Energy management in hypertrophic chondrocyte under ER stress

Abstract

Perturbation of endoplasmic reticulum (ER) homeostasis is associated with various diseases. In a mouse model (13del-tg) for metaphyseal chondrodysplasia type Schmid (MCDS), misfolded mutant type X collagen proteins accumulate in hypertrophic chondrocytes (HCs) triggering ER stress, and was shown as the molecular basis of the disease. Interestingly, under ER stress, HCs activate a survival signal that also alters the chondrocyte differentiation program in the growth plate, impairing endochondral ossification causing dwarfism. How HCs survive ER stress in this context is unknown. Previously, an altered metabolic machinery favoring glycolysis in 13del-tg HCs was shown using a label-free quantitative proteomic approach. Recently, transcriptome analysis identified a marked upregulated expression of FGF21, a novel ER stress target and a key metabolic regulator involved in various metabolic diseases, in 13del HCs. However, the role of FGF21 in energy flux maintenance for HCs survival under ER stress has not been studied. In this study I employed Label-free Quantitative proteomics to obtain insights of FGF21’s role in energy management in both normal and ER stress experiencing HCs through systematic comparison between the proteomes in WT and FGF21null HCs, and 13del and 13delFGF21null HCs. A total of 2,106 and 2,369 proteins were identified in the proteome comparison in WT vs FGF21null, and 13del vs 13delFGF21null respectively. In normal HCs, my data analysis showed that systemic FGF21 maintains fatty acid synthesis protein levels, for instance Fatty acid synthase and Acetyl-CoA Carboxylase; normal HCs proteomic analysis rated “Fatty acyl-CoA biosynthesis” the top altered pathway suggesting fatty acid biosynthesis capacity was reduced in FGF21null HCs, but FGF21 deficiency does not affect chondrocyte differentiation as confirmed by histological and immunohistochemical analysis. In 13del HCs, my data analysis showed that FGF21 upregulates glycolytic proteins, for instance, Glyceraldehyde-3-phosphate dehydrogenase, Phosphoglycerate kinase and Phosphoglycerate mutase, and pro-survival protein 14-3-3 protein family; 13del HCs proteomic analysis indicated “Glycolysis” and “Activation of BAD and translocation to mitochondria” as significantly altered pathway suggesting glycolytic capacity was reduced while apoptotic program was enhanced in 13delFGF21null HCs. This suggests that FGF21 has a key role in the switch in activating and/or sustaining an up-regulated glycolytic process under ER stress for survival, and built evidence to previous hypothesis that 13del HCs capture rapid energy flux from glycolysis to survive ER stress. Altogether this study provides evidence that rapid energy supply is necessary to sustain pro-survival pathways for HCs to survive ER stress, and FGF21 mediates the survival strategy in part through glycolytic protein upregulation. My data also provide evidence that FGF21 functions are context dependent. Integrating previous data with my findings suggest a possible non-canonical FGF21 signaling pathway in HCs under ER stress. In summary, my findings support a switch to rapid energy supply is necessary to sustain pro-survival pathways in HCs to survive under ER stress, and it is in part mediated through FGF21’s metabolic regulatory function, suggesting a potential implication of FGF21 to ER stress related diseases.