Allergen-specific immunotherapy (AIT) is the only curative treatment for allergic diseases. However, the long desensitization phase and potentially dangerous allergic side effects limit its broad ...application. Therefore, safer and more effective vaccines are required. Targeting dendritic cells (DCs) with novel allergen conjugates is a promising strategy for AIT. In this study, a novel vaccine with a DC-targeting effect for AIT was constructed. Liposomes were used as vehicles, and a targeted nanovaccine (Lex-lip-Der f 2) was constructed by loading the recombinant group 2 allergen of Dermatophagoides farinae (Der f 2) and conjugating with the DC-SIGN ligand Lewis X. The effect of the vaccine on DCs and T cell responses and the safety of the vaccine were investigated in vitro. The results showed that the Lex-lip-Der f 2 vaccine was spherical, with size of approximately 128 nm. The protein-loading capacity of the vaccine was 0.106 ± 0.001 mg per mg liposome and protein was gradually released from the liposomes during the first 12 h. Lex-lip-Der f 2 was taken up more efficiently by DCs than non-targeted liposomes or free Der f 2. Besides, Lex-lip-Der f 2 significantly inhibited the release of IL-4, IL-6, and TNF-a from DCs. Accordingly, Der f 2-lip loaded DCs significantly decreased IL-4 levels in autologous naïve CD4+T cells. Moreover, Lex-lip-Der f 2-treated basophils showed lower activation levels. These results suggest that DC-SIGN targeting mediated by Lewis X could inhibit the Th2 cell response and improve vaccine safety, and may be a novel vaccination strategy.
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Multivalency plays a major role in biological processes and particularly in the relationship between pathogenic microorganisms and their host that involves protein-glycan recognition. These ...interactions occur during the first steps of infection, for specific recognition between host and bacteria, but also at different stages of the immune response. The search for high-affinity ligands for studying such interactions involves the combination of carbohydrate head groups with different scaffolds and linkers generating multivalent glycocompounds with controlled spatial and topology parameters. By interfering with pathogen adhesion, such glycocompounds including glycopolymers, glycoclusters, glycodendrimers and glyconanoparticles have the potential to improve or replace antibiotic treatments that are now subverted by resistance. Multivalent glycoconjugates have also been used for stimulating the innate and adaptive immune systems, for example with carbohydrate-based vaccines. Bacteria present on their surfaces natural multivalent glycoconjugates such as lipopolysaccharides and S-layers that can also be exploited or targeted in anti-infectious strategies.
Dendritic cell‐specific intercellular adhesion molecule‐3‐grabbing non‐integrin (DC‐SIGN), a C‐type lectin expressed on the plasma membrane by human immature dendritic cells, is a receptor for ...numerous viruses including Ebola, SARS and dengue. A controversial question has been whether DC‐SIGN functions as a complete receptor for both binding and internalization of dengue virus (DENV) or whether it is solely a cell surface attachment factor, requiring either hand‐off to another receptor or a co‐receptor for internalization. To examine this question, we used 4 cell types: human immature dendritic cells and NIH3T3 cells expressing either wild‐type DC‐SIGN or 2 internalization‐deficient DC‐SIGN mutants, in which either the 3 cytoplasmic internalization motifs are silenced by alanine substitutions or the cytoplasmic region is truncated. Using confocal and super‐resolution imaging and high content single particle tracking, we investigated DENV binding, DC‐SIGN surface transport, endocytosis, as well as cell infectivity. DC‐SIGN was found colocalized with DENV inside cells suggesting hand‐off at the plasma membrane to another receptor did not occur. Moreover, all 3 DC‐SIGN molecules on NIH3T3 cells supported cell infection. These results imply the involvement of a co‐receptor because cells expressing the internalization‐deficient mutants could still be infected.
Whether dendritic cell‐specific intercellular adhesion molecule‐3‐grabbing non‐integrin (DC‐SIGN) functions as merely an attachment factor for dengue virus (DENV) or whether DC‐SIGN plays further roles beyond attachment has been controversial. We use mammalian cell culture models, as well as primary dendritic cells, and high resolution, quantitative fluorescence microscopy to track the movements of DC‐SIGN and DENV during viral entry. Our results support a model in which DC‐SIGN captures DENV and participates, along with a co‐receptor, in DENV internalization via clathrin‐coated structures and subsequent trafficking to early endosomes.
