Nuclear reprogramming sets in motion dedifferentiation processes leading to acquisition of pluripotency. This study resolved the metabolic attributes underpinning iPSC generation. Metaboproteome restructuring was demonstrated to underlie conversion from somatic oxidative bioenergetics to glycolysis of pluripotent progeny. Promotion of a glycolytic flux-dependent, mitochondria-independent metabotype fostered nuclear reprogramming.
Regression of the parental somatic network of abundant tubular and cristae-rich mitochondria into sparse spherical and cristae-poor structures characterized iPSC progeny. A mitochondria-poor capacity would enable iPSC to revert reactive oxygen species-dependent telomere shortening associated with cellular aging (Passos et al., 2007
; Marion et al., 2009
). In fact, mitochondrial regression and reduction of mtDNA as a result of nuclear reprogramming is consistent with the undeveloped mitochondrial morphology of ESC (Kondoh et al., 2007
; Armstrong et al., 2010
; Prigione et al., 2010
; Zeuschner et al., 2010
). Induced reduction in mitochondria number and maturity suggests metamorphosis of the bioenergetic infrastructure underlying dedifferentiation.
Metabolomic footprinting and fingerprinting indeed demonstrated distinct features of iPSC metabolism. The key metabolic rates contributing to the iPSC phenotype were consistent among multiple clones, and included elevated utilization of glucose and accumulation of intracellular and extracellular lactate (via lactate dehydrogenase) and acetate (via acetyl-CoA synthase or ATP-citrate lyase), suggesting that accelerated glycolysis results in accumulation of metabolic byproducts within the cell, followed by metabolite exportation. iPSC demonstrated diminished basal oxygen consumption and uncoupled oxidative capacity, indicating departure from oxidative metabolism. ESC also rely on glycolytic ATP generation (Cho et al., 2006
; Facucho-Oliveira et al., 2007
), with pluripotency maintenance sustained under hypoxic conditions (Ezashi et al., 2005
). Acquisition of an embryonic-like metabolic state is concomitant with global transcriptional and epigenetic reorganization that includes demethylation and activation of the promoter region of pluripotency inductors (Bock et al., 2011
; Plath and Lowry, 2011
). A glycolytic burst could promote linkage of energy metabolism with nuclear reprogramming-induced epigenetic resetting (Huangfu et al., 2008
; Friis et al., 2009
; Wellen et al., 2009
; Teperino et al., 2010
), priming the cell for pluripotent induction. Indeed, expression of glycolytic genes preceded pluripotent gene expression, implicating metabolic remodeling as a facilitator of nuclear reprogramming.
Complete oxidation of glucose limits generation of substrates for biosynthetic pathways. Transition to glycolytic metabolism would fuel specific cellular processes, such as ATP consumption in the cell nucleus and provide the necessary balance between ATP generation and production of metabolic precursors to meet anabolic requirements of dedifferentiation and associated proliferation (DeBerardinis et al., 2008
; Chung et al., 2010
). Strategies that in part stimulate glycolysis, including induction of hypoxia or inhibition of the p53 pathway, increase nuclear reprogramming efficiency (Kruse and Gu, 2006
; Krizhanovsky and Lowe, 2009
; Yoshida et al., 2009
). Consistent with the ability of small molecules with known metabolic targets to modulate the appearance of pluripotent colonies (Zhu et al., 2010
), here stimulation of glycolytic flux potentiated reprogramming whereas oxidative metabolism promotion preserved the somatic cytotype.
c-Myc, through its regulation of both glycolysis and mitochondrial metabolism, is a key contributor to the metabolic Warburg effect, reflecting the tendency of highly proliferative cell types to convert glucose to lactate despite sufficient oxygen for mitochondrial oxidative metabolism (Dang, 2007
). Here 3F iPSC displayed equivalent metabolic profiles to 4F iPSC counterparts, verifying that nuclear reprogramming-induced metabolic remodeling did not require c-Myc transduction. Proteomic dissection resolved a metabolic infrastructure that distinguished 3F and 4F iPSC from fibroblasts. Specifically, reprogramming induced reciprocal changes in glycolytic enzymes and electron transport chain components, indicating a molecular substrate contributing to the switch from oxidative to glycolytic metabolism. The observed decreased expression of complex I and II subunits would uncouple oxidative metabolism and ATP generation, favoring glycolytic dependent ATP production.
In summary, dedifferentiation of parental somatic cells induced mitochondrial regression, electron transport chain downregulation, and glycolysis enzyme upregulation, revealing a pluripotent glycolytic signature with limited dependence on mitochondrial metabolism. Induction of the glycolytic metabotype support anabolic and catabolic requirements for bona fide pluripotency to regulate cell fate.