eIF4A inhibition changes the polysomal association of certain mRNAs.
Cultured cells (SH-SY5Y, N2a or HeLa) were treated with 10 µM hippuristanol (+) or DMSO control (−) for 10 minutes, lysed and mRNA fractionated on a 10%–50% sucrose gradient. OD260nm absorbance was determined with simultaneous 1 ml fraction collection to determine subpolysomal and polysomal fractions (fraction number and subpolysomal or polysomal association indicated). Northern analysis was performed to determine the positions of mRNAs for: A Actin and PABP which do not change in polysome associations; B APP and tau which become less polysomally associated; C TXN, NDUFB2 and SOD1 which become more polysomally associated. Relative levels of RNA are indicated by dark grey (DMSO control) and light grey (10 µM hippuristanol) bars beneath each fraction, as percentages of the total amount RNA in each gradient.