A modified Chromium 10x droplet-based protocol that subsamples cells for both short-read and long-read (nanopore) sequencing together with a new computational pipeline (FLAMES) is developed to enable isoform discovery, splicing analysis, and mutation detection in single cells. We identify thousands of unannotated isoforms and find conserved functional modