RESUMEN
Our understanding of muscle glycosylation to date has derived from studies in mouse models and a limited number of human lectin histochemistry studies. As various therapeutic approaches aimed at treating patients with muscular dystrophies are being translated from rodent models to human, it is critical to better understand human muscle glycosylation and relevant disease-specific differences between healthy and dystrophic muscle. Here, we report the first quantitative characterization of human muscle glycosylation, and identify differentiation- and disease-specific differences in human muscle glycosylation. Utilizing a panel of 13 lectins with varying glycan specificities, we surveyed lectin binding to primary and immortalized myoblasts and myotubes from healthy and dystrophic sources. Following differentiation of primary and immortalized healthy human muscle cells, we observed increased binding of Narcissus pseudonarcissus agglutinin (NPA), PNA, MAA-II and WFA to myotubes compared to myoblasts. Following differentiation of immortalized healthy and dystrophic human muscle cells, we observed disease-specific differences in binding of NPA, Jac and Tricosanthes japonica agglutinin-I (TJA-I) to differentiated myotubes. We also observed differentiation- and disease-specific differences in binding of NPA, Jac, PNA, TJA-I and WFA to glycoprotein receptors in muscle cells. Additionally, Jac, PNA and WFA precipitated functionally glycosylated α-DG, that bound laminin, while NPA and TJA-I did not. Lectin histochemistry of healthy and dystrophic human muscle sections identified disease-specific differences in binding of O-glycan and sialic acid-specific lectins between healthy and dystrophic muscle. These results indicate that specific and discrete changes in glycosylation occur following differentiation, and identify specific lectins as potential biomarkers sensitive to changes in healthy human muscle glycosylation.
Asunto(s)
Glicoproteínas/metabolismo , Proteínas Musculares/metabolismo , Distrofias Musculares/metabolismo , Mioblastos Esqueléticos/metabolismo , Narcissus/química , Lectinas de Plantas/farmacología , Línea Celular Transformada , Glicoproteínas/química , Humanos , Proteínas Musculares/química , Distrofias Musculares/patología , Mioblastos Esqueléticos/química , Mioblastos Esqueléticos/patología , Lectinas de Plantas/químicaRESUMEN
Changes in the T cell surface redox environment regulate critical cell functions, such as cell migration, viral entry and cytokine production. Cell surface protein disulfide isomerase (PDI) contributes to the regulation of T cell surface redox status. Cell surface PDI can be released into the extracellular milieu or can be internalized by T cells. We have found that galectin-9, a soluble lectin expressed by T cells, endothelial cells and dendritic cells, binds to and retains PDI on the cell surface. While endogenous galectin-9 is not required for basal cell surface PDI expression, exogenous galectin-9 mediated retention of cell surface PDI shifted the disulfide/thiol equilibrium on the T cell surface. O-glycans on PDI are required for galectin-9 binding, and PDI recognition appears to be specific for galectin-9, as galectin-1 and galectin-3 do not bind PDI. Galectin-9 is widely expressed by immune and endothelial cells in inflamed tissues, suggesting that T cells would be exposed to abundant galectin-9, in cis and in trans, in infectious or autoimmune conditions.
