Dataset Information

Analysis of human sperm surface glycoproteins which can interact with galectin-1

ABSTRACT: Herein, we constructed a human lectin microarray with 60 human lectin and lectin-like proteins. All of the lectins and lectin-like proteins were purified from yeast, and most showed binding to human glycans. To demonstrate the applicability of the human lectin microarray, human sperm were probed on the microarray and strong bindings were observed for several lectins, including galectin-1, 7, 8, GalNAc-T6, and ERGIC-53 (LMAN1). These bindings were validated by flow cytometry and fluorescence immunostaining. Further, mass spectrometry analysis showed that galectin-1 binds several membrane proteins including HSP90. Finally, functional assays showed that binding of galectin-8 could significantly enhance the acrosome reaction within human sperms. To our knowledge, this is the first construction of a human lectin microarray, and we anticipate it will find wide use for a range of human or mammalian studies, alone or in combination with plant lectin microarrays.

INSTRUMENT(S): maXis

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Testis, Sperm

SUBMITTER: Yangyang Sun

LAB HEAD: Sheng-ce Tao

PROVIDER: PXD003785 | Pride | 2018-10-26

REPOSITORIES: Pride

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Publications

A Human Lectin Microarray for Sperm Surface Glycosylation Analysis.

Sun Yangyang Y Cheng Li L Gu Yihua Y Xin Aijie A Wu Bin B Zhou Shumin S Guo Shujuan S Liu Yin Y Diao Hua H Shi Huijuan H Wang Guangyu G Tao Sheng-Ce SC

Molecular & cellular proteomics : MCP 20160630 9

Glycosylation is one of the most abundant and functionally important protein post-translational modifications. As such, technology for efficient glycosylation analysis is in high demand. Lectin microarrays are a powerful tool for such investigations and have been successfully applied for a variety of glycobiological studies. However, most of the current lectin microarrays are primarily constructed from plant lectins, which are not well suited for studies of human glycosylation because of the ext ...[more]

PMID: 27364157

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Project description:Objectives: Hashimoto’s thyroiditis (HT) is one of the most common autoimmune disorders; however, its underlying pathological mechanisms remain unclear. Although aberrant glycosylation has been implicated in the N-glycome of immunoglobulin G (IgG), changes in serum proteins have not been comprehensively characterized. This study aimed to investigate the glycosylation profile of serum samples of HT patients that were depleted of highly abundant proteins by and propose the potential functions of glycoproteins for the further study of pathological mechanisms of HT . Methods: A lectin microarray containing 70 lectins was used to detect and analyze glycosylation of serum proteins using serum samples (N=27 HT; N=26 healthy control [HC]) depleted of abundant proteins. Significant differences in glycosylation status between HT patients and the HC group were verified by lectin blot analysis. A lectin-based pull-down assay combined with mass spectrometry was used to investigate potential glycoproteins combined with differentially present lectins, and an enzyme-linked immunosorbent assay (ELISA) was used to identify the expression of targeted glycoproteins in 131 patients with papillary thyroid carcinoma (PTC), 131 patients with benign thyroid nodules (BTN) patients, 130 patients with HT, and 128 HCs. Results: Compared with the HC group, the majority of the lectin binding signals in HT group were weakened, while the Vicia villosa agglutinin (VVA) binding signal was increased. The difference in VVA binding signals verified by lectin blotting was consistent with the results of the lectin microarray. A total of 113 potential VVA-binding glycoproteins were identified by mass spectrometry and classified by gene ontology (GO) and kyoto encyclopedia of genes and genomes (KEGG) analyses . Using ELISA, we confirmed that lactoferrin (LTF) and mannan-binding lectin-associated serine protease 1 (MASP-1) levels were elevated in the serum of patients with HT and PTC. Conclusions: Following depletion of abundant proteins, remaining serum proteins in HT patients exhibited lower glycosylation levels than those observed in HCs. An increased level of potential VVA-binding glycoproteins may play an important role in HT development. LTF and MASP-1 expression was significantly higher in the serum of HT and PTC patients, suggesting key roles played by glycosylation in pathological mechanisms underlying HT and PTC.

Project description:Human monocyte derived dendritic cells matured via galectin-1 or LPS. Experiment Overall Design: 4 conditions with 3 replicates of each include control or intreated immature MDDCS, galectin-1 treated MDDCs, LPS treated MDDCs, and vehicle control MDDCs. (each replicate is from a distinct DONOR). Dendritic cells (DCs) are potent mediators of the immune response, and can be activated by exogenous pathogen components. Galectin-1 is a member of the conserved beta-galactoside-binding lectin family that binds galactoside residues on cell surface glycoconjugates. Galectin-1 is known to play a role in immune regulation via action on multiple immune cells. However, its effects on human DCs are unknown. In this study, we show that galectin-1 induces a phenotypic and functional maturation in human monocyte-derived DCs (MDDCs) similar to but distinct from the activity of the exogenous pathogen stimuli, LPS. Immature human MDDCs exposed to galectin-1 up-regulated cell surface markers characteristic of DC maturation (CD40, CD83, CD86, and HLA-DR), secreted high levels of IL-6 and TNF-alpha, stimulated T cell proliferation, and showed reduced endocytic capacity, similar to LPS-matured MDDCs. However, unlike LPS-matured DCs, galectin-1-treated MDDCs did not produce the Th1-polarizing cytokine IL-12. Microarray analysis revealed that in addition to modulating many of the same DC maturation genes as LPS, galectin-1 also uniquely up-regulated a significant subset of genes related to cell migration through the extracellular matrix (ECM). Indeed, compared with LPS, galectin-1-treated human MDDCs exhibited significantly better chemotactic migration through Matrigel, an in vitro ECM model. Our findings show that galectin-1 is a novel endogenous activator of human MDDCs that up-regulates a significant subset of genes distinct from those regulated by a model exogenous stimulus (LPS). One unique effect of galectin-1 is to increase DC migration through the ECM, suggesting that galectin-1 may be an important component in initiating an immune response.

Dataset Information

Analysis of human sperm surface glycoproteins which can interact with galectin-1

Publications

A Human Lectin Microarray for Sperm Surface Glycosylation Analysis.

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