Cell Surface Heparan Sulfate Proteoglycans Research Articles

Sdc4 deletion perturbs intervertebral disc matrix homeostasis and promotes early osteopenia in the aging mouse spine

Syndecan 4 (SDC4), a cell surface heparan sulfate proteoglycan, is known to regulate matrix catabolism by nucleus pulposus cells in an inflammatory milieu. However, the role of SDC4 in the aging spine has never been explored. Here we analyzed the spinal phenotype of Sdc4 global knockout (KO) mice as a function of age. Micro-computed tomography showed that Sdc4 deletion severely reduced vertebral trabecular and cortical bone mass, and biomechanical properties of vertebrae were significantly altered in Sdc4 KO mice. These changes in vertebral bone were likely due to elevated osteoclastic activity. The histological assessment showed subtle phenotypic changes in the intervertebral disc. Imaging-Fourier transform-infrared analyses showed a reduced relative ratio of mature collagen crosslinks in young adult nucleus pulposus (NP) and annulus fibrosus (AF) of KO compared to wildtype discs. Additionally, relative chondroitin sulfate levels increased in the NP compartment of the KO mice. Transcriptomic analysis of NP tissue using CompBio, an AI-based tool showed biological themes associated with prominent dysregulation of heparan sulfate GAG degradation, mitochondria metabolism, autophagy, endoplasmic reticulum (ER)-associated misfolded protein processes and ER to Golgi protein processing. Overall, this study highlights the important role of SDC4 in fine-tuning vertebral bone homeostasis and extracellular matrix homeostasis in the mouse intervertebral disc.

Myosin 9 and N-glycans jointly regulate human papillomavirus entry

Persistent high-risk HPV infection is closely associated with cervical cancer development, and there is no drug targeting HPV on the market at present, so it is particularly important to understand the interaction mechanism between HPV and the host which may provide the novel strategies for treating HPV diseases. HPV can hijack cell surface heparan sulfate proteoglycans (HSPGs) as primary receptors. However, the secondary entry receptors for HPV remain elusive. We identify myosin-9 (NMHC-IIA) as a host factor that interacts with HPV L1 protein and mediates HPV internalization. Efficient HPV entry required myosin-9 redistribution to the cell surface regulated by HPV-hijacked MEK-MLCK signaling. Myosin-9 maldistribution by ML-7 or ML-9 significantly inhibited HPV pseudoviruses infection invitro and invivo. Meanwhile, N-glycans, especially the galactose chains, may act as the decoy receptors for HPV, which can block the interaction of HPV to myosin-9 and influence the way of HPV infection. Taken together, we identify myosin-9 as a novel functional entry receptor for high-risk HPV both invitro and invivo, and unravel the new roles of myosin-9 and N-glycans in HPV entry, which provides the possibilities for host targets of antiviral drugs.

On-demand chlorine dioxide solution enhances odontoblast differentiation through desulfation of cell surface heparan sulfate proteoglycan and subsequent activation of canonical Wnt signaling.

Heparan sulfate proteoglycans (HSPGs) surround the surface of odontoblasts, and their modification affects their affinity for Wnt ligands. This study proposes applying Matching Transformation System® (MA-T), a novel chlorinated oxidant, to enhance dentinogenesis. MA-T treatment in odontoblasts decreased sulfation of HSPG and upregulated the expression of dentin sialophosphoprotein (Dspp) and Dentin Matrix Protein 1 (Dmp1) via activation of canonical Wnt signaling in vitro. Ex vivo application of MA-T also enhanced dentin matrix formation in developing tooth explants. Reanalysis of a public single-cell RNA-seq dataset revealed significant Wnt activity in the odontoblast population, with enrichment for Wnt10a and Wnt6. Silencing assays showed that Wnt10a and Wnt6 were redundant in inducing Dspp and Dmp1 mRNA expression. These Wnt ligands' expression was upregulated by MA-T treatment, and TCF/LEF binding sites are present in their promoters. Furthermore, the Wnt inhibitors Notum and Dkk1 were enriched in odontoblasts, and their expression was also upregulated by MA-T treatment, together suggesting autonomous maintenance of Wnt signaling in odontoblasts. This study provides evidence that MA-T activates dentinogenesis by modifying HSPG and through subsequent activation of Wnt signaling.

