Biology subjects

Costello, C. E.

Publications and source records attributed to Costello, C. E..

7 recordsLinked to original sources

bioRxiv2026

The Human Contraceptive Antibody Targets A Semenogelin Galectin-3 N-Linked Glycan Complex on the Sperm Surface

The human contraception antibody (HCA) is a non-hormonal topical contraceptive entering Phase II clinical trials that functions by agglutinating and immobilizing sperm. To date, the HCA epitope has been described as an N-linked polylactosamine glycan motif on CD52, but this assignment has been inferred only by colocalization with anti-CD52 and S19 antibodies in western blots. Here, we report that HCA may instead bind semenogelin-galectin-3 complexes associated with the sperm surface. Semenogelins are gel forming, highly glycosylated proteins, with molecular weights of 50-70 kDa that are cleaved into smaller segments that adhere to sperm during semen liquefaction. In this study, we isolated the HCA antigen from seminal plasma and sperm using an HCA affinity column and identified semenogelin by mass spectrometry. Western blot analysis of seminal plasma showed that HCA binds to a large smear of proteins with a higher apparent molecular mass than CD52, and that these may correspond to semenogelin sequences. Flow cytometric analysis of enzymatically deglycosylated sperm demonstrated that HCA binding requires sperm surface N-linked glycans. However, despite this apparent glycan dependence, HCA failed to bind any glycans represented on a comprehensive glycan microarray. In addition to semenogelin, Galectin-3 was identified in the HCA purified sperm lysates, supporting a model in which HCA recognizes semenogelin-derived glycopeptides that are tethered to the sperm surface via a galectin mediated interaction with an N-linked glycan. We found that O-linked glycan modifications on the semenogelin proteins contain LacNAc motifs that can bind to galectins. Finally, we found that depletion of galectins bound to the sperm surface by competitive inhibitors reduced HCA-mediated agglutination, and this could be rescued by addition of recombinant galectin-3/semenogelin-2 complexes to sperm cultures. Collectively, our findings suggest that HCA's binding site involves a semenogelin-galectin-3 complex anchored to the sperm surface via an N-linked glycan.

immunology↗

bioRxiv2026

Glycoconjugate diversification in Campylobacter concisus is determined by two glycosyltransferases

Bacterial glycoconjugates are structurally diverse, with enormous variation in sugar identity, modifications and linkages. Glycoconjugates play key roles in numerous cell functions, acting as the primary interface with the environment. Asparagine (N)-linked glycosylation has been extensively studied in the pathogenic Campylobacter genus, due to the availability of numerous genome sequences and the highly conserved pathway logic, despite the final N-linked glycan product diversity. We recently reported on a partitioning of N-linked glycan structures between the Campylobacter species, focused on the inclusion of a C6-carboxyl-sugar in the third position of the growing glycan in Campylobacter concisus. However, at the time, the final glycan was not fully defined. Here, we identify the final glycan product in C. concisus, demonstrating surprising substrate promiscuity of the GT-A enzyme, PglI, which adds the penultimate sugar, and uncover a previously uncharacterized enzyme (GT-25) that unexpectedly adds the final sugar to complete the heptasaccharide product. Through a detailed study of these two C. concisus pathway enzymes, the intermediate and final glycans were defined, with determination of linkage positions of major and minor isomeric products following each glycan addition, through high-resolution electronic excitation dissociation tandem mass spectrometry. These findings on the Group II C. concisus N-linked glycan highlight the diversification of the glycan and the utilization, at the non-reducing end, of GlcNAc over GalNAc, which is dominant in the Group I species.

