Fig. 10

Polysome profile analysis revealed that eIF4E3_A associates with translation intiation complexes and light polysome fractions. GFP-eIF4E1 (a) and GFP-eIF4E3_A (c) stably expressed in HEK293 cells are distributed along polysome profiles similarly to endogenous eIF4E1 (b). To evaluate possible influence of a GFP-fusion tag, a polysome profile analysis from HEK293 cells stably expressing GFP alone was performed (b). Western blots of the first nine fractions were probed either with anti-eIF4E1 antibody (a, b) or anti-GFP antibody (b, c). Highest amounts of GFP-eIF4E1, GFP-eIF4E3_A and endogenous eIF4E1 were detected from the end of the loading peak to the light polysomes, whereas GFP protein alone was detected in the early loading peak exclusively. Loading peak (≈fractions 1–3); 40S, 60S and 80S (≈fractions 4–6 in a, b and 3–5 in c); light and heavy polysomes (≈fractions 7–10 in a, b and 6–10 in c)