Clinical/prognostic significance of Syndecan-1 expression in invasive breast carcinoma with distant metastasis and its correlation with tumor immunity

ObjectiveBreast Cancer (BC) is the most common malignant tumor for women in the world. 90% of BC-associated deaths are attributed to distant metastasis (DM). Therefore, there is an urgent need for a novel molecular target for the treatment of distant metastatic breast cancer (DMBC). Syndecan-1 (SDC-1) is a cell surface heparan sulfate proteoglycan (HSPG). This study aims to study the expression patterns of SDC-1 in invasive breast carcinoma (IBC) with DM and to analyze its relationship with different clinicopathologic features, stromal tumor infiltrating lymphocytes (sTILs) status and the clinical outcomes. MethodsA total of 50 DM breast cancer and 100 non-distant metastasis (non-DM) breast cancer patients in West China Hospital, Sichuan University from January 1, 2011 to December 31, 2011 were collected. Immunohistochemical (IHC) method was used to detect the expression of SDC-1, estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor-2 (HER2), and Ki-67 in 150 specimens of patients with IBC. STILs were used to evaluate immune cells in the stromal tissue within the tumor. Various clinicopathologic characteristics were retrospectively analyzed, and follow-up information were collected for prognosis analyses. The expression pattern difference of SDC-1 in the DM group and the non-DM group and its correlation with clinicopathologic characteristics of IBC were analyzed. ResultsCompared with the non-DM group, SDC-1 had higher cytoplasmic (90.0%) and stromal diffuse (70.0%) expressions and lower stromal peritumoral (18.0%) expression in the DM group. SDC-1 cytoplasmic expression was significantly associated with HER2-positive and high Ki-67 index in DM group, and with high histological grade and lymph node (LN) metastasis in non-DM group (P < 0.05). Compared with the non-DM group, the membranous expression of SDC-1 in the DM group was related to higher histological grade and T stage, higher frequency of LN involvement. Meanwhile, the expression pattern of SDC-1 in tumor stroma was associated with sTILs status (P < 0.05). The different combinations of SDC-1 staining patterns were correlated with clinicopathological features, biomarkers and sTILs status between DM group and non-DM group.There was no significant difference in overall survival between DMBC with different expression patterns of SDC-1. ConclusionThe cytoplasmic and stromal expressions of SDC-1 in the primary lesion of IBC are closely associated with DM, and the stromal expression of SDC-1 is correlated with tumor immune microenvironment. SDC-1 is expected to be a potential new marker for predicting the risk of DM in IBC.

SMOC-1 interacts with both BMP and glypican to regulate BMP signaling in C. elegans.

Secreted modular calcium-binding proteins (SMOCs) are conserved matricellular proteins found in organisms from Caenorhabditis elegans to humans. SMOC homologs characteristically contain 1 or 2 extracellular calcium-binding (EC) domain(s) and 1 or 2 thyroglobulin type-1 (TY) domain(s). SMOC proteins in Drosophila and Xenopus have been found to interact with cell surface heparan sulfate proteoglycans (HSPGs) to exert both positive and negative influences on the conserved bone morphogenetic protein (BMP) signaling pathway. In this study, we used a combination of biochemical, structural modeling, and molecular genetic approaches to dissect the functions of the sole SMOC protein in C. elegans. We showed that CeSMOC-1 binds to the heparin sulfate proteoglycan GPC3 homolog LON-2/glypican, as well as the mature domain of the BMP2/4 homolog DBL-1. Moreover, CeSMOC-1 can simultaneously bind LON-2/glypican and DBL-1/BMP. The interaction between CeSMOC-1 and LON-2/glypican is mediated specifically by the EC domain of CeSMOC-1, while the full interaction between CeSMOC-1 and DBL-1/BMP requires full-length CeSMOC-1. We provide both in vitro biochemical and in vivo functional evidence demonstrating that CeSMOC-1 functions both negatively in a LON-2/glypican-dependent manner and positively in a DBL-1/BMP-dependent manner to regulate BMP signaling. We further showed that in silico, Drosophila and vertebrate SMOC proteins can also bind to mature BMP dimers. Our work provides a mechanistic basis for how the evolutionarily conserved SMOC proteins regulate BMP signaling.