Asunto(s)
Membrana Celular/metabolismo , Galectina 1/metabolismo , Galectinas/metabolismo , Proteína Disulfuro Isomerasas/metabolismo , Linfocitos T/metabolismo , Sitios de Unión , Línea Celular , Membrana Celular/química , Membrana Celular/efectos de los fármacos , Membrana Celular/inmunología , Clonación Molecular , Escherichia coli/genética , Escherichia coli/metabolismo , Galectina 1/genética , Galectina 3/genética , Galectina 3/metabolismo , Galectinas/antagonistas & inhibidores , Galectinas/genética , Galectinas/farmacología , Expresión Génica , Regulación de la Expresión Génica , Glicosilación , Humanos , Modelos Moleculares , Oxidación-Reducción , Polisacáridos/química , Polisacáridos/metabolismo , Unión Proteica , Proteína Disulfuro Isomerasas/química , Proteína Disulfuro Isomerasas/genética , Proteína Disulfuro Isomerasas/inmunología , Transporte de Proteínas , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Proteínas Recombinantes de Fusión/farmacología , Transducción de Señal , Linfocitos T/química , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunologíaRESUMEN
The biological impact of glycans is as diverse and complex as the impact of proteins on biology. Familiar roles include those as a protein folding checkpoint in the endoplasmic reticulum and as a modulator of the serum half-life of secreted glycoproteins, but it has become clear over the last several decades that glycans are key signaling moieties, participate in cell-cell interactions and modulate the function of individual proteins, to name but a few examples. In the immune system, the majority of microbial "patterns" are glycans or glycoconjugates, while virtually all cell surface receptors are glycoproteins, and antibody glycosylation critically influences antibody function. In order to provide a simple contextual framework to understand the myriad roles, glycans play in immunity, we propose that glycan effects are considered direct or indirect, depending on their direct participation or their indirect effects on other components in a given biological process or pathway. Here, we present the published evidence that supports this framework, which ultimately leads to the conclusion that we should learn to embrace the complexity inherent to the glycome and its potential as a largely uncharted but target rich area of new therapeutic investigation.
Asunto(s)
Inmunidad , Polisacáridos/metabolismo , Procesamiento Proteico-Postraduccional , Animales , Glicosilación , HumanosRESUMEN
The neuromuscular junction (NMJ) is enriched with glycoproteins modified with N-acetylgalactosamine (GalNAc) residues, and four nominally GalNAc-specific plant lectins have historically been used to identify the NMJ and the utrophin-glycoprotein complex. However, little is known about the specific glycan epitopes on skeletal muscle that are bound by these lectins, the glycoproteins that bear these epitopes or how creation of these glycan epitopes is regulated. Here, we profile changes in cell surface glycosylation during muscle cell differentiation and identify distinct differences in the binding preferences of GalNAc-specific lectins, Wisteria floribunda agglutinin (WFA), Vicia villosa agglutinin (VVA), soybean agglutinin (SBA) and Dolichos biflorus agglutinin (DBA). While we find that all four GalNAc binding lectins specifically label the NMJ, each of the four lectins binds distinct sets of muscle glycoproteins; furthermore, none of the major adhesion complexes are required for binding of any of the four GalNAc-specific lectins. Analysis of glycosylation-related transcripts identified target glycosyltransferases and glycosidases that could potentially create GalNAc-containing epitopes; reducing expression of these transcripts by siRNA highlighted differences in lectin binding specificities. In addition, we found that complex N-glycans are required for binding of WFA and SBA to murine C2C12 myotubes and for WFA binding to wild-type skeletal muscle, but not for binding of VVA or DBA. These results demonstrate that muscle cell surface glycosylation is finely regulated during muscle differentiation in a domain- and acceptor-substrate-specific manner, suggesting that temporal- and site-specific glycosylation are important for skeletal muscle cell function.
Asunto(s)
Epítopos/inmunología , Glicocálix/metabolismo , Músculo Esquelético/metabolismo , Polisacáridos/inmunología , Animales , Diferenciación Celular , Línea Celular , Pollos , Glicocálix/química , Glicocálix/inmunología , Ratones , Ratones Noqueados , Músculo Esquelético/química , Músculo Esquelético/citología , Músculo Esquelético/inmunologíaRESUMEN
Galectins are a family of mammalian carbohydrate-binding proteins expressed by many cell types. Galectins can function intracellularly and can also be secreted to bind to cell surface glycoconjugate counterreceptors. Some galectins are made by immune cells, whereas other galectins are secreted by different cell types, such as endothelial or epithelial cells, and bind to immune cells to regulate immune responses. Galectin binding to a single glycan ligand is a low-affinity interaction, but the multivalency of galectins and the glycan ligands presented on cell surface glycoproteins results in high-avidity binding that can reversibly scaffold or cluster these glycoproteins. Galectin binding to a specific glycoprotein counterreceptor is regulated in part by the repertoire of glycosyltransferase enzymes (which make the glycan ligands) expressed by that cell, and the effect of galectin binding results from clustering or retention of specific glycoprotein counterreceptors bearing these specific ligands.
