The transmembrane domains of the prM and E proteins of yellow fever virus are endoplasmic reticulum localization signals.

The immature flavivirus particle contains two envelope proteins, prM and E, that are associated as a heterodimer. Virion morphogenesis of the flaviviruses occurs in association with endoplasmic reticulum (ER) membranes, suggesting that there should be accumulation of the virion components in this compartment. This also implies that ER localization signals must be present in the flavivirus envelope proteins. In this work, we looked for potential subcellular localization signals in the yellow fever virus envelope proteins. Confocal immunofluorescence analysis of the subcellular localization of the E protein in yellow fever virus-infected cells indicated that this protein accumulates in the ER. Similar results were obtained with cells expressing only prM and E. Chimeric proteins containing the ectodomain of CD4 or CD8 fused to the transmembrane domains of prM or E were constructed, and their subcellular localization was studied by confocal immunofluorescence and by analyzing the maturation of their associated glycans. Although a small fraction was detected in the ER-to-Golgi intermediate and Golgi compartments, these chimeric proteins were located mainly in the ER. The C termini of prM and E form two antiparallel transmembrane alpha-helices. Interestingly, the first transmembrane passage contains enough information for ER localization. Taken altogether, these data indicate that, besides their role as membrane anchors, the transmembrane domains of yellow fever virus envelope proteins are ER retention signals. In addition, our data show that the mechanisms of ER retention of the flavivirus and hepacivirus envelope proteins are different.