Moniezia benedeni drives the SNAP-25 expression of the enteric nerves in sheep's small intestine.

BACKGROUND: The neuroimmune network plays a crucial role in regulating mucosal immune homeostasis within the digestive tract. Synaptosome-associated protein 25 (SNAP-25) is a presynaptic membrane-binding protein that activates ILC2s, initiating the host's anti-parasitic immune response. METHODS: To investigate the effect of Moniezia benedeni (M. benedeni) infection on the distribution of SNAP-25 in the sheep's small intestine, the recombinant plasmid pET-28a-SNAP-25 was constructed and expressed in BL21, yielding the recombinant protein. Then, the rabbit anti-sheep SNAP-25 polyclonal antibody was prepared and immunofluorescence staining was performed with it. The expression levels of SNAP-25 in the intestines of normal and M. benedeni-infected sheep were detected by ELISA. RESULTS: The results showed that the SNAP-25 recombinant protein was 29.3 KDa, the titer of the prepared immune serum reached 1:128,000. It was demonstrated that the rabbit anti-sheep SNAP-25 polyclonal antibody could bind to the natural protein of sheep SNAP-25 specifically. The expression levels of SNAP-25 in the sheep's small intestine revealed its primary presence in the muscular layer and lamina propria, particularly around nerve fibers surrounding the intestinal glands. Average expression levels in the duodenum, jejunum, and ileum were 130.32 pg/mg, 185.71 pg/mg, and 172.68 pg/mg, respectively. Under conditions of M. benedeni infection, the spatial distribution of SNAP-25-expressing nerve fibers remained consistent, but its expression level in each intestine segment was increased significantly (P < 0.05), up to 262.02 pg/mg, 276.84 pg/mg, and 326.65 pg/mg in the duodenum, jejunum, and ileum, and it was increased by 101.06%, 49.07%, and 89.16% respectively. CONCLUSIONS: These findings suggest that M. benedeni could induce the SNAP-25 expression levels in sheep's intestinal nerves significantly. The results lay a foundation for further exploration of the molecular mechanism by which the gastrointestinal nerve-mucosal immune network perceives parasites in sheep.

BDNF/TrkB signalling, in cooperation with muscarinic signalling, retrogradely regulates PKA pathway to phosphorylate SNAP-25 and Synapsin-1 at the neuromuscular junction.

BACKGROUND: Protein kinase A (PKA) enhances neurotransmission at the neuromuscular junction (NMJ), which is retrogradely regulated by nerve-induced muscle contraction to promote Acetylcholine (ACh) release through the phosphorylation of molecules involved in synaptic vesicle exocytosis (SNAP-25 and Synapsin-1). However, the molecular mechanism of the retrograde regulation of PKA subunits and its targets by BDNF/TrkB pathway and muscarinic signalling has not been demonstrated until now. At the NMJ, retrograde control is mainly associated with BDNF/TrkB signalling as muscle contraction enhances BDNF levels and controls specific kinases involved in the neurotransmission. Neurotransmission at the NMJ is also highly modulated by muscarinic receptors M1 and M2 (mAChRs), which are related to PKA and TrkB signallings. Here, we investigated the hypothesis that TrkB, in cooperation with mAChRs, regulates the activity-dependent dynamics of PKA subunits to phosphorylate SNAP-25 and Synapsin-1. METHODS: To explore this, we stimulated the rat phrenic nerve at 1Hz (30 minutes), with or without subsequent contraction (abolished by µ-conotoxin GIIIB). Pharmacological treatments were conducted with the anti-TrkB antibody clone 47/TrkB for TrkB inhibition and exogenous h-BDNF; muscarinic inhibition with Pirenzepine-dihydrochloride and Methoctramine-tetrahydrochloride for M1 and M2 mAChRs, respectively. Diaphragm protein levels and phosphorylation' changes were detected by Western blotting. Location of the target proteins was demonstrated using immunohistochemistry. RESULTS: While TrkB does not directly impact the levels of PKA catalytic subunits Cα and Cß, it regulates PKA regulatory subunits RIα and RIIß, facilitating the phosphorylation of critical exocytotic targets such as SNAP-25 and Synapsin-1. Furthermore, the muscarinic receptors pathway maintains a delicate balance in this regulatory process. These findings explain the dynamic interplay of PKA subunits influenced by BDNF/TrkB signalling, M1 and M2 mAChRs pathways, that are differently regulated by pre- and postsynaptic activity, demonstrating the specific roles of the BDNF/TrkB and muscarinic receptors pathway in retrograde regulation. CONCLUSION: This complex molecular interplay has the relevance of interrelating two fundamental pathways in PKA-synaptic modulation: one retrograde (neurotrophic) and the other autocrine (muscarinic). This deepens the fundamental understanding of neuromuscular physiology of neurotransmission that gives plasticity to synapses and holds the potential for identifying therapeutic strategies in conditions characterized by impaired neuromuscular communication.

