Relationship between HLA genetic variations, COVID-19 vaccine antibody response, and risk of breakthrough outcomes.

The rapid global distribution of COVID-19 vaccines, with over a billion doses administered, has been unprecedented. However, in comparison to most identified clinical determinants, the implications of individual genetic factors on antibody responses post-COVID-19 vaccination for breakthrough outcomes remain elusive. Here, we conducted a population-based study including 357,806 vaccinated participants with high-resolution HLA genotyping data, and a subset of 175,000 with antibody serology test results. We confirmed prior findings that single nucleotide polymorphisms associated with antibody response are predominantly located in the Major Histocompatibility Complex region, with the expansive HLA-DQB1*06 gene alleles linked to improved antibody responses. However, our results did not support the claim that this mutation alone can significantly reduce COVID-19 risk in the general population. In addition, we discovered and validated six HLA alleles (A*03:01, C*16:01, DQA1*01:02, DQA1*01:01, DRB3*01:01, and DPB1*10:01) that independently influence antibody responses and demonstrated a combined effect across HLA genes on the risk of breakthrough COVID-19 outcomes. Lastly, we estimated that COVID-19 vaccine-induced antibody positivity provides approximately 20% protection against infection and 50% protection against severity. These findings have immediate implications for functional studies on HLA molecules and can inform future personalised vaccination strategies.

Arsenic removal in flue gas through anaerobic denitrification and sulfate reduction cocoupled arsenic oxidation.

Arsenic in flue gas from municipal solid waste incineration can damage to human health and ecological environment. A sulfate-nitrate-reducing bioreactor (SNRBR) for flue gas arsenic removal was investigated. Arsenic removal efficiency attained 89.4%. An integrated metagenomic and metaproteomic investigation showed that three nitrate reductases (NapA, NapB and NarG), three sulfate reductases (Sat, AprAB and DsrAB), and arsenite oxidase (ArxA) regulated nitrate reduction, sulfate reduction and bacterial As(III)-oxidation, respectively. Citrobacter and Desulfobulbus could synthetically regulate the expression of arsenite-oxidizing gene, nitrate reductases and sulfate reducatases, which involved in As(III) oxidation, nitrate and sulfate reduction. A bacterial consortium containing Citrobacter, UG_Enterobacteriaceas, Desulfobulbus and Desulfovibrio could capable of simultaneously arsenic oxidation, sulfate reduction and denitrification. Anaerobic denitrification and sulfate reduction were cocoupled to arsenic oxidation. The biofilm was characterized by FTIR, XPS, XRD, EEM, and SEM. XRD and XPS spectra verified the formation of aarsenic species (As(V)) from flue gas As(III) conversion. Arsenic speciation in biofilms of SNRBR consisted of 77% residual arsenic, 15.9% organic matter-bound arsenic, and 4.3% strongly absorbed arsenic. Flue gas arsenic was bio-stabilized in the form of Fe-As-S and As-EPS through biodeposition, biosorption and biocomplexation. This provides a new way of flue gas arsenic removal using the sulfate-nitrate-reducing bioreactor.

Pb0 flue gas biosorption in denitrifying MBfR: Characteristics and binding ability of extracellular polymeric substances.

Pb0 in flue gas which is ubiquitous in the environment, poses a certain threat to human and ecology, but the study on EPS-dependent stabilization of lead to remove Pb0 from flue gas remains insufficient. In this investigation, the characteristics and heavy metals-binding ability of four EPS fractions were evaluated. The EPS were extracted from denitrifying membrane biofilm reactor (MBfR) and divided into slime EPS (S-EPS), loosely-bound EPS (LB-EPS), tightly-bound EPS (TB-EPS) and EPS in circulating flow (Y-EPS). The S, LB, TB-EPS related to Pb stabilization on biofilm need more attention. Compared to Pb-S-EPS (0.013 mg g-1) and Pb-LB-EPS (0.13 mg g-1), the Pb-TB-EPS (0.26 mg g-1) was mainly stable form of vapor Pb0, since TB-EPS's higher content (30.67-82.44 mg g-1 VSS), proteins (13.47-36.32 mg g-1 VSS) and polysaccharides (9.37-32.48 mg g-1 VSS) concentration. Particularly, proteins related ligands were more effective in S, LB, TB-EPS dependent adsorption of Pb, complexing with hydrophobic acid ligands further strengthened in TB-EPS adsorption. The Pb-EPS complex formed via binding with functional groups (such as O-H, N-H, C-H and CC) on EPS, also facilitated by loose structure of proteins. This study enlightens the researchers on the bio-treatment and EPS-dependent biosorption of Pb0 in flue gas in denitrifying MBfR.

