Pivotal role of the carbohydrate recognition domain in self-interaction of CLEC4A to elicit the ITIM-mediated inhibitory function in murine conventional dendritic cells in vitro

Abstract C-type lectin receptors (CLRs), pattern recognition receptors (PRRs) with a characteristic carbohydrate recognition domain (CRD) in the extracellular portion, mediate crucial cellular functions upon recognition of glycosylated pathogens and self-glycoproteins. CLEC4A is the only classical CLR that possesses an intracellular immunoreceptor tyrosine-based inhibitory motif (ITIM), which possibly transduces negative signals. However, how CLEC4A exerts cellular inhibition remains unclear. Here, we report that the self-interaction of CLEC4A through the CRD is required for the ITIM-mediated suppressive function in conventional dendritic cells (cDCs). Human type 2 cDCs (cDC2) and monocytes display a higher expression of CLEC4A than cDC1 and plasmacytoid DCs (pDCs) as well as B cells. The extracellular portion of CLEC4A specifically binds to a murine cDC cell line expressing CLEC4A, while its extracellular portion lacking the N-glycosylation site or the EPS motif within the CRD reduces their association. Furthermore, the deletion of the EPS motif within the CRD or ITIM in CLEC4A almost completely impairs its suppressive effect on the activation of the murine cDC cell line, whereas the absence of the N-glycosylation site within the CRD exhibits partial inhibition on their activation. On the other hand, antagonistic monoclonal antibody (mAb) to CLEC4A, which inhibits the self-interaction of CLEC4A and its downstream signaling in murine transfectants, enhances the activation of monocytes and monocyte-derived immature DCs upon stimulation with a Toll-like receptor (TLR) ligand. Thus, our findings suggest a pivotal role of the CRD in self-interaction of CLEC4A to elicit the ITIM-mediated inhibitory signal for the control of the function of cDCs.

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Role of conserved and nonconserved residues in the Ca(2+)-dependent carbohydrate-recognition domain of a rat mannose-binding protein. Analysis by random cassette mutagenesis.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(19)50094-1 ◽

1992 ◽

Vol 267 (15) ◽

pp. 10831-10841

Author(s):

M.S. Quesenberry ◽

K Drickamer

Keyword(s):

Binding Protein ◽

Protein Analysis ◽

Carbohydrate Recognition Domain ◽

Carbohydrate Recognition ◽

Mannose Binding Protein ◽

Mannose Binding

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Analysis of a Preliminary microRNA Expression Signature in a Human Telangiectatic Osteogenic Sarcoma Cancer Cell Line

International Journal of Molecular Sciences ◽

10.3390/ijms22031163 ◽

2021 ◽

Vol 22 (3) ◽

pp. 1163

Author(s):

Gaia Palmini ◽

Cecilia Romagnoli ◽

Simone Donati ◽

Roberto Zonefrati ◽

Gianna Galli ◽

...

Keyword(s):

Cell Line ◽

Molecular Mechanisms ◽

Osteogenic Sarcoma ◽

Cancer Cell Line ◽

Microscopic Features ◽

In Vitro Establishment ◽

Profile Characteristic ◽

First Time

Telangiectatic osteosarcoma (TOS) is an aggressive variant of osteosarcoma (OS) with distinctive radiographic, gross, microscopic features, and prognostic implications. Despite several studies on OS, we are still far from understanding the molecular mechanisms of TOS. In recent years, many studies have demonstrated not only that microRNAs (miRNAs) are involved in OS tumorigenesis, development, and metastasis, but also that the presence in high-grade types of OS of cancer stem cells (CSCs) plays an important role in tumor progression. Despite these findings, nothing has been described previously about the expression of miRNAs and the presence of CSCs in human TOS. Therefore, we have isolated/characterized a putative CSC cell line from human TOS (TOS-CSCs) and evaluated the expression levels of several miRNAs in TOS-CSCs using real-time quantitative assays. We show, for the first time, the existence of CSCs in human TOS, highlighting the in vitro establishment of this unique stabilized cell line and an identification of a preliminary expression of the miRNA profile, characteristic of TOS-CSCs. These findings represent an important step in the study of the biology of one of the most aggressive variants of OS and the role of miRNAs in TOS-CSC behavior.

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Pivotal role of MUC1 glycosylation by cigarette smoke in modulating disruption of airway adherens junctions in vitro

The Journal of Pathology ◽

10.1002/path.4375 ◽

2014 ◽

Vol 234 (1) ◽

pp. 60-73 ◽

Cited By ~ 21

Author(s):

Lili Zhang ◽

Marianne Gallup ◽

Lorna Zlock ◽

Yu Ting Feeling Chen ◽

Walter E Finkbeiner ◽

...

