Abstract
Collagen VI (ColVI) is an abundant and distinctive extracellular matrix protein secreted by fibroblasts in different tissues. Human diseases linked to mutations on ColVI genes are primarily affecting skeletal muscle due to non-cell autonomous myofiber defects. To date, it is not known whether and how fibroblast homeostasis is affected by ColVI deficiency, a critical missing information as this may strengthen the use of patients’ fibroblasts for preclinical purposes. Here, we established primary and immortalized fibroblast cultures from ColVI null (Col6a1-/-) mice, the animal model of ColVI-related diseases. We found that, under nutrient-stringent condition, lack of ColVI affects fibroblast survival, leading to increased apoptosis. Moreover, Col6a1-/- fibroblasts display defects in the autophagy/lysosome machinery, with impaired clearance of autophagosomes and failure of Parkin-dependent mitophagy. Col6a1-/- fibroblasts also show an increased activation of the Akt/mTOR pathway, compatible with the autophagy impairment, and adhesion onto purified ColVI elicits a major effect on the autophagic flux. Our findings reveal that ColVI ablation in fibroblasts impacts on autophagy regulation and cell survival, thus pointing at the new concept that this cell type may contribute to the pathological features of ColVI-related diseases.
Highlights
The extracellular matrix (ECM) microenvironment plays a crucial role for tissue homeostasis and ensures proper functionality through physical interactions and structural support
Recent studies revealed that some ECM proteins are Abbreviations: 3-MA, 3-methyladenine; AMPK, AMP-activated protein kinase; BM, Bethlem myopathy; CCCP, carbonyl cyanide m-chlorophenylhydrazone; Collagen VI (ColVI), collagen VI; CQ, chloroquine; ECM, extracellular matrix; Erk1/2, extracellular signal-regulated kinases 1 and 2; LAMP-2, lysosomal-associated membrane protein 2; MAP1LC3B/LC3, microtubuleassociated protein 1 light chain 3B; murine embryonic fibroblasts (MEF), mouse embryonic fibroblasts; mTOR, mammalian target of rapamycin; PI, propidium iodide; Raptor, regulatory-associated protein of TOR; rpS6, ribosomal protein S6; SQSTM1/p62, sequestosome 1; TFEB, transcription factor EB; UCMD, Ullrich congenital muscular dystrophy; Ulk1, Unc-51 like autophagy activating kinase 1
In skeletal muscle ColVI is produced by fibroblasts, and not by myofibers (Braghetta et al, 2008; Zou et al, 2008), muscles represent the most affected tissue in human pathologies caused by mutations of COL6 genes, such as BM and UCMD (Bönnemann, 2011; Bushby et al, 2014)
Summary
The ECM microenvironment plays a crucial role for tissue homeostasis and ensures proper functionality through physical interactions and structural support. Several ECM components are involved in key cellular pathways other than the mere cell-matrix adhesion, such as activation and presentation of soluble factors, preservation of the stem cell niche, and signal transduction into cells (Hynes, 2009; Gattazzo et al, 2014). Extra- and intracellular nutrient and growth factor availability regulates autophagy by means of a complex machinery of interactors that enable a balanced adaptive response (Feng et al, 2014). Both defective and excessive activation of autophagy are detrimental for cell homeostasis and result in human degenerative pathologies (Levine and Kroemer, 2008; Schneider and Cuervo, 2014). The roles of the ECM on autophagy regulation are increasingly evident, still under dissection (Neill et al, 2014; Buraschi et al, 2017)
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