple processes that are dysregulated in cancer; however, anti-cancer therapies targeting NAD+ synthesis have had limited success due to insufficient mechanistic understanding. Here, we adapted a Drosophila glial neoplasia model and discovered the genetic requirement for NAD+ synthase nicotinamide mononucleotide adenylyltransferase (NMNAT) in glioma progression in vivo and in human glioma cells. Overexpressing enzymatically active NMNAT significantly promotes glial neoplasia growth and reduces animal viability. Mechanistic analysis suggests that NMNAT interferes with DNA damage-p53-caspase-3 apoptosis signaling pathway by enhancing NAD+-dependent posttranslational modifications (PTMs) poly(ADP-ribosyl)ation (PARylation) and deacetylation of p53. Since PARylation and deacetylation reduce p53 pro-apoptotic activity, modulating p53 PTMs could be a key mechanism by which NMNAT promotes glioma growth. Our findings reveal a novel tumorigenic mechanism involving protein complex formation of p53 with NAD+ synthetic enzyme NMNAT and NAD+-dependent PTM enzymes that regulates glioma growth.

One of the most common types of brain cancer, glioma, emerges when harmful mutations take place in the 'glial' cells tasked with supporting neurons. When these genetically damaged cells are not fixed or eliminated, they can go on to multiply uncontrollability. A protein known as p53 can help to repress emerging tumors by stopping mutated cells in their tracks. Glioma is a highly deadly cancer, and treatments are often ineffective. Some of these approaches have focused on a protein involved in the creation of the coenzyme NAD+, which is essential to the life processes of all cells. However, these drugs have had poor outcomes. Instead, Liu et al. focused on NMNAT, the enzyme that participates in the final stage of the creation of NAD+. NMNAT is known to protect neurons, but it is unclear how it involved in cancer. Experiments in fruit flies which were then validated in human glioma cells showed that increased NMNAT activity allowed glial cells with harmful mutations to survive and multiply. Detailed molecular analysis showed that NMNAT orchestrates chemical modifications that inactivate p53. It does so by working with other molecular actors to direct NAD+ to add and remove chemical groups that control the activity of p53. Taken together, these results show how NMNAT can participate in the emergence of brain cancers. They also highlight the need for further research on whether drugs that inhibit this enzyme could help to suppress tumors before they become deadly.

Development of a Redox-Sensitive Spermine Prodrug for the Potential Treatment of Snyder Robinson Syndrome.

Snyder Robinson Syndrome (SRS) is a rare disease associated with a defective spermine synthase gene and low intracellular spermine levels. In this study, a spermine replacement therapy was developed using a spermine prodrug that enters cells via the polyamine transport system. The prodrug was comprised of three components: a redox-sensitive quinone "trigger", a "trimethyl lock (TML)" aryl "release mechanism", and spermine. The presence of spermine in the design facilitated uptake by the polyamine transport system. The quinone-TML motifs provided a redox-sensitive agent, which upon intracellular reduction generated a hydroquinone, which underwent intramolecular cyclization to release free spermine and a lactone byproduct. Rewardingly, most SRS fibroblasts treated with the prodrug revealed a significant increase in intracellular spermine. Administering the spermine prodrug through feeding in a Drosophila model of SRS showed significant beneficial effects. In summary, a spermine prodrug is developed and provides a lead compound for future spermine replacement therapy experiments.

Exposure to Aerosolized Algal Toxins in South Florida Increases Short- and Long-Term Health Risk in Drosophila Model of Aging.

Harmful algal blooms (HABs) are a rising health and environmental concern in the United States, particularly in South Florida. Skin contact and the ingestion of contaminated water or fish and other seafood have been proven to have severe toxicity to humans in some cases. However, the impact of aerosolized HAB toxins is poorly understood. In particular, knowledge regarding either the immediate or long-term effects of exposure to aerosolized cyanotoxins produced by freshwater blue-green algae does not exist. The aim of this study was to probe the toxicity of aerosolized cyanobacterial blooms using Drosophila melanogaster as an animal model. The exposure of aerosolized HABs at an early age leads to the most severe long-term impact on health and longevity among all age groups. Young groups and old males showed a strong acute response to HAB exposure. In addition, brain morphological analysis using fluorescence imaging reveals significant indications of brain degeneration in females exposed to aerosolized HABs in early or late stages. These results indicate that one-time exposure to aerosolized HAB particles causes a significant health risk, both immediately and in the long-term. Interestingly, age at the time of exposure plays an important role in the specific nature of the impact of aerosol HABs. As BMAA and microcystin have been found to be the significant toxins in cyanobacteria, the concentration of both toxins in the water and aerosols was examined. BMAA and microcystin are consistently detected in HAB waters, although their concentrations do not always correlate with the severity of the health impact, suggesting the potential contribution from additional toxins present in the aerosolized HAB. This study demonstrates, for the first time, the health risk of exposure to aerosolized HAB, and further highlights the critical need and importance of understanding the toxicity of aerosolized cyanobacteria HAB particles and determining the immediate and long-term health impacts of HAB exposure.

microRNA-92a regulates the expression of aphid bacteriocyte-specific secreted protein 1.

OBJECTIVE: Aphids harbor a nutritional obligate endosymbiont in specialized cells called bacteriocytes, which aggregate to form an organ known as the bacteriome. Aphid bacteriomes display distinct gene expression profiles that facilitate the symbiotic relationship. Currently, the mechanisms that regulate these patterns of gene expression are unknown. Recently using computational pipelines, we identified miRNAs that are conserved in expression in the bacteriomes of two aphid species and proposed that they function as important regulators of bacteriocyte gene expression. Here using a dual luciferase assay in mouse NIH/3T3 cell culture, we aimed to experimentally validate the computationally predicted interaction between Myzus persicae miR-92a and the predicted target region of M. persicae bacteriocyte-specific secreted protein 1 (SP1) mRNA. RESULTS: In the dual luciferase assay, miR-92a interacted with the SP1 target region resulting in a significant downregulation of the luciferase signal. Our results demonstrate that miR-92a interacts with SP1 to alter expression in a heterologous expression system, thereby supporting our earlier assertion that miRNAs are regulators of the aphid/Buchnera symbiotic interaction.

Nmnat restores neuronal integrity by neutralizing mutant Huntingtin aggregate-induced progressive toxicity.

Accumulative aggregation of mutant Huntingtin (Htt) is a primary neuropathological hallmark of Huntington's disease (HD). Currently, mechanistic understanding of the cytotoxicity of mutant Htt aggregates remains limited, and neuroprotective strategies combating mutant Htt-induced neurodegeneration are lacking. Here, we show that in Drosophila models of HD, neuronal compartment-specific accumulation of mutant Htt aggregates causes neurodegenerative phenotypes. In addition to the increase in the number and size, we discovered an age-dependent acquisition of thioflavin S+, amyloid-like adhesive properties of mutant Htt aggregates and a concomitant progressive clustering of aggregates with mitochondria and synaptic proteins, indicating that the amyloid-like adhesive property underlies the neurotoxicity of mutant Htt aggregation. Importantly, nicotinamide mononucleotide adenylyltransferase (NMNAT), an evolutionarily conserved nicotinamide adenine dinucleotide (NAD+) synthase and neuroprotective factor, significantly mitigates mutant Htt-induced neurodegeneration by reducing mutant Htt aggregation through promoting autophagic clearance. Additionally, Nmnat overexpression reduces progressive accumulation of amyloid-like Htt aggregates, neutralizes adhesiveness, and inhibits the clustering of mutant Htt with mitochondria and synaptic proteins, thereby restoring neuronal function. Conversely, partial loss of endogenous Nmnat exacerbates mutant Htt-induced neurodegeneration through enhancing mutant Htt aggregation and adhesive property. Finally, conditional expression of Nmnat after the onset of degenerative phenotypes significantly delays the progression of neurodegeneration, revealing the therapeutic potential of Nmnat-mediated neuroprotection at advanced stages of HD. Our study uncovers essential mechanistic insights to the neurotoxicity of mutant Htt aggregation and describes the molecular basis of Nmnat-mediated neuroprotection in HD.