Signalling

Signalling by the TCR controls T cell activation. We focus on the signalling mechanisms by the TCR.

The scaffold protein Kidins220

In two independent proteomics screens we have found the transmembrane scaffold protein Kidins220/ARMS to bind to the TCR and to the BCR. So far Kidins220 was mostly studied in the nervous system and is a very large scaffold protein. We could show that Kidins220 binds to the resting TCR and BCR and couples these receptors to the Ras-Raf-Erk signalling pathway (Fiala et al, 2015; Deswal et al, 2013). A B cell-specific knock out has shown that Kidins220 promotes all developmental steps where pre-BCR or BCR signalling is required and that it enhances B cell activation. Rearrangement of the lambda light chain locus is particularly sensitive to the absence of Kidins220 (manuscript accepted in eLife).

Very recently we have shown that Kidins220 also controls TCR signalling strength and thus development of invariant natural killer T (iNKT) cells (manuscript accepted in Science Advances).

CD3 conformational changes

In cooperation with the lab of Balbino Alarcon in Madrid we have discovered that the TCR undergoes a conformational change at the CD3 subunits upon peptide-MHC binding  (Gil et al, 2002). This CD3 conformational change (CD3 CC) exposes the proline-rich region in CD3epsilon, so that it is able to bind to the adapter protein Nck. In cooperation with the Pongcharoen group we have shown that Nck is a crucial protein in TCR signalling (Hartl et al, 2021; Paensuwan et al, 2016).

We could further show that the CD3 CC is necessary for T cell activation and that only dimeric antigens can induce this change (Minguet et al, 2007). Mechanistically the TCR spontaneously switches its conformation, also in the absence of any ligand. Cholesterol keeps the inactive-active conformation equilibrium on the side of the inactive TCRs, by only binding and thus stabilizing the inactive TCR. In contrast, antigen only binds bivalently to the TCRs in the active conformation, thus stabilising active TCRs. This is a new model of TCR functioning (Schamel et al, 2019; Minguet et al, 2007).

Finally, we discovered how the kinase LCK is recruited to the TCR. In the active, lignad-bound conformation a new motif, the RK motif, is exposed. This allows binding of LCK with the LCK SH3 domain. Appending the RK motif to a 2nd generation chimeric antigen receptor (CAR) improved the anti-tumoral activity of the CAR-expressing T cells in a pre-clinical mouse model (Hartl et al, 2020).

References

Hartl FA, Ngoenkam J, Beck-Garcia E, Cerqueira L, Wipa P, Paensuwan P, Suriyaphol P, Mishra P, Schraven B, Günther S, et al (2021) Cooperative Interaction of Nck and Lck Orchestrates Optimal TCR Signaling. Cells 10

Hartl FA, Beck-Garcìa E, Woessner NM, Flachsmann LJ, Cárdenas RMHV, Brandl SM, Taromi S, Fiala GJ, Morath A, Mishra P, et al (2020) Noncanonical binding of Lck to CD3ε promotes TCR signaling and CAR function. Nat Immunol 21: 902–913

Schamel WW, Alarcon B & Minguet S (2019) The TCR is an allosterically regulated macromolecular machinery changing its conformation while working. Immunol Rev 291: 8–25

Paensuwan P, Hartl FA, Yousefi OS, Ngoenkam J, Wipa P, Beck-Garcia E, Dopfer EP, Khamsri B, Sanguansermsri D, Minguet S, et al (2016) Nck Binds to the T Cell Antigen Receptor Using Its SH3.1 and SH2 Domains in a Cooperative Manner, Promoting TCR Functioning. J Immunol 196: 448–58

Fiala GJ, Janowska I, Prutek F, Hobeika E, Satapathy A, Sprenger A, Plum T, Seidl M, Dengjel J, Reth M, et al (2015) Kidins220/ARMS binds to the B cell antigen receptor and regulates B cell development and activation. J Exp Med 212: 1693–708

Deswal S, Meyer A, Fiala GJ, Eisenhardt AE, Schmitt LC, Salek M, Brummer T, Acuto O & Schamel WWA (2013) Kidins220/ARMS associates with B-Raf and the TCR, promoting sustained Erk signaling in T cells. J Immunol 190: 1927–35

Minguet S, Swamy M, Alarcón B, Luescher IF & Schamel WW (2007) Full activation of the T cell receptor requires both clustering and conformational changes at CD3. Immunity 26: 43–54

Gil D, Schamel WW, Montoya M, Sánchez-Madrid F & Alarcón B (2002) Recruitment of Nck by CD3 epsilon reveals a ligand-induced conformational change essential for T cell receptor signaling and synapse formation. Cell 109: 901–12