Asunto(s)
Galectinas/metabolismo , Glicosiltransferasas/metabolismo , Inmunidad , Animales , Carbohidratos/inmunología , Citoesqueleto , Galectinas/inmunología , Glicoproteínas/metabolismo , Humanos , Unión Proteica , Agregación de ReceptoresRESUMEN
O-linked glycosylation is a ubiquitous protein modification in organisms belonging to several kingdoms. Both microbial and host protein glycans are used by many pathogens for host invasion and immune evasion, yet little is known about the roles of O-glycans in viral pathogenesis. Reportedly, there is no single function attributed to O-glycans for the significant paramyxovirus family. The paramyxovirus family includes many important pathogens, such as measles, mumps, parainfluenza, metapneumo- and the deadly Henipaviruses Nipah (NiV) and Hendra (HeV) viruses. Paramyxoviral cell entry requires the coordinated actions of two viral membrane glycoproteins: the attachment (HN/H/G) and fusion (F) glycoproteins. O-glycan sites in HeV G were recently identified, facilitating use of the attachment protein of this deadly paramyxovirus as a model to study O-glycan functions. We mutated the identified HeV G O-glycosylation sites and found mutants with altered cell-cell fusion, G conformation, G/F association, viral entry in a pseudotyped viral system, and, quite unexpectedly, pseudotyped viral F protein incorporation and processing phenotypes. These are all important functions of viral glycoproteins. These phenotypes were broadly conserved for equivalent NiV mutants. Thus our results identify multiple novel and pathologically important functions of paramyxoviral O-glycans, paving the way to study O-glycan functions in other paramyxoviruses and enveloped viruses.
Asunto(s)
Virus Nipah/fisiología , Polisacáridos/metabolismo , Proteínas Virales de Fusión/metabolismo , Internalización del Virus , Línea Celular , Glicosilación , Humanos , Proteínas del Envoltorio Viral/genética , Proteínas del Envoltorio Viral/metabolismo , Acoplamiento ViralRESUMEN
During inflammation, dendritic cells emigrate from inflamed tissue across the lymphatic endothelium into the lymphatic vasculature and travel to regional lymph nodes to initiate immune responses. However, the processes that regulate dendritic cell tissue egress and migration across the lymphatic endothelium are not well defined. The mammalian lectin galectin-1 is highly expressed by vascular endothelial cells in inflamed tissue and has been shown to regulate immune cell tissue entry into inflamed tissue. Here, we show that galectin-1 is also highly expressed by human lymphatic endothelial cells, and deposition of galectin-1 in extracellular matrix selectively regulates migration of specific human dendritic cell subsets. The presence of galectin-1 inhibits migration of immunogenic dendritic cells through the extracellular matrix and across lymphatic endothelial cells, but it has no effect on migration of tolerogenic dendritic cells. The major galectin-1 counter-receptor on both dendritic cell populations is the cell surface mucin CD43; differential core 2 O-glycosylation of CD43 between immunogenic dendritic cells and tolerogenic dendritic cells appears to contribute to the differential effect of galectin-1 on migration. Binding of galectin-1 to immunogenic dendritic cells reduces phosphorylation and activity of the protein-tyrosine kinase Pyk2, an effect that may also contribute to reduced migration of this subset. In a murine lymphedema model, galectin-1(-/-) animals had increased numbers of migratory dendritic cells in draining lymph nodes, specifically dendritic cells with an immunogenic phenotype. These findings define a novel role for galectin-1 in inhibiting tissue emigration of immunogenic, but not tolerogenic, dendritic cells, providing an additional mechanism by which galectin-1 can dampen immune responses.
Asunto(s)
Movimiento Celular/inmunología , Células Dendríticas/inmunología , Células Endoteliales/inmunología , Galectina 1/inmunología , Animales , Línea Celular , Movimiento Celular/genética , Células Dendríticas/patología , Modelos Animales de Enfermedad , Células Endoteliales/patología , Matriz Extracelular/genética , Matriz Extracelular/inmunología , Quinasa 2 de Adhesión Focal/genética , Quinasa 2 de Adhesión Focal/inmunología , Galectina 1/genética , Glicosilación , Humanos , Leucosialina/genética , Leucosialina/inmunología , Linfedema/genética , Linfedema/inmunología , Linfedema/patología , Ratones , Ratones NoqueadosRESUMEN
UNLABELLED: Nipah virus (NiV) is a deadly emerging enveloped paramyxovirus that primarily targets human endothelial cells. Endothelial cells express the innate immune effector galectin-1 that we have previously shown can bind to specific N-glycans on the NiV envelope fusion glycoprotein (F). NiV-F mediates fusion of infected endothelial cells into syncytia, resulting in endothelial disruption and hemorrhage. Galectin-1 is an endogenous carbohydrate-binding protein that binds to specific glycans on NiV-F to reduce endothelial cell fusion, an effect that may reduce pathophysiologic sequelae of NiV infection. However, galectins play multiple roles in regulating host-pathogen interactions; for example, galectins can promote attachment of HIV to T cells and macrophages and attachment of HSV-1 to keratinocytes but can also inhibit influenza entry into airway epithelial cells. Using live Nipah virus, in the present study, we demonstrate that galectin-1 can enhance NiV attachment to and infection of primary human endothelial cells by bridging glycans on the viral envelope to host cell glycoproteins. In order to exhibit an enhancing effect, galectin-1 must be present during the initial phase of virus attachment; in contrast, addition of galectin-1 postinfection results in reduced production of progeny virus and syncytium formation. Thus, galectin-1 can have dual and opposing effects on NiV infection of human endothelial cells. While various roles for galectin family members in microbial-host interactions have been described, we report opposing effects of the same galectin family member on a specific virus, with the timing of exposure during the viral life cycle determining the outcome. IMPORTANCE: Nipah virus is an emerging pathogen that targets endothelial cells lining blood vessels; the high mortality rate (up to 70%) in Nipah virus infections results from destruction of these cells and resulting catastrophic hemorrhage. Host factors that promote or prevent Nipah virus infection are not well understood. Endogenous human lectins, such as galectin-1, can function as pattern recognition receptors to reduce infection and initiate immune responses; however, lectins can also be exploited by microbes to enhance infection of host cells. We found that galectin-1, which is made by inflamed endothelial cells, can both promote Nipah virus infection of endothelial cells by "bridging" the virus to the cell, as well as reduce production of progeny virus and reduce endothelial cell fusion and damage, depending on timing of galectin-1 exposure. This is the first report of spatiotemporal opposing effects of a host lectin for a virus in one type of host cell.
Asunto(s)
Células Endoteliales/fisiología , Células Endoteliales/virología , Galectina 1/metabolismo , Células Gigantes/virología , Interacciones Huésped-Patógeno , Virus Nipah/fisiología , Internalización del Virus , Células Cultivadas , Células Endoteliales/inmunología , Galectina 1/inmunología , Humanos , Virus Nipah/inmunologíaRESUMEN
Leukocyte migration from the bloodstream into tissues, and from tissues to lymph nodes, depends on expression of specific adhesion and signaling molecules by vascular endothelial cells and lymphatic endothelial cells. Tissue damage and microbial infection induce vascular endothelial cells to up-regulate expression of adhesion molecules to facilitate entry of several leukocyte populations from blood into tissues. Many of these cells then leave inflamed tissue and migrate to regional lymph nodes. A critical population that emigrates from inflamed tissue is dendritic cells. Dendritic cells in tissue have to migrate through extracellular matrix and across a layer of lymphatic endothelial cells to enter the lymphatic vasculature. Little is known about the adhesion molecules expressed by lymphatic endothelial cells or the processes required for the critical step of dendritic cell exit from tissues, specifically migration through the extracellular matrix and basal-to-apical migration across the lymphatic endothelial cell layer into lymphatic vasculature.Members of the galectin family of carbohydrate binding proteins are expressed in both vascular and lymphatic endothelial cells. Dynamic changes in galectin expression during inflammation are known to regulate leukocyte tissue entry during inflammation. However, the roles of galectin family members expressed by lymphatic endothelial cells in leukocyte tissue exit remain to be explored.Here, we describe an in vitro transmigration assay that mimics dendritic cell tissue exit in the presence and absence of galectin protein. Fluorescently labeled human dendritic cell migration through extracellular matrix and across human lymphatic endothelial cells is examined in the presence and absence of recombinant human galectin protein.
Asunto(s)
Movimiento Celular , Células Dendríticas/citología , Células Endoteliales/citología , Matriz Extracelular/metabolismo , Galectinas/metabolismo , Técnicas de Cultivo de Célula , Separación Celular , Fluoresceínas/metabolismo , Humanos , Membranas Artificiales , Proteínas Recombinantes/metabolismo , Coloración y Etiquetado , Succinimidas/metabolismoRESUMEN
Microbe-host interactions are complex processes that are directly and indirectly regulated by a variety of factors, including microbe presentation of specific molecular signatures on the microbial surface, as well as host cell presentation of receptors that recognize these pathogen signatures. Cell surface glycans are one important class of microbial signatures that are recognized by a variety of host cell lectins. Host cell lectins that recognize microbial glycans include members of the galectin family of lectins that recognize specific glycan ligands on viruses, bacteria, fungi, and parasites. In this review, we will discuss the ways that the interactions of microbial glycans with host cell galectins positively and negatively regulate pathogen attachment, invasion, and survival, as well as regulate host responses that mitigate microbial pathogenesis.
RESUMEN
Three adhesion complexes span the sarcolemma and facilitate critical connections between the extracellular matrix and the actin cytoskeleton: the dystrophin- and utrophin-glycoprotein complexes and α7ß1 integrin. Loss of individual protein components results in a loss of the entire protein complex and muscular dystrophy. Muscular dystrophy is a progressive, lethal wasting disease characterized by repetitive cycles of myofiber degeneration and regeneration. Protein-replacement therapy offers a promising approach for the treatment of muscular dystrophy. Recently, we demonstrated that sarcospan facilitates protein-protein interactions amongst the adhesion complexes and is an important potential therapeutic target. Here, we review current protein-replacement strategies, discuss the potential benefits of sarcospan expression, and identify important experiments that must be addressed for sarcospan to move to the clinic.
Asunto(s)
Proteínas Portadoras/metabolismo , Distrofina/metabolismo , Integrinas/metabolismo , Proteínas de la Membrana/metabolismo , Distrofias Musculares/terapia , Proteínas de Neoplasias/metabolismo , Sarcolema/metabolismo , Utrofina/metabolismo , Animales , Humanos , Distrofias Musculares/metabolismo , Transducción de SeñalRESUMEN
Diffuse large B-cell lymphoma (DLBCL) is the most common non-Hodgkin lymphoma and an aggressive malignancy. Galectin-3 (gal-3), the only antiapoptotic member of the galectin family, is overexpressed in DLBCL. While gal-3 can localize to intracellular sites, gal-3 is secreted by DLBCL cells and binds back to the cell surface in a carbohydrate-dependent manner. The major counterreceptor for gal-3 on DLBCL cells was identified as the transmembrane tyrosine phosphatase CD45. Removal of cell-surface gal-3 from CD45 with the polyvalent glycan inhibitor GCS-100 rendered DLBCL cells susceptible to chemotherapeutic agents. Binding of gal-3 to CD45 modulated tyrosine phosphatase activity; removal of endogenous cell-surface gal-3 from CD45 with GCS-100 increased phosphatase activity, while addition of exogenous gal-3 reduced phosphatase activity. Moreover, the increased susceptibility of DLBCL cells to chemotherapeutic agents after removal of gal-3 by GCS-100 required CD45 phosphatase activity. Gal-3 binding to a subset of highly glycosylated CD45 glycoforms was regulated by the C2GnT-1 glycosyltransferase, indicating that specific glycosylation of CD45 is important for regulation of gal-3-mediated signaling. These data identify a novel role for cell-surface gal-3 and CD45 in DLBCL survival and suggest novel therapeutic targets to sensitize DLBCL cells to death.