Revisiting the Syndecans: Master Signaling Regulators with Prognostic and Targetable Therapeutic Values in Breast Carcinoma

Syndecans (SDC1 to 4), a family of cell surface heparan sulfate proteoglycans, are frequently expressed in mammalian tissues. SDCs are aberrantly expressed either on tumor or stromal cells, influencing cancer initiation and progression through their pleiotropic role in different signaling pathways relevant to proliferation, cell-matrix adhesion, migration, invasion, metastasis, cancer stemness, and angiogenesis. In this review, we discuss the key roles of SDCs in the pathogenesis of breast cancer, the most common malignancy in females worldwide, focusing on the prognostic significance and molecular regulators of SDC expression and localization in either breast tumor tissue or its microenvironmental cells and the SDC-dependent epithelial-mesenchymal transition program. This review also highlights the molecular mechanisms underlying the roles of SDCs in regulating breast cancer cell behavior via modulation of nuclear hormone receptor signaling, microRNA expression, and exosome biogenesis and functions, as well as summarizing the potential of SDCs as promising candidate targets for therapeutic strategies against breast cancer.

Conditional ablation of heparan sulfate expression in stromal fibroblasts promotes tumor growth in vivo.

Heparan sulfate (HS) is a glycocalyx component present in the extracellular matrix and cell-surface HS proteoglycans (HSPGs). Although HSPGs are known to play functional roles in multiple aspects of tumor development and progression, the effect of HS expression in the tumor stroma on tumor growth in vivo remains unclear. We conditionally deleted Ext1, which encodes a glycosyltransferase essential for the biosynthesis of HS chains, using S100a4-Cre (S100a4-Cre; Ext1f/f) to investigate the role of HS in cancer-associated fibroblasts, which is the main component of the tumor microenvironment. Subcutaneous transplantation experiments with murine MC38 colon cancer and Pan02 pancreatic cancer cells demonstrated substantially larger subcutaneous tumors in S100a4-Cre; Ext1f/f mice. Additionally, the number of myofibroblasts observed in MC38 and Pan02 subcutaneous tumors of S100a4-Cre; Ext1f/f mice decreased. Furthermore, the number of intratumoral macrophages decreased in MC38 subcutaneous tumors in S100a4-Cre; Ext1f/f mice. Finally, the expression of matrix metalloproteinase-7 (MMP-7) markedly increased in Pan02 subcutaneous tumors in S100a4-Cre; Ext1f/f mice, suggesting that it may contribute to rapid growth. Therefore, our study demonstrates that the tumor microenvironment with HS-reduced fibroblasts provides a favorable environment for tumor growth by affecting the function and properties of cancer-associated fibroblasts, macrophages, and cancer cells.

Galectin-14 promotes hepatocellular carcinoma tumor growth via enhancing heparan sulfate proteoglycan modification.

Hepatocellular carcinoma (HCC) is a highly heterogeneous malignancy and lacks effective treatment. Bulk-sequencing of different gene transcripts by comparing HCC tissues and adjacent normal tissues provides some clues for investigating the mechanisms or identifying potential targets for tumor progression. However, genes that are exclusively expressed in a subpopulation of HCC may not be enriched or detected through such a screening. In the current study, we performed a single cell-clone-based screening and identified galectin-14 as an essential molecule in the regulation of tumor growth. The aberrant expression of galectin-14 was significantly associated with a poor overall survival of liver cancer patients with database analysis. Knocking down galectin-14 inhibited the proliferation of tumor growth, whereas overexpressing galectin-14 promoted tumor growth in vivo. Non-targeted metabolomics analysis indicated that knocking down galectin-14 decreased glycometabolism; specifically that glycoside synthesis was significantly changed. Further study found that galectin-14 promoted the expression of cell surface heparan sulfate proteoglycans (HSPGs) that functioned as co-receptors, thereby increasing the responsiveness of HCC cells to growth factors, such as epidermal growth factor and transforming growth factor-alpha. In conclusion, the current study identifies a novel HCC-specific molecule galectin-14, which increases the expression of cell surface HSPGs and the uptake of growth factors to promote HCC cell proliferation.

Small-molecule compound from AlphaScreen disrupts tau-glycan interface.

