Neem leaf glycoprotein binding to Dectin-1 receptors on dendritic cell induces type-1 immunity through CARD9 mediated intracellular signal to NFκB

• Published in: Cell communication and signaling 2024-04, Vol.22 (1)
• Main Authors: Ganguly, Nilanjan, Das, Tapasi, Bhuniya, Avishek, Guha, Ipsita, Chakravarti, Mohona, Dhar, Sukanya, Sarkar, Anirban, Bera, Saurav, Dhar, Jesmita, Dasgupta, Shayani, Saha, Akata, Ghosh, Tithi, Das, Juhina, Sk, Ugir Hossain, Banerjee, Saptak, Laskar, Subrata, Bose, Anamika, Baral, Rathindranath
• Format: Article
• Language: English
• Subjects: Botanical research; Dendritic cells; Glycoproteins; Immune response; Margosa; Physiological aspects; Plant proteins
• ISSN: 1478-811X; 1478-811X
• DOI: 10.1186/s12964-024-01576-z

A water-soluble ingredient of mature leaves of the tropical mahogany 'Neem' (Azadirachta indica), was identified as glycoprotein, thus being named as 'Neem Leaf Glycoprotein' (NLGP). This non-toxic leaf-component regressed cancerous murine tumors (melanoma, carcinoma, sarcoma) recurrently in different experimental circumstances by boosting prime antitumor immune attributes. Such antitumor immunomodulation, aid cytotoxic T cell (T.sub.c)-based annihilation of tumor cells. This study focused on identifying and characterizing the signaling gateway that initiate this systemic immunomodulation. In search of this gateway, antigen-presenting cells (APCs) were explored, which activate and induce the cytotoxic thrust in T.sub.c cells. Six glycoprotein-binding C-type lectins found on APCs, namely, MBR, Dectin-1, Dectin-2, DC-SIGN, DEC205 and DNGR-1 were screened on bone marrow-derived dendritic cells from C57BL/6 J mice. Fluorescence microscopy, RT-PCR, flow cytometry and ELISA revealed Dectin-1 as the NLGP-binding receptor, followed by verifications through RNAi. Following detection of [beta]-Glucans in NLGP, their interactions with Dectin-1 were explored in silico. Roles of second messengers and transcription factors in the downstream signal were studied by co-immunoprecipitation, western blotting, and chromatin-immunoprecipitation. Intracellularization of FITC-coupled NLGP was observed by processing confocal micrographs of DCs. Considering extents of hindrance in NLGP-driven transcription rates of the cytokines IL-10 and IL-12p35 by receptor-neutralization, Dectin-1 receptors on dendritic cells were found to bind NLGP through the ligand's peripheral [beta]-Glucan chains. The resulting signal phosphorylates PKC[delta], forming a trimolecular complex of CARD9, Bcl10 and MALT1, which in turn activates the canonical NFκB-pathway of transcription-regulation. Consequently, the NFκB-heterodimer p65:p50 enhances Il12a transcription and the p50:p50 homodimer represses Il10 transcription, bringing about a cytokine-based systemic-bias towards type-1 immune environment. Further, NLGP gets engulfed within dendritic cells, possibly through endocytic activities of Dectin-1. NLGP's binding to Dectin-1 receptors on murine dendritic cells, followed by the intracellular signal, lead to NFκB-mediated contrasting regulation of cytokine-transcriptions, initiating a pro-inflammatory immunopolarization, which amplifies further by the responding immune cells including T.sub.c cells, a