A novel homozygous mutation in the human ALG12 gene results in an aberrant profile of oligomannose N-glycans in patient's serum.
Ontology highlight
ABSTRACT:
SUBMITTER: Ziburova J
PROVIDER: S-EPMC9291070 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:OBJECTIVE:Autosomal-recessive mutations in TBCK cause intellectual disability of variable severity. Although the physiological function of TBCK remains unclear, loss-of-function mutations are associated with inhibition of mechanistic target of rapamycin complex 1 (mTORC1) signaling. Given that mTORC1 signaling is known to regulate autophagy, we hypothesized that TBCK-encephalopathy patients with a neurodegenerative course have defects in autophagic-lysosomal dysfunction. METHODS:Children (n?=?8) of Puerto Rican (Boricua) descent affected with homozygous TBCK p.R126X mutations underwent extensive neurological phenotyping and neurophysiological studies. We quantified autophagosome content in TBCK-/- patient-derived fibroblasts by immunostaining and assayed autophagic markers by western assay. Free sialylated oligosaccharide profiles were assayed in patient's urine and fibroblasts. RESULTS:The neurological phenotype of children with TBCK p.R126X mutations, which we call TBCK-encephaloneuronopathy (TBCKE), include congenital hypotonia, progressive motor neuronopathy, leukoencephalopathy, and epilepsy. Systemic features include coarse facies, dyslipidemia, and osteoporosis. TBCK-/- fibroblasts in vitro exhibit increased numbers of LC3+ autophagosomes and increased autophagic flux by immunoblots. Free oligosaccharide profiles in fibroblasts and urine of TBCKE patients differ from control fibroblasts and are ameliorated by treatment with the mTORC1 activator leucine. INTERPRETATION:TBCKE is a clinically distinguishable syndrome with progressive central and peripheral nervous system dysfunction, consistently observed in patients with the p.R126X mutation. We provide evidence that inappropriate autophagy in the absence of cellular stressors may play a role in this disorder, and that mTORC1 activation may ameliorate the autophagic-lysosomal system dysfunction. Free oligosaccharide profiles could serve as a novel biomarker for this disorder as well as a tool to evaluate potential therapeutic interventions. Ann Neurol 2018;83:153-165.
| S-EPMC5876123 | biostudies-literature
Project description:The envelope spike of HIV is one of the most highly N-glycosylated structures found in nature. However, despite extensive research revealing essential functional roles in infection and immune evasion, the chemical structures of the glycans on the native viral envelope glycoprotein gp120--as opposed to recombinantly generated gp120--have not been described. Here, we report on the identity of the N-linked glycans from primary isolates of HIV-1 (clades A, B, and C) and from the simian immunodeficiency virus. MS analysis reveals a remarkably simple and highly conserved virus-specific glycan profile almost entirely devoid of medial Golgi-mediated processing. In stark contrast to recombinant gp120, which shows extensive exposure to cellular glycosylation enzymes (>70% complex type glycans), the native envelope shows barely detectable processing beyond the biosynthetic intermediate Man5GlcNAc2 (<2% complex type glycans). This oligomannose (Man5-9GlcNAc2) profile is conserved across primary isolates and geographically divergent clades but is not reflected in the current generation of gp120 antigens used for vaccine trials. In the context of vaccine design, we also note that Manalpha1-->2Man-terminating glycans (Man6-9GlcNAc2) of the type recognized by the broadly neutralizing anti-HIV antibody 2G12 are 3-fold more abundant on the native envelope than on the recombinant monomer and are also found on isolates not neutralized by 2G12. The Manalpha1-->2Man residues of gp120 therefore provide a vaccine target that is physically larger and antigenically more conserved than the 2G12 epitope itself. This study revises and extends our understanding of the glycan shield of HIV with implications for AIDS vaccine design.
