Biallelic variants in the transcription factor PAX7 are a new genetic cause of Myopathy

Orfi, Z¹ ; Dort, J¹ ; Fabre, P¹ ; Molina, T¹ ; Conte, T¹ ; Pellerito, O¹ ; Campeau, P¹ ; and Dumont, N^1,2.

1-University of Montreal, CHU St Justine Research Center, Montreal, QC, Canada
2-University of Montreal, School of Rehabilitation, Faculty of Medicine, Montreal, QC, Canada

Genetic defect on satellite cells will trigger their self-renewal capacity and their differentiation ability, which in turn will have drastic consequences on skeletal muscle growth, regeneration and development. Specific transcription factors, particularly PAX7, are key regulators of the function of these cells. The consequences of lack of PAX7 in humans have not been yet established. Here we generated iPSC-derived myoblast from a patient with homozygous nonsense variants in the PAX7gene, and their homologue controls. Using a well-established in-vitro differentiation protocol, we have assessed the function of iPSC-derived myoblasts (proliferation, differentiation, and fusion) by immunofluorescence for different myogenic markers (PAX3, PAX7, MYOD1, MYOG, MYH3) to determine how myogenesis is affected by PAX7 variants. Moreover, single cell RNA sequencing analysis was performed tocomprehend the mechanism of PAX7 action on human muscle stem cell self-renewal ability and identity process. Although PAX7 wasnot mandatory for myogenesis during in-vitro iPSC to myoblast differentiation, its implication on muscle stem cell self-renewal and identity is more significant during regeneration. Therefore, we propose to generate 3D engineered skeletal muscle tissue from iPSC and induce cardiotoxin injury in order to monitor muscle stem cell self-renewal capacity during regeneration in ex-vivo model. Finally, we have also transplanted iPSC-derived myoblasts from patient and controls in immuno-suppressed mdx mice to assess how PAX7 is implicated in regeneration and muscle stem cell identity in-vivo. Our findings are the first to clearly establish causality between pathogenic variants in PAX7 and a new genetic cause of myopathy.