Isoform localization of Dectin‐1 regulates the signaling quality of anti‐fungal immunity

• Published in: European journal of immunology 2017-05, Vol.47 (5), p.848-859
• Main Authors: Fischer, Mike, Müller, Jörg P., Spies‐Weisshart, Bärbel, Gräfe, Christine, Kurzai, Oliver, Hünniger, Kerstin, Hochhaus, Andreas, Scholl, Sebastian, Schnetzke, Ulf
• Format: Article
• Language: English
• Subjects: Cell Line; Cell lines; Cell surface; Confocal microscopy; Cytokine secretion; Cytokines; Cytokines - biosynthesis; Cytokines - immunology; Dectin‐1 isoforms; Flow Cytometry; Fungal diseases; Fungicides; Glucans; Glycosylation; Humans; Immune responses; Immunity; Immunoblotting; Infectious diseases; Intracellular Signaling Peptides and Proteins - metabolism; Intracellular signalling; Isoforms; Lectins, C-Type - chemistry; Lectins, C-Type - genetics; Lectins, C-Type - metabolism; Localization; Lymphocytes B; Macrophages - immunology; Membrane Proteins - genetics; Membrane Proteins - immunology; Microscopy; Microscopy, Confocal; Monocytes; Monocytes - immunology; Monocytes - metabolism; Monocytes - physiology; Mycoses - immunology; Mycoses - metabolism; Protein Isoforms - chemistry; Protein Isoforms - metabolism; Scanning microscopy; Signal Transduction; Stimulation
• ISSN: 0014-2980; 1521-4141
• DOI: 10.1002/eji.201646849

Dectin‐1 is recognized as a major receptor for fungal ß‐glucans and contributes to anti‐fungal immunity. Human monocyte populations express Dectin‐1 isoforms A and B, which differ by the presence of a stalk region and its N‐linked glycosylation site. Here, we analyzed the expression of both isoforms in human monocyte‐derived cells. The cellular localization on cell lines stably expressing either Dectin‐1 isoform A or B was studied by flow cytometry and confocal laser scanning microscopy. Intracellular protein signaling and cytokine production were analyzed by immunoblotting and cytometric bead array, respectively. Monocyte‐derived cells showed cell type‐specific expression of the two isoforms. Glycosylated Dectin‐1 isoform A was predominantly localized at the cell surface, non‐glycosylated isoform B was retained intracellularly. Inhibition of glycosylation resulted in efficient abrogation of cell surface expression of isoform A. Signaling quality following Dectin‐1 stimulation was reduced in isoform B cells. Differential isoform specific cytokine secretion was observed by cytometric bead array. We show here that n‐glycosylation of Dectin‐1 is crucial for its cell surface expression and consequently signal transduction. Taken together, unique cytokine secretion and varying expression levels of human Dectin‐1 isoforms on monocyte‐derived cells may indicate distinct isoform usage as a cell type‐specific mechanism of regulating anti‐fungal immunity. On the basis of cell lines stably expressing Dectin‐1 isoforms A or B we provide insight into its isoform‐specific cellular compartmentation and ability to bind ß‐glucan. Furthermore, isoform‐dependent protein signaling and cytokine secretion were revealed which may contribute to regulation of antifungal immunity.