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PLoS Computational Biology Jun 2024Carbohydrates and glycoproteins modulate key biological functions. However, experimental structure determination of sugar polymers is notoriously difficult....
Carbohydrates and glycoproteins modulate key biological functions. However, experimental structure determination of sugar polymers is notoriously difficult. Computational approaches can aid in carbohydrate structure prediction, structure determination, and design. In this work, we developed a glycan-modeling algorithm, GlycanTreeModeler, that computationally builds glycans layer-by-layer, using adaptive kernel density estimates (KDE) of common glycan conformations derived from data in the Protein Data Bank (PDB) and from quantum mechanics (QM) calculations. GlycanTreeModeler was benchmarked on a test set of glycan structures of varying lengths, or "trees". Structures predicted by GlycanTreeModeler agreed with native structures at high accuracy for both de novo modeling and experimental density-guided building. We employed these tools to design de novo glycan trees into a protein nanoparticle vaccine to shield regions of the scaffold from antibody recognition, and experimentally verified shielding. This work will inform glycoprotein model prediction, glycan masking, and further aid computational methods in experimental structure determination and refinement.
PubMed: 38913746
DOI: 10.1371/journal.pcbi.1011895 -
PLoS Pathogens Jun 2024COVID-associated coagulopathy seemly plays a key role in post-acute sequelae of SARS- CoV-2 infection. However, the underlying pathophysiological mechanisms are poorly...
COVID-associated coagulopathy seemly plays a key role in post-acute sequelae of SARS- CoV-2 infection. However, the underlying pathophysiological mechanisms are poorly understood, largely due to the lack of suitable animal models that recapitulate key clinical and pathological symptoms. Here, we fully characterized AC70 line of human ACE2 transgenic (AC70 hACE2 Tg) mice for SARS-CoV-2 infection. We noted that this model is highly permissive to SARS-CoV-2 with values of 50% lethal dose and infectious dose as ~ 3 and ~ 0.5 TCID50 of SARS-CoV-2, respectively. Mice infected with 105 TCID50 of SARS-CoV-2 rapidly succumbed to infection with 100% mortality within 5 days. Lung and brain were the prime tissues harboring high viral titers, accompanied by histopathology. However, viral RNA and inflammatory mediators could be detectable in other organs, suggesting the nature of a systemic infection. Lethal challenge of AC70 hACE2 Tg mice caused acute onset of leukopenia, lymphopenia, along with an increased neutrophil-to-lymphocyte ratio (NLR). Importantly, infected animals recapitulated key features of COVID-19-associated coagulopathy. SARS-CoV-2 could induce the release of circulating neutrophil extracellular traps (NETs), along with activated platelet/endothelium marker. Immunohistochemical staining with anti-platelet factor-4 (PF4) antibody revealed profound platelet aggregates especially within blocked veins of the lungs. We showed that acute SARS-CoV-2 infection triggered a hypercoagulable state coexisting with ill-regulated fibrinolysis. Finally, we highlighted the potential role of Annexin A2 (ANXA2) in fibrinolytic failure. ANXA2 is a calcium-dependent phospholipid-binding protein that forms a heterotertrameric complexes localized at the extracellular membranes with two S100A10 small molecules acting as a co-receptor for tissue-plasminogen activator (t-PA), tightly involved in cell surface fibrinolysis. Thus, our results revealing elevated IgG type anti-ANXA2 antibody production, downregulated de novo ANXA2/S100A10 synthesis, and reduced ANXA2/S100A10 association in infected mice, this protein might serve as druggable targets for development of antithrombotic and/or anti-fibrinolytic agents to attenuate pathogenesis of COVID-19.
PubMed: 38913740
DOI: 10.1371/journal.ppat.1011777 -
BMJ Neurology Open 2024Dystonia is a genetic or non-genetic movement disorder with typical patterned and twisting movements due to abnormal muscle contractions that may be associated with...
BACKGROUND
Dystonia is a genetic or non-genetic movement disorder with typical patterned and twisting movements due to abnormal muscle contractions that may be associated with tremor. Genetic and phenotypic heterogeneity leads to variable clinical presentation.
METHODOLOGY
Next-generation sequencing technologies are being currently used in the workup of patients with inherited dystonia to determine the specific cause in the individuals with autosomal dominant, recessive, X-linked or mitochondrial inheritance patterns. Calcium voltage-gated channel subunit alpha1 A (CACNA1A) gene variants are rare in dystonias.