Tauopathies are neurodegenerative diseases characterized by intracellular abnormal tau deposits in the brain. Tau aggregates can propagate from one neuron to another in a prion-like manner, mediated by the interaction between tau and cell surface heparan sulfate proteoglycans. We developed an AlphaScreen assay, with His-tagged tau and biotinylated heparin, to represent the tau-HS interface to target the tau-glycan interface. Using our AlphaScreen assay, with a Z-factor of 0.65, we screened ∼300 compounds and discovered a small-molecule compound (herein referred to as A9), which can disrupt the tau-heparin interaction with micromolar efficacy. A9 also effectively inhibited heparin-induced tau aggregation in Thioflavin T fluorescence assays and attenuated tau internalization by H4 neuroglioma cells. These results strongly suggest that A9 can disrupt the tau-glycan interface in both in vitro molecular and cellular environments. We further determined that A9 interacts with heparin rather than tau and does so with micromolar binding affinity as shown by nuclear magnetic resonance and surface plasmon resonance experiments. A9 binds to heparin in a manner that blocks the sites where tau binds to heparin on the cell surface. These results demonstrate our AlphaScreen method as an effective method for targeting the tau-glycan interface in drug discovery and A9 as a promising lead compound for tauopathies, including Alzheimer's disease.

Role of syndecan-4 in breast cancer pathophysiology.

Expression of the cell surface heparan sulfate proteoglycan syndecan-4 is dysregulated in breast cancer, the most frequent malignancy in women. High expression of syndecan-4 correlates with a worse survival in the subgroup of estrogen receptor negative and estrogen/progesterone-receptor negative patients. Aberrant expression of syndecan-4 in breast cancer involves both transcriptional and posttranscriptional mechanisms, including estrogen- and growth factor-dependent regulation, mutations in GAPVD1, NUP153, PDE4DIP, and RREB1, as well as targeting by microRNAs. At the functional level, syndecan-4 plays an important role in various stages of breast cancer progression by interacting with ligands as diverse as plasma proteins, extracellular matrix proteins, growth factors, and surface receptors, as well as members of the integrin family. Mechanisms including integrin recycling, ectodomain shedding, and crosstalk with other syndecans expand the repertoire of syndecan-4 function. Through these interactions, syndecan-4 regulates cellular processes such as adhesion, migration, and invasion. Additional possible functions of syndecan-4 in cells of the microenvironment contribute to the complexity of its pathophysiology. Notably, syndecan-4 expression is modulated by drugs used in breast cancer treatment, such as trastuzumab and zoledronate. Overall, these findings mark syndecan-4 as a novel pathogenesis factor and promising target for therapeutic interventions in breast cancer.

Interaction of recombinant human lactoferrin and SARS-CoV-2 virus to heparin-protein conjugate

The advantages of the complex of recombinant human lactoferrin (rhLF) with europium ions have been used to establish quantitative parameters of specific interaction of rhLF with immobilized heparin-protein conjugate as a model of cell-surface heparan sulfate proteoglycans. Heparin coupled through terminal formyl by reductive amination to an inert protein was adsorbed through the protein part in the wells of a polystyrene microplate. The rhLF–Eu3+ complex obtained from native rhLF contains 0.8 mol of lanthanide ion per mol of protein (40 % saturation level). Equilibrium in the heterophase binding system is established within 1 min at room temperature, and the calculated association constant of the rhLF-heparin complex is 2.1 × 107 M–1. The reversible and saturable character of binding rhLF labeled by Eu3+ at the active site to heparin was confirmed by the transition of rhLF–Eu3+ into the liquid phase when a 1000-fold molar excess of unlabeled rhLF was added to the system. Based on the affinity of rhLF for glycosaminoglycan, a blocking effect of this protein on the binding of the SARS-CoV-2 virus to the immobilized heparin-protein conjugate that imitates proteoglycan on the host cell surface was revealed. Pretreatment of the adsorbed conjugate with a solution of rhLF (10 µg per well) reduces the specific binding of 100 ng of viral particles added to the well by approximately 80 %. The presented results allow one, in particular, to evaluate the integrity of the structure and activity of rhLF as a possible substance in food supplements and pharmaceuticals and may be useful in developing combined drugs for corona virus infection.

