Alzheimer\'s disease (AD) is the leading form of dementia, characterized by the accumulation and aggregation of amyloid in brain. Transient receptor potential vanilloid 2 (TRPV2) is an ion channel involved in diverse physiopathological processes, including microglial phagocytosis. Previous studies suggested that cannabidiol (CBD), an activator of TRPV2, improves microglial amyloid-β (Aβ) phagocytosis by TRPV2 modulation. However, the molecular mechanism of TRPV2 in microglial Aβ phagocytosis remains unknown. In this study, we aimed to investigate the involvement of TRPV2 channel in microglial Aβ phagocytosis and the underlying mechanisms. Utilizing human datasets, mouse primary neuron and microglia cultures, and AD model mice, to evaluate TRPV2 expression and microglial Aβ phagocytosis in both in vivo and in vitro. TRPV2 was expressed in cortex, hippocampus, and microglia.Cannabidiol (CBD) could activate and sensitize TRPV2 channel. Short-term CBD (1 week) injection intraperitoneally (i.p.) reduced the expression of neuroinflammation and microglial phagocytic receptors, but long-term CBD (3 week) administration (i.p.) induced neuroinflammation and suppressed the expression of microglial phagocytic receptors in APP/PS1 mice. Furthermore, the hyper-sensitivity of TRPV2 channel was mediated by tyrosine phosphorylation at the molecular sites Tyr(338), Tyr(466), and Tyr(520) by protein tyrosine kinase JAK1, and these sites mutation reduced the microglial Aβ phagocytosis partially dependence on its localization. While TRPV2 was palmitoylated at Cys 277 site and blocking TRPV2 palmitoylation improved microglial Aβ phagocytosis. Moreover, it was demonstrated that TRPV2 palmitoylation was dynamically regulated by ZDHHC21. Overall, our findings elucidated the intricate interplay between TRPV2 channel regulated by tyrosine phosphorylation/dephosphorylation and cysteine palmitoylation/depalmitoylation, which had divergent effects on microglial Aβ phagocytosis. These findings provide valuable insights into the underlying mechanisms linking microglial phagocytosis and TRPV2 sensitivity, and offer potential therapeutic strategies for managing AD.

BACKGROUND: Breast cancer is a prevalent public health concern affecting numerous women globally and is associated with palmitoylation, a post-translational protein modification. Despite increasing focus on palmitoylation, its specific implications for breast cancer prognosis remain unclear. The work aimed to identify prognostic factors linked to palmitoylation in breast cancer and assess its effectiveness in predicting responses to chemotherapy and immunotherapy.
METHODS: We utilized the \"limma\" package to analyze the differential expression of palmitoylation-related genes between breast cancer and normal tissues. Hub genes were identified using the \"WGCNA\" package. Using the least absolute shrinkage and selection operator (LASSO) Cox regression analysis, we identified a prognostic feature associated with palmitoylation and developed a prognostic nomogram with the \"regplot\" package. The predictive values of the model for chemotherapy and immunotherapy responses were assessed using immunophenoscore (IPS) and the \"pRophetic\" package.
RESULTS: We identified 211 differentially expressed genes related to palmitoylation, among which 44 demonstrated prognostic potential. Subsequently, a predictive model comprising eleven palmitoylation-related genes was developed. Patients were classified into high-risk and low-risk groups based on the median risk score. The findings revealed that individuals in the high-risk group exhibited lower survival rates, while those in the low-risk group showed increased immune cell infiltration and improved responses to chemotherapy and immunotherapy. Moreover, the BC-Palmitoylation Tool website was established.
CONCLUSIONS: This study developed the first machine learning-based predictive model for palmitoylation-related genes and created a corresponding website, providing clinicians with a valuable tool to improve patient outcomes.

N-Nitrosamine disinfection by-products (NAs-DBPs) have been well proven for its role in esophageal carcinogenesis. However, the role of intratumoral microorganisms in esophageal squamous cell carcinoma (ESCC) has not yet been well explored in the context of exposure to NAs-DBPs. Here, the multi-omics integration reveals F. periodonticum (Fp) as \"facilitators\" is highly enriched in cancer tissues and promotes the epithelial mesenchymal transition (EMT)-like subtype formation of ESCC. We demonstrate that Fp potently drives de novo synthesis of fatty acids, migration, invasion and EMT phenotype through its unique FadAL adhesin. However, N-nitrosomethylbenzylamine upregulates the transcription level of FadAL. Mechanistically, co-immunoprecipitation coupled to mass spectrometry shows that FadAL interacts with FLOT1. Furthermore, FLOT1 activates PI3K-AKT/FASN signaling pathway, leading to triglyceride and palmitic acid (PA) accumulation. Innovatively, the results from the acyl-biotin exchange demonstrate that FadAL-mediated PA accumulation enhances Wnt3A palmitoylation on a conserved cysteine residue, Cys-77, and promotes Wnt3A membrane localization and the translocation of β-catenin into the nucleus, further activating Wnt3A/β-catenin axis and inducing EMT phenotype. We therefore propose a \"microbiota-cancer cell subpopulation\" interaction model in the highly heterogeneous tumor microenvironment. This study unveils a mechanism by which Fp can drive ESCC and identifies FadAL as a potential diagnostic and therapeutic target for ESCC.

Click chemistry, also known as \"link chemistry,\" is an important molecular connection method that can achieve simple and efficient connections between specific small molecular groups at the molecular level. Click chemistry offers several advantages, including high efficiency, good selectivity, mild conditions, and few side reactions. These features make it a valuable tool for in-depth analysis of various protein posttranslational modifications (PTMs) caused by changes in cell metabolism during viral infection. This chapter considers the palmitoylation, carbonylation, and alkylation of STING and presents detailed information and experimental procedures for measuring PTMs using click chemistry.

NLRP3 inflammasome activation, essential for cytokine secretion and pyroptosis in response to diverse stimuli, is closely associated with various diseases. Upon stimulation, NLRP3 undergoes subcellular membrane trafficking and conformational rearrangements, preparing itself for inflammasome assembly at the microtubule-organizing center (MTOC). Here, we elucidate an orchestrated mechanism underlying these ordered processes using human and murine cells. Specifically, NLRP3 undergoes palmitoylation at two sites by palmitoyl transferase zDHHC1, facilitating its trafficking between subcellular membranes, including the mitochondria, trans-Golgi network (TGN), and endosome. This dynamic trafficking culminates in the localization of NLRP3 to the MTOC, where LATS1/2, pre-recruited to MTOC during priming, phosphorylates NLRP3 to further facilitate its interaction with NIMA-related kinase 7 (NEK7), ultimately leading to full NLRP3 activation. Consistently, Zdhhc1-deficiency mitigated LPS-induced inflammation and conferred protection against mortality in mice. Altogether, our findings provide valuable insights into the regulation of NLRP3 membrane trafficking and inflammasome activation, governed by palmitoylation and phosphorylation events.

BACKGROUND: Breast cancer (BC) ranks as the third most fatal malignant tumor worldwide, with a strong reliance on fatty acid metabolism. CLDN6, a candidate BC suppressor gene, was previously identified as a regulator of fatty acid biosynthesis; however, the underlying mechanism remains elusive. In this research, we aim to clarify the specific mechanism through which CLDN6 modulates fatty acid anabolism and its impact on BC growth and metastasis.
METHODS: Cell function assays, tumor xenograft mouse models, and lung metastasis mouse models were conducted to evaluate BC growth and metastasis. Human palmitic acid assay, triglyceride assay, Nile red staining, and oil red O staining were employed to investigate fatty acid anabolism. Reverse transcription polymerase chain reaction (RT-PCR), western blot, immunohistochemistry (IHC) assay, nuclear fractionation, immunofluorescence (IF), immunoprecipitation and acyl-biotin exchange (IP-ABE), chromatin immunoprecipitation (ChIP), dual luciferase reporter assay, and co-immunoprecipitation (Co-IP) were applied to elucidate the underlying molecular mechanism. Moreover, tissue microarrays of BC were analyzed to explore the clinical implications.
RESULTS: We identified that CLDN6 inhibited BC growth and metastasis by impeding RAS palmitoylation both in vitro and in vivo. We proposed a unique theory suggesting that CLDN6 suppressed RAS palmitoylation through SREBP1-modulated de novo palmitic acid synthesis. Mechanistically, CLDN6 interacted with MAGI2 to prevent KLF5 from entering the nucleus, thereby restraining SREBF1 transcription. The downregulation of SREBP1 reduced de novo palmitic acid synthesis, hindering RAS palmitoylation and subsequent endosomal sorting complex required for transport (ESCRT)-mediated plasma membrane localization required for RAS oncogenic activation. Besides, targeting inhibition of RAS palmitoylation synergized with CLDN6 to repress BC progression.
CONCLUSIONS: Our findings provide compelling evidence that CLDN6 suppresses the palmitic acid-induced RAS palmitoylation through the MAGI2/KLF5/SREBP1 axis, thereby impeding BC malignant progression. These results propose a new insight that monitoring CLDN6 expression alongside targeting inhibition of palmitic acid-mediated palmitoylation could be a viable strategy for treating oncogenic RAS-driven BC.