Cerebrospinal fluid biomarkers and cognitive trajectories in patients with Alzheimer's disease and a history of traumatic brain injury.

Traumatic brain injury (TBI) and Alzheimer's disease (AD) have overlapping mechanisms but it remains unknown if pathophysiological characteristics and cognitive trajectories in AD patients are influenced by TBI history. Here, we studied AD patients (stage MCI or dementia) with TBI history (ADTBI+, n=110), or without (ADTBI-, n=110) and compared baseline CSF concentrations of amyloid beta 1-42 (Aß42), phosphorylated tau181 (pTau181), total tau, neurofilament light chain (NfL), synaptosomal associated protein-25kDa (SNAP25), neurogranin (Ng), neuronal pentraxin-2 (NPTX2) and glutamate receptor-4 (GluR4), as well as differences in cognitive trajectories using linear mixed models. Explorative, analyses were repeated within stratified TBI groups by TBI characteristics (timing, severity, number). We found no differences in baseline CSF biomarker concentrations nor in cognitive trajectories between ADTBI+ and ADTBI- patients. TBI >5 years ago was associated with higher NPTX2 and a tendency for higher SNAP25 concentrations compared to TBI ≤ 5 years ago, suggesting that TBI may be associated with long-term synaptic dysfunction only when occurring before onset or in a pre-clinical disease stage of AD.

Immunohistochemical localization of proteins involved in exocytosis of glutamate from P2X3 purinoceptor-expressing subserosal afferent nerve endings in the rat gastric antrum.

We previously reported the presence of P2X3 purinoceptors (P2X3)-expressing subserosal afferent nerve endings consisting of net- and basket-like nerve endings in the rat gastric antrum. These nerve endings may morphologically be vagal mechanoreceptors activated by antral peristalsis. The present study investigated immunoreactivities for vesicular glutamate transporter (VGLUT) 1 and VGLUT2 as well as exocytosis-related proteins, i.e., core components of the SNARE complex (SNAP25, Stx1, and VAMP2) and synaptotagmin-1 (Syt1), in whole-mount preparations of the rat gastric antrum using double immunofluorescence. VGLUT1 immunoreactivity was not detected, whereas VGLUT2 immunoreactivity was observed in P2X3-immunoreactive subserosal nerve endings composed of both net- and basket-like endings. In net-like nerve endings, intense VGLUT2 immunoreactivity was localized in polygonal bulges of reticular nerve fibers and peripheral axon terminals. Furthermore, intense immunoreactivities for SNAP25, Stx1, and VAMP2 were localized in net-like nerve endings. Intense immunoreactivities for VAMP2 and Syt1 were observed in VGLUT2-immunoreactive net-like nerve endings. In basket-like nerve endings, VGLUT2 immunoreactivity was localized in pleomorphic terminal structures and small bulges surrounding the subserosal ganglion, whereas immunoreactivities for SNAP25, Stx1, and VAMP2 were weak in these nerve endings. VGLUT2-immunoreactive basket-like nerve endings were weakly immunoreactive for VAMP2 and Syt1. These results suggest that subserosal afferent nerve endings release glutamate by exocytosis mainly from net-like nerve endings to modulate their mechanoreceptor function.

PTPRS is a novel marker for early Tau pathology and synaptic integrity in Alzheimer's disease.

We examined the role of protein tyrosine phosphatase receptor sigma (PTPRS) in the context of Alzheimer's disease and synaptic integrity. Publicly available datasets (BRAINEAC, ROSMAP, ADC1) and a cohort of asymptomatic but "at risk" individuals (PREVENT-AD) were used to explore the relationship between PTPRS and various Alzheimer's disease biomarkers. We identified that PTPRS rs10415488 variant C shows features of neuroprotection against early Tau pathology and synaptic degeneration in Alzheimer's disease. This single nucleotide polymorphism correlated with higher PTPRS transcript abundance and lower p(181)Tau and GAP-43 levels in the CSF. In the brain, PTPRS protein abundance was significantly correlated with the quantity of two markers of synaptic integrity: SNAP25 and SYT-1. We also found the presence of sexual dimorphism for PTPRS, with higher CSF concentrations in males than females. Male carriers for variant C were found to have a 10-month delay in the onset of AD. We thus conclude that PTPRS acts as a neuroprotective receptor in Alzheimer's disease. Its protective effect is most important in males, in whom it postpones the age of onset of the disease.

SYT4 binds to SNAP25 to facilitate exosomal secretion and prostate cancer enzalutamide resistance.

Prostate carcinoma represents a predominant malignancy affecting the male population, with androgen deprivation therapy (ADT) serving as a critical therapeutic modality for advanced disease states, but it often leads to the development of resistance. Enzalutamide (Enz), a second-generation antiandrogen drug, initially offers substantial therapeutic benefit, but its efficacy wanes as drug resistance ensues. In this study, we found that synaptotagmin 4 (SYT4) is an upregulated gene in enzalutamide-resistant (EnzR) cell lines. The downregulation of SYT4, in combination with enzalutamide therapy, substantially enhances the antiproliferative effect on resistant prostate cancer cells beyond the capacity of enzalutamide monotherapy. SYT4 promotes vesicle efflux by binding to the synaptosome-associated protein 25 (SNAP25), thereby contributing to cell resistance against enzalutamide. The elevated expression of SYT4 is mediated by bromodomain-containing protein 4 (BRD4), and BRD4 inhibition effectively suppressed the expression of SYT4. Treatment with a therapeutic dose of enzalutamide combined with ASO-1, an antisense oligonucleotide drug targeting SYT4, shows promising results in reversing the resistance of prostate cancer to enzalutamide.

The impact of the migraine treatment onabotulinumtoxinA on inflammatory and pain responses: Insights from an animal model.

OBJECTIVE: Migraine, a prevalent and debilitating disease, involves complex pathophysiology possibly including inflammation and heightened pain sensitivity. The current study utilized the complete Freund's adjuvant (CFA) model of inflammation, with onabotulinumtoxinA (BoNT/A) as a treatment of interest due to its use in clinical migraine management. Using an animal model, the study sought to investigate the role of BoNT/A in modulating CFA-induced inflammation, alterations in pain sensitivity, and the regulation of calcitonin gene-related peptide (CGRP) release. Further, we aimed to assess the changes in SNAP-25 through western blot analysis to gain insights into the mechanistic action of BoNT/A. METHODS: BoNT/A or control was administered subcutaneously at the periorbital region of rats 3 days before the induction of inflammation using CFA. Periorbital mechanical sensitivity was assessed post-inflammation, and alterations in CGRP release were evaluated. Changes in SNAP-25 levels were determined using western blot analysis. RESULTS: Upon CFA-induced inflammation, there was a marked increase in periorbital mechanical sensitivity, with the inflammation side showing increased sensitivity compared to other periorbital areas. BoNT/A did decrease the withdrawal thresholds in the electronic von Frey test. Despite not being able to observe differences in pain thresholds or CGRP release, BoNT/A reduced baseline release under CFA inflamed conditions. Analysis of SNAP-25 levels in the trigeminal ganglion revealed both intact and cleaved forms that were notably elevated in BoNT/A-treated animals. These findings, derived from western blot analysis, suggest an effect on neurotransmitter release. CONCLUSION: Our investigation highlights the role of BoNT/A in reducing baseline CGRP in the context of inflammation and its involvement in SNAP-25 cleavage. In contrast, BoNT/A did not appear to alter facial pain sensitivity induced by inflammation, suggesting that mechanisms other than baseline CGRP could be implicated in the elevated thresholds in the CFA model.

Brain Expression Levels of Commonly Measured Blood Biomarkers of Neurological Damage Differ with Respect to Sex, Race, and Age.

It is increasingly evident that blood biomarkers have potential to improve the diagnosis and management of both acute and chronic neurological conditions. The most well-studied candidates, and arguably those with the broadest utility, are proteins that are highly enriched in neural tissues and released into circulation upon cellular damage. It is currently unknown how the brain expression levels of these proteins is influenced by demographic factors such as sex, race, and age. Given that source tissue abundance is likely a key determinant of the levels observed in the blood during neurological pathology, understanding such influences is important in terms of identifying potential clinical scenarios that could produce diagnostic bias. In this study, we leveraged existing mRNA sequencing data originating from 2,642 normal brain specimens harvested from 382 human donors to examine potential demographic variability in the expression levels of genes which code for 28 candidate blood biomarkers of neurological damage. Existing mass spectrometry data originating from 26 additional normal brain specimens harvested from 26 separate human donors was subsequently used to tentatively assess whether observed transcriptional variance was likely to produce corresponding variance in terms of protein abundance. Genes associated with several well-studied or emerging candidate biomarkers including neurofilament light chain (NfL), ubiquitin carboxyl-terminal hydrolase isozyme L1 (UCH-L1), neuron-specific enolase (NSE), and synaptosomal-associated protein 25 (SNAP-25) exhibited significant differences in expression with respect to sex, race, and age. In many instances, these differences in brain expression align well with and provide a mechanistic explanation for previously reported differences in blood levels.

Gypenoside XVII Reduces Synaptic Glutamate Release and Protects against Excitotoxic Injury in Rats.

Excitotoxicity is a common pathological process in neurological diseases caused by excess glutamate. The purpose of this study was to evaluate the effect of gypenoside XVII (GP-17), a gypenoside monomer, on the glutamatergic system. In vitro, in rat cortical nerve terminals (synaptosomes), GP-17 dose-dependently decreased glutamate release with an IC50 value of 16 µM. The removal of extracellular Ca2+ or blockade of N-and P/Q-type Ca2+ channels and protein kinase A (PKA) abolished the inhibitory effect of GP-17 on glutamate release from cortical synaptosomes. GP-17 also significantly reduced the phosphorylation of PKA, SNAP-25, and synapsin I in cortical synaptosomes. In an in vivo rat model of glutamate excitotoxicity induced by kainic acid (KA), GP-17 pretreatment significantly prevented seizures and rescued neuronal cell injury and glutamate elevation in the cortex. GP-17 pretreatment decreased the expression levels of sodium-coupled neutral amino acid transporter 1, glutamate synthesis enzyme glutaminase and vesicular glutamate transporter 1 but increased the expression level of glutamate metabolism enzyme glutamate dehydrogenase in the cortex of KA-treated rats. In addition, the KA-induced alterations in the N-methyl-D-aspartate receptor subunits GluN2A and GluN2B in the cortex were prevented by GP-17 pretreatment. GP-17 also prevented the KA-induced decrease in cerebral blood flow and arginase II expression. These results suggest that (i) GP-17, through the suppression of N- and P/Q-type Ca2+ channels and consequent PKA-mediated SNAP-25 and synapsin I phosphorylation, reduces glutamate exocytosis from cortical synaptosomes; and (ii) GP-17 has a neuroprotective effect on KA-induced glutamate excitotoxicity in rats through regulating synaptic glutamate release and cerebral blood flow.

Intramuscular Botulinum Neurotoxin Serotypes E and A Elicit Distinct Effects on SNAP25 Protein Fragments, Muscular Histology, Spread and Neuronal Transport: An Integrated Histology-Based Study in the Rat.

Botulinum neurotoxins E (BoNT/E) and A (BoNT/A) act by cleaving Synaptosome-Associated Protein 25 (SNAP25) at two different C-terminal sites, but they display very distinct durations of action, BoNT/E being short acting and BoNT/A long acting. We investigated the duration of action, spread and neuronal transport of BoNT/E (6.5 ng/kg) and BoNT/A (125 pg/kg) after single intramuscular administrations of high equivalent efficacious doses, in rats, over a 30- or 75-day periods, respectively. To achieve this, we used (i) digit abduction score assay, (ii) immunohistochemistry for SNAP25 (N-ter part; SNAP25N-ter and C-ter part; SNAP25C-ter) and its cleavage sites (cleaved SNAP25; c-SNAP25E and c-SNAP25A) and (iii) muscular changes in histopathology evaluation. Combined in vivo observation and immunohistochemistry analysis revealed that, compared to BoNT/A, BoNT/E induces minimal muscular changes, possesses a lower duration of action, a reduced ability to spread and a decreased capacity to be transported to the lumbar spinal cord. Interestingly, SNAP25C-ter completely disappeared for both toxins during the peak of efficacy, suggesting that the persistence of toxin effects is driven by the persistence of proteases in tissues. These data unveil some new molecular mechanisms of action of the short-acting BoNT/E and long-acting BoNT/A, and reinforce their overall safety profiles.

Theobromine improves hyperactivity, inattention, and working memory via modulation of dopaminergic neural function in the frontal cortex of spontaneously hypertensive rats.

