Structural basis for the recognition of IFNAR1 by the humanized therapeutic monoclonal antibody QX006N for the treatment of systemic lupus erythematosus.

Interferon (IFN) alpha/beta receptor 1 (IFNAR1) is indispensable for antiviral responses and the immune regulation. Dysregulation of the IFNAR1-mediaetd signaling pathways leads to deleterious autoimmune diseases such as systemic lupus erythematosus (SLE). QX006N, a humanized therapeutic monoclonal antibody, specifically targets human IFNAR1 and is in the clinical trial phase for treating SLE, but the molecular mechanism underlying the QX006N-mediated recognition of IFNAR1 remains unclear. Here, we report the high neutralization activities of QX006N against IFNAR1-mediated signal transduction. Meanwhile, we determine the structures of the fragment antigen-binding domain (Fab) of QX006N (QX006N-Fab) and QX006N-Fab in complex with the subdomains 1-3 of IFNAR1 (IFNAR1-SD123) at 2.87 Šand 2.68 Šresolutions, respectively. In the structure of the QX006N-Fab/IFNAR1-SD123 complex, QX006N-Fab only recognizes the SD3 subdomain of IFNAR1 by the hydrophobic, hydrogen-bonding and electrostatic interactions. Compared with the structure of the IFN/IFNAR1/IFNAR2 complex, the binding of QX006N-Fab to IFNAR1-SD3 blocks its association with IFN due to steric hindrance, which inhibits the IFN/IFNAR1/IFNAR2 complex formation for signal transduction. The results of this study provide the structural evidence for the specific targeting of IFNAR1 by the therapeutic antibody QX006N and pave the way for the rational design of antibody drugs to combat IFNAR1-related autoimmune diseases.

Complement C1q/C3-CR3 signaling pathway mediates abnormal microglial phagocytosis of synapses in a mouse model of depression.

BACKGROUND: Both functional brain imaging studies and autopsy reports have indicated the presence of synaptic loss in the brains of depressed patients. The activated microglia may dysfunctionally engulf neuronal synapses, leading to synaptic loss and behavioral impairments in depression. However, the mechanisms of microglial-synaptic interaction under depressive conditions remain unclear. METHODS: We utilized lipopolysaccharide (LPS) to induce a mouse model of depression, examining the effects of LPS on behaviors, synapses, microglia, microglial phagocytosis of synapses, and the C1q/C3-CR3 complement signaling pathway. Additionally, a C1q neutralizing antibody was employed to inhibit the C1q/C3-CR3 signaling pathway and assess its impact on microglial phagocytosis of synapses and behaviors in the mice. RESULTS: LPS administration resulted in depressive and anxiety-like behaviors, synaptic loss, and abnormal microglial phagocytosis of synapses in the hippocampal dentate gyrus (DG) of mice. We found that the C1q/C3-CR3 signaling pathway plays a crucial role in this abnormal microglial activity. Treatment with the C1q neutralizing antibody moderated the C1q/C3-CR3 pathway, leading to a decrease in abnormal microglial phagocytosis, reduced synaptic loss, and improved behavioral impairments in the mice. CONCLUSIONS: The study suggests that the C1q/C3-CR3 complement signaling pathway, which mediates abnormal microglial phagocytosis of synapses, presents a novel potential therapeutic target for depression treatment.

Repressing miR-23a promotes the transdifferentiation of pancreatic α cells to ß cells via negatively regulating the expression of SDF-1α.

Pancreatic ß-cell failure is a pathological feature in type 1 diabetes. One promising approach involves inducing transdifferentiation of related pancreatic cell types, specifically α cells that produce glucagon. The chemokine stromal cell-derived factor-1 alpha (SDF-1α) is implicated in pancreatic α-to-ß like cell transition. Here, the serum level of SDF-1α was lower in T1D with C-peptide loss, the miR-23a was negatively correlated with SDF-1α. We discovered that exosomal miR-23a, secreted from ß cells, functionally downregulates the expression of SDF-1α, leading to increased Pax4 expression and decreased Arx expression in vivo. Adenovirus-vectored miR-23a sponge and mimic were constructed to further explored the miR-23a on pancreatic α-to-ß like cell transition in vitro, which yielded results consistent with our cell-based assays. Suppression of miR-23a upregulated insulin level and downregulated glucagon level in STZ-induced diabetes mice models, effectively promoting α-to-ß like cell transition. Our findings highlight miR-23a as a new therapeutic target for regenerating pancreatic ß cells from α cells.