RESULTS
We here present a 20-year-old man with a history of delayed milestones, flexor posturing, dysarthria, dysphagia and a negative family history from consanguineous parents. Neurological examination revealed right lateral scoliosis of the neck and generalised dystonic posturing affecting both upper and lower limbs. MRI of the brain was unremarkable. Molecular genetic results revealed a heterozygous variant in the CACNA1A gene (CHR19: NM_023035.2, c. 1602G>A; p. Met534Ile). Segregation analyses in both the parents revealed wild-type CACNA1A gene suggesting de novo nature of the variant with a likely pathogenic classification.
CONCLUSION
Dystonia is one of the clinical phenotypes that can be associated with CACNA1A gene mutations and we recommend that this gene either be included in the dystonia panel offered or tested when the initial primary genetic result is negative.
PubMed: 38912174
DOI: 10.1136/bmjno-2024-000710 -
BMC Genomic Data Jun 2024The sweet chestnut Castanea sativa Mill. is the only native Castanea species in Europe, and it is a tree of high economic value that provides appreciated fruits and...
OBJECTIVES
The sweet chestnut Castanea sativa Mill. is the only native Castanea species in Europe, and it is a tree of high economic value that provides appreciated fruits and valuable wood. In this study, we assembled a high-quality nuclear genome of the ancient Italian chestnut variety 'Marrone di Chiusa Pesio' using a combination of Oxford Nanopore Technologies long reads, whole-genome and Omni-C Illumina short reads.
DATA DESCRIPTION
The genome was assembled into 238 scaffolds with an N50 size of 21.8 Mb and an N80 size of 7.1 Mb for a total assembled sequence of 750 Mb. The BUSCO assessment revealed that 98.6% of the genome matched the embryophyte dataset, highlighting good completeness of the genetic space. After chromosome-level scaffolding, 12 chromosomes with a total length of 715.8 and 713.0 Mb were constructed for haplotype 1 and haplotype 2, respectively. The repetitive elements represented 37.3% and 37.4% of the total assembled genome in haplotype 1 and haplotype 2, respectively. A total of 57,653 and 58,146 genes were predicted in the two haplotypes, and approximately 73% of the genes were functionally annotated using the EggNOG-mapper. The assembled genome will be a valuable resource and reference for future chestnut breeding and genetic improvement.
Topics: Fagaceae; Genome, Plant; Chromosomes, Plant; Haplotypes; Molecular Sequence Annotation
PubMed: 38909221
DOI: 10.1186/s12863-024-01245-7 -
Cell Death Discovery Jun 2024The physiological quantum of stress-inducible transcriptional protein, Lens Epithelium-Derived Growth Factor (LEDGF), is vital for the maintenance of cellular...
The physiological quantum of stress-inducible transcriptional protein, Lens Epithelium-Derived Growth Factor (LEDGF), is vital for the maintenance of cellular physiology. Erratic epigenetic reprogramming in response to oxidative stress or with advancing age is found to be a major cause in the gene silencing, leading to pathobiologies. Using aging human (h) eye lens/lens epithelial cells (LECs) coupled with redox-active Peroxiredoxin 6 (Prdx6)-deficient (Prdx6) mLECs as model systems, herein, we showed that in aging/oxidative stress, the human LEDGF gene was regulated by unique methylation patterns of CGs nucleotides within and around the Sp1 binding site(s) of CpG island of the LEDGF promoter (-170 to -27nts). The process caused the repression of LEDGF and its target, Hsp27, resulting in reactive oxygen species (ROS) amplification and cellular insults. This phenomenon was opposed to the unmethylated promoter in LECs. Clinically, we observed that the loss of LEDGF in the Prdx6 mLECs or aging lenses/LECs, correlating with increased expression of DNMT1, DNMT3a, and DNMT3b along with the methyl CpG binding protein 2 (MeCP2). Upon oxidative stress, the expression of these molecules was increased with the dramatic reduction in LEDGF expression. While demethylating agent, 5-Aza deoxycytidine (5-AzaC) transposed the aberrant methylation status, and revived LEDGF and Hsp27 expression. Mechanistically, the chloramphenicol acetyltransferase (CAT) reporter gene driven by the LEDGF promoter (-170/ + 35) and ChIP assays uncovered that 5-AzaC acted on GC/Sp1 sites to release LEDGF transcription. The data argued, for the first time, that de novo methylation of CGs around and within Sp1 sites of the CpG island directly disrupted Sp1 activity, which ensued in LEDGF repression and its biological functions. The findings should improve our understanding of cellular insults-associated with aberrant DNMTs-mediated LEDGF's activity, and can offer strategies for therapeutic intervention to halt aging/oxidative stress-induced abnormalities.
