Galectin-9 binding to HLA-DR in dendritic cells controls immune synapse formation and T cell proliferation

To initiate T cell-mediated immunity, dendritic cells (DCs) present antigens to specific T cells through the establishment of an immune synapse (IS). While the molecular mechanisms behind the formation of the IS on the T cell side are well understood, how IS components are organized at the DC membrane remain ill-defined. Galectins, a family of β-galactoside binding proteins, modulate immune cell function via the establishment of specific glycan-dependent or independent interactions. Nonetheless, the molecular mechanisms that underlie galectin function are poorly described and very little is known regarding their contribution to DC-mediated T cell activation. Here, we demonstrate that intracellular galectin-9 (gal9) in DCs is required for T cell activation. Murine and human DCs lacking gal9 showed impaired induction of CD4 ⁺ , but not CD8 ⁺ , T cell proliferation, suggesting a conserved function for gal9 in modulating DC–T cell interactions. Live-cell imaging revealed that galectin-9-depleted DCs fail to establish stable ISs with T cells, resulting in reduced T cell activation and proliferation. Unbiased co-immunoprecipitation and mass spectrometry identified HLA-II as a gal9 binding partner in DCs, and we observed a marked reduction of HLA-II recruitment to the immune synapse in DCs lacking gal9. Conditional gal9 knockout in DCs led to enhanced tumor growth in vivo , compared to their wild-type (WT) counterparts, underscoring a role for gal9 in T cell-dependent anti-tumor immunity. Collectively, this study provides the first detailed account of gal9-mediated HLA-II organization at the synaptic site of DCs, revealing a novel mechanism by which galectins orchestrate immune receptor positioning from within the cytoplasm to enhance CD4 ⁺ T cell activation.