Alternative polyadenylation profiles of susceptible and resistant rice (Oryza sativa L.) in response to bacterial leaf blight using RNA-seq.

BACKGROUND: Alternative polyadenylation (APA) is an important pattern of post-transcriptional regulation of genes widely existing in eukaryotes, involving plant physiological and pathological processes. However, there is a dearth of studies investigating the role of APA profile in rice leaf blight. RESULTS: In this study, we compared the APA profile of leaf blight-susceptible varieties (CT 9737-613P-M) and resistant varieties (NSIC RC154) following bacterial blight infection. Through gene enrichment analysis, we found that the genes of two varieties typically exhibited distal poly(A) (PA) sites that play different roles in two kinds of rice, indicating differential APA regulatory mechanisms. In this process, many disease-resistance genes displayed multiple transcripts via APA. Moreover, we also found five polyadenylation factors of similar expression patterns of rice, highlighting the critical roles of these five factors in rice response to leaf blight about PA locus diversity. CONCLUSION: Notably, the present study provides the first dynamic changes of APA in rice in early response to biotic stresses and proposes a possible functional conjecture of APA in plant immune response, which lays the theoretical foundation for in-depth determination of the role of APA events in plant stress response and other life processes.

HES4 is a potential biomarker for bladder cancer: a Mendelian randomization study.

Background: Patients with bladder cancer (BLCA) have a poor prognosis and little progress has been made in treatment. Therefore, the purpose of this work was to employ Mendelian randomization (MR) and transcriptome analysis to identify a novel biomarker that could be used to reliably diagnose BLCA. Methods: TCGA-BLCA and GSE121711 datasets were obtained from public databases. Genome-wide association study (GWAS) data of BLCA outcome (373,295 samples containing 9,904,926 single nucleotide polymorphisms) were obtained through the IEU OpenGWAS database. Differentially expressed genes were applied as exposure factors, and MR analysis was performed to identify genes that had a causal relationship with BLCA. Then, the patients were divided into high and low expression groups according to the expression levels of candidate genes, and genes with survival differences were identified. Univariate and multivariate Cox regression were used to investigate the prognostic value of the expression of these genes. A nomogram was constructed based on independent prognostic factors, and we analyzed the functions and pathways associated with the identified genes as well as their relationship with the immune microenvironment. Results: HES4 was identified as a biomarker. HES4 status, age, and stage were identified as independent prognostic factors, and an excellent nomogram was established. Bioinformatic analysis suggested that HES4 might be associated with the activation of the immune response, bone development, and cancer pathways. The BLCA samples were divided into high and low HES4 groups. The stromal score and 33 immune cells were remarkably different between the two groups, with HES4 expression being negatively correlated with macrophages and mast cells, and positively correlated with eosinophils and central memory CD4+ T cells. Finally, HES4 was up-regulated in cancer samples in both TCGA-BLCA and GSE121711 datasets. Conclusion: This study identified HES4 as an independent prognostic factor for BLCA outcome based on MR and transcriptome analysis, which provides useful information for future research on and treatment of BLCA.

A Secreted BBE-Like Enzyme Acting as a Drug-Binding Efflux Carrier Confers Microbial Self-Resistance to Mitomycin C.

The berberine bridge enzyme (BBE)-like flavoproteins have attracted continuous attention for their capability to catalyze various oxidative reactions. Here we demonstrate that MitR, a secreted BBE-like enzyme, functions as a special drug-binding efflux protein evolved from quinone reductase. Moreover, this protein provides self-resistance to its hosts toward the DNA-alkylating agent mitomycin C with a distinctive strategy, featured by independently performing drug binding and efflux.

Clinical and multiparametric MRI features for differentiating uterine carcinosarcoma from endometrioid adenocarcinoma.