Attention-deficit/hyperactivity disorder (ADHD) is a developmental disorder and dopaminergic dysfunction in the prefrontal cortex (PFC) may play a role. Our previous research indicated that theobromine (TB), a methylxanthine, enhances cognitive function in rodents via the PFC. This study investigates TB's effects on hyperactivity and cognitive function in stroke-prone spontaneously hypertensive rats (SHR), an ADHD animal model. Male SHRs (6-week old) received a diet containing 0.05% TB for 40 days, while control rats received normal diets. Age-matched male Wistar-Kyoto rats (WKY) served as genetic controls. During the TB administration period, we conducted open-field tests and Y-maze tasks to evaluate hyperactivity and cognitive function, then assessed dopamine concentrations and tyrosine hydroxylase (TH), dopamine receptor D1-5 (DRD1-5), dopamine transporter (DAT), vesicular monoamine transporter-2 (VMAT-2), synaptosome-associated protein-25 (SNAP-25), and brain-derived neurotrophic factor (BDNF) expressions in the PFC. Additionally, the binding affinity of TB for the adenosine receptors (ARs) was evaluated. Compared to WKY, SHR exhibited hyperactivity, inattention and working memory deficits. However, chronic TB administration significantly improved these ADHD-like behaviors in SHR. TB administration also normalized dopamine concentrations and expression levels of TH, DRD2, DRD4, SNAP-25, and BDNF in the PFC of SHR. No changes were observed in DRD1, DRD3, DRD5, DAT, and VMAT-2 expression between SHR and WKY rats, and TB intake had minimal effects. TB was found to have affinity binding to ARs. These results indicate that long-term TB supplementation mitigates hyperactivity, inattention and cognitive deficits in SHR by modulating dopaminergic nervous function and BDNF levels in the PFC, representing a potential adjunctive treatment for ADHD.

A hydrophobic groove in secretagogin allows for alternate interactions with SNAP-25 and syntaxin-4 in endocrine tissues.

Vesicular release of neurotransmitters and hormones relies on the dynamic assembly of the exocytosis/trans-SNARE complex through sequential interactions of synaptobrevins, syntaxins, and SNAP-25. Despite SNARE-mediated release being fundamental for intercellular communication in all excitable tissues, the role of auxiliary proteins modulating the import of reserve vesicles to the active zone, and thus, scaling repetitive exocytosis remains less explored. Secretagogin is a Ca2+-sensor protein with SNAP-25 being its only known interacting partner. SNAP-25 anchors readily releasable vesicles within the active zone, thus being instrumental for 1st phase release. However, genetic deletion of secretagogin impedes 2nd phase release instead, calling for the existence of alternative protein-protein interactions. Here, we screened the secretagogin interactome in the brain and pancreas, and found syntaxin-4 grossly overrepresented. Ca2+-loaded secretagogin interacted with syntaxin-4 at nanomolar affinity and 1:1 stoichiometry. Crystal structures of the protein complexes revealed a hydrophobic groove in secretagogin for the binding of syntaxin-4. This groove was also used to bind SNAP-25. In mixtures of equimolar recombinant proteins, SNAP-25 was sequestered by secretagogin in competition with syntaxin-4. Kd differences suggested that secretagogin could shape unidirectional vesicle movement by sequential interactions, a hypothesis supported by in vitro biological data. This mechanism could facilitate the movement of transport vesicles toward release sites, particularly in the endocrine pancreas where secretagogin, SNAP-25, and syntaxin-4 coexist in both α- and ß-cells. Thus, secretagogin could modulate the pace and fidelity of vesicular hormone release by differential protein interactions.

SNAP-25, but not SNAP-23, is essential for photoreceptor development, survival, and function in mice.

SNARE-mediated vesicular transport is thought to play roles in photoreceptor glutamate exocytosis and photopigment delivery. However, the functions of Synaptosomal-associated protein (SNAP) isoforms in photoreceptors are unknown. Here, we revisit the expression of SNAP-23 and SNAP-25 and generate photoreceptor-specific knockout mice to investigate their roles. Although we find that SNAP-23 shows weak mRNA expression in photoreceptors, SNAP-23 removal does not affect retinal morphology or vision. SNAP-25 mRNA is developmentally regulated and undergoes mRNA trafficking to photoreceptor inner segments at postnatal day 9 (P9). SNAP-25 knockout photoreceptors develop normally until P9 but degenerate by P14 resulting in severe retinal thinning. Photoreceptor loss in SNAP-25 knockout mice is associated with abolished electroretinograms and vision loss. We find mistrafficked photopigments, enlarged synaptic vesicles, and abnormal synaptic ribbons which potentially underlie photoreceptor degeneration. Our results conclude that SNAP-25, but not SNAP-23, mediates photopigment delivery and synaptic functioning required for photoreceptor development, survival, and function.