biochemistry↗

bioRxiv2025

Robust Serum Proteomic Signatures of APOE2

We previously identified a signature of 16 serum proteins that highlighted a role of the e2 allele of APOE in lipid regulation via apolipoprotein B (APOB) and apolipoprotein E (APOE), and in inflammation. The serum proteins were profiled using the aptamer-based Somalogic technology. Here, we validate and expand the serum protein signature of APOE using a combination of mass-spectrometry, ELISA, Luminex, antibody-based Olink proteomics, and blood transcriptomics. We replicate the association between APOB and the e2 allele of APOE, we correct the pattern of association between APOE genotypes and serum level of APOE, and we detect new associations between APOE genotypes and the complex of apolipoproteins APOC1, APOC4, APOC2, APOC3, APOE, APOF and APOL1. In addition, we discover 13 new proteins that correlate with APOE genotypes. This extended signature includes granule proteins CAMP, CTSG, DEFA3, and MPO secreted from neutrophils and points to olfactomedin 4 (OLFM4) as a new target for the prevention of Alzheimers disease.

genetics↗

bioRxiv2025

Sialidases derived from Gardnerella vaginalis remodel the sperm glycocalyx and impair sperm function

Bacterial vaginosis (BV), a dysbiosis of the vaginal microbiome, affects approximately 30 percent of women worldwide (up to 50% in some regions) and is associated with several adverse health outcomes including preterm birth and increased incidence of sexually transmitted infections (STIs). BV-associated bacteria such as Gardnerella vaginalis and Prevotella timonensis damage the vaginal mucosa through the activity of sialidase enzymes that remodel the epithelial glycocalyx and degrade mucin glycoproteins. This damage may contribute to adverse health outcomes. However, whether BV-associated glycolytic enzymes also damage sperm has not yet been determined. Here, we show that sialidase-mediated glycocalyx remodeling of human sperm increases sperm susceptibility to damage and adversely affects their function in vitro. Specifically, we report that sperm motility was not adversely affected by sialidase treatment, but desialylated human sperm demonstrate increased susceptibility to agglutination and complement-mediated cytotoxicity as well as impaired transit through cervical mucus. Our results demonstrate mechanisms by which BV-associated sialidases affect sperm survival and function and potentially contribute to adverse reproductive outcomes such as infertility.

immunology↗

bioRxiv2024

Cross-platform proteomics signatures of extreme old age

In previous work we used a Somalogic platform targeting approximately 5000 proteins to generate a serum protein signature of centenarians that we validated in independent studies that used the same technology. We set here to validate and possibly expand the results by profiling the serum proteome of a subset of individuals included in the original study using liquid chromatography tandem mass spectrometry (LC-MS/MS). Following pre-processing, the LC-MS/MS data provided quantification of 398 proteins, with only 266 proteins shared by both platforms. At 1% FDR statistical significance threshold, the analysis of LC-MS/MS data detected 44 proteins associated with extreme old age, including 23 of the original analysis. To identify proteins for which associations between expression and extreme-old age were conserved across platforms, we performed inter-study conservation testing of the 266 proteins quantified by both platforms using a method that accounts for the correlation between the results. From these tests, a total of 80 proteins reached 5% FDR statistical significance, and 26 of these proteins had concordant pattern of gene expression in whole blood. This signature of 80 proteins points to blood coagulation, IGF signaling, extracellular matrix (ECM) organization, and complement cascade as important pathways whose protein level changes provide evidence for age-related adjustments that distinguish centenarians from younger individuals.

bioinformatics↗

bioRxiv2018

The most abundant cyst wall proteins of Acanthamoeba castellanii are cellulose-binding lectins from three gene families that localize to distinct structures in cyst walls

