PAB/WT polysome loading and transcript levels (Arabidopsis thaliana)

To understand the contribution of the poly(A)binding protein to the translation of specific mRNAs, we compared the ribosome occupancy of mRNAs in wild type Arabidopsis and pab2 pab8 double mutant seedlings. The mutants continue to express the PAB4 paralog of PABP. RNA was fractionated using sucrose gradients into polysomal and nonpolysomal RNAs. We also determined overall total transcript levels. We used Affymetrix ATH1 microarrays. Each plant sample was analyzed for the mRNA abundance in total mRNA (T), polysomal mRNA (PL), and nonpolysomal mRNA (NP). Four biological replicates were collected for polysomes and three for total RNA. The pab2 pab8 double mutant was compared with wild type.