Background. Chikungunya infections range from subclinical infection to debilitating arthralgia and to chronic inflammatory rheumatism. Tumor necrosis factor (TNF) alpha, DC-SIGN (dendritic ...cell-specific intercellular adhesion molecule 3-grabbing nonintegrin), Toll-like receptor (TLR) 3, and blood groups have been directly or indirectly implicated in the susceptibility and pathogenesis of chikungunya. Methods. To test the hypothesis that polymorphisms in genes coding for these molecules determine clinical outcomes of chikungunya infection, a retrospective case-control study was performed in Leon, Nicaragua. The study included 132 case patients and 132 controls, matched for age, sex and neighborhood. Case patients had clinical symptoms of chikungunya, which was diagnosed by means of polymerase chain reaction. Controls were individuals not reporting abrupt presentation of clinical chikungunya-like symptoms. Polymorphisms were identified by TaqMan single-nucleotide polymorphism genotyping assays. Results. After adjustment for sociodemographic risk factors, chikungunya disease was associated with polymorphism in DC-SIGN and TLR3 genes (odds ratios, 5.2 and 3.3, respectively), and TNF-alpha with reduced persistent joint pain (0.24). Persistent joint pain was also associated with age, female sex and other comorbid conditions. Most interestingly, the Lewis-negative phenotype was strongly associated with both symptomatic chikungunya and immunoglobulin G seropositivity (odds ratios, 2.7, and 3.3, respectively). Conclusion. This study identified polymorphisms in DC-SIGN, TLR3, and TNF-alpha genes as well as Lewis-negative phenotype as risk factors for chikungunya infection and disease progression.
Zika virus (ZIKV) and dengue virus (DENV) share a lot of similarities being both phylogenetically closely related, share the same insect vector passage for reaching the host, affinity for the same ...carbohydrate receptor domains (CRDs), indicating feasible competition between them on the natural field. Here, we prospected interactions of both envelope proteins with a DC-SIGN, a transmembrane c-type lectine receptor with the most implicated CRD with the Flavivirus infection presents on dendritic cells involved in viruses replication processes into the host, and among rares CRD receptors susceptible to interacting with a broad of subtypes of DENV. Protein-protein docking procedures produced structures for molecular dynamics experiments, suggesting the most energetically favorable complex. The difference found in the deltaG results prompted the experimentation with molecular dynamics. To investigate further specific residues involved with such interactions we produced a decomposition analysis using molecular dynamics of the docked proteins evaluated afterward with the Generalized Born Surface Area method. Solvent-accessible surface area (SASA) analysis for both showed very similar but with a slight reduction for ZIKV_E, which agreed with residues SASA analysis highlighting regions more exposed in the ZIVK protein than in DENV. Despite residues PHE313 is reponsible for most of the interactions with the envelope of these arboviruses, ZIKV interacted with this residue in DC-SIGN with lower energies and using more interactions with not expexted residues GLU241 and ARG386. Taken together these results suggest better competitive interaction of ZIKV with the DC-SIGN receptor, particularly in the CRD portion.