PubMed: 38909054
DOI: 10.1038/s41420-024-02076-2 -
Journal For Immunotherapy of Cancer Jun 2024The immunosuppressive tumor microenvironment (TME) plays an essential role in cancer progression and immunotherapy response. Despite the considerable advancements in...
BACKGROUND AND AIMS
The immunosuppressive tumor microenvironment (TME) plays an essential role in cancer progression and immunotherapy response. Despite the considerable advancements in cancer immunotherapy, the limited response to immune checkpoint blockade (ICB) therapies in patients with hepatocellular carcinoma (HCC) remains a major challenge for its clinical implications. Here, we investigated the molecular basis of the protein O-fucosyltransferase 1 (POFUT1) that drives HCC immune evasion and explored a potential therapeutic strategy for enhancing ICB efficacy.
METHODS
De novo liver tumor and the xenograft tumor models were used to evaluate the function of POFUT1 in immune evasion. Biochemical assays were performed to elucidate the underlying mechanism of POFUT1-mediated immune evasion.
RESULTS
We identified POFUT1 as a crucial promoter of immune evasion in liver cancer. Notably, POFUT1 promoted HCC progression and inhibited T-cell infiltration in the xenograft tumor and de novo MYC/Trp53 mouse liver tumor models. Mechanistically, we demonstrated that POFUT1 stabilized programmed death ligand 1 (PD-L1) protein by preventing tripartite motif containing 21-mediated PD-L1 ubiquitination and degradation independently of its protein-O-fucosyltransferase activity. In addition, we further demonstrated that PD-L1 was required for the tumor-promoting and immune evasion effects of POFUT1 in HCC. Importantly, inhibition of POFUT1 could synergize with anti-programmed death receptor 1 therapy by remodeling TME in the xenograft tumor mouse model. Clinically, POFUT1 high expression displayed a lower response rate and worse clinical outcome to ICB therapies.
CONCLUSIONS
Our findings demonstrate that POFUT1 functions as a novel regulator of tumor immune evasion and inhibition of POFUT1 may be a potential therapeutic strategy to enhance the efficacy of immune therapy in HCC.
Topics: Fucosyltransferases; Liver Neoplasms; Humans; Mice; Animals; B7-H1 Antigen; Immunotherapy; Carcinoma, Hepatocellular; Tumor Escape; Tumor Microenvironment; Immune Evasion; Cell Line, Tumor
PubMed: 38908854
DOI: 10.1136/jitc-2024-008917 -
Molecular Cell Jun 2024Protein folding is assisted by molecular chaperones that bind nascent polypeptides during mRNA translation. Several structurally distinct classes of chaperones promote...
Protein folding is assisted by molecular chaperones that bind nascent polypeptides during mRNA translation. Several structurally distinct classes of chaperones promote de novo folding, suggesting that their activities are coordinated at the ribosome. We used biochemical reconstitution and structural proteomics to explore the molecular basis for cotranslational chaperone action in bacteria. We found that chaperone binding is disfavored close to the ribosome, allowing folding to precede chaperone recruitment. Trigger factor recognizes compact folding intermediates that expose an extensive unfolded surface, and dictates DnaJ access to nascent chains. DnaJ uses a large surface to bind structurally diverse intermediates and recruits DnaK to sequence-diverse solvent-accessible sites. Neither Trigger factor, DnaJ, nor DnaK destabilize cotranslational folding intermediates. Instead, the chaperones collaborate to protect incipient structure in the nascent polypeptide well beyond the ribosome exit tunnel. Our findings show how the chaperone network selects and modulates cotranslational folding intermediates.
PubMed: 38908370
DOI: 10.1016/j.molcel.2024.06.002 -
Journal of Cheminformatics Jun 2024Generative models have demonstrated substantial promise in Natural Language Processing (NLP) and have found application in designing molecules, as seen in General...