Lead removal in flue gas from sludge incineration by denitrification: Insights from metagenomics and metaproteomics.

Flue gas lead emission during sludge incineration damages to human health and ecological environment seriously. Therefore, a denitrifying bio-trickling filter (DNBTF) for lead removal in flue gas from sludge incineration was investigated. Lead removal efficiency was up to 90.7% in 60 days' operation. Lead speciation in biofilms of DNBTF consists of 84.27% residue lead, 15.18% organic bound lead, and less than 1% exchangeable and reducible lead. Lead resistant bacteria and lead resistant-denitrifying bacteria accounted for 85.04% and 58.25%, respectively. Lead resistant microorganisms(Pseudomonas, Azoarcus, Stappia, Pararhodobacter, Paracoccus, Azospirillum, Hyphomonas, Rhodobacter, Polymorphum, Brevunimonas, Stenotrophomonas) could resist the toxicity of Pb2+ in flue gas by transport protein and binding protein, and detoxify Pb2+ in flue gas by extracellular polymeric substances (EPS) adsorption, protein binding and precipitation under the action of resistance genes, such as pbrAB, golT, troABCD, znuABC, czcABCD, pcoB, copA, as shown by integrated metagenomic and metaproteomic analyses. The biofilm was characterized by FTIR, XRD, 3D-EEM, and SEM-EDS. XRD and SEM-EDS spectra indicated the formation of pyromorphite from bioconversion of lead in flue gas. Lead-containing flue gas was bio-stabilized in the form of pyromorphite and HA-Pb via complexation of humic acids in extracellular polymeric substances (EPS), biosorption and biodeposition. This provides a new way of sludge incineration flue gas lead removal using a denitrifying biotricking filter.

Microwave Irradiation: Effects on the Change of Colour Characteristics and Main Phenolic Compounds of Cabernet Gernischt Dry Red Wine during Storage.

Wine colour is an essential organoleptic property considered by consumers. In this paper, the potential effects on colour characteristics and the content of main phenolic compounds in red wine under microwave irradiation were investigated during wine storage. The results showed that the changing trend of colour characteristics of microwave-treated and untreated wines was very similar. Moreover, total phenolic compounds, total monomeric anthocyanins, main anthocyanins, main flavonoids, and main phenolic acids (gallic acid; caffeic acid; syringic acid; (+)-catechin; Cy-3-glu; Mv-3-glu) also showed similar change trends during storage. In other words, microwave irradiation had a long-term effect on the colour properties and main phenolic compounds of red wine, changes that require long-time aging in traditional processing. In terms of the studied parameters, the changes in microwave-treated wine were faster than those in untreated wine. These results showed that microwave technology, as a promising artificial aging technology, could in a short time produce red wine of similar quality to traditional aging.

Disease-associated KBTBD4 mutations in medulloblastoma elicit neomorphic ubiquitylation activity to promote CoREST degradation.