Interleukin‐34 (IL‐34) is a cytokine that supports the viability and differentiation of macrophages. An important cytokine for the development of epidermal immunity, IL‐34, is present and plays a ...role in the immunity of the oral environment. IL‐34 has been linked to inflammatory periodontal diseases, which involve innate phagocytes, including macrophages. Whether IL‐34 can alter the ability of macrophages to effectively interact with oral microbes is currently unclear. Using macrophages derived from human blood monocytes with either the canonical cytokine colony‐stimulating factor (CSF)1 or IL‐34, we compared the ability of the macrophages to phagocytose, kill, and respond through the production of cytokines to the periodontal keystone pathogen Porphyromonas gingivalis. While macrophages derived from both cytokines were able to engulf the bacterium equally, IL‐34‐derived macrophages were much less capable of killing internalized P. gingivalis. Of the macrophage cell surface receptors known to interact with P. gingivalis, dendritic cell‐specific intercellular adhesion molecule‐grabbing nonintegrin was found to have the largest variation between IL‐34‐ and CSF1‐derived macrophages. We also found that upon interaction with P. gingivalis, IL‐34‐derived macrophages produced significantly less of the neutrophil chemotactic factor IL‐8 than macrophages derived in the presence of CSF1. Mechanistically, we identified that the levels of IL‐8 corresponded with P. gingivalis survival and dephosphorylation of the major transcription factor NF‐κB p65. Overall, we found that macrophages differentiated in the presence of IL‐34, a dominant cytokine in the oral gingiva, have a reduced ability to kill the keystone pathogen P. gingivalis and may be susceptible to specific bacteria‐mediated cytokine modification.
Overview of changes within Interleukin‐34 (IL‐34)‐differentiated macrophages leading to altered interactions with Porphyromonas gingivalis. IL‐34 differentiation leads to increased dendritic cell‐specific intercellular adhesion molecule‐grabbing nonintegrin, compared to CSF1‐matured macrophages, as well as increased P. gingivalis survival, decreased NF‐κB p65 phosphorylation and subsequent decrease of IL‐8 production.
Dendritic cells (DC) are critical cellular mediators of host immunity, notably by expressing a broad panel of pattern recognition receptors. One of those receptors, the C-type lectin receptor ...DC-SIGN, was previously reported as a regulator of endo/lysosomal targeting through functional connections with the autophagy pathway. Here, we confirmed that DC-SIGN internalization intersects with LC3
autophagy structures in primary human monocyte-derived dendritic cells (MoDC). DC-SIGN engagement promoted autophagy flux which coincided with the recruitment of ATG-related factors. As such, the autophagy initiation factor ATG9 was found to be associated with DC-SIGN very early upon receptor engagement and required for an optimal DC-SIGN-mediated autophagy flux. The autophagy flux activation upon DC-SIGN engagement was recapitulated using engineered DC-SIGN-expressing epithelial cells in which ATG9 association with the receptor was also confirmed. Finally, Stimulated emission depletion (STED) microscopy performed in primary human MoDC revealed DC-SIGN-dependent submembrane nanoclusters formed with ATG9, which was required to degrade incoming viruses and further limit DC-mediated transmission of HIV-1 infection to CD4
T lymphocytes. Our study unveils a physical association between the Pattern Recognition Receptor DC-SIGN and essential components of the autophagy pathway contributing to early endocytic events and the host's antiviral immune response.
The receptors and cellular factors used by phleboviruses (Bunyaviridae) to enter host cells remain largely unidentified. In addition to the C‐type lectin DC‐SIGN (Dendritic cell‐specific ...intercellular adhesion molecule 3‐grabbing non‐integrin), we report here that several phleboviruses subvert the closely related protein L‐SIGN for infection. Together, our results establish that L‐SIGN is an attachment factor not required for virus internalization whereas DC‐SIGN is an authentic entry receptor required for both binding and endocytosis. Our study underlines the importance of the subsequent entry processes in virus–receptor interactions beyond attachment.
Bunyaviruses represent a growing threat to humans and livestock globally. The receptors, cellular factors and endocytic pathways used by these emerging pathogens to infect cells remain largely unidentified and poorly characterized. DC‐SIGN is a C‐type lectin highly expressed on dermal dendritic cells that has been found to act as an authentic entry receptor for many phleboviruses (Bunyaviridae), including Rift Valley fever virus (RVFV), Toscana virus (TOSV) and Uukuniemi virus (UUKV). We found that these phleboviruses can exploit another C‐type lectin, L‐SIGN, for infection. L‐SIGN shares 77% sequence homology with DC‐SIGN and is expressed on liver sinusoidal endothelial cells. L‐SIGN is required for UUKV binding but not for virus internalization. An endocytosis‐defective mutant of L‐SIGN was still able to mediate virus uptake and infection, indicating that L‐SIGN acts as an attachment receptor for phleboviruses rather than an endocytic receptor. Our results point out a fundamental difference in the use of the C‐type lectins L‐SIGN and DC‐SIGN by UUKV to enter cells, although both proteins are closely related in terms of molecular structure and biological function. This study sheds new light on the molecular mechanisms by which phleboviruses target the liver and also highlights the added complexity in virus–receptor interactions beyond attachment.