INTRODUCTION: The purpose of our study was to differentiate uterine carcinosarcoma (UCS) from endometrioid adenocarcinoma (EAC) by the multiparametric magnetic resonance imaging (MRI) features. METHODS: We retrospectively evaluated clinical and MRI findings in 17 patients with UCS and 34 patients with EAC proven by histologically. The following clinical and pathological features were evaluated: post- or pre-menopausal, clinical presentation, invasion depth, FIGO stage, lymphaticmetastasis. The following MRI features were evaluated: tumor dimension, cystic degeneration or necrosis, hemorrhage, signal intensity (SI) on T2-weighted images (T2WI), relative SI of lesion to myometrium on T2WI, T1WI, DWI, ADCmax, ADCmin, ADCmean (RSI-T2, RSI-T1, RSI-DWI, RSI-ADCmax, RSI-ADCmin, RSI-ADCmean), ADCmax, ADCmin, ADCmean, the maximum, minimum and mean relative enhancement (RE) of lesion to myometrium on the arterial and venous phases (REAmax, REAmin, REAmean, REVmax, REVmin, REVmean). Receiver operating characteristic (ROC) analysis and the area under the curve (AUC) were used to evaluate prediction ability. RESULTS: The mean age of UCS was higher than EAC. UCS occurred more often in the postmenopausal patients. UCS and EAC did not significantly differ in depth of myometrial invasion, FIGO stage and lymphatic metastasis. The anterior-posterior and transverse dimensions were significantly larger in UCS than EAC. Cystic degeneration or necrosis and hemorrhage were more likely occurred in UCS. The SI of tumor on T2WI was more heterogeneous in UCS. The RSI-T2, ADCmax, ADCmean, RSI-ADCmax and RSI-ADCmean of UCS were significantly higher than EAC. The REAmax, REAmin, REAmean, REVmax, REVmin and REVmean of UCS were all higher than EAC. The AUCs were 0.72, 0.71, 0.86, 0.96, 0.89, 0.84, 0.73, 0.97, 0.88, 0.94, 0.91, 0.69 and 0.80 for the anterior-posterior dimension, transverse dimension, RSI-T2, ADCmax, ADCmean, RSI-ADCmax, RSI-ADCmean, REAmax, REAmin, REAmean, REVmax, REVmin and REVmean, respectively. The AUC was 0.997 of the combined of ADCmax, REAmax and REVmax. Our study showed that ADCmax threshold value of 789.05 (10-3mm2/s) can differentiate UCS from EAC with 100% sensitivity, 76.5% specificity, and 0.76 AUC, REAmax threshold value of 0.45 can differentiate UCS from EAC with 88.2% sensitivity, 100% specificity, and 0.88 AUC. CONCLUSION: Multiparametric MRI features may be utilized as a biomarker to distinguish UCS from EAC.

Microglial NLRP3 inflammasome-mediated neuroinflammation and therapeutic strategies in depression.

Previous studies have demonstrated a bidirectional relationship between inflammation and depression. Activation of the nucleotide-binding oligomerization domain, leucine-rich repeat, and NLR family pyrin domain-containing 3 (NLRP3) inflammasomes is closely related to the pathogenesis of various neurological diseases. In patients with major depressive disorder, NLRP3 inflammasome levels are significantly elevated. Understanding the role that NLRP3 inflammasome-mediated neuroinflammation plays in the pathogenesis of depression may be beneficial for future therapeutic strategies. In this review, we aimed to elucidate the mechanisms that lead to the activation of the NLRP3 inflammasome in depression as well as to provide insight into therapeutic strategies that target the NLRP3 inflammasome. Moreover, we outlined various therapeutic strategies that target the NLRP3 inflammasome, including NLRP3 inflammatory pathway inhibitors, natural compounds, and other therapeutic compounds that have been shown to be effective in treating depression. Additionally, we summarized the application of NLRP3 inflammasome inhibitors in clinical trials related to depression. Currently, there is a scarcity of clinical trials dedicated to investigating the applications of NLRP3 inflammasome inhibitors in depression treatment. The modulation of NLRP3 inflammasomes in microglia holds promise for the management of depression. Further investigations are necessary to ascertain the efficacy and safety of these therapeutic approaches as potential novel antidepressant treatments.

Cannabinoid type 2 receptors play a crucial role in social defeat-induced depression.