Medulloblastoma is the most common malignant brain tumour in children. Genomic studies have identified distinct disease subgroups: wnt/wingless (WNT), sonic hedgehog (SHH), and non-WNT/non-SHH, comprising group 3 and group 4. Alterations in WNT and SHH signalling form the pathogenetic basis for their subgroups, whereas those for non-WNT/non-SHH tumours remain largely elusive. Recent analyses have revealed recurrent in-frame insertions in the E3 ubiquitin ligase adaptor Kelch Repeat and BTB Domain Containing 4 (KBTBD4) in cases of group 3/4 medulloblastoma. Critically, group 3/4 tumours with KBTBD4 mutations typically lack other gene-specific alterations, such as MYC amplification, indicating KBTBD4 insertion mutations as the primary genetic driver. Delineating the role of KBTBD4 mutations thus offers significant opportunities to understand tumour pathogenesis and to exploit the underpinning mechanisms therapeutically. Here, we show a novel mechanism in cancer pathogenesis whereby indel mutations in KBTBD4 drive its recognition of neo-substrates for degradation. We observe that KBTBD4 mutants promote the recruitment and ubiquitylation of the REST Corepressor (CoREST), which forms a complex to modulate chromatin accessibility and transcriptional programmes. The degradation of CoREST promoted by KBTBD4 mutation diverts epigenetic programmes inducing significant alterations in transcription to promote increased stemness of cancer cells. Transcriptional analysis of >200 human group 3 and 4 medulloblastomas by RNA-seq, highlights the presence of CoREST and stem-like signatures in tumours with KBTBD4 mutations, which extend to a further sub-set of non-mutant tumours, suggesting CoREST alterations as a novel pathogenetic mechanism of wide relevance in groups 3 and 4. Our findings uncover KBTBD4 mutation as a novel driver of epigenetic reprogramming in non-WNT/non-SHH medulloblastoma, establish a novel mode of tumorigenesis through gain-of-function mutations in ubiquitin ligases (neo-substrate recruitment) and identify both mutant KBTBD4 and CoREST complexes as new druggable targets for improved tumour-specific therapies.

Sequence and structural variations determining the recruitment of WNK kinases to the KLHL3 E3 ligase.

The BTB-Kelch protein KLHL3 is a Cullin3-dependent E3 ligase that mediates the ubiquitin-dependent degradation of kinases WNK1-4 to control blood pressure and cell volume. A crystal structure of KLHL3 has defined its binding to an acidic degron motif containing a PXXP sequence that is strictly conserved in WNK1, WNK2 and WNK4. Mutations in the second proline abrograte the interaction causing the hypertension syndrome pseudohypoaldosteronism type II. WNK3 shows a diverged degron motif containing four amino acid substitutions that remove the PXXP motif raising questions as to the mechanism of its binding. To understand this atypical interaction, we determined the crystal structure of the KLHL3 Kelch domain in complex with a WNK3 peptide. The electron density enabled the complete 11-mer WNK-family degron motif to be traced for the first time revealing several conserved features not captured in previous work, including additional salt bridge and hydrogen bond interactions. Overall, the WNK3 peptide adopted a conserved binding pose except for a subtle shift to accommodate bulkier amino acid substitutions at the binding interface. At the centre, the second proline was substituted by WNK3 Thr541, providing a unique phosphorylatable residue among the WNK-family degrons. Fluorescence polarisation and structural modelling experiments revealed that its phosphorylation would abrogate the KLHL3 interaction similarly to hypertension-causing mutations. Together, these data reveal how the KLHL3 Kelch domain can accommodate the binding of multiple WNK isoforms and highlight a potential regulatory mechanism for the recruitment of WNK3.

Ubiquitin ligases: guardians of mammalian development.

Mammalian development demands precision. Millions of molecules must be properly located in temporal order, and their function regulated, to orchestrate important steps in cell cycle progression, apoptosis, migration and differentiation, to shape developing embryos. Ubiquitin and its associated enzymes act as cellular guardians to ensure precise spatio-temporal control of key molecules during each of these important cellular processes. Loss of precision results in numerous examples of embryological disorders or even cancer. This Review discusses the crucial roles of E3 ubiquitin ligases during key steps of early mammalian development and their roles in human disease, and considers how new methods to manipulate and exploit the ubiquitin regulatory machinery - for example, the development of molecular glues and PROTACs - might facilitate clinical therapy.

The role of E3 ubiquitin ligases in the development and progression of glioblastoma.