Keyword(s):

Cigarette Smoke ◽

Adherens Junctions ◽

Pivotal Role

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Homeopathic potencies of Arnica montana L. change gene expression in a Tamm-Horsfall protein-1 cell line in vitro model: the role of ethanol as a possible confounder and statistical bias

Journal of Integrative Medicine ◽

10.1016/s2095-4964(17)60346-7 ◽

2017 ◽

Vol 15 (4) ◽

pp. 255-264 ◽

Cited By ~ 4

Author(s):

Salvatore Chirumbolo ◽

Geir Bjørklund

Keyword(s):

Gene Expression ◽

Cell Line ◽

In Vitro Model ◽

Statistical Bias ◽

Arnica Montana ◽

Vitro Model

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Exploring the role of dosing procedure and chemical properties in in vitro assays using a fish gill cell line

Comparative Biochemistry and Physiology Part A Molecular & Integrative Physiology ◽

10.1016/j.cbpa.2009.04.629 ◽

2009 ◽

Vol 153 (2) ◽

pp. S89

Author(s):

Kristin Schirmer ◽

Katrin Tanneberger ◽

Nynke I. Kramer ◽

Frans J.M. Busser ◽

Joop L.M. Hermens ◽

...

Keyword(s):

Cell Line ◽

Chemical Properties ◽

In Vitro Assays ◽

Fish Gill ◽

Gill Cell ◽

And Chemical Properties

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In Vivo Role of Dendritic Cells in a Murine Model of Pulmonary Cryptococcosis

Infection and Immunity ◽

10.1128/iai.00317-06 ◽

2006 ◽

Vol 74 (7) ◽

pp. 3817-3824 ◽

Cited By ~ 58

Author(s):

Karen L. Wozniak ◽

Jatin M. Vyas ◽

Stuart M. Levitz

Keyword(s):

T Cells ◽

Dendritic Cells ◽

T Cell Activation ◽

Cell Activation ◽

Ex Vivo ◽

Interleukin 2 ◽

Mouse Lung

ABSTRACT Dendritic cells (DC) have been shown to phagocytose and kill Cryptococcus neoformans in vitro and are believed to be important for inducing protective immunity against this organism. Exposure to C. neoformans occurs mainly by inhalation, and in this study we examined the in vivo interactions of C. neoformans with DC in the lung. Fluorescently labeled live C. neoformans and heat-killed C. neoformans were administered intranasally to C57BL/6 mice. At specific times postinoculation, mice were sacrificed, and lungs were removed. Single-cell suspensions of lung cells were prepared, stained, and analyzed by microscopy and flow cytometry. Within 2 h postinoculation, fluorescently labeled C. neoformans had been internalized by DC, macrophages, and neutrophils in the mouse lung. Additionally, lung DC from mice infected for 7 days showed increased expression of the maturation markers CD80, CD86, and major histocompatibility complex class II. Finally, ex vivo incubation of lung DC from infected mice with Cryptococcus-specific T cells resulted in increased interleukin-2 production compared to the production by DC from naïve mice, suggesting that there was antigen-specific T-cell activation. This study demonstrated that DC in the lung are capable of phagocytosing Cryptococcus in vivo and presenting antigen to C. neoformans-specific T cells ex vivo, suggesting that these cells have roles in innate and adaptive pulmonary defenses against cryptococcosis.

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Inhibition of O-GlcNAc Transferase Alters the Differentiation and Maturation Process of Human Monocyte Derived Dendritic Cells

Cells ◽

10.3390/cells10123312 ◽

2021 ◽

Vol 10 (12) ◽

pp. 3312

Author(s):

Matjaž Weiss ◽

Marko Anderluh ◽

Martina Gobec

Keyword(s):

Dendritic Cells ◽

Posttranslational Modification ◽

Human Monocyte ◽

T Cell Differentiation ◽

Maturation Process ◽

Hla Dr ◽

Glcnac Transferase ◽

And Function

The O-GlcNAcylation is a posttranslational modification of proteins regulated by O-GlcNAc transferase (OGT) and O-GlcNAcase. These enzymes regulate the development, proliferation and function of cells, including the immune cells. Herein, we focused on the role of O-GlcNAcylation in human monocyte derived dendritic cells (moDCs). Our study suggests that inhibition of OGT modulates AKT and MEK/ERK pathways in moDCs. Changes were also observed in the expression levels of relevant surface markers, where reduced expression of CD80 and DC-SIGN, and increased expression of CD14, CD86 and HLA-DR occurred. We also noticed decreased IL-10 and increased IL-6 production, along with diminished endocytotic capacity of the cells, indicating that inhibition of O-GlcNAcylation hampers the transition of monocytes into immature DCs. Furthermore, the inhibition of OGT altered the maturation process of immature moDCs, since a CD14medDC-SIGNlowHLA-DRmedCD80lowCD86high profile was noticed when OGT inhibitor, OSMI-1, was present. To evaluate DCs ability to influence T cell differentiation and polarization, we co-cultured these cells. Surprisingly, the observed phenotypic changes of mature moDCs generated in the presence of OSMI-1 led to an increased proliferation of allogeneic T cells, while their polarization was not affected. Taken together, we confirm that shifting the O-GlcNAcylation status due to OGT inhibition alters the differentiation and function of moDCs in in vitro conditions.

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