Ovalbumin antigen-specific activation of T cell receptor closely resembles soluble antibody stimulation as revealed by BOOST phosphotyrosine proteomics
Activation of T cell receptors (TCR) leads to a network of early signaling predominantly orchestrated by tyrosine phosphorylation in T cells. TCR are commonly activated using soluble anti-TCR antibodies, but this approach is not antigen-specific. Alternatively, activating the TCR using specific antigens of a range of binding affinities in the form of peptide-major histocompatibility complex (pMHC) is presumed to be more physiological. However, due to the lack of wide-scale phosphotyrosine (pTyr) proteomic studies directly comparing anti-TCR antibodies and pMHC, a comprehensive definition of these activated states remains enigmatic. Elucidation of the tyrosine phosphoproteome using quantitative pTyr proteomics enables a better understanding of the unique features of these activating agents and the role of ligand binding affinity on signaling. Here, we apply the recently established Broad-spectrum Optimization Of Selective Triggering (BOOST) to examine perturbations in tyrosine phosphorylation of TCR triggered by anti-TCR antibodies and pMHC. Our data reveals that high-affinity ovalbumin (OVA) pMHC activation of the TCR triggers a largely similar, albeit potentially stronger, pTyr-mediated signaling regulatory axis compared to anti-TCR antibody. Signaling output resulting from OVA pMHC variants correlates well with their weaker affinities, enabling affinity-tunable control of signaling strength. Collectively, we provide a framework for applying BOOST to compare pTyr-mediated signaling pathways of T cells activated in an antigen-independent and antigen-specific manner. Abstract Graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=108 SRC="FIGDIR/small/436968v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@152f03aorg.highwire.dtl.DTLVardef@19a6e2org.highwire.dtl.DTLVardef@1b48b6org.highwire.dtl.DTLVardef@142ced6_HPS_FORMAT_FIGEXP M_FIG C_FIG