Efficient production of inhibitor-free foamy virus glycoprotein-containing retroviral vectors by proteoglycan-deficient packaging cells

Foamy viruses (FVs) or heterologous retroviruses pseudotyped with FV glycoprotein enable transduction of a great variety of target tissues of disparate species. Specific cellular entry receptors responsible for this exceptionally broad tropism await their identification. Though, ubiquitously expressed heparan sulfate proteoglycan (HS-PG) is known to serve as an attachment factor of FV envelope (Env)-containing virus particles, greatly enhancing target cell permissiveness. Production of high-titer, FV Env-containing retroviral vectors is strongly dependent on the use of cationic polymer-based transfection reagents like polyethyleneimine (PEI). We identified packaging cell-surface HS-PG expression to be responsible for this requirement. Efficient release of FV Env-containing virus particles necessitates neutralization of HS-PG binding sites by PEI. Remarkably, remnants of PEI in FV Env-containing vector supernatants, which are not easily removable, negatively impact target cell transduction, in particular those of myeloid and lymphoid origin. To overcome this limitation for production of FV Env-containing retrovirus supernatants, we generated 293T-based packaging cell lines devoid of HS-PG by genome engineering. This enabled, for the first, time production of inhibitor-free, high-titer FV Env-containing virus supernatants by non-cationic polymer-mediated transfection. Depending on the type of virus, produced titers were 2- to 10-fold higher compared with those obtained by PEI transfection.

TNF-α Inhibitors in Combination with MTX Reduce Circulating Levels of Heparan Sulfate/Heparin and Endothelial Dysfunction Biomarkers (sVCAM-1, MCP-1, MMP-9 and ADMA) in Women with Rheumatoid Arthritis.

Sulfated glycosaminoglycans (sGAGs) are likely to play an important role in the development and progression of rheumatoid arthritis (RA)-associated atherosclerosis. The present study investigated the effect of anti-tumor necrosis factor-α (anti-TNF-α) therapy in combination with methotrexate on plasma sGAG levels and serum markers of endothelial dysfunction. Among sGAG types, plasma chondroitin/dermatan sulfate (CS/DS) and heparan sulfate/heparin (HS/H) were characterized using electrophoretic fractionation. Serum levels of soluble vascular cell adhesion molecule-1 (sVCAM-1), monocyte chemoattractant protein-1 (MCP-1), matrix metalloproteinase-9 (MMP-9) and asymmetric dimethylarginine (ADMA) were measured by immunoassays. The measurements were carried out four times: at baseline and after 3, 9 and 15 months of anti-TNF-α therapy. All analyzed parameters, excluding ADMA, were significantly elevated in patients with RA before the implementation of biological therapy compared to healthy subjects. Performed anti-TNF-α treatment led to a successive decrease in HS/H levels toward normal values, without any effect on CS/DS levels in female RA patients. The treatment was also effective at lowering the serum levels of sVCAM-1, MCP-1, MMP-9 and ADMA. Moreover, a significant positive correlation was found between the circulating HS/H and the 28 joint disease activity score based on the erythrocyte sedimentation rate (DAS28-ESR, r = 0.408; p <0.05), MCP-1 (r = 0.398; p <0.05) and ADMA (r = 0.396; p <0.05) in patients before the first dose of TNF-α inhibitor. In conclusion, a beneficial effect of anti-TNF-α therapy on cell-surface heparan sulfate proteoglycans (HSPGs)/HS turnover and endothelial dysfunction was observed in this study. This was manifested by a decrease in blood HS/H levels and markers of endothelial activation, respectively. Moreover, the decrease in the concentration of HS/H in the blood of patients during treatment, progressing with the decline in disease activity, indicates that the plasma HS/H profile may be useful for monitoring the efficacy of anti-TNF-α treatment in patients with RA.

Inhibition of SARS-CoV-2 wild-type (Wuhan-Hu-1) and Delta (B.1.617.2) strains by marine sulfated glycans.

