Figure 4. eIF4E-dependent rescue of yeast translation.

Yeast strain Jo55 lacks a chromosomal eIF4E gene and only survives on galactose-containing media due to the presence of vector YCp33supex-h4E URA3, which encodes human eIF4E under the control of a glucose-repressible, galactose-dependent promoter. This strain was subsequently transformed with empty vector YCpTRP-GW (negative control), or the same vector containing the coding sequence of human eIF4E (positive control) and pea eIF4E derivatives under the control of the constitutive TEF1 promoter. Transformed cells were grown in selective drop-out media (SD −Trp −Ura+galactose) to an optical density (OD₆₀₀) of 1.0 before being serially diluted and assessed for growth on glucose- and galactose-containing media. Three independent experiments were performed for each eIF4E variant and translation complementation was scored as the relative level of cell growth. This ranged in amount from, for example, (++) for eIF4E^S, (+) for eIF4E^R to no observable growth (−) for eIF4E^W75A.