The mitogen-activated protein kinase JNK1 suppresses interleukin-3 (IL-3) withdrawal-induced cell death through phosphorylation of the BH3-only pro-apoptotic Bcl-2 family protein Bad at threonine 201 (Thr201). Yet, it is unknown whether JNK1 regulates glycolysis, an important metabolic process that is involved in cell survival and if so, whether the regulation depends on Thr201 phosphorylation of Bad. Here we report that phosphorylation of Bad by JNK1 is required for glycolysis through activation of phosphofructokinase-1 (PFK-1), one of the key enzymes that catalyze glycolysis. Genetic disruption of Jnk1 alleles or silencing of Jnk1 by siRNA abrogates glycolysis induced by growth/survival factors like serum or IL-3. Proteomic analysis identifies PFK-1 as a novel Bad-associated protein. Although the interaction between PFK-1 and Bad is independent of JNK1, Thr201 phosphorylation of Bad by JNK1 is required for PFK-1 activation. Thus, our results provide a novel molecular mechanism by which JNK1 promotes glycolysis for cell survival.