GLP-1 signaling-regulated SNAP25 is involved in improved insulin secretion in diabetic GK rats after Roux-en-Y gastric bypass surgery.

BACKGROUND: Roux-en-Y gastric bypass surgery (RYGB) improves glucose-stimulated insulin secretion (GSIS) in type 2 diabetes (T2D) patients. SNAP25 plays an essential role in GSIS. Clinical studies indicate that enhanced GLP-1 signaling is an important contributor to the improved ß-cell function in T2D. We aimed to explore whether GLP-1-regulated SNAP25 is involved in the enhanced secretory function of ß-cells in diabetic Goto-Kakizaki (GK) rats after RYGB. METHODS AND RESULTS: RYGB or sham surgery was conducted in GK rats. mRNA and protein expression of SNAP25 was assessed by qPCR and Western blot, respectively. Occupancy of CREB and acetyltransferase CBP and acetylation of histone H3 (ACH3) at the Snap25 promoter were determined using ChIP assay. RYGB led to increased SNAP25 expression and CREB phosphorylation in islets from GK rats. Increased SNAP25 improved GSIS in ß-cells cultured in high glucose conditions. Consistent with increased plasma GLP-1 after RYGB, GLP-1R agonist exendin4 increased SNAP25 expression and CREB phosphorylation in ß-cells. Mechanistically, exendin4 promoted the recruitment of CREB and CBP, thereby increasing ACH3 at the Snap25 promoter. Consistently, inhibition of CBP attenuated the effect of exendin4 on SNAP25 expression. Furthermore, the knockdown of SNAP25 diminished the increase of GSIS potentiated by chronic GLP-1 culture in INS-1 832/13 cells. CONCLUSIONS: Our findings unravel the novel mechanisms of RYGB-enhanced SNAP25 expression in ß-cells, and SNAP25 may contribute to the improved ß-cell secretory function induced by RYGB.

All SNAP25 molecules in the vesicle-plasma membrane contact zone change conformation during vesicle priming.

In neuronal cell types, vesicular exocytosis is governed by the SNARE (soluble NSF attachment receptor) complex consisting of synaptobrevin2, SNAP25, and syntaxin1. These proteins are required for vesicle priming and fusion. We generated an improved SNAP25-based SNARE COmplex Reporter (SCORE2) incorporating mCeruelan3 and Venus and overexpressed it in SNAP25 knockout embryonic mouse chromaffin cells. This construct rescues vesicle fusion with properties indistinguishable from fusion in wild-type cells. Combining electrochemical imaging of individual release events using electrochemical detector arrays with total internal reflection fluorescence resonance energy transfer (TIR-FRET) imaging reveals a rapid FRET increase preceding individual fusion events by 65 ms. The experiments are performed under conditions of a steady-state cycle of docking, priming, and fusion, and the delay suggests that the FRET change reflects tight docking and priming of the vesicle, followed by fusion after ~65 ms. Given the absence of wt SNAP25, SCORE2 allows determination of the number of molecules at fusion sites and the number that changes conformation. The number of SNAP25 molecules changing conformation in the priming step increases with vesicle size and SNAP25 density in the plasma membrane and equals the number of copies present in the vesicle-plasma membrane contact zone. We estimate that in wt cells, 6 to 7 copies of SNAP25 change conformation during the priming step.

SNAP25 is a potential target for early stage Alzheimer's disease and Parkinson's disease.

BACKGROUND: Alzheimer's disease (AD) and Parkinson's disease (PD), two common irreversible neurodegenerative diseases, share similar early stage syndromes, such as olfaction dysfunction. Yet, the potential comorbidity mechanism of AD and PD was not fully elucidated. METHODS: The gene expression profiles of GSE5281 and GSE8397 were downloaded from the Gene Expression Omnibus (GEO) database. We utilized a series of bioinformatics analyses to screen the overlapped differentially expressed genes (DEGs). The hub genes were further identified by the plugin CytoHubba of Cytoscape and validated in the hippocampus (HIP) samples of APP/PS-1 transgenic mice and the substantial nigra (SN) samples of A53T transgenic mice by real-time quantitative polymerase chain reaction (RT-qPCR). Meanwhile, the expression of the target genes in the olfactory epithelium/bulb was detected by RT-qPCR. Finally, molecular docking was used to screen potential compounds for the target gene. RESULTS: One hundred seventy-four overlapped DEGs were identified in AD and PD. Five of the top ten enrichment pathways mainly focused on the synapse. Five hub genes were identified and further validated. As a common factor in AD and PD, the changes of synaptosomal-associated protein 25 (SNAP25) mRNA in olfactory epithelium/bulb were significantly decreased and had a strong association with those in the HIP and SN samples. Pazopanib was the optimal compound targeting SNAP25, with a binding energy of - 9.2 kcal/mol. CONCLUSIONS: Our results provided a theoretical basis for understanding the comorbidity mechanism of AD and PD and highlighted that SNAP25 in the olfactory epithelium may serve as a potential target for early detection and intervention in both AD and PD.