Asunto(s)
Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Galectina 3/metabolismo , Antígenos Comunes de Leucocito/metabolismo , Linfoma de Células B Grandes Difuso/metabolismo , Anticuerpos Monoclonales de Origen Murino/farmacología , Línea Celular Tumoral , Membrana Celular/efectos de los fármacos , Membrana Celular/metabolismo , Supervivencia Celular/efectos de los fármacos , Dexametasona/farmacología , Relación Dosis-Respuesta a Droga , Etopósido/farmacología , Citometría de Flujo , Glicosilación/efectos de los fármacos , Humanos , Immunoblotting , Inmunohistoquímica , Antígenos Comunes de Leucocito/genética , Linfoma de Células B Grandes Difuso/genética , Linfoma de Células B Grandes Difuso/patología , N-Acetilglucosaminiltransferasas/genética , N-Acetilglucosaminiltransferasas/metabolismo , Polisacáridos/farmacología , Unión Proteica/efectos de los fármacos , Interferencia de ARN , Rituximab , Análisis de Matrices TisularesRESUMEN
Duchenne muscular dystrophy is an X-linked disorder characterized by loss of dystrophin, a cytoskeletal protein that connects the actin cytoskeleton in skeletal muscle cells to extracellular matrix. Dystrophin binds to the cytoplasmic domain of the transmembrane glycoprotein ß-dystroglycan (ß-DG), which associates with cell surface α-dystroglycan (α-DG) that binds laminin in the extracellular matrix. ß-DG can also associate with utrophin, and this differential association correlates with specific glycosylation changes on α-DG. Genetic modification of α-DG glycosylation can promote utrophin binding and rescue dystrophic phenotypes in mouse dystrophy models. We used high throughput screening with the plant lectin Wisteria floribunda agglutinin (WFA) to identify compounds that altered muscle cell surface glycosylation, with the goal of finding compounds that increase abundance of α-DG and associated sarcolemmal glycoproteins, increase utrophin usage, and increase laminin binding. We identified one compound, lobeline, from the Prestwick library of Food and Drug Administration-approved compounds that fulfilled these criteria, increasing WFA binding to C2C12 cells and to primary muscle cells from wild type and mdx mice. WFA binding and enhancement by lobeline required complex N-glycans but not O-mannose glycans that bind laminin. However, inhibiting complex N-glycan processing reduced laminin binding to muscle cell glycoproteins, although O-mannosylation was intact. Glycan analysis demonstrated a general increase in N-glycans on lobeline-treated cells rather than specific alterations in cell surface glycosylation, consistent with increased abundance of multiple sarcolemmal glycoproteins. This demonstrates the feasibility of high throughput screening with plant lectins to identify compounds that alter muscle cell glycosylation and identifies a novel role for N-glycans in regulating muscle cell function.
Asunto(s)
Glicómica/métodos , Laminina/metabolismo , Mioblastos Esqueléticos/metabolismo , Polisacáridos/metabolismo , Músculo Cuádriceps/metabolismo , Sarcolema/metabolismo , Animales , Células Cultivadas , Glicómica/instrumentación , Glicosilación , Lobelina/farmacología , Masculino , Manosiltransferasas/genética , Manosiltransferasas/metabolismo , Ratones , Ratones Endogámicos mdx , Fibras Musculares Esqueléticas/citología , Fibras Musculares Esqueléticas/efectos de los fármacos , Fibras Musculares Esqueléticas/metabolismo , Proteínas Musculares/metabolismo , Distrofia Muscular de Duchenne/metabolismo , Distrofia Muscular de Duchenne/patología , Mioblastos Esqueléticos/citología , Mioblastos Esqueléticos/efectos de los fármacos , Agonistas Nicotínicos/farmacología , Lectinas de Plantas/farmacología , Músculo Cuádriceps/citología , ARN Interferente Pequeño/genética , Receptores N-Acetilglucosamina , Sarcolema/efectos de los fármacosRESUMEN
Glycosylation affects many essential T cell processes and is intrinsically controlled throughout the lifetime of a T cell. CD43 and CD45 are the two most abundant glycoproteins on the T cell surface and are decorated with O- and N-glycans. Global T cell glycosylation and specific glycosylation of CD43 and CD45 are modulated during thymocyte development and T cell activation; T cells control the type and abundance of glycans decorating CD43 and CD45 by regulating expression of glycosyltransferases and glycosidases. Additionally, T cells regulate glycosylation of CD45 by expressing alternatively spliced isoforms of CD45 that have different glycan attachment sites. The glycophenotype of CD43 and CD45 on T cells influences how T cells interact with the extracellular environment, including how T cells interact with endogenous lectins. This review focuses on changes in glycosylation of CD43 and CD45 occurring throughout T cell development and activation and the role that glycosylation plays in regulating T cell processes, such as migration, T cell receptor signaling, and apoptosis.
