The transmembrane domain of the adenovirus E3/19K protein acts as an ER retention signal and contributes to intracellular sequestration of MHC class I molecules

摘要

The human adenovirus E3/19K-protein is a type-I transmembrane glycoprotein of the endoplasmic reticulum (ER) that abrogates cell-surface transport of MHC class-I (MHC-I) and MICA/B-molecules. Previous data suggested that E3/19K comprises two functional modules: a luminal domain for interaction with MHC-I and MICA/B-molecules, and a di-lysine motif in the cytoplasmic tail that confers retrieval from the Golgi back to the ER. This study was prompted by the unexpected phenotype of an E3/19K-molecule that was largely retained intracellularly despite having a mutated ER-retrieval motif. To identify additional structural determinants responsible for ER-localization, chimeric molecules were generated containing the luminal E3/19K-domain and the cytoplasmic and/or transmembrane domain (TMD) of the cell-surface protein MHC-I Kd. These were analysed for transport, cell-surface expression and impact on MHC-I/MICA/B down-regulation. Similar to the retrieval mutant, replacing the cytoplasmic tail of E3/19K allowed only limited transport of the chimera to the cell surface. Efficient cell-surface expression was only achieved by additionally replacing the TMD of E3/19K with that of MHC-I, suggesting that the E3/19K-TMD may confer static ER-retention. This was verified by ER-retention of an MHC-I Kd molecule with the TMD replaced by that of E3/19K. Thus, we have identified the E3/19K TMD as a novel functional element that mediates static ER-retention, thereby increasing its ER-concentration. Remarkably, the ER-retrieval signal alone without E3/19K-TMD did not mediate efficient HLA-down regulation, even in the context of infection. This suggests that the TMD is required together with the ER-retrieval function to ensure efficient ER-localization and transport inhibition of MHC-I and MIC-A/B-molecules.