Abstract
The unfolded protein response (UPR) is a conserved cellular response to the accumulation of proteinaceous material in endoplasmic reticulum (ER), active both in health and disease to alleviate cellular stress and improve protein folding. Multiple epiphyseal dysplasia (EDM5) is a genetic skeletal condition and a classic example of an intracellular protein aggregation disease, whereby mutant matrilin-3 forms large insoluble aggregates in the ER lumen, resulting in a specific ‘disease signature’ of increased expression of chaperones and foldases, and alternative splicing of the UPR effector XBP1. Matrilin-3 is expressed exclusively by chondrocytes thereby making EDM5 a perfect model system to study the role of protein aggregation in disease. In order to dissect the role of XBP1 signalling in aggregation-related conditions we crossed a p.V194D Matn3 knock-in mouse model of EDM5 with a mouse line carrying a cartilage specific deletion of XBP1 and analysed the resulting phenotype. Interestingly, the growth of mice carrying the Matn3 p.V194D mutation compounded with the cartilage specific deletion of XBP1 was severely retarded. Further phenotyping revealed increased intracellular retention of amyloid-like aggregates of mutant matrilin-3 coupled with dramatically decreased cell proliferation and increased apoptosis, suggesting a role of XBP1 signalling in protein accumulation and/or degradation. Transcriptomic analysis of chondrocytes extracted from wild type, EDM5, Xbp1-null and compound mutant lines revealed that the alternative splicing of Xbp1 is crucial in modulating levels of protein aggregation. Moreover, through detailed transcriptomic comparison with a model of metaphyseal chondrodysplasia type Schmid (MCDS), an UPR-related skeletal condition in which XBP1 was removed without overt consequences, we show for the first time that the differentiation-state of cells within the cartilage growth plate influences the UPR resulting from retention of a misfolded mutant protein and postulate that modulation of XBP1 signalling pathway presents a therapeutic target for aggregation related conditions in cells undergoing proliferation.
Highlights
The unfolded protein response (UPR) is one of the canonical cellular stress pathways that is triggered by unfolded proteins accumulating in the endoplasmic reticulum (ER) lumen
UPR signals are conveyed through three effector molecules, IRE1, ATF6 and PERK, understanding of which is of utmost importance for the development of novel therapeutic approaches
Using deep phenotyping and extensive transcriptomic analysis we demonstrate that whilst the IRE1/XBP1 pathway is redundant in hypertrophic chondrocyte pathology, it is essential for UPR responses in proliferating cells, making this highly conserved pathway an attractive therapeutic target for a broad range of UPR related conditions
Summary
The unfolded protein response (UPR) is one of the canonical cellular stress pathways that is triggered by unfolded proteins accumulating in the ER lumen. The canonical UPR response is initiated when the chaperone protein BiP dissociates from its three membrane bound receptors, PERK, ATF6 and IRE1 [7]. This dissociation is triggered by the exposed hydrophobic residues of misfolded proteins in the ER lumen and the UPR proceeds along the three signalling pathways, which are further modulated by the levels and duration of the stress [8]. Following dissociation of BiP, PERK dimerises and autophosphorylates, which in turn triggers the phosphorylation of elongation factor eIF2ɑ This leads to attenuation of general protein translation allowing the cell to recover from an abnormal protein load [9]. It is not surprising that the XBP1 branch of the UPR pathway has been
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