Glycolytic flux-signaling controls mouse embryo mesoderm development

Abstract

How cellular metabolic state impacts cellular programs is a fundamental, unresolved question. Here we investigated how glycolytic flux impacts embryonic development, using presomitic mesoderm (PSM) patterning as the experimental model. First, we identified fructose 1,6-bisphosphate (FBP) as an in vivo sentinel metabolite that mirrors glycolytic flux within PSM cells of post-implantation mouse embryos. We found that medium-supplementation with FBP, but not with other glycolytic metabolites, such as fructose 6-phosphate and 3-phosphoglycerate, impaired mesoderm segmentation. To genetically manipulate glycolytic flux and FBP levels, we generated a mouse model enabling the conditional overexpression of dominant active, cytoplasmic PFKFB3 (cytoPFKFB3). Overexpression of cytoPFKFB3 indeed led to increased glycolytic flux/FBP levels and caused an impairment of mesoderm segmentation, paralleled by the downregulation of Wnt-signaling, reminiscent of the effects seen upon FBP-supplementation. To probe for mechanisms underlying glycolytic flux-signaling, we performed subcellular proteome analysis and revealed that cytoPFKFB3 overexpression altered subcellular localization of certain proteins, including glycolytic enzymes, in PSM cells. Specifically, we revealed that FBP supplementation caused depletion of Pfkl and Aldoa from the nuclear-soluble fraction. Combined, we propose that FBP functions as a flux-signaling metabolite connecting glycolysis and PSM patterning, potentially through modulating subcellular protein localization.

Glycolysis–Wnt signaling axis tunes developmental timing of embryo segmentation

“…Increasing glucose led to a slower segmentation clock (Fig. 1C), which we also observed when fructose 1,6-bisphosphate (FBP), a glycolytic sentinel metabolite [6, 27], was supplemented to the medium (Fig. 1D).…”

“…We limited our analysis to the tailbud region, where the clock period phenotype is most apparent. Combined with the dataset from our previous study [6], this analysis revealed 617 flux-responsive differentially expressed genes (DEGs) that were either upregulated (Cluster (C) 1 and C3) or downregulated (C2, C4, and C5) by increasing glycolytic flux (Fig. 2A, Supplementary Table 1).…”

“…In the PSM, changes in central carbon metabolism impact Wnt signaling [3–6] and the period of the segmentation clock [7]. In particular, glycolysis has been shown to establish an activity gradient from the posterior to anterior PSM [3, 4], being functionally linked to graded signaling activity within the mouse PSM [4–6]. Furthermore, it has been shown that active glycolysis is required for maintaining the segmentation clock oscillation [3].…”

“…We tackled this question in the context of mouse embryonic axis segmentation. Previous studies have shown that changes in central carbon metabolism impact Wnt signaling [3–6] and the period of the segmentation clock [7], which controls the timing of axis segmentation. Here, we reveal that glycolysis tunes the segmentation clock period in an anti-correlated manner: higher glycolytic flux slows down the clock, and vice versa.…”

Integrated Molecular-Phenotypic Profiling Reveals Metabolic Control of Morphological Variation in Stembryos

“…On the contrary, in mouse embryo explants glycolysis was reported to negatively regulate WNT signalling. 38,45 Although the authors suggested that the discrepancy could be rooted in short-term negative 38 versus long-term positive regulation 34 , our data show that -at the same time-point -higher glycolytic activity is positively associated with expression of WNTs and their targets. Our discovery that the FGF/WNT/glycolysis connection is conserved in the scalable and tractable stembryos presents an exciting opportunity to unravel the intricate spatiotemporal dynamics of feedback.…”