Despite recent advances in our understanding of the disease, glioblastoma (GB) continues to have limited treatment options and carries a dismal prognosis for patients. Efforts to stratify this heterogeneous malignancy using molecular classifiers identified frequent alterations in targetable proteins belonging to several pathways including the receptor tyrosine kinase (RTK) and mitogen-activated protein kinase (MAPK) signalling pathways. However, these findings have failed to improve clinical outcomes for patients. In almost all cases, GB becomes refractory to standard-of-care therapy, and recent evidence suggests that disease recurrence may be associated with a subpopulation of cells known as glioma stem cells (GSCs). Therefore, there remains a significant unmet need for novel therapeutic strategies. E3 ubiquitin ligases are a family of >700 proteins that conjugate ubiquitin to target proteins, resulting in an array of cellular responses, including DNA repair, pro-survival signalling and protein degradation. Ubiquitin modifications on target proteins are diverse, ranging from mono-ubiquitination through to the formation of polyubiquitin chains and mixed chains. The specificity in substrate tagging and chain elongation is dictated by E3 ubiquitin ligases, which have essential regulatory roles in multiple aspects of brain cancer pathogenesis. In this review, we begin by briefly summarising the histological and molecular classification of GB. We comprehensively describe the roles of E3 ubiquitin ligases in RTK and MAPK, as well as other, commonly altered, oncogenic and tumour suppressive signalling pathways in GB. We also describe the role of E3 ligases in maintaining glioma stem cell populations and their function in promoting resistance to ionizing radiation (IR) and chemotherapy. Finally, we consider how our knowledge of E3 ligase biology may be used for future therapeutic interventions in GB, including the use of blood-brain barrier permeable proteolysis targeting chimeras (PROTACs).

BiVO4 prepared by the sol-gel doped on graphite felt cathode for ciprofloxacin degradation and mechanism in solar-photo-electro-Fenton.

In this research, bismuth vanadate-doped graphite felt (GF-BiVO4) was successfully prepared by sol-gel method, in which BiVO4 owned superior electro-Fenton (EF) and solar-photo-electro-Fenton (SPEF) performance. Combined with the analysis by X-ray diffractometer (XRD), field emission transmission electron microscopy (FE-TEM), nitrogen adsorption-desorption isotherms and cyclic voltammetry (CV), the changes of electrodes were reflected in structure and physicochemical properties. The doping of monoclinic BiVO4 endued GF with a higher surface area and more electro-active sites and better electrode activity in comparison to Raw-GF. Then, the GFs were used as cathodes to detect •OH concentration with coumarin (COU) as probe molecule and to evaluate photoelectric performance with ciprofloxacin (CIP) in photocatalysis, EF and SPEF processes. The results demonstrated that the concentration of •OH followed an order of SPEF> EF> photocatalysis, which was consistent with the removal rate of CIP (99.8%, 99.4% and 21.2%, respectively) on GF-BiVO4 at 5 min. Further, five degradation pathways of CIP in SPEF system were proposed including the attack on piperazine ring, oxidation on cyclopropyl group, decarboxylation and hydroxyl radical addition, oxidation on benzene group and defluorination. The study provides insights into the enhancement of EF and SPEF performance and the degradation pathway of CIP in SPEF.

Anodic oxidation of ciprofloxacin using different graphite felt anodes: Kinetics and degradation pathways.

Ciprofloxacin (CIP) is ubiquitous in the environment which poses a certain threat to human and ecology. In this investigation, the physical and electrochemical properties of graphite felt (GF) anodes which affected the anodic oxidation (AO) performance, and the CIP removal effect of GF were evaluated. The GFs were used as anodes for detection of ·OH with coumarin (COU) as molecule probe and removal of CIP in a 150 mL electrolytic cell with Pt cathode (AO-GF/Pt system). The results showed that hydrophilic GF (B-GF) owned higher sp3/sp2 and more oxygen-containing and nitrogen-containing functional groups than the hydrophobic GF (A-GF). Moreover, B-GF possessed higher oxygen evolution potential (1.12 V), more active sites and stronger ·OH generation capacity. Above mentioned caused that B-GF exhibited more superior properties for CIP removal. The best efficiencies (96.95%, 99.83%) were obtained in the AO-B-GF/Pt system at 6.25 mAcm-2 after 10 min (k1, 0.356 min-1) and 60 min (k2, 0.224 min-1), respectively. Furthermore, nine degradation pathways of CIP in AO-B-GF/Pt system were summarized as the cleavage of the piperazine ring, cyclopropyl group, quinolone ring and F atom by ·OH. It provides new insights into the removal and degradation pathways of CIP with GF in AO system.

