Mitochondrial respiratory chain function promotes extracellular matrix integrity in cartilage

Kristina Bubb,Tatjana Holzer,Janica L Nolte,Marcus Krüger,Richard Wilson,Ursula Schlötzer-Schrehardt,Jürgen Brinckmann,Janine Altmüller,Attila Aszodi,Lutz Fleischhauer,Hauke Clausen-Schaumann,Kristina Probst,Bent Brachvogel

doi:10.1016/j.jbc.2021.101224

Abstract

Energy metabolism and extracellular matrix (ECM) function together orchestrate and maintain tissue organization, but crosstalk between these processes is poorly understood. Here, we used single-cell RNA-Seq (scRNA-Seq) analysis to uncover the importance of the mitochondrial respiratory chain for ECM homeostasis in mature cartilage. This tissue produces large amounts of a specialized ECM to promote skeletal growth during development and maintain mobility throughout life. A combined approach of high-resolution scRNA-Seq, mass spectrometry/matrisome analysis, and atomic force microscopy was applied to mutant mice with cartilage-specific inactivation of respiratory chain function. This genetic inhibition in cartilage results in the expansion of a central area of 1-month-old mouse femur head cartilage, showing disorganized chondrocytes and increased deposition of ECM material. scRNA-Seq analysis identified a cell cluster–specific decrease in mitochondrial DNA–encoded respiratory chain genes and a unique regulation of ECM-related genes in nonarticular chondrocytes. These changes were associated with alterations in ECM composition, a shift in collagen/noncollagen protein content, and an increase of collagen crosslinking and ECM stiffness. These results demonstrate that mitochondrial respiratory chain dysfunction is a key factor that can promote ECM integrity and mechanostability in cartilage and presumably also in many other tissues.

Highlights

A large amount of energy is required to drive skeletal growth and synthesize the structural components of the cartilage extracellular matrix (ECM) that withstand mechanical forces and maintain lifelong mobility
The results indicate that distinct chondrocyte subpopulations show a higher degree of mitochondrial respiratory chain (mtRC) deficiency, leading to subpopulationspecific histomorphological changes in proximal femoral epiphysis (PFE) cartilage of CreTW mice
Impaired mtRC leads to a cell subpopulation–specific ECMdamage response and an altered mechanostability in PFE cartilage

Summary

Introduction

A large amount of energy is required to drive skeletal growth and synthesize the structural components of the cartilage extracellular matrix (ECM) that withstand mechanical forces and maintain lifelong mobility. We recently induced the expression of the mitochondrial (mt) DNA Twinkle (TW) helicase mutant K320E [2, 3] in growth plate cartilage by Col2a1-directed expression of Cre recombinase (Cre; [4, 5]) to generate mice with impaired mtRC function in chondrocytes (CreTW) (Fig. 1A). ScRNA-Seq analysis could confirm the cluster-specific expression of genes in growth plate cartilage [11] originally defined by oligoarraytranscriptome profiling [12] This technology could be suited to correlate activated ECM-related genetic programs in chondrocyte subpopulations with impaired mtRC activity, but to our knowledge, scRNA-Seq was not yet applied to highresolution transcriptome analysis of mutant mice with a cartilage phenotype. The aim of this study was to define the individual cellular load of mtRC dysfunction and define the impact on ECM homeostasis and cartilage stability using a combined approach of high-resolution scRNA-Seq, mass spectrometry/matrisome analysis, and atomic force microscopy (AFM) in mutant mice with cartilage-specific inactivation of mtRC function

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Conclusion