BACKGROUND: The endocannabinoid system plays a crucial role in regulating mood, but the specific involvement of cannabinoid receptor type 2 (CB2R) in depression remains poorly understood. Similarly, the mechanisms by which electroacupuncture (EA) provides therapeutic benefits for depression are not clearly defined. This research aims to explore the function of CB2R in depression and examine if the therapeutic effects of EA are associated with the hippocampal CB2R system. METHODS: Mice experiencing social defeat stress (SDS) were used to model depression and anxiety behaviors. We quantified hippocampal CB2R and N-arachidonoylethanolamide (AEA) levels. The efficacy of a CB2R agonist, JWH133, in mitigating SDS-induced behaviors was evaluated. Additionally, EA's impact on CB2R and AEA was assessed, along with the influence of CB2R antagonist AM630 on EA's antidepressant effects. RESULTS: SDS led to depressive and anxiety-like behaviors, with corresponding decreases in hippocampal CB2R and AEA. Treatment with JWH133 ameliorated these behaviors. EA treatment resulted in increased CB2R and AEA levels, while AM630 blocked these antidepressant effects. LIMITATIONS: The study mainly focused on the SDS model, which may not entirely reflect other depression models. Besides, further investigation is needed to understand the precise mechanisms by which CB2R and AEA contribute to EA's effects. CONCLUSIONS: The study suggests hippocampal downregulation of CB2R and AEA contributes to depression. Upregulation of CB2R and AEA in response to EA suggests their involvement in EA's antidepressant effects. These findings provide insights into the role of the hippocampal CB2R system in depression and the potential mechanisms underlying EA's therapeutic effects.

The Neuroprotective Effects of Electroacupuncture on Parkinson's Disease and the Underlying Molecular Mechanisms.

Parkinson's disease (PD) is a chronic neurodegenerative disease whose main pathological features are the degeneration of dopamine neurons and deposition of α-synuclein in neurons. At present, the most important treatment strategy for PD is drugs, and one of the most used drugs is levodopa. However, this therapy shows many problems, such as tolerance and long-term effects, so other treatment strategies need to be explored. As a traditional Chinese medicine treatment method with effective and few side effects, electroacupuncture is considered a non-drug therapy. It serves as a novel, promising therapeutic approach for the treatment of PD. In this review, the application and the effects of electroacupuncture on PD have been described. Besides, the underlying molecular mechanisms of electroacupuncture on PD that contribute to protecting dopaminergic neurons and reducing α-synuclein levels have been illustrated, including ① anti-oxidant stress response, ② anti-neuroinflammatory response, ③ up-regulation of neurotrophic factors and reduction of nerve cell apoptosis, ④ down-regulation of endoplasmic reticulum stress and improvement of mitochondrial function, ⑤ improvement of the function of the ubiquitin-proteasome system, ⑥ anti-excitatory toxicity response, ⑦ activation of autophagy, and ⑧ modulation of gut microbiota. Achieving a better understanding of the neuroprotective effects of electroacupuncture on PD will provide a theoretical basis and facilitate the application of electroacupuncture on PD.

Economic burden attributable to healthcare-associated infections at western China hospitals: 6 Year, prospective cohort study.

OBJECTIVES: Healthcare-associated infections (HAIs) represent a major threat to patient safety and are associated with significant economic burden. Calculating the costs attributable to HAIs is challenging given the various sources of bias. Although HAIs as a reasonably preventable medical harm should have been closely linked to medical insurance incentives, there was little linkage between HAIs and medicare in western China owing to the lack of economic evaluation data. The present study aimed to generate estimates of the attributable costs associated with HAIs and the magnitude of costs growth. METHODS: In this cohort study designed horizontally and vertically from 2016 to 2022, we compared outcomes of randomly sampling patients with HAIs and individually matched patients without HAIs in two cohorts at a 6-year interval at 34 hospitals in western China. The primary outcome was the direct medical cost for the entire hospital stay, converted to US dollars ($ for the benchmark year), discounted at 3% annually, and estimated separately in the full analysis set (FAS) and the per protocol set (PPS). We used multiple linear regression to adjust the discounted costs and to assess subgroups effects within each cohort. We nested a dynamic vertical comparison of costs attributable to HAIs between the front and rear cohorts. RESULTS: A total of 230 patients with HAIs in 2016 and 204 patients with HAIs in 2022 were enrolled. After a 1:1 match, all 431 pairs were recruited as FAS, of which 332 pairs as PPS met all matching restrictions. Compared to the 2016 cohort in FAS, the patients with HAIs in 2022 had a significantly older age (64.40 ± 16.45 years), higher repeat hospitalization rate (65 [32.02%] of 203), and lower immune function (69 [33.99%] of 203). The discounted costs and adjusted-discounted costs for patients with HAIs in the 2022 cohort were found to be significantly higher than those of patients without HAIs (discounted costs: $5484.60 [IQR 8426.03] vs $2554.04(4530.82), P < 0.001; adjusted-discounted costs: $5235.90 [3772.12] vs $3040.21(1823.36), P < 0.001, respectively), and also higher than those of patients with HAIs in the 2016 cohort (discounted costs: $5484.60 [8426.03] vs $3553.00 [6127.79], P < 0.001; adjusted-discounted costs: $5235.90 [3772.12] vs $3703.82 [3159.14], P < 0.001, respectively). In vertical comparison of PPS, the incremental costs of the 2022 cohort are 1.48 times higher than those of the 2016 cohort ($964.63(4076.15) vs $652.43 [2533.44], P = 0.084). CONCLUSIONS: This meticulously designed study in western China has successfully and accurately examined the economic burden attributable to HAIs. Their rapidly increasing tendency poses a serious challenge to patients, hospitals, and the medical insurance. A closer linkage between HAIs and ongoing motivating system changes is urgently needed in western China.

Elucidation of the Mechanism of Rapid Growth Recovery in Rice Seedlings after Exposure to Low-Temperature Low-Light Stress: Analysis of Rice Root Transcriptome, Metabolome, and Physiology.

Late spring cold is a disastrous weather condition that often affects early rice seedlings in southern China, limiting the promotion of direct seeding cultivation. However, there are few reports on the effect of these events and on the growth recovery mechanism of rice root systems after rice seedlings are exposed to this stress. This study selected the strong-growth-recovery variety B116 (R310/R974, F17) and the slow-recovery variety B811 (Zhonghui 286) for direct seeding cultivation and exposed them to low temperature and low-light stress to simulate a late spring cold event in an artificial climate chamber. The treatment consisted of 4 days of exposure to a day/night temperature of 14/10 °C and a light intensity of 266 µmol m-2s-1 while the control group was kept at a day/night temperature of 27/25 °C and light intensity of 533 µmol m-2s-1. The results showed that 6 days after stress, the total length, surface area, and volume of B116 roots increased by 335.5%, 290.1%, and 298.5%, respectively, while those of B811 increased by 228.8%, 262.0%, and 289.1%, respectively. In B116, the increase in root fresh weight was 223.1%, and that in B811 was 165.6%, demonstrating rapid root recovery after stress and significant differences among genotypes. The content of H2O2 and MDA in the B116 roots decreased faster than that in the B811 roots after normal light intensity and temperature conditions were restored, and the activity of ROS metabolism enzymes was stronger in B116 roots than in B811 roots. The correlation analysis between the transcriptome and metabolome showed that endogenous signal transduction and starch and sucrose metabolism were the main metabolic pathways affecting the rapid growth of rice seedling roots after exposure to combined stress from low temperature and low light intensities. The levels of auxin and sucrose in the roots of the strong-recovery variety B116 were higher, and this variety's metabolism was downregulated significantly faster than that of B811. The auxin response factor and sucrose synthesis-related genes SPS1 and SUS4 were significantly upregulated. This study contributes to an understanding of the rapid growth recovery mechanism in rice after exposure to combined stress from low-temperature and low-light conditions.

Design and evaluation of tadpole-like conformational antimicrobial peptides.

Antimicrobial peptides are promising alternatives to conventional antibiotics. Herein, we report a class of "tadpole-like" peptides consisting of an amphipathic α-helical head and an aromatic tail. A structure-activity relationship (SAR) study of "tadpole-like" temporin-SHf and its analogs revealed that increasing the number of aromatic residues in the tail, introducing Arg to the α-helical head and rearranging the peptide topology dramatically increased antimicrobial activity. Through progressive structural optimization, we obtained two peptides, HT2 and RI-HT2, which exhibited potent antimicrobial activity, no hemolytic activity and cytotoxicity, and no propensity to induce resistance. NMR and molecular dynamics simulations revealed that both peptides indeed adopted "tadpole-like" conformations. Fluorescence experiments and electron microscopy confirmed the membrane targeting mechanisms of the peptides. Our studies not only lead to the discovery of a series of ultrashort peptides with potent broad-spectrum antimicrobial activities, but also provide a new strategy for rational design of novel "tadpole-like" antimicrobial peptides.

Reduction of p11 in dorsal raphe nucleus serotonergic neurons mediates depression-like behaviors.

The pathology of depression is related to the imbalance of various neurotransmitters. The dorsal raphe nucleus (DRN), the main brain region producing 5-HT, is crucially involved in the pathophysiology of depression. It contains several neuron types, in which GABAergic neurons are activated by stimuli associated with negative experiences and 5-HT neurons are activated by reward signals. However, little is known about its underlying molecular mechanisms. Here, we found that p11, a multifunctional protein associated with depression, was down-regulated by chronic social defeat stress in 5-HTDRN neurons. Knockdown of p11 in DRN induced depression-like behaviors, while its overexpression in 5-HTDRN neurons alleviated depression-like behavior caused by chronic social defeat stress. Further, p11 regulates membrane trafficking of glutamate receptors in 5-HTDRN neurons, suggesting a possible molecular mechanism underlying the participation of p11 in the pathological process of depression. This may facilitate the understanding of the molecular and cellular basis of depression.

S-acylation of p62 promotes p62 droplet recruitment into autophagosomes in mammalian autophagy.

p62 is a well-characterized autophagy receptor that recognizes and sequesters specific cargoes into autophagosomes for degradation. p62 promotes the assembly and removal of ubiquitinated proteins by forming p62-liquid droplets. However, it remains unclear how autophagosomes efficiently sequester p62 droplets. Herein, we report that p62 undergoes reversible S-acylation in multiple human-, rat-, and mouse-derived cell lines, catalyzed by zinc-finger Asp-His-His-Cys S-acyltransferase 19 (ZDHHC19) and deacylated by acyl protein thioesterase 1 (APT1). S-acylation of p62 enhances the affinity of p62 for microtubule-associated protein 1 light chain 3 (LC3)-positive membranes and promotes autophagic membrane localization of p62 droplets, thereby leading to the production of small LC3-positive p62 droplets and efficient autophagic degradation of p62-cargo complexes. Specifically, increasing p62 acylation by upregulating ZDHHC19 or by genetic knockout of APT1 accelerates p62 degradation and p62-mediated autophagic clearance of ubiquitinated proteins. Thus, the protein S-acylation-deacylation cycle regulates p62 droplet recruitment to the autophagic membrane and selective autophagic flux, thereby contributing to the control of selective autophagic clearance of ubiquitinated proteins.

Combining Anti-epidermal Growth Factor Receptor (EGFR) Therapy with Immunotherapy in Metastatic Colorectal Cancer (mCRC).

INTRODUCTION: Monoclonal antibodies binding the EGFR, such as cetuximab and panitumumab, have been extensively used as targeted therapy for the treatment of mCRC. However, in clinical practice, it has been found that these treatment options have some limitations and fail to fully exploit their immunoregulatory activities. Meanwhile, because of the limited effects of current treatments, immunotherapy is being widely studied for patients with mCRC. However, previous immunotherapy trials in mCRC patients have had unsatisfactory outcomes as monotherapy. Thus, combinatorial treatment strategies are being researched. AREAS COVERED: The authors retrieved relevant documents of combination therapy for mCRC from PubMed and Medline. This review elaborates on the knowledge of immunomodulatory effects of anti-EGFR therapy alone and in combination with immunotherapy for mCRC. EXPERT OPINION: Although current treatment options have improved median overall survival (OS) for advanced disease to 30 months, the prognosis remains challenging for those with metastatic disease. More recently, the combination of anti-EGFR therapy with immunotherapy has been shown activity with complementary mechanisms. Hence, anti-EGFR therapy in combination with immunotherapy may hold the key to improving the therapeutic effect of refractory mCRC.

Integrating Bulk and Single-Cell RNA Sequencing Reveals Heterogeneity, Tumor Microenvironment, and Immunotherapeutic Efficacy Based on Sialylation-Related Genes in Bladder Cancer.

Background: As known abnormal sialylation exerts crucial roles in the growth, metastasis, and immune evasion of cancers, but the molecular characteristics and roles in bladder cancer (BLCA) remain unclear. This study intends to establish BLCA risk stratification based on sialylation-related genes and elucidate its role in prognosis, tumor microenvironment, and immunotherapy of BLCA. Methods: Bulk RNA-seq and scRNA-seq data were downloaded from open-access databases. The scRNA-seq data were processed using the R package "Seurat" to identify the core cell types. The tumor sub-typing of BLCA samples was performed by the R package "ConsensusClusterPlus" in the bulk RNA-seq data. Signature genes were identified by the R package "limma" and univariate regression analysis to calculate risk scores using the R package "GSVA" and establish risk stratification of BLCA patients. Finally, the differences in clinicopathological characteristics, tumor microenvironment, and immunotherapy efficacy between the different groups were investigated. Results: 5 core cell types were identified in the scRNA-seq dataset, with monocytes and macrophages presenting the greatest percentage, sialylation-related gene expression, and sialylation scores. The bulk RNA-seq samples were classified into 3 tumor subtypes based on 19 prognosis-related sialylation genes. The 10 differential expressed genes (DEGs) with the smallest p-values were collected as signature genes, and the risk score was calculated, with the samples divided into high and low-risk score groups. The results showed that patients in the high-risk score group exhibited worse survival outcomes, higher tumor grade, more advanced stage, more frequency of gene mutations, higher expression levels of immune checkpoints, and lower immunotherapy response. Conclusion: We established a novel risk stratification of BLCA from a glycomics perspective, which demonstrated good accuracy in determining the prognostic outcome, clinicopathological characteristics, immune microenvironment, and immunotherapy efficacy of patients, and we are proposing to apply it to direct the choice of clinical treatment options for patients.