Identification of a PGXPP degron motif in dishevelled and structural basis for its binding to the E3 ligase KLHL12.

Wnt signalling is dependent on dishevelled proteins (DVL1-3), which assemble an intracellular Wnt signalosome at the plasma membrane. The levels of DVL1-3 are regulated by multiple Cullin-RING E3 ligases that mediate their ubiquitination and degradation. The BTB-Kelch protein KLHL12 was the first E3 ubiquitin ligase to be identified for DVL1-3, but the molecular mechanisms determining its substrate interactions have remained unknown. Here, we mapped the interaction of DVL1-3 to a 'PGXPP' motif that is conserved in other known partners and substrates of KLHL12, including PLEKHA4, PEF1, SEC31 and DRD4. To determine the binding mechanism, we solved a 2.4 Å crystal structure of the Kelch domain of KLHL12 in complex with a DVL1 peptide that bound with low micromolar affinity. The DVL1 substrate adopted a U-shaped turn conformation that enabled hydrophobic interactions with all six blades of the Kelch domain ß-propeller. In cells, the mutation or deletion of this motif reduced the binding and ubiquitination of DVL1 and increased its stability confirming this sequence as a degron motif for KLHL12 recruitment. These results define the molecular mechanisms determining DVL regulation by KLHL12 and establish the KLHL12 Kelch domain as a new protein interaction module for a novel proline-rich motif.

Screening of cardioprotective diseases bioactive components from Danshen extracts and LC-MS analysis.

A rapid and efficient analysis and screening method is adopted for cell affinity capture coupled with HPLC-MS (CAC-HPLC-MS) analysis of bioactive components that have possible efficiency against cardiovascular diseases. This method involves affinity capture, concentration, and separation of bioactive components from Danshen library using oxidatively damaged endothelial cells induced by H2 O2 , as well as analysis and identification of targeted compounds with HPLC and MS. It combines the specific interaction between cell membrane receptors and bioactive components with the powerful analysis and identification function of HPLC-MS. The CAC-HPLC-MS method was also used for analysis and screening of bioactive components from crude extracts of Danshen. A total of 19 components were found to be bound to oxidatively damaged endothelial cells with seven of these identified. Existing literature confirms that these seven components have many activities related to cardioprotective diseases. Therefore, the combination of biological affinity capture with HPLC-MS should be regarded as an attractive method with great potential for rapid and efficient screening of bioactive components related to anti-cardiovascular diseases from natural product libraries.

Kinetics and mechanisms of oxytetracycline degradation in an electro-Fenton system with a modified graphite felt cathode.

The removal of trace antibiotics from the aquatic environment has received great interest. In this investigation, NaOH activated graphite felt (NaOH-GF) was characterized by multiple-methods, including scanning electron microscopy (SEM), X-ray photoelectron spectroscopy (XPS), contact angle, linear sweep voltammetry (LSV) and electron paramagnetic resonance (EPR). The NaOH-GF was then used as the cathode in the electro-Fenton process for oxytetracycline (OTC) degradation, the experiment was carried out in an undivided and light-proof beaker with a Pt anode and a NaOH-GF cathode at pH 3. The results showed that the modification with NaOH enhanced the antibiotics degradation efficiency of graphite felt by increasing the oxygen reduction capacity and hydroxyl radicals yielding rate. Complete OTC removal was achieved at 5.17 mA cm-2 after 40, 60 and 90 s with initial OTC concentration of 22, 44, and 66 µM, respectively. With an initial OTC concentration of 44 µM, after 30 min the removal rates of chemical oxygen demand (COD) by Raw-GF and NaOH-GF were 59.18% and 83.75%, respectively. The proposed degradation mechanism of OTC was an EF process, which consisted of hydroxylation, secondary alcohol oxidation, demethylation, decarbonylation, dehydration and deamination. This study demonstrates that NaOH activated GF cathode possesses high degradation capacity and good stability. It provides insight into the removal of non-biodegradable antibiotics and may shed light on future to its practical application.

