Spatial distribution of surface proteins in single T cells and in T-B cell conjugates 3523

Abstract Description Surface proteins mediate cellular functions, migration and cell-cell interactions, shaping cellular behavior. Investigating their spatial distribution and receptor colocalization (proximity to one another) can provide key insights into these processes. However, studies at single-cell resolution have previously been limited by low-dimensional assays. Here, we applied the recently developed Molecular Pixelation technique to study spatial distributions, abundances and colocalizations of 74 surface proteins in 9,324 single CD4+ T cells during differentiation (Th1, Th2, Th17 and Treg) and T-B cell conjugates during adaptive responses following vaccination in lymphoid organoids. We find consistent colocalization of tetraspanins CD37 and CD82 across all T cell subsets, with enhanced colocalization in T-B conjugates. These tetraspanins colocalize with different partner receptors depending on T cell subset, i.e. CD40L in Th1 cells and CTLA4 in Treg. Furthermore, CD82 localizes to specific membrane regions, suggesting the CD37-CD82 platform resides in distinct areas of the cell surface to regulate cell function. On T-B conjugates, T cells show increased colocalization between CD5 and CD7, indicating a coordinated interaction between CD5-mediated inhibition and CD7-mediated stimulation to ensure controlled T and B cell activation. Our study provides novel insights into context dependent spatial distributions of receptors that suggest an additional layer of T cell biology to study further. Topic Categories Computational and Systems Immunology (COMP)