Palmitoylation is a type of lipid modification that plays an important role in various aspects of neuronal function. Over the past few decades, several studies have shown that the palmitoylation of synaptic proteins is involved in neurotransmission and synaptic functions. Palmitoyl acyltransferases (PATs), which belong to the DHHC family, are major players in the regulation of palmitoylation. Dysregulated palmitoylation of synaptic proteins and mutated/dysregulated DHHC proteins are associated with several neurodegenerative diseases, such as Alzheimer\'s disease (AD), Huntington\'s disease (HD), and Parkinson\'s disease (PD). In this review, we summarize the recent discoveries on the subcellular distribution of DHHC proteins and analyze their expression patterns in different brain cells. In particular, this review discusses how palmitoylation of synaptic proteins regulates synaptic vesicle exocytotic fusion and the localization, clustering, and transport of several postsynaptic receptors, as well as the role of palmitoylation of other proteins in regulating synaptic proteins. Additionally, some of the specific known associations of these factors with neurodegenerative disorders are explored, with a few suggestions for the development of therapeutic strategies. Finally, this review provides possible directions for future research to reveal detailed and specific mechanisms underlying the roles of synaptic protein palmitoylation.

Recent studies have highlighted palmitoylation, a novel protein post-translational modification, as a key player in various signaling pathways that contribute to tumorigenesis and drug resistance. Despite this, its role in bladder cancer (BCa) development remains inadequately understood. In this study, ZDHHC9 emerged as a significantly upregulated oncogene in BCa. Functionally, ZDHHC9 knockdown markedly inhibited tumor proliferation, promoted tumor cell apoptosis, and enhanced the efficacy of gemcitabine (GEM) and cisplatin (CDDP). Mechanistically, SP1 was found to transcriptionally activate ZDHHC9 expression. ZDHHC9 subsequently bound to and palmitoylated the Bip protein at cysteine 420 (Cys420), thereby inhibiting the unfolded protein response (UPR). This palmitoylation at Cys420 enhanced Bip\'s protein stability and preserved its localization within the endoplasmic reticulum (ER). ZDHHC9 might become a novel therapeutic target for BCa and could also contribute to combination therapy with GEM and CDDP.

Gasdermin D (GSDMD)-mediated pyroptotic cell death drives inflammatory cytokine release and downstream immune responses upon inflammasome activation, which play important roles in host defense and inflammatory disorders. Upon activation by proteases, the GSDMD N-terminal domain (NTD) undergoes oligomerization and membrane translocation in the presence of lipids to assemble pores. Despite intensive studies, the molecular events underlying the transition of GSDMD from an autoinhibited soluble form to an oligomeric pore form inserted into the membrane remain incompletely understood. Previous work characterized S-palmitoylation for gasdermins from bacteria, fungi, invertebrates, as well as mammalian gasdermin E (GSDME). Here, we report that a conserved residue Cys191 in human GSDMD was S-palmitoylated, which promoted GSDMD-mediated pyroptosis and cytokine release. Mutation of Cys191 or treatment with palmitoyltransferase inhibitors cyano-myracrylamide (CMA) or 2-bromopalmitate (2BP) suppressed GSDMD palmitoylation, its localization to the membrane and dampened pyroptosis or IL-1β secretion. Furthermore, Gsdmd-dependent inflammatory responses were alleviated by inhibition of palmitoylation in vivo. By contrast, coexpression of GSDMD with palmitoyltransferases enhanced pyroptotic cell death, while introduction of exogenous palmitoylation sequences fully restored pyroptotic activities to the C191A mutant, suggesting that palmitoylation-mediated membrane localization may be distinct from other molecular events such as GSDMD conformational change during pore assembly. Collectively, our study suggests that S-palmitoylation may be a shared regulatory mechanism for GSDMD and other gasdermins, which points to potential avenues for therapeutically targeting S-palmitoylation of gasdermins in inflammatory disorders.

Aberrant activity of NLRP3 has been shown associations with severe diseases. Palmitoylation is a kind of protein post-translational modification, which has been shown to regulate cancer development and the innate immune system. Here, we showed that NLRP3 is palmitoylated at Cys419 and that palmitoyltransferase ZDHHC17 is the predominant enzyme that mediates NLRP3 palmitoylation and promotes NLRP3 activation by interacting with NLRP3 and facilitating NIMA-related kinase 7 (NEK7)-NLRP3 interactions. Blockade of NLRP3 palmitoylation by a palmitoylation inhibitor, 2-bromopalmitate, effectively inhibited NLRP3 activation in vitro. Also, in a dextran sulfate sodium-induced colitis model in mice, 2-bromopalmitate application could attenuate weight loss, improve the survival rate, and rescue pathological changes in the colon of mice. Overall, our study reveals that palmitoylation of NLPR3 modulates inflammasome activation and inflammatory bowel disease development. We propose that drugs targeting NLRP3 palmitoylation could be promising candidates in the treatment of NLRP3-mediated inflammatory diseases.