p21 induction under amino acid deprivation requires GCN2 and eIF2α phosphorylation.

<p>A) The 5’ regions of the two known mouse transcript variants of p21 are shown. The start codon for p21 is indicated in green capital letters. Variant 2 contains three 5’ uORFs; the start and stop codons for each are indicated in red, blue, and purple, respectively. B) Western blot analysis of p21 induction in leucine-deprived GCN2^+/+, GCN2^-/-, and eIF2α S51A MEFs. β-actin was used as a loading control. All samples were collected at the same time and the blots were run in parallel. C) Western blot analysis of p21 induction in GCN2^+/+, GCN2^-/-, and eIF2α S51A MEFs irradiated with a dose of 4 Gy. Total eIF2α was used as a loading control. D) Western blot analysis of the reversibility of p21 induction in GCN2^+/+ MEFs. β-tubulin was used as a loading control. E) Western blot analysis of p21 induction in leucine-deprived SQ20B cells. Cells were transfected with either non-targeting siRNA (siNT) or siRNA against GCN2 (siGCN2). β-tubulin was used as a loading control. F) Western blot analysis of p27 induction in leucine-deprived GCN2^+/+, GCN2^-/-, and eIF2α S51A MEFs. β-tubulin was used as a loading control. Data information: Values below blot represent the fold change in total pixel intensity over control of p21, GCN2, or p27 normalized to the loading control for each lane.