| S-EPMC2922250 | biostudies-literature
Project description:The high mannose patch (HMP) of the HIV envelope protein (Env) is the structure most frequently targeted by broadly neutralizing antibodies; therefore, many researchers have attempted to use mimics of this region as a vaccine immunogen. In our previous efforts, vaccinating rabbits with evolved HMP mimic glycopeptides containing Man9 resulted in an overall antibody response targeting the glycan core and linker rather than the full glycan or Manα1→2Man tips of Man9 glycans. A possible reason could be processing of our immunogen by host serum mannosidases. We sought to test whether more prolonged dosing could increase the antibody response to intact glycans, possibly by increasing the availability of intact Man9 to germinal centers. Here, we describe a study investigating the impact of immunization regimen on antibody response by testing immunogen delivery through bolus, an exponential series of mini doses, or a continuously infusing mini-osmotic pump. Our results indicate that, with our glycopeptide immunogens, standard bolus immunization elicited the strongest HIV Env-binding antibody response, even though higher overall titers to the glycopeptide were elicited by the exponential and pump regimens. Antibody selectivity for intact glycan was, if anything, slightly better in the bolus-immunized animals.
| S-EPMC7091532 | biostudies-literature
Project description:Hemagglutinins (HAs) from human influenza viruses adapt to bind α2-6-linked sialosides, overcoming a receptor-defined species barrier distinct from the α2-3 specificity of avian virus progenitors. Additionally, human-adapted HAs gain glycosylation sites over time, although their biological function is poorly defined. Using quantitative glycomic analysis, we show that HAs from human pandemic viruses exhibit significant proportions of high-mannose type N-linked glycans throughout the head domain. By contrast, poorly adapted avian-origin HAs contain predominately complex-type glycans, which have greater structural diversity. Although oligomannose levels vary, they are present in all tested recombinant HAs and whole viruses and can be specifically targeted for universal detection. The positions of high-mannose glycosites on the HA of human H1N1 and H3N2 strains are conserved. Additionally, high-mannose-binding lectins possess a broad capacity to neutralize and prevent infection with contemporary H3N2 strains. These findings reveal the biological significance of HA glycosylation and therapeutic potential of targeting these structures.
| S-EPMC7158820 | biostudies-literature
Project description:Oligomannoses are evolutionarily the oldest class of N-glycans, where the arms of the common pentasaccharide unit, i.e., Manα(1-6)-[Manα(1-3)]-Manβ(1-4)-GlcNAcβ(1-4)-GlcNAcβ1-Asn, are functionalized exclusively with branched arrangements of mannose (Man) monosaccharide units. In mammalian species oligomannose N-glycans can have up to 9 Man; meanwhile structures can grow to over 200 units in yeast mannan. The highly dynamic nature, branching complexity, and 3D structure of oligomannoses have been recently highlighted for their roles in immune escape and infectivity of enveloped viruses, such as HIV-1 and SARS-CoV2. The architectural features that allow these N-glycans to perform their functions are yet unclear, due to their intrinsically disordered nature that hinders their structural characterization. In this work we will discuss the results of over 54 μs of cumulative sampling by molecular dynamics (MD) simulations of differently processed, free (not protein-linked) oligomannose N-glycans common in vertebrates. We then discuss the effects of a protein surface on their structural equilibria based on over 4 μs cumulative MD sampling of the fully glycosylated CD16a Fc γ receptor (FcγRIIIa), where the type of glycosylation is known to modulate its binding affinity for IgG1s, regulating the antibody-dependent cellular cytotoxicity (ADCC). Our results show that the protein's structural constraints shift the oligomannoses conformational ensemble to promote conformers that satisfy the steric requirements and hydrogen bonding networks demanded by the protein's surface landscape. More importantly, we find that the protein does not actively distort the N-glycans into structures not populated in the unlinked forms in solution. Ultimately, the highly populated conformations of the Man5 linked glycans support experimental evidence of high levels of hybrid complex forms at N45 and show a specific presentation of the arms at N162, which may be involved in mediating binding affinity to the IgG1 Fc.
