Translational deregulation in PDK-1-/- embryonic stem cells

PDK-1 is a protein kinase that is critical for the activation of many downstream protein kinases in the AGC superfamily, through phosphorylation of the activation loop site on these substrates. Cells lacking PDK- I show decreased activity of these protein kinases, including protein kinase B (PKB) and p70(S6K), whereas mTOR activity remains largely unaffected. Here we show, by assessing both association of cellular RNAs with polysomes and by metabolic labeling, that PDK-1(-/-) embryonic stem (ES) cells exhibit defects in mRNA translation. We identify which mRNAs are most dramatically translationally regulated in cells lacking PDK-1 expression by performing microarray analysis of total and polysomal RNA in these cells. In addition to the decreased translation of many RNAs, a smaller number of RNAs show increased association with polyribosomes in PDK-1(-/-) ES cells relative to PDK-1(+/+) ES cells. We show that PKB activity is a critical downstream component of PDK-1 in mediating translation of cystatin C, RANKL, and Rab11a, whereas mTOR activity is less important for effective translation of these targets.