Abstract
Mesencephalic astrocyte-derived neurotrophic factor (MANF) is an endoplasmic reticulum (ER) resident protein that can be secreted due to an imperfect KDEL motif. MANF plays a cytoprotective role in several soft tissues and is upregulated in conditions resulting from intracellular retention of mutant protein, including two skeletal diseases, metaphyseal chondrodysplasia, Schmid type (MCDS) and multiple epiphyseal dysplasia (MED). The role of MANF in skeletal tissue homeostasis is currently unknown. Interestingly, cartilage-specific deletion of Manf in a mouse model of MED resulted in increased disease severity, suggesting its upregulation may be chondroprotective. Treatment of MED chondrocytes with exogenous MANF led to a decrease in the cellular levels of BiP (GRP78), confirming MANF’s potential to modulate ER stress responses. However, it did not alleviate the intracellular retention of mutant matrilin-3, suggesting that it is the intracellular MANF that is of importance in the pathobiology of skeletal dysplasias. The Col2Cre-driven deletion of Manf from mouse cartilage resulted in a chondrodysplasia-like phenotype. Interestingly, ablation of MANF in cartilage did not have extracellular consequences but led to an upregulation of several ER-resident chaperones including BiP. This apparent induction of ER stress in turn led to dysregulated chondrocyte apoptosis and decreased proliferation, resulting in reduced long bone growth. We have previously shown that ER stress is an underlying disease mechanism for several skeletal dysplasias. The cartilage-specific deletion of Manf described in this study phenocopies our previously published chondrodysplasia models, further confirming that ER stress itself is sufficient to disrupt skeletal growth and thus represents a potential therapeutic target.
Highlights
Mesencephalic astrocyte-derived neurotrophic factor (MANF) is an endoplasmic reticulum (ER)-stress associated protein that was initially identified as a neuroprotective factor in dopaminergic neurons (Hellman et al 2011; Petrova et al 2003)
Global deletion of Manf leads to perinatal lethality due to breathing difficulties induced by lung malformation
The clones were injected into C57BL/6 blastocysts resulting in a extracellular matrix components, type II collagen, type X collagen and matrilin-3 in Manffl/fl Col2Cre− and Col2Cre+ growth plates at P21. b 2 h BrdU labelling of proliferation in Manffl/fl Col2Cre− and Col2Cre+ growth plates at P21 showing a 29% decrease in chondrocyte proliferation upon deletion of MANF (n = 5). c TUNEL assay per zone in Manffl/fl Col2Cre− and Col2Cre+ growth plates at P21 (n = 3). d Total apoptosis in Manffl/fl Col2Cre− and Col2Cre+ growth plates at P21 (n = 3)
Summary
Mesencephalic astrocyte-derived neurotrophic factor (MANF) is an endoplasmic reticulum (ER)-stress associated protein that was initially identified as a neuroprotective factor in dopaminergic neurons (Hellman et al 2011; Petrova et al 2003). MANF is an ER-resident protein; it contains an imperfect KDEL sequence that allows it to be secreted under ER stress, suggesting it could be used as a biomarker for a subset of ER stress-related conditions (Henderson et al 2013; Oh-Hashi et al 2012). Exogenous MANF has been shown to have cytoprotective effects, potentially exerted via cell surface KDEL receptors (Henderson et al 2013), and works in conjunction with the canonical ER chaperone BiP (GRP78). Data on the effect of exogenous MANF on BiP expression suggests that this relationship might be cell or stress-type specific (Apostolou et al 2008; Huang et al 2016; Zhao et al 2013)
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