Defective autophagy and increased apoptosis contribute toward the pathogenesis of FKRP-associated muscular dystrophies

Abstract

Fukutin-related protein (FKRP) is a glycosyltransferase involved in glycosylation of alpha-dystroglycan (a-DG). Mutations in FKRP are associated with muscular dystrophies (MD) ranging from limb-girdle LGMDR9 to Walker-Warburg Syndrome (WWS), a severe type of congenital MD. Although hypoglycosylation of a-DG is the main hallmark of this group of diseases, a full understanding of the underlying pathophysiology is still missing. Here, we investigated molecular mechanisms impaired by FKRP mutations in pluripotent stem (PS) cell-derived myotubes. FKRP-deficient myotubes show transcriptome alterations in genes involved in extracellular matrix receptor interactions, calcium signaling, PI3K-Akt pathway, and lysosomal function. Accordingly, using a panel of patient-specific LGMDR9 and WWS induced PS cell-derived myotubes, we found a significant reduction in the autophagy-lysosome pathway for both disease phenotypes. In addition, we show that WWS myotubes display decreased ERK1/2 activity and increased apoptosis, which were restored in gene edited myotubes. Our results suggest the autophagy-lysosome pathway and apoptosis may contribute to the FKRP-associated MD pathogenesis.

Modeling Patient-Specific Muscular Dystrophy Phenotypes and Therapeutic Responses in Reprogrammed Myotubes Engineered on Micromolded Gelatin Hydrogels

“…DMD 1023 and DMD 1003 iDRM also differentiated into multi-nucleated myotubes with near-typical myogenic index and myotube width. These results are similar to previous studies that have shown relatively unimpaired fusion in human iPSC–derived myoblasts from DMD ( Caputo et al, 2020 ) or LGMDR9 ( Ortiz-Cordero et al, 2021 ) patients. However, striations in DMD 1023 and DMD 1003 myotubes were qualitatively impaired relative to healthy 1001 myotubes.…”

“…Similar issues related to myofibril immaturity have routinely been observed in myotubes derived from a variety of reprogrammed ( Boularaoui et al, 2018 ) and iPSC-derived myoblasts ( Lainé et al, 2018 ; Rao et al, 2018 ) and likely reduce the baseline and drug-induced differences in the phenotypes of healthy and diseased cells. Overall, the maturity of iDRM-derived myotubes observed in this study was similar or weaker than iPSC-derived myotubes ( Caputo et al, 2020 ; Ortiz-Cordero et al, 2021 ). Of note, however, iDRMs require less time, cost, and expertise to generate compared to iPSC-derived myoblasts, which may be especially beneficial for generating patient-specific muscle tissues in time-sensitive or resource-limited settings.…”

“…LMGD2A/R1 has a more variable presentation, with onset of muscle weakness ranging between 0 and 40 years of age and different rates of progression ( Kramerova et al, 2007 ). DMD -encoded dystrophin and CAPN3 -encoded calpain proteins have both been implicated in contributing to the structural integrity and maintenance of the sarcomere and muscle health ( Blau et al, 1983 ; Muntoni, 2001 ; Kramerova et al, 2007 ; van der Wal et al, 2018 ; Pakula et al, 2019 ; Verhaart et al, 2019 ; Caputo et al, 2020 ; Ortiz-Cordero et al, 2021 ). Therefore, culture platforms that enable sarcomere maturation may help in elucidating the molecular basis of dystrophin and calpain 3 contributions to the organization and maturation of striated skeletal muscle.…”

“…Alternatively, dermal fibroblasts derived from patient skin punches can be reprogrammed to induced pluripotent stem cells (iPSCs) and subsequently differentiated to myoblasts ( van der Wal et al, 2018 ). Although iPSC technology has led to useful models for DMD and other muscle disorders ( Caputo et al, 2020 ; Ortiz-Cordero et al, 2021 ), iPSCs can be difficult and costly to derive and expand ( Speciale et al, 2020 ). A simpler approach is to reprogram dermal fibroblasts directly to myoblasts by expression of MyoD, a master regulator of muscle cell development.…”

Dystroglycanopathy: From Elucidation of Molecular and Pathological Mechanisms to Development of Treatment Methods

“…In addition, a recent study using induced pluripotent stem (iPS) cells derived from FKRP-deficient LGMD2I or WWS patients revealed decreased autophagy and increased apoptosis in myotubes, indicating that alterations in cell homeostasis may be involved in DGpathy pathogenesis [ 53 ]. Effective biomarkers of DGpathy, including miRNA and lncRNA, have not been established to date.…”