| S-EPMC8279474 | biostudies-literature
Project description:[Table: see text] This study bridges a carbohydrate microarray discovery and a large-scale serological validation of anti-oligomannose antibodies as novel serum biomarkers of aggressive prostate cancer (PCa). Experimentally, a Man9-cluster-specific enzyme-linked immunosorbent assay was established to enable sensitive detection of anti-Man9 antibodies in human sera. A large-cohort of men with PCa or benign prostatic hyperplasia (BPH) whose sera were banked at Stanford University was characterized using this assay. Subjects included patients with 100% Gleason grade 3 cancer (n = 84), with Gleason grades 4 and/or 5 cancer (n = 204), and BPH controls (n = 135). Radical prostatectomy Gleason grades and biochemical (PSA) recurrence served as key parameters for serum biomarker evaluation. It was found that IgGMan9 and IgMMan9 were widely present in the sera of men with BPH, as well as those with cancer. However, these antibody reactivities were significantly increased in the subjects with the largest volumes of high grade cancer. Detection of serum IgGMan9 and IgMMan9 significantly predicted the clinical outcome of PCa post-radical prostatectomy. Given these results, we suggest that IgGMan9 and IgMMan9 are novel serum biomarkers for monitoring aggressive progression of PCa. The potential of oligomannosyl antigens as targets for PCa subtyping and targeted immunotherapy is yet to be explored.
| S-EPMC4139012 | biostudies-literature
Project description:The role of Fc glycans on clearance of IgG molecule has been examined by various groups in experiments where specific glycans have been enriched or the entire spectrum of glycans was studied after administration in pre-clinical or clinical pharmacokinetic (PK) studies. The overall conclusions from these studies are inconsistent, which may result from differences in antibody structure or experimental design. In the present study a well-characterized recombinant monoclonal IgG1 molecule (mAb-1) was analyzed from serum samples obtained from a human PK study. mAb-1 was recovered from serum using its ligand cross-linked to Sepharose beads. The overall purity and recovery of all isoforms were carefully evaluated using a variety of methods. Glycans were then enzymatically cleaved, labeled using 2-aminobenzamide and analyzed by normal phase high performance liquid chromatography. The assays for recovering mAb-1 from serum and subsequent glycan analysis were rigorously qualified at a lower limit of quantitation of 15 ?g/mL, thus permitting analysis to day 14 of the clinical PK study. Eight glycans were monitored and classified into two groups: (1) the oligomannose type structures (M5, M6 and M7) and (2) fucosylated biantennary oligosaccharides (FBO) structures (NGA2F, NA1F, NA2F, NA1F-GlcNAc and NGA2F-GlcNAc). We observed that the oligomannose species were cleared at a much faster rate (40%) than FBOs and conclude that high mannose species should be carefully monitored and controlled as they may affect PK of the therapeutic; they should thus be considered an important quality attribute. These observations were only possible through the application of rigorous analytical methods that we believe will need to be employed when comparing innovator and biosimilar molecules.
| S-EPMC3499345 | biostudies-literature
Project description:BackgroundWnt signaling pathway plays an important role in promoting ostergenesis. WNT1 mutations have been considered as a major cause of ostergenesis imperfect (OI). We identified an OI patient with pathogenic consanguineous-derived homozygous WNT1 missense mutation.MethodsWe designed and applied a panel of known 261 genes associated with hereditary bone diseases for targeted next-generation sequencing to examine clinically diagnosed OI patients. Detected mutations were confirmed by Sanger sequencing.ResultsThe female proband presented with severe OI with low bone density, multiple long bone fractures, short stature, and absence of dentinogenesis imperfect and brain malformation. She had congenital ptosis and exotropia with her left eye, and absence of blue sclera. The proband came from a consanguineous family and had a homozygous WNT1 missense mutation (c.677C>T, (p.S226L)). In addition, three other compound heterozygous mutations (c.1729C>T in FKBP10, c.1958A>C in FGFR3, c.760G>C in TRPV4) were also detected in her family members.ConclusionWe report the first identified case of consanguineous derived homozygous WNT1 mutation leading to severe osteogenesis imperfecta with congenital ptosis and exotropia.