Tethering ATG16L1 or LC3 induces targeted autophagic degradation of protein aggregates and mitochondria.

Proteolysis-targeting chimeras (PROTACs) based on the ubiquitin-proteasome system have made great progress in the field of drug discovery. There is mounting evidence that the accumulation of aggregation-prone proteins or malfunctioning organelles is associated with the occurrence of various age-related neurodegenerative disorders and cancers. However, PROTACs are inefficient for the degradation of such large targets due to the narrow entrance channel of the proteasome. Macroautophagy (hereafter referred to as autophagy) is known as a self-degradative process involved in the degradation of bulk cytoplasmic components or specific cargoes that are sequestered into autophagosomes. In the present study, we report the development of a generalizable strategy for the targeted degradation of large targets. Our results suggested that tethering large target models to phagophore-associated ATG16L1 or LC3 induced targeted autophagic degradation of the large target models. Furthermore, we successfully applied this autophagy-targeting degradation strategy to the targeted degradation of HTT65Q aggregates and mitochondria. Specifically, chimeras consisting of polyQ-binding peptide 1 (QBP) and ATG16L1-binding peptide (ABP) or LC3-interacting region (LIR) induced targeted autophagic degradation of pathogenic HTT65Q aggregates; and the chimeras consisting of mitochondria-targeting sequence (MTS) and ABP or LIR promoted targeted autophagic degradation of dysfunctional mitochondria, hence ameliorating mitochondrial dysfunction in a Parkinson disease cell model and protecting cells from apoptosis induced by the mitochondrial stress agent FCCP. Therefore, this study provides a new strategy for the selective proteolysis of large targets and enrich the toolkit for autophagy-targeting degradation.Abbreviations: ABP: ATG16L1-binding peptide; ATG16L1: autophagy related 16 like 1; ATTEC: autophagy-tethering compound; AUTAC: autophagy-targeting chimera; AUTOTAC: autophagy-targeting chimera; Baf A1: bafilomycin A1; BCL2: BCL2 apoptosis regulator; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; CASP3: caspase 3; CPP: cell-penetrating peptide; CQ: chloroquine phosphate; DAPI: 4',6-diamidino-2-phenylindole; DCM: dichloromethane; DMF: N,N-dimethylformamide; DMSO: dimethyl sulfoxide; EBSS: Earle's balanced salt solution; FCCP: carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone; FITC: fluorescein-5-isothiocyanate; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GFP: green fluorescent protein; HEK293: human embryonic kidney 293; HEK293T: human embryonic kidney 293T; HPLC: high-performance liquid chromatography; HRP: horseradish peroxidase; HTT: huntingtin; LIR: LC3-interacting region; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MFF: mitochondrial fission factor; MTS: mitochondria-targeting sequence; NBR1: NBR1 autophagy cargo receptor; NLRX1: NLR family member X1; OPTN: optineurin; P2A: self-cleaving 2A peptide; PB1: Phox and Bem1p; PBS: phosphate-buffered saline; PE: phosphatidylethanolamine; PINK1: PTEN induced kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; PROTACs: proteolysis-targeting chimeras; QBP: polyQ-binding peptide 1; SBP: streptavidin-binding peptide; SDS-PAGE: sodium dodecyl sulfate-polyacrylamide gel electrophoresis; SPATA33: spermatogenesis associated 33; TIMM23: translocase of inner mitochondrial membrane 23; TMEM59: transmembrane protein 59; TOMM20: translocase of outer mitochondrial membrane 20; UBA: ubiquitin-associated; WT: wild type.