Glyceraldehyde 3-phosphate dehydrogenase depletion induces cell cycle arrest and resistance to antimetabolites in human carcinoma cell lines.
Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) is a multifunctional protein that acts at the intersection of energy metabolism and stress response in tumor cells. To elucidate the role of GAPDH in chemotherapy-induced stress, we analyzed its activity, protein level, intracellular distribution, and intranuclear mobility in human carcinoma cells A549 and UO31 after treatment with cytarabine, doxorubicin, and mercaptopurine. After treatment with cytosine arabinoside (araC), enzymatically inactive GAPDH accumulated in the nucleus. Experiments on fluorescence recovery after photobleaching with green fluorescent protein-GAPDH fusion protein in the live cells treated with araC demonstrated reduced mobility of green fluorescent protein-GAPDH inside the nucleus, indicative of interactions with nuclear macromolecular components after genotoxic stress. Depletion of GAPDH with RNA interference stopped cell proliferation, and induced cell cycle arrest in G(1) phase via p53 stabilization, and accumulation of p53-inducible CDK inhibitor p21. Neither p21 accumulation nor cell cycle arrest was detected in GAPDH-depleted p53-null NCI-H358 cells. GAPDH-depleted A549 cells were 50-fold more resistant to treatment with cytarabine (1.68 +/- 0.182 microM versus 0.03 +/- 0.015 microM in control). Depletion of GAPDH did not significantly alter cellular sensitivity to doxorubicin (0.05 +/- 0.023 microM versus 0.035 +/- 0.0154 microM in control). Induction of cell cycle arrest in p53-proficient carcinoma cells via GAPDH abrogation suggests that GAPDH-depleting agents may have a cytostatic effect in cancer cells. Our results define GAPDH as an important determinant of cellular sensitivity to antimetabolite chemotherapy because of its regulatory functions.
Phadke
Manali S
MS
Temple University School of Pharmacy, Philadelphia, PA 19140, USA.
Krynetskaia
Natalia F
NF
Mishra
Anurag K
AK
Krynetskiy
Evgeny
E
D000964:Antimetabolites, Antineoplastic / Q000494:pharmacology*; D001374:Azacitidine / Q000031:analogs & derivatives / Q000494:pharmacology; D002277:Carcinoma / Q000188:drug therapy / Q000201:enzymology* / Q000473:pathology; D002453:Cell Cycle / Q000187:drug effects* / Q000235:genetics; D016923:Cell Death / Q000187:drug effects / Q000235:genetics; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D019008:Drug Resistance, Neoplasm / Q000187:drug effects* / Q000235:genetics; D055785:Gene Knockdown Techniques / Q000379:methods; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008175:Lung Neoplasms / Q000188:drug therapy / Q000201:enzymology / Q000473:pathology
D000964:Antimetabolites, Antineoplastic; C022121:fazarabine; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D001374:Azacitidine