Neuronal-specific TNFAIP1 ablation attenuates postoperative cognitive dysfunction via targeting SNAP25 for K48-linked ubiquitination.

BACKGROUND: Synaptosomal-associated protein 25 (SNAP25) exerts protective effects against postoperative cognitive dysfunction (POCD) by promoting PTEN-induced kinase 1 (PINK1)/Parkin-mediated mitophagy and repressing caspase-3/gasdermin E (GSDME)-mediated pyroptosis. However, the regulatory mechanisms of SNAP25 protein remain unclear. METHODS: We employed recombinant adeno-associated virus 9 (AAV9)-hSyn to knockdown tumor necrosis factor α-induced protein 1 (TNFAIP1) or SNAP25 and investigate the role of TNFAIP1 in POCD. Cognitive performance, hippocampal injury, mitophagy, and pyroptosis were assessed. Co-immunoprecipitation (co-IP) and ubiquitination assays were conducted to elucidate the mechanisms by which TNFAIP1 stabilizes SNAP25. RESULTS: Our results demonstrated that the ubiquitin ligase TNFAIP1 was upregulated in the hippocampus of mice following isoflurane (Iso) anesthesia and laparotomy. The N-terminal region (residues 1-96) of TNFAIP1 formed a conjugate with SNAP25, leading to lysine (K) 48-linked polyubiquitination of SNAP25 at K69. Silencing TNFAIP1 enhanced SH-SY5Y cell viability and conferred antioxidant, pro-mitophagy, and anti-pyroptosis properties in response to Iso and lipopolysaccharide (LPS) challenges. Conversely, TNFAIP1 overexpression reduced HT22 cell viability, increased reactive oxygen species (ROS) accumulation, impaired PINK1/Parkin-dependent mitophagy, and induced caspase-3/GSDME-dependent pyroptosis by suppressing SNAP25 expression. Neuron-specific knockdown of TNFAIP1 ameliorated POCD, restored mitophagy, and reduced pyroptosis, which was reversed by SNAP25 depletion. CONCLUSIONS: In summary, our findings demonstrated that inhibiting TNFAIP1-mediated degradation of SNAP25 might be a promising therapeutic approach for mitigating postoperative cognitive decline. Video Abstract.

Diagnostic and prognostic value of cerebrospinal fluid SNAP-25 and neurogranin in Creutzfeldt-Jakob disease in a clinical setting cohort of rapidly progressive dementias.

BACKGROUND: The levels of synaptic markers synaptosomal-associated protein 25 (SNAP-25) and neurogranin (Ng) have been shown to increase early in the cerebrospinal fluid (CSF) of patients with Creutzfeldt-Jakob disease (CJD) and to have prognostic potential. However, no validation studies assessed these biomarkers' diagnostic and prognostic value in a large clinical setting cohort of rapidly progressive dementia. METHODS: In this retrospective study, using commercially available immunoassays, we measured the levels of SNAP-25, Ng, 14-3-3, total-tau (t-tau), neurofilament light chain (NfL), and phospho-tau181 (p-tau) in CSF samples from consecutive patients with CJD (n = 220) or non-prion rapidly progressive dementia (np-RPD) (n = 213). We evaluated and compared the diagnostic accuracy of each CSF biomarker and biomarker combination by receiver operating characteristics curve (ROC) analyses, studied SNAP-25 and Ng CSF concentrations distribution across CJD subtypes, and estimated their association with survival using multivariable Cox regression analyses. RESULTS: CSF SNAP-25 and Ng levels were higher in CJD than in np-RPD (SNAP-25: 582, 95% CI 240-1250 vs. 115, 95% CI 78-157 pg/ml, p < 0.0001; Ng: 841, 95% CI 411-1473 vs. 390, 95% CI 260-766 pg/ml, p < 0.001). SNAP-25 diagnostic accuracy (AUC 0.902, 95% CI 0.873-0.931) exceeded that of 14-3-3 (AUC 0.853, 95% CI 0.816-0.889), t-tau (AUC 0.878, 95% CI 0.845-0.901), and the t-tau/p-tau ratio (AUC 0.884, 95% CI 0.851-0.916). In contrast, Ng performed worse (AUC 0.697, 95% CI 0.626-0.767) than all other surrogate biomarkers, except for NfL (AUC 0.649, 95% CI 0.593-0.705). SNAP-25 maintained a relatively high diagnostic value even for atypical CJD subtypes (AUC 0.792, 95% CI 0.729-0.854). In Cox regression analyses, SNAP-25 levels were significantly associated with survival in CJD (hazard ratio [HR] 1.71 95% CI 1.40-2.09). Conversely, Ng was associated with survival only in the most rapidly progressive CJD subtypes (sCJD MM(V)1 and gCJD M1) (HR 1.81 95% CI 1.21-2.93). CONCLUSIONS: In the clinical setting, CSF SNAP-25 is a viable alternative to t-tau, 14-3-3, and the t-tau/p-tau ratio in discriminating the CJD subtypes from other RPDs. Additionally, SNAP-25 and, to a lesser extent, Ng predict survival in CJD, showing prognostic power in the range of CSF t-tau/14-3-3 and NfL, respectively.

