Molecular modelling and endoplasmic reticulum retention of mutated TCR/CD3 complexes

Area of Science:

• Immunology
• Molecular Biology
• Cell Biology

Background:

• T cell receptor (TCR)/CD3 complex assembly initiates in the endoplasmic reticulum (ER).
• Proper TCR/CD3 complex transport to the cell surface involves interaction with zeta2 homodimers in the cis-Golgi.
• Mutations can impact TCR/CD3 expression and cell surface transport.

Purpose of the Study:

• To investigate the molecular basis of Phe195/216 --> Val mutations on TCR/CD3 expression in Jurkat cells.
• To determine why mutated TCRalphabeta/CD3 hexamers in J79 cells fail to interact with zeta2 homodimers.
• To elucidate the mechanism of ER retention for mutated TCR/CD3 complexes.

Main Methods:

• Analysis of hydrophobic interactions within TCR/CD3 domains affected by Phe --> Val mutations.
• Confocal microscopy to assess colocalization of TCR/CD3 hexamers and zeta2 homodimers in various cell lines.
• Investigation of ER retention mechanisms for mutated TCR/CD3 complexes.

Main Results:

• Phe --> Val mutations significantly alter a hydrophobic region in Calpha and Cbeta domains of TCR/CD3.
• Mutated TCR/CD3 hexamers in J79 cells do not colocalize with zeta2 homodimers, unlike in normal or rescued cells.
• Mutated TCR/CD3 complexes are actively retained within the ER in J79 cells via a nondominant mechanism.

Conclusions:

• The identified mutations disrupt critical hydrophobic interactions necessary for TCR/CD3 complex assembly and zeta2 homodimer binding.
• Mutated TCR/CD3 complexes are retained in the ER, preventing their transport to the cell surface.
• A novel, undefined ER-retention molecule likely controls TCR/CD3 complex structure and ER egress in both alphabeta and gammadelta T cells.