The Coronavirus disease pandemic has steered the global therapeutic research efforts toward the discovery of potential anti-severe acute respiratory syndrome coronavirus (SARS-CoV-2) molecules. The role of the viral spike glycoprotein (S-protein) has been clearly established in SARS-CoV-2 infection through its capacity to bind to the host cell surface heparan sulfate proteoglycan (HSPG) and angiotensin-converting enzyme-2. The antiviral strategies targeting these 2 virus receptors are currently under intense investigation. However, the rapid evolution of the SARS-CoV-2 genome has resulted in numerous mutations in the S-protein posing a significant challenge for the design of S-protein-targeted inhibitors. As an example, the 2 key mutations in the S-protein receptor-binding domain (RBD), L452R, and T478K in the SARS-CoV-2 Delta variant (B.1.617.2) confer tighter binding to the host epithelial cells. Marine sulfated glycans (MSGs) demonstrate excellent inhibitory activity against SARS-CoV-2 via competitive disruption of the S-protein RBD-HSPG interactions and thus have the potential to be developed into effective prophylactic and therapeutic molecules. In this study, 7 different MSGs were evaluated for their anti-SARS-CoV-2 activity in a virus entry assay utilizing a SARS-CoV-2 pseudovirus coated with S-protein of the wild-type (Wuhan-Hu-1) or the Delta (B.1.617.2) strain. Although all tested MSGs showed strong inhibitory activity against both strains, no correlations between MSG structural features and virus inhibition could be drawn. Nevertheless, the current study provides evidence for the maintenance of inhibitory activity of MSGs against evolving SARS-CoV-2 strains.

The dual fates of exogenous tau seeds: Lysosomal clearance versus cytoplasmic amplification

Tau assembly movement from the extracellular to intracellular space may underlie transcellular propagation of neurodegenerative tauopathies. This begins with tau binding to cell surface heparan sulfate proteoglycans, which triggers macropinocytosis. Pathological tau assemblies are proposed then to exit the vesicular compartment as “seeds” for replication in the cytoplasm. Tau uptake is highly efficient, but only ∼1 to 10% of cells that endocytose aggregates exhibit seeding. Consequently, we studied fluorescently tagged full-length (FL) tau fibrils added to native U2OS cells or “biosensor” cells expressing FL tau or repeat domain. FL tau fibrils bound tubulin. Seeds triggered its aggregation in multiple locations simultaneously in the cytoplasm, generally independent of visible exogenous aggregates. Most exogenous tau trafficked to the lysosome, but fluorescence imaging revealed a small percentage that steadily accumulated in the cytosol. Intracellular expression of Gal3-mRuby, which binds intravesicular galactosides and forms puncta upon vesicle rupture, revealed no evidence of vesicle damage following tau exposure, and most seeded cells had no evidence of endolysosome rupture. However, live-cell imaging indicated that cells with pre-existing Gal3-positive puncta were seeded at a slightly higher rate than the general population, suggesting a potential predisposing role for vesicle instability. Clearance of tau seeds occurred rapidly in both vesicular and cytosolic fractions. The lysosome/autophagy inhibitor bafilomycin inhibited vesicular clearance, whereas the proteasome inhibitor MG132 inhibited cytosolic clearance. Tau seeds that enter the cell thus have at least two fates: lysosomal clearance that degrades most tau, and entry into the cytosol, where seeds amplify, and are cleared by the proteasome.

Graph Convolutional Network-Based Screening Strategy for Rapid Identification of SARS-CoV-2 Cell-Entry Inhibitors.

The cell entry of SARS-CoV-2 has emerged as an attractive drug development target. We previously reported that the entry of SARS-CoV-2 depends on the cell surface heparan sulfate proteoglycan (HSPG) and the cortex actin, which can be targeted by therapeutic agents identified by conventional drug repurposing screens. However, this drug identification strategy requires laborious library screening, which is time consuming, and often limited number of compounds can be screened. As an alternative approach, we developed and trained a graph convolutional network (GCN)-based classification model using information extracted from experimentally identified HSPG and actin inhibitors. This method allowed us to virtually screen 170,000 compounds, resulting in ∼2000 potential hits. A hit confirmation assay with the uptake of a fluorescently labeled HSPG cargo further shortlisted 256 active compounds. Among them, 16 compounds had modest to strong inhibitory activities against the entry of SARS-CoV-2 pseudotyped particles into Vero E6 cells. These results establish a GCN-based virtual screen workflow for rapid identification of new small molecule inhibitors against validated drug targets.

Glypican-1 drives unconventional secretion of fibroblast growth factor 2.

