Expression of eIF4AI and eIF4AII during C2C12 differentiation.

<p>(<b>A</b>) Phase contrast images of C2C12 cells grown in the presence of DM for the indicated number of days (d). Scale bars represent 50 µm. (<b>B</b>) Western Blot analysis documenting expression levels of the indicated proteins during C2C12 cell differentiation. Long (l.e.) and short (s.e.) exposures of the eIF4AII Western blot are presented. (<b>C</b>) Quantification of changes in eIF4AI and eIF4AII protein levels relative to those obtained on day 0. n = 3±SEM. (<b>D</b>) ³⁵S-methionine/cysteine incorporation into TCA-insoluble protein. C2C12 cells were induced to differentiate and protein extracts were prepared at the indicated time points. Cells were labeled for 30 min and the amount of radiolabeled protein quantitated by TCA precipitation. Values are standardized against total protein content. n = 3±SEM. (<b>E</b>) eIF4AI/II are efficiently incorporated into the eIF4F complex during C2C12 differentiation. m⁷GTP affinity purification of the eIF4F complex from C2C12 cells at the indicated days following induction of differentiation. Western blots to the indicated proteins were performed on an aliquot of input extract (lanes 1–4), GDP eluents (lanes 5–8), and m⁷GTP eluents (lanes 9–12).