Altered glycosylation in epithelial cancers may play an important role in tumour progression, as it may affect tumour cell migration and antigen presentation by antigen presenting cells. We ...specifically characterise the glycosylation patterns of two tumour antigens that are highly expressed in cancer tissue and often detected in their secreted form in serum: the epithelial mucin MUC1 and carcinoembryonic antigen (CEA, also called CEACAM5). We analysed 48 colorectal cancer patients, comparing normal colon and tumour epithelium within each patient. Lectin binding was studied by a standardised CEA/MUC1 capture ELISA, using several plant lectins, and the human C‐type lectins MGL and DC‐SIGN, and Galectin‐3. Peanut agglutinin (PNA) bound to MUC1 from tumour tissue in particular, suggests increased expression of the Thomsen‐Friedenreich antigen (TF‐antigen) (Core 1, Galβ1‐3GalNAc‐Ser/Thr). Only small amounts of Tn‐antigen (GalNAcα‐Ser/Thr) expression was observed, but the human C‐type lectin MGL showed increased binding to tumour‐associated MUC1. Furthermore, sialylation was greatly enhanced. In sharp contrast, tumour‐associated CEA (CEACAM5) contained high levels of the blood‐group related carbohydrates, Lewis X and Lewis Y. This correlated strongly with the interaction of the human C‐type lectin DC‐SIGN to tumour‐associated CEA, suggesting that CEA can be recognized and taken up by antigen presenting cells. In addition, increased mannose expression was observed and branched N‐glycans were prominent, and this correlated well with human Galectin‐3 binding. These data demonstrate that individual tumour antigens contain distinct glycan structures associated with cancer and, since glycans affect cellular interactions with its microenvironment, this may have consequences for progression of the disease.
Although COVID-19 transmission has been reduced by the advent of vaccinations and a variety of rapid monitoring techniques, the SARS-CoV-2 virus itself has shown a remarkable ability to mutate and ...persist. With this long track record of immune escape, researchers are still exploring prophylactic treatments to curtail future SARS-CoV-2 variants. Specifically, much focus has been placed on the antiviral lectin Griffithsin in preventing spike protein-mediated infection via the hACE2 receptor (direct infection). However, an oft-overlooked aspect of SARS-CoV-2 infection is viral capture by attachment receptors such as DC-SIGN, which is thought to facilitate the initial stages of COVID-19 infection in the lung tissue (called trans-infection). In addition, while immune escape is dictated by mutations in the spike protein, coronaviral virions also incorporate M, N, and E structural proteins within the particle. In this paper, we explored how several structural facets of both the SARS-CoV-2 virion and the antiviral lectin Griffithsin can affect and attenuate the infectivity of SARS-CoV-2 pseudovirus. We found that Griffithsin was a better inhibitor of hACE2-mediated direct infection when the coronaviral M protein is present compared to when it is absent (possibly providing an explanation regarding why Griffithsin shows better inhibition against authentic SARS-CoV-2 as opposed to pseudotyped viruses, which generally do not contain M) and that Griffithsin was not an effective inhibitor of DC-SIGN-mediated trans-infection. Furthermore, we found that DC-SIGN appeared to mediate trans-infection exclusively via binding to the SARS-CoV-2 spike protein, with no significant effect observed when other viral proteins (M, N, and/or E) were present. These results provide etiological data that may help to direct the development of novel antiviral treatments, either by leveraging Griffithsin binding to the M protein as a novel strategy to prevent SARS-CoV-2 infection or by narrowing efforts to inhibit trans-infection to focus on DC-SIGN binding to SARS-CoV-2 spike protein.
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