Acanthamoeba castellanii, cause of keratitis and blindness, is an emerging pathogen because of its association with contact lens use. The cyst wall contributes to pathogenesis as cysts are resistant to sterilizing reagents in lens solutions and to antibiotics applied to the eye. Here we used structured illumination microscopy (SIM) and probes for glycopolymers to show that purified cyst walls of A. castellanii retain endocyst and ectocyst layers and conical structures (ostioles) that connect them. Mass spectrometry showed candidate cyst wall proteins (CWPs) are dominated by three families of lectins (named here Luke, Leo, and Jonah), because each binds to microcrystalline cellulose +/- chitin. Luke lectins contain two or three carbohydrate-binding modules (CBM49), which were first identified in a tomato cellulase. Leo lectins have two unique domains with eight Cys residues each (8-Cys) +/- a Thr-, Lys-, and His-rich spacer. Jonah lectins contain one or three choice-of-anchor A (CAA) domains previously of unknown function. Representative members of each family were tagged with green fluorescent protein (GFP) and expressed under their own promoters in transfected parasites. A representative Jonah lectin with one CAA domain is made early during encystation and localizes to the ectocyst layer. In contrast, Leo and Luke lectins are made later and localize to the endocyst layer and ostioles. Probes for CWPs (anti-GFP antibodies) and for glycopolymers (maltose-binding protein-fusions with CWPs) suggest Jonah lectin and the glycopolymers to which it binds are accessible in the ectocyst layer, while Luke and Leo lectins and their epitopes are mostly inaccessible in the ectocyst layer and ostioles. In summary, the most abundant A. castellanii CWPs are three sets of lectins, which have conserved (CBM49s of Luke), newly characterized (CAA of Jonah), or unique carbohydrate-binding modules (8-Cys of Jonah).\n\nAuthor summaryFifty years ago, the cyst wall of Acanthamoeba castellanii was shown to contain cellulose and have an ectocyst layer, an endocyst layer, and conical ostioles that attach them. The goals here were to identify abundant cyst wall proteins (CWPs) and begin to determine how the wall is assembled. We used wheat germ agglutinin to show cyst walls also contain chitin fibrils. When trophozoites are starved of nutrients, they become immotile and make CWPs and glycopolymers in dozens of small vesicles. The primordial cyst wall is composed of a single, thin layer containing cellulose, chitin, and an abundant CWP we called Jonah. The primordial wall also has small, flat ostioles that contain another abundant CWP we called Luke. Jonah (the best candidate for diagnostic antibodies) is accessible in the ectocyst layer of mature cyst walls, while Luke and a third abundant CWP we termed Leo are present but mostly inaccessible in the endocyst layer and ostioles. While A. castellanii cyst walls contain cellulose (like plants) and chitin (like fungi), the glycopolymers are made in vesicles rather than at the plasma membrane, and the CWPs (Luke, Leo, and Jonah lectins) are unique to the protist.

microbiology↗

bioRxiv2018

O-fucosylation of thrombospondin-like repeats is required for processing of MIC2 and for efficient host cell invasion by Toxoplasma gondii tachyzoites

Toxoplasma gondii is an intracellular parasite that causes disseminated infections which can lead to neurological damage in fetuses and immunocompromised individuals. Microneme protein 2 (MIC2)2, a member of the thrombospondin-related anonymous protein (TRAP) family, is a secreted protein important for motility, host cell attachment, invasion, and egress. MIC2 contains six thrombospondin type I repeats (TSRs) that are modified by C-mannose and O-fucose in Plasmodium spp. and mammals.\n\nHere we used mass spectrometry to show that the four TSRs in T. gondii MIC2 with protein O-fucosyltransferase 2 (POFUT2) acceptor sites are modified by a dHexHex disaccharide, while Trp residues within three TSRs are also modified with C-mannose. Disruption of genes encoding either pofut2 or nucleotide sugar transporter 2 (nst2), the putative GDP-fucose transporter, results in loss of MIC2 O-fucosylation, as detected by an antibody against the GlcFuc disaccharide, and markedly reduced cellular levels of MIC2. Furthermore, in 10-15% of the{Delta} pofut2 or{Delta} nst2 vacuoles, MIC2 accumulates earlier in the secretory pathway rather than localizing to micronemes. Dissemination of tachyzoites in human foreskin fibroblasts is reduced in these knockouts, which both show defects in attachment to and invasion of host cells comparable to the phenotype observed in the Bmic2.\n\nThese results, which show O-fucosylation of TSRs is required for efficient processing of MIC2 and for normal parasite invasion, are consistent with the recent demonstration that P. falciparum {Delta}pofut2 has decreased virulence and support a conserved role for this glycosylation pathway in quality control of TSR-containing proteins in eukaryotes.

microbiology↗