Overexpression of GAPDH followed by its translocation into particulate fractions and the protective effect of antisense oligonucleotides to GAPDH against cell death in primary thymocytes. (A) After Dex stimulation (3 μM), GAPDH levels in total extracts are greatly elevated as early as 3 hr after stimulation. Modest cytosolic overexpression of GAPDH at 3 hr is followed at 6 hr by particulate augmentation, especially in LP. Twenty micrograms of proteins were loaded on each lane. (B) Antisense oligonucleotides (5 μM) to GAPDH reduce the augmented GAPDH protein level after Dex. The antisense oligonucleotide was added 6 hr before Dex stimulation, and cell extracts were taken 12 hr after treatment. Five micrograms of proteins were loaded on each lane. Con, without treatment; Dex, only Dex addition; Dex+AS, antisense treatment prior to Dex addition. (C) Antisense oligonucleotides to GAPDH protect, with levels increased half of the way to control levels. By contrast, GAPDH sense oligonucleotides have no effect.