| S-EPMC7434602 | biostudies-literature
Project description:Glycosylation is an essential form of post-translational modification that regulates intracellular and extracellular processes. Regrettably, conventional biochemical and genetic methods often fall short for the study of glycans, because their structures are often not precisely defined at the genetic level. To address this deficiency, chemists have developed technologies to perturb glycan biosynthesis, profile their presentation at the systems level, and perceive their spatial distribution. These tools have identified potential disease biomarkers and ways to monitor dynamic changes to the glycome in living organisms. Still, glycosylation remains the underexplored frontier of many biological systems. In this Account, we focus on research in our laboratory that seeks to transform the study of glycan function from a challenge to routine practice. In studies of proteins and nucleic acids, functional studies have often relied on genetic manipulations to perturb structure. Though not directly subject to mutation, we can determine glycan structure-function relationships by synthesizing defined glycoconjugates or by altering natural glycosylation pathways. Chemical syntheses of uniform glycoproteins and polymeric glycoprotein mimics have facilitated the study of individual glycoconjugates in the absence of glycan microheterogeneity. Alternatively, selective inhibition or activation of glycosyltransferases or glycosidases can define the biological roles of the corresponding glycans. Investigators have developed tools including small molecule inhibitors, decoy substrates, and engineered proteins to modify cellular glycans. Current approaches offer a precision approaching that of genetic control. Genomic and proteomic profiling form a basis for biological discovery. Glycans also present a rich matrix of information that adapts rapidly to changing environs. Glycomic and glycoproteomic analyses via microarrays and mass spectrometry are beginning to characterize alterations in glycans that correlate with disease. These approaches have already identified several cancer biomarkers. Metabolic labeling can identify recently synthesized glycans and thus directly track glycan dynamics. This approach can highlight changes in physiology or environment and may be more informative than steady-state analyses. Together, glycomic and metabolic labeling techniques provide a comprehensive description of glycosylation as a foundation for hypothesis generation. Direct visualization of proteins via the green fluorescent protein (GFP) and its congeners has revolutionized the field of protein dynamics. Similarly, the ability to perceive the spatial organization of glycans could transform our understanding of their role in development, infection, and disease progression. Fluorescent tagging in cultured cells and developing organisms has revealed important insights into the dynamics of these structures during growth and development. These results have highlighted the need for additional imaging probes.
| S-EPMC2697281 | biostudies-other
Project description:Immunologically, "self" carbohydrates protect the HIV-1 surface glycoprotein, gp120, from antibody recognition. However, one broadly neutralizing antibody, 2G12, neutralizes primary viral isolates by direct recognition of Manalpha1-->2Man motifs formed by the host-derived oligomannose glycans of the viral envelope. Immunogens, capable of eliciting antibodies of similar specificity to 2G12, are therefore candidates for HIV/AIDS vaccine development. In this context, it is known that the yeast mannan polysaccharides exhibit significant antigenic mimicry with the glycans of HIV-1. Here, we report that modulation of yeast polysaccharide biosynthesis directly controls the molecular specificity of cross-reactive antibodies to self oligomannose glycans. Saccharomyces cerevisiae mannans are typically terminated by alpha1-->3-linked mannoses that cap a Manalpha1-->2Man motif that otherwise closely resembles the part of the oligomannose epitope recognized by 2G12. Immunization with S. cerevisiae deficient for the alpha1-->3 mannosyltransferase gene (DeltaMnn1), but not with wild-type S. cerevisiae, reproducibly elicited antibodies to the self oligomannose glycans. Carbohydrate microarray analysis of DeltaMnn1 immune sera revealed fine carbohydrate specificity to Manalpha1-->2Man units, closely matching that of 2G12. These specificities were further corroborated by enzyme-linked immunosorbent assay with chemically defined glycoforms of gp120. These antibodies exhibited remarkable similarity in the carbohydrate specificity to 2G12 and displayed statistically significant, albeit extremely weak, neutralization of HIV-1 compared to control immune sera. These data confirm the Manalpha1-->2Man motif as the primary carbohydrate neutralization determinant of HIV-1 and show that the genetic modulation of microbial polysaccharides is a route towards immunogens capable of eliciting antibody responses to the glycans of HIV-1.
| S-EPMC2900896 | biostudies-literature