Dissecting the structure and function of the macrophage testraspan MS4A4A

Membrane-spanning 4A (MS4A) proteins are emerging as a new class of immune cell regulators, but the molecular mechanisms underpinning their biological functions are currently poorly characterized. Our lab has focused its attention on MS4A4A which is induced along macrophage differentiation and is detected in tissue resident macrophages (e.g. CNS, colon, lungs, skin). In vitro MS4A4A expression is increased in “alternative activated” macrophages (e.g. IL-4, dexamethasone) and is found highly expressed in some pathological settings (like rheumatoid arthritis) and in tumor-associated macrophages (e.g. colon carcinoma, lung adenocarcinoma, melanoma). Here, we aim to investigate the role of MS4A4A achieving a deeper mechanistic understanding of its function in macrophage biology and disclosing its contribution to immune responses. Our data showed that in macrophages MS4A4A deficiency impaired the expression of anti-inflammatory genes (Arg1) after IL-4 treatment and the production of IL-10 after LPS stimulation. suggesting that this protein might be involved in macrophage anti-inflammatory response. To well define MS4A4A contribution to macrophage biology, RNA sequencing of Wild Type (WT) and Ms4a4a Knock Out (KO) bone marrow derived macrophages (BMDM) has been performed and analysis is ongoing. Like other tetraspan proteins, MS4A4A forms omo- and hetero-complexes with other protein partners (MS4A4A, MS4A6A, MS4A7, Dectin-1, TREM2). Using a split-ubiquitin two-hybrid approach and mass spectrometry-based immunoprecipitation proteomics, we identified new possible partners for MS4A4A, including the γ signaling chain of Fcγ receptors, Toll-Like Receptor 2, and calcium/calmodulin-dependent 3',5'-cyclic nucleotide phosphodiesterase 1A. Furthermore, we are computationally characterizing MS4A4A structure and dynamics through an extensive molecular docking calculations campaign with some families of endogenous and exogenous bioactive compounds.