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Antígenos Comunes de Leucocito/inmunología , Leucosialina/inmunología , Polisacáridos/inmunología , Linfocitos T/inmunología , Animales , Apoptosis , Diferenciación Celular , Movimiento Celular , Supervivencia Celular , Glicosilación , Humanos , Antígenos Comunes de Leucocito/química , Leucosialina/química , Activación de Linfocitos , Ratones , Polisacáridos/química , Transducción de Señal , Linfocitos T/citología , Linfocitos T/fisiologíaRESUMEN
Interaction of cell surface glycoproteins with endogenous lectins on the cell surface regulates formation and maintenance of plasma membrane domains, clusters signaling complexes, and controls the residency time of glycoproteins on the plasma membrane. Galectin-9 is a soluble, secreted lectin that binds to glycoprotein receptors to form galectin-glycoprotein lattices on the cell surface. Whereas galectin-9 binding to specific glycoprotein receptors induces death of CD4 Th1 cells, CD4 Th2 cells are resistant to galectin-9 death due to alternative glycosylation. On Th2 cells, galectin-9 binds cell surface protein disulfide isomerase (PDI), increasing retention of PDI on the cell surface and altering the redox status at the plasma membrane. Cell surface PDI regulates integrin function on platelets and also enhances susceptibility of T cells to infection with HIV. We find that galectin-9 binding to PDI on Th2 cells results in increased cell migration through extracellular matrix via ß3 integrins, identifying a unique mechanism to regulate T-cell migration. In addition, galectin-9 binding to PDI on T cells potentiates infection with HIV. We identify a mechanism for regulating cell surface redox status via a galectin-glycoprotein lattice, to regulate distinct T-cell functions.
Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Movimiento Celular , Galectinas/metabolismo , VIH-1/fisiología , Fusión de Membrana , Proteína Disulfuro Isomerasas/metabolismo , Animales , Western Blotting , Línea Celular , Membrana Celular/metabolismo , Electroforesis en Gel de Poliacrilamida , Humanos , Ratones , Ratones Endogámicos C57BL , Oxidación-Reducción , Unión ProteicaRESUMEN
By definition, adhesion/growth-regulatory galectins are known for their ability to bind ß-galactosides such as Galß(1 â 4)Glc (lactose). Indications for affinity of human galectin-1 to α-linked digalactosides pose questions on the interaction profile with such bound ligands and selection of the galactose moiety for CH-π stacking. These issues are resolved by a combination of (15)N-(1)H heteronuclear single quantum coherence (HSQC) chemical shift and saturation transfer difference nuclear magnetic resonance (STD NMR) epitope mappings with docking analysis, using the α(1 â 3/4)-linked digalactosides and also Galα(1 â 6)Glc (melibiose) as test compounds. The experimental part revealed interaction with the canonical lectin site, and this preferentially via the non-reducing-end galactose moiety. Low-energy conformers appear to be selected without notable distortion, as shown by molecular dynamics simulations. With the α(1 â 4) disaccharide, however, the typical CH-π interaction is significantly diminished, yet binding appears to be partially compensated for by hydrogen bonding. Overall, these findings reveal that the type of α-linkage in digalactosides has an impact on maintaining CH-π interactions and the pattern of hydrogen bonding, explaining preference for the α(1 â 3) linkage. Thus, this lectin is able to accommodate both α- and ß-linked galactosides at the same site, with major contacts to the non-reducing-end sugar unit.