SIX1+PAX3+ Identify a Progenitor for Myogenic Lineage Commitment from hPSCs
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE234616 | GEO | 2023/06/14
REPOSITORIES: GEO
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Project description:SIX1+PAX3+ Identify a Progenitor for Myogenic Lineage Commitment from hPSCs
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Project description:Pathogenic mutations in alpha kinase 3 (ALPK3) cause cardiomyopathy and a range of musculoskeletal defects. How ALPK3 mutations result in disease remains unclear and little is known about this atypical kinase. Using a suite of engineered human pluripotent stem cells (hPSCs) we show that ALPK3 localizes to the sarcomere, specifically at the M-Band. Both sarcomeric organization and calcium kinetics were disrupted in ALPK3 deficient hPSC derived cardiomyocytes. Further, cardiac organoids derived from ALPK3 knockout hPSCs displayed reduced force generation. Phosphoproteomic profiling of wildtype and ALPK3 null hPSC derived cardiomyocytes revealed ALPK3-dependant phospho-peptides were enriched for proteins involved in sarcomere function and protein quality control. We demonstrate that ALPK3 binds to the selective autophagy receptor SQSTM1 (Sequestome 1) and is required for the sarcomeric localization of SQSTM1. We propose that ALPK3 is a myogenic kinase with an integral role in the intracellular signaling networks underlying sarcomere maintenance required for continued cardiac contractility.
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Project description:Genome-wide approaches allow investigating the molecular circuitry wiring the genetic and epigenetic programs of human somatic stem cells. Hematopoietic stem/progenitor cells (HSPC) give rise to the different blood cell types; however, the molecular basis of human hematopoietic lineage commitment is poorly characterized. Here, we define the transcriptional and epigenetic profile of human HSPC and early myeloid and erythroid progenitors by a combination of Cap Analysis of Gene Expression (CAGE), ChIP-seq and Moloney leukemia virus (MLV) integration site mapping. Most promoters and transcripts were shared by HSPC and committed progenitors, while enhancers and super-enhancers consistently changed upon differentiation, indicating that lineage commitment is essentially regulated by enhancer elements. A significant fraction of CAGE promoters differentially expressed upon commitment were novel, harbored a chromatin enhancer signature, and may identify promoters and transcribed enhancers driving cell commitment. MLV-targeted genomic regions co-mapped with cell-specific active enhancers and super-enhancers. Expression analyses, together with an enhancer functional assay, indicate that MLV integration can be used to identify bona fide developmentally regulated enhancers. Overall, this study provides an overview of transcriptional and epigenetic changes associated to HSPC lineage commitment, and a novel signature for regulatory elements involved in cell identity.
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Project description:Abstract: Human pluripotent stem cells (hPSCs) can be directed to differentiate into skeletal muscle progenitor cells (SMPCs). However, the myogenic potential of hPSC-SMPCs compared to human fetal or adult satellite cells (Scs) remains unclear. This study demonstrates hPSC-SMPCs derived by commonly used protocols are functionally less mature than freshly-isolated human fetal or adult Scs. We utilized RNA-SEQ of human fetal Scs to identify differentially expressed genes including NGFR, ERBB3, which enriched for MYOD+ or PAX7+ cells. Furthermore, inhibition of TGFβ (TGFβi) improved hPSC-muscle maturation in three independent lines as measured by TEM and expression of myosins. Engraftment of CRISPR/Cas9-corrected Duchenne Muscular Dystrophy (DMD) hiPSC-SMPC subpopulations treated with TGFβi in vivo, increased dystrophin-positive fibers to levels of engrafted cultured fetal Scs in mdx mouse models of DMD. This work provides the first characterization of the developmental status of hPSC-SMPCs and identifies candidates to enable robust myogenic activity in vitro and in vivo.
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