Quantitative Interactomics of Lck-TurboID in Living Human T Cells Unveils T Cell Receptor Stimulation-Induced Proximal Lck Interactors

While
Lck has been widely recognized to play a pivotal role in
the initiation of the T cell receptor (TCR) signaling pathway, an
understanding of the precise regulation of Lck in T cells upon TCR
activation remains elusive. Investigation of protein–protein
interaction (PPI) using proximity labeling techniques such as TurboID
has the potential to provide valuable molecular insights into Lck
regulatory networks. By expressing Lck-TurboID in Jurkat T cells,
we have uncovered a dynamic, short-range Lck protein interaction network
upon 30 min of TCR stimulation. In this novel application of TurboID,
we detected 27 early signaling-induced Lck-proximal interactors in
living T cells, including known and novel Lck interactors, validating
the discovery power of this tool. Our results revealed previously
unappreciated Lck PPI which may be associated with cytoskeletal rearrangement,
ubiquitination of TCR signaling proteins, activation of the mitogen-activated
protein kinase cascade, coalescence of the LAT signalosome, and formation
of the immunological synapse. In this study, we demonstrated for the
first time in immune cells and for the kinase Lck that TurboID can
be utilized to unveil PPI dynamics in living cells at a time scale
consistent with early TCR signaling. Data are available via ProteomeXchange with identifier PXD020759.