Phosphorylated Ser187-SNAP25-modulated hyperfunction of glutamatergic system in the vmPFC mediates depressive-like behaviors in male mice.

Dysfunctional glutamatergic neurotransmission contributes importantly to the pathophysiology of depression. However, the underlying neural mechanisms of glutamatergic dysfunction remain poorly understood. Here, we employed chronic unpredictable mild stress (CUMS) to induce depression-like behavior in male mice and to assess the alterations of glutamatergic system within the ventromedial prefrontal cortex (vmPFC). Male mice subjected to CUMS showed an increase in levels of glutamate content, synaptosomal GluN2B-NMDA receptors (GluN2B-NMDARs) and phosphorylated synaptosomal associated protein 25 KD of Ser187 (pSer187-SNAP25), which is involved in synaptic vesicular fusion processes in the vmPFC. Downregulation of pSer187-SNAP25 via the TAT-S187 fusion peptide efficiently alleviated CUMS-induced depressive-like behaviors in male mice by reversing the increase of glutamate content and synaptosomal GluN2B-NMDARs. These findings demonstrated a critical role for pSer187-SNAP25-mediated glutamatergic dysfunction in CUMS-induced depressive-like behaviors, suggesting the potential of pS187-SNAP25 inhibitors for further investigation on depression management.

Role of SNAP25 on the occurrence and development of eosinophilic gastritis.

Eosinophilic gastritis is characterized by gastrointestinal symptoms accompanied by peripheral eosinophilia. This study aims to explore the association between eosinophilic gastritis and Synaptosome Associated Protein 25 (SNAP25), and provide a new direction for the diagnosis and treatment of eosinophilic gastritis. GSE54043 was downloaded from the gene expression omnibus database. Differentially expressed genes (DEGs) were screened. The functions of common DEGs were annotated by Database for Annotation, Visualization and Integrated Discovery and Metascape. The protein-protein interaction network of common DEGs was obtained by Search Tool for the Retrieval of Interacting Genes and visualized by Cytoscape. Significant modules were identified from the protein-protein interaction network. A total of 186 patients with eosinophilic gastritis were recruited. The clinical data were recorded and the expression levels of CPE, SST, PCSK2, SNAP25, and SYT4 were detected. Pearson chi-square test and Spearman correlation coefficient were used to analyze the relationship between eosinophilic gastritis and related parameters. Univariate and multivariate Logistic regression were used for further analysis. 353 DEGs were presented. The top 10 genes screened by cytoHubb were shown, and Veen diagram figured out 5 mutual genes. Pearson's chi-square test showed that SNAP25 (P < .001) was significantly associated with eosinophilic gastritis. Spearman correlation coefficient showed a significant correlation between eosinophilic gastritis and SNAP25 (ρ = -0.569, P < .001). Univariate logistic regression analysis showed that SNAP25 (OR = 0.046, 95% CI: 0.018-0.116, P < .001) was significantly associated with eosinophilic gastritis. Multivariate logistic regression analysis showed that SNAP25 (OR = 0.024, 95% CI: 0.007-0.075, P < .001) was significantly associated with eosinophilic gastritis. The low expression of SNAP25 gene in eosinophilic gastritis is associated with a higher risk of eosinophilic gastritis.