Fibroblast growth factor 2 (FGF2) is a tumor cell survival factor that is transported into the extracellular space by an unconventional secretory mechanism. Cell surface heparan sulfate proteoglycans are known to play an essential role in this process. Unexpectedly, we found that among the diverse subclasses consisting of syndecans, perlecans, glypicans, and others, Glypican-1 (GPC1) is the principle and rate-limiting factor that drives unconventional secretion of FGF2. By contrast, we demonstrate GPC1 to be dispensable for FGF2 signaling into cells. We provide first insights into the structural basis for GPC1-dependent FGF2 secretion, identifying disaccharides with N-linked sulfate groups to be enriched in the heparan sulfate chains of GPC1 to which FGF2 binds with high affinity. Our findings have broad implications for the role of GPC1 as a key molecule in tumor progression.

Coreceptor functions of cell surface heparan sulfate proteoglycans.

Receptor-ligand interactions play an important role in many biological processes by triggering specific cellular responses. These interactions are frequently regulated by coreceptors that facilitate, alter, or inhibit signaling. Coreceptors work in parallel with other specific and accessory molecules to coordinate receptor-ligand interactions. Cell surface heparan sulfate proteoglycans (HSPGs) function as unique coreceptors because they can bind to many ligands and receptors through their HS and core protein motifs. Cell surface HSPGs are typically expressed in abundance of the signaling receptors and, thus, are capable of mediating the initial binding of ligands to the cell surface. HSPG coreceptors do not possess kinase domains or intrinsic enzyme activities and, for the most part, binding to cell surface HSPGs does not directly stimulate intracellular signaling. Because of these features, cell surface HSPGs primarily function as coreceptors for many receptor-ligand interactions. Given that cell surface HSPGs are widely conserved, they likely serve fundamental functions to preserve basic physiological processes. Indeed, cell surface HSPGs can support specific cellular interactions with growth factors, morphogens, chemokines, extracellular matrix (ECM) components, and microbial pathogens and their secreted virulence factors. Through these interactions, HSPG coreceptors regulate cell adhesion, proliferation, migration, and differentiation, and impact the onset, progression, and outcome of pathophysiological processes, such as development, tissue repair, inflammation, infection, and tumorigenesis. This review seeks to provide an overview of the various mechanisms of how cell surface HSPGs function as coreceptors.

Hedgehog pathway modulation by glypican 3-conjugated heparan sulfate.

Glypicans are a family of cell surface heparan sulfate proteoglycans that play critical roles in multiple cell signaling pathways. Glypicans consist of a globular core, an unstructured stalk modified with sulfated glycosaminoglycan chains, and a glycosylphosphatidylinositol anchor. Though these structural features are conserved, their individual contribution to glypican function remains obscure. Here, we investigate how glypican 3 (GPC3), which is mutated in Simpson-Golabi-Behmel tissue overgrowth syndrome, regulates Hedgehog signaling. We find that GPC3 is necessary for the Hedgehog response, surprisingly controlling a downstream signal transduction step. Purified GPC3 ectodomain rescues signaling when artificially recruited to the surface of GPC3-deficient cells but has dominant-negative activity when unattached. Strikingly, the purified stalk, modified with heparan sulfate but not chondroitin sulfate, is necessary and sufficient for activity. Our results demonstrate a novel function for GPC3-associated heparan sulfate and provide a framework for the functional dissection of glycosaminoglycans by in vivo biochemical complementation. This article has an associated First Person interview with the first author of the paper.

Regulation of Wnt distribution and function by Drosophila glypicans.

The extracellular distribution of secreted Wnt proteins is crucial for their ability to induce a response in target cells at short and long ranges to ensure proper development. Wnt proteins are evolutionarily conserved ligands that are lipid-modified, and their hydrophobic nature interferes with their solubility in the hydrophilic extracellular environment. This raises the question of how Wnt proteins spread extracellularly despite their lipid modifications, which are essential for both their secretion and function. Seminal studies on Drosophila Wingless (Wg), a prototypical Wnt, have discovered multiple mechanisms by which Wnt proteins spread. A central theme emerges from these studies: the Wnt lipid moiety is shielded from the aqueous environment, allowing the ligands to spread and remain viable for signaling. Wnt distribution in vivo is primarily facilitated by glypicans, which are cell-surface heparan sulfate proteoglycans, and recent studies have further provided mechanistic insight into how glypicans facilitate Wnt distribution. In this Review, we discuss the many diverse mechanisms of Wnt distribution, with a particular focus on glypican-mediated mechanisms.