Structural Basis for Recruitment of DAPK1 to the KLHL20 E3 Ligase.

BTB-Kelch proteins form the largest subfamily of Cullin-RING E3 ligases, yet their substrate complexes are mapped and structurally characterized only for KEAP1 and KLHL3. KLHL20 is a related CUL3-dependent ubiquitin ligase linked to autophagy, cancer, and Alzheimer's disease that promotes the ubiquitination and degradation of substrates including DAPK1, PML, and ULK1. We identified an "LPDLV"-containing motif in the DAPK1 death domain that determines its recruitment and degradation by KLHL20. A 1.1-Å crystal structure of a KLHL20 Kelch domain-DAPK1 peptide complex reveals DAPK1 binding as a loose helical turn that inserts deeply into the central pocket of the Kelch domain to contact all six blades of the ß propeller. Here, KLHL20 forms salt-bridge and hydrophobic interactions including tryptophan and cysteine residues ideally positioned for covalent inhibitor development. The structure highlights the diverse binding modes of ß-propeller domains versus linear grooves and suggests a new target for structure-based drug design.

The Arabidopsis RING-Type E3 Ligase TEAR1 Controls Leaf Development by Targeting the TIE1 Transcriptional Repressor for Degradation.

The developmental plasticity of leaf size and shape is important for leaf function and plant survival. However, the mechanisms by which plants form diverse leaves in response to environmental conditions are not well understood. Here, we identified TIE1-ASSOCIATED RING-TYPE E3 LIGASE1 (TEAR1) and found that it regulates leaf development by promoting the degradation of TCP INTERACTOR-CONTAINING EAR MOTIF PROTEIN1 (TIE1), an important repressor of CINCINNATA (CIN)-like TEOSINTE BRANCHED1/CYCLOIDEA/PCF (TCP) transcription factors, which are key for leaf development. TEAR1 contains a typical C3H2C3-type RING domain and has E3 ligase activity. We show that TEAR1 interacts with the TCP repressor TIE1, which is ubiquitinated in vivo and degraded by the 26S proteasome system. We demonstrate that TEAR1 is colocalized with TIE1 in nuclei and negatively regulates TIE1 protein levels. Overexpression of TEAR1 rescued leaf defects caused by TIE1 overexpression, whereas disruption of TEAR1 resulted in leaf phenotypes resembling those caused by TIE1 overexpression or TCP dysfunction. Deficiency in TEAR partially rescued the leaf defects of TCP4 overexpression line and enhanced the wavy leaf phenotypes of jaw-5D We propose that TEAR1 positively regulates CIN-like TCP activity to promote leaf development by mediating the degradation of the TCP repressor TIE1.

The molecular mechanism of sporocyteless/nozzle in controlling Arabidopsis ovule development.

Ovules are essential for plant reproduction and develop into seeds after fertilization. Sporocyteless/nozzle (SPL/NZZ) has been known for more than 15 years as an essential factor for ovule development in Arabidopsis, but the biochemical nature of SPL function has remained unsolved. Here, we demonstrate that SPL functions as an adaptor-like transcriptional repressor. We show that SPL recruits topless/topless-related (TPL/TPR) co-repressors to inhibit the Cincinnata (CIN)-like Teosinte branched1/cycloidea/PCF (TCP) transcription factors. We reveal that SPL uses its EAR motif at the C-terminal end to recruit TPL/TPRs and its N-terminal part to bind and inhibit the TCPs. We demonstrate that either disruption of TPL/TPRs or overexpression of TCPs partially phenocopies the defects of megasporogenesis in spl. Moreover, disruption of TCPs causes phenotypes that resemble spl-D gain-of-function mutants. These results define the action mechanism for SPL, which along with TPL/TPRs controls ovule development by repressing the activities of key transcription factors. Our findings suggest that a similar gene repression strategy is employed by both plants and fungi to control sporogenesis.