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Imaging of T-cell receptor fused to CD3ζ reveals enhanced expression and improved pairing in living cells

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T cell receptor (TCR) gene adoptive therapy is a promising clinical approach for the treatment of malignant tumors and viral diseases. However, the effectiveness of this strategy is hampered by the generation of mixed TCR heterodimers comprising both exogenous and endogenous TCR chains (i.e., mispairing of TCR chains). In the present study, we constructed genetically encoded reporters fused to a pair of fluorescent proteins [enhanced cyan fluorescent protein (ECFP)/enhanced yellow fluorescent protein (EYFP)] to monitor the expression of TCRαζβζ and pairing between TCRαζ and TCRβζ. We demonstrate that these reporters provide accurate images of TCRαζβζ expression, which is markedly stronger with evident microclusters accumulated at the plasma membrane compared to wild-type (wt)TCR. Using fluorescence resonance energy transfer (FRET) analysis, we demonstrate that, in addition to the improved pairing, the expression and assembly of TCRαζβζ are independent of endogenous CD3 subunits. These results suggest that the fusion genes, TCRαζ and TCRβζ, coupled to ECFP and EYFP, respectively can effectively monitor the expression and interaction in cells. Our data suggest a novel strategy with which it is possible to effectively express and pair TCRαζβζ, thus making TCR gene adoptive therapy more effective.
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Document Type: Research Article

Affiliations: 1: Institute of Genetic Engineering, Southern Medical University, Guangzhou, Guangdong 510515, P.R. China 2: Guangdong Province Key Laboratory for Biotechnology Drug Candidates, School of Life Science and Biopharmaceutics, Guangdong Pharmaceutical University, Guangzhou, Guangdong 510006, P.R. China

Publication date: January 1, 2014

More about this publication?
  • The International Journal of Molecular Medicine is a monthly, peer-reviewed journal devoted to the publication of high quality studies related to the molecular mechanisms of human disease. The journal welcomes research on all aspects of molecular and clinical research, ranging from biochemistry to immunology, pathology, genetics, human genomics, microbiology, molecular pathogenesis, molecular cardiology, molecular surgery and molecular psychology.

    The International Journal of Molecular Medicine aims to provide an insight for researchers within the community in regard to developing molecular tools and identifying molecular targets for the diagnosis and treatment of a diverse number of human diseases.
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