Generative models have demonstrated substantial promise in Natural Language Processing (NLP) and have found application in designing molecules, as seen in General Pretrained Transformer (GPT) models. In our efforts to develop such a tool for exploring the organic chemical space in search of potentially electro-active compounds, we present Llamol, a single novel generative transformer model based on the Llama 2 architecture, which was trained on a 12.5M superset of organic compounds drawn from diverse public sources. To allow for a maximum flexibility in usage and robustness in view of potentially incomplete data, we introduce Stochastic Context Learning (SCL) as a new training procedure. We demonstrate that the resulting model adeptly handles single- and multi-conditional organic molecule generation with up to four conditions, yet more are possible. The model generates valid molecular structures in SMILES notation while flexibly incorporating three numerical and/or one token sequence into the generative process, just as requested. The generated compounds are very satisfactory in all scenarios tested. In detail, we showcase the model's capability to utilize token sequences for conditioning, either individually or in combination with numerical properties, making Llamol a potent tool for de novo molecule design, easily expandable with new properties. SCIENTIFIC CONTRIBUTION: We developed a novel generative transformer model, Llamol, based on the Llama 2 architecture that was trained on a diverse set of 12.5 M organic compounds. It introduces Stochastic Context Learning (SCL) as a new training procedure, allowing for flexible and robust generation of valid organic molecules with up to multiple conditions that can be combined in various ways, making it a potent tool for de novo molecular design.
PubMed: 38907298
DOI: 10.1186/s13321-024-00863-8 -
EMBO Reports Jun 2024Extracellular matrix (ECM) is a major component of the tumor environment, promoting the establishment of a pro-invasive behavior. Such environment is supported by both...
Extracellular matrix (ECM) is a major component of the tumor environment, promoting the establishment of a pro-invasive behavior. Such environment is supported by both tumor- and stromal-derived metabolites, particularly lactate. In prostate cancer (PCa), cancer-associated fibroblasts (CAFs) are major contributors of secreted lactate, able to impact on metabolic and transcriptional regulation in cancer cells. Here, we describe a mechanism by which CAF-secreted lactate promotes in PCa cells the expression of genes coding for the collagen family. Lactate-exploiting PCa cells rely on increased α-ketoglutarate (α-KG) which activates the α-KG-dependent collagen prolyl-4-hydroxylase (P4HA1) to support collagen hydroxylation. De novo synthetized collagen plays a signaling role by activating discoidin domain receptor 1 (DDR1), supporting stem-like and invasive features of PCa cells. Inhibition of lactate-induced collagen hydroxylation and DDR1 activation reduces the metastatic colonization of PCa cells. Overall, these results provide a new understanding of the link between collagen remodeling/signaling and the nutrient environment exploited by PCa.
PubMed: 38907027
DOI: 10.1038/s44319-024-00180-z -
European Journal of Pharmaceutical... Jun 2024The hepatitis B virus (HBV) capsid or core protein is a promising drug target currently being investigated for potential curative therapies for chronic HBV infection. In...
The hepatitis B virus (HBV) capsid or core protein is a promising drug target currently being investigated for potential curative therapies for chronic HBV infection. In this study, we performed extensive in vitro and in vivo characterization of a novel and potent HBV core protein assembly modulator (CpAM), CU15, for both anti-HBV activity and druggability properties. CU15 potently inhibited HBV DNA replication in in vitro HBV-infected HepG2.2.15 cells (EC of 8.6 nM), with a low serum shift. It was also effective in inhibiting HBV DNA and cccDNA formation in de novo HBV-infected primary human hepatocytes. Furthermore, CU15 was active across several HBV genotypes and across clinically relevant core protein variants. After oral administration to an in vivo HBV mouse model, CU15 significantly reduced plasma HBV DNA and RNA levels, at plasma exposure consistent with the estimated in vitro potency. In vitro, CU15 exhibited excellent passive permeability and relatively high metabolic stability in liver preparations across species (human > dog> rat). In vitro human liver microsomal studies suggest that the compound's major metabolic pathway is CYP3A-mediated oxidation. Consistent with the in vitro findings, CU15 is a compound with a low-to-moderate clearance and high oral bioavailability in rats and dogs. Based on the apparent in vitro-in vivo correlation observed, CU15 has the potential to exhibit low clearance and high oral bioavailability in humans. In addition, CU15 also showed low drug-drug interaction liability with an acceptable in vitro safety profile (IC > 10 µM).
PubMed: 38906232
DOI: 10.1016/j.ejps.2024.106834