Abstract
T-cell receptor-CD3 complex (TCR) is a versatile signaling machine that can initiate antigen-specific immune responses based on various biochemical changes of CD3 cytoplasmic domains, but the underlying structural basis remains elusive. Here we developed biophysical approaches to study the conformational dynamics of CD3ε cytoplasmic domain (CD3εCD). At the single-molecule level, we found that CD3εCD could have multiple conformational states with different openness of three functional motifs, i.e., ITAM, BRS and PRS. These conformations were generated because different regions of CD3εCD had heterogeneous lipid-binding properties and therefore had heterogeneous dynamics. Live-cell imaging experiments demonstrated that different antigen stimulations could stabilize CD3εCD at different conformations. Lipid-dependent conformational dynamics thus provide structural basis for the versatile signaling property of TCR.
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Acknowledgements
CX is funded by the Chinese Academy of Sciences (Strategic Priority Research Program XDB08020100), and the National Natural Science Foundation of China (31370860, 31425009, 31530022 and 31621003). HL is funded by the Ministry of Science and Technoloy of China (2014CB541903) and the National Natural Science Foundation of China (31470734). HW is funded by the Ministry of Science and Technoloy of China (2011CB933600) and the National Natural Science Foundation of China (21373200). XG is funded by China Postdoctoral Science Foundation (2015M580357). NMR experiments, part of AFM and imaging experiments were performed at the National Center for Protein Science Shanghai. Part of imaging experiments was performed at the core facility for cell biology of Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences.
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( Supplementary information is linked to the online version of the paper on the Cell Research website.)
Supplementary information
Supplementary information, Figure S1
Plasma membrane sheet (PMS) preparation and rupture force measurements. (PDF 287 kb)
Supplementary information, Figure S2
The mechanical signature of the PEG linker and the polypeptide. (PDF 190 kb)
Supplementary information, Figure S3
AFM data analysis with different data filtering criteria. (PDF 281 kb)
Supplementary information, Figure S4
Analysis of the two-peak events when pulling from the N-terminus of the hCD3εCD WT peptide. (PDF 195 kb)
Supplementary information, Figure S5
Histograms of the rupture force values obtained for peptides shown in Figure 3F, respectively. (PDF 169 kb)
Supplementary information, Figure S6
Multiple kinetic intermediates between the closed and open conformations of the CD3ε cytoplasmic domain. (PDF 182 kb)
Supplementary information, Figure S7
The estimation of the free energy barrier with different theoretical models. (PDF 271 kb)
Supplementary information, Figure S8
The expression, purification of hCD3εTMCD proteins and reconstitution of hCD3εTMCD into lipid bicelle. (PDF 450 kb)
Supplementary information, Figure S9
Quenching of the PRE effect of TEMPOL by Ascorbic Acid. (PDF 114 kb)
Supplementary information, Figure S10
Expression and localization of HA-mCD3ε (YY-FF)-mTFP in mouse OT-I T cells. (PDF 206 kb)
Supplementary information, Figure S11
T-cell receptor activation model. (PDF 76 kb)
Supplementary information, Data S1
Materials and Methods (PDF 161 kb)
Supplementary information, Table S1
The fitting parameters for the three types of potential shapes with different scaling factors. (PDF 97 kb)
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Guo, X., Yan, C., Li, H. et al. Lipid-dependent conformational dynamics underlie the functional versatility of T-cell receptor. Cell Res 27, 505–525 (2017). https://doi.org/10.1038/cr.2017.42
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DOI: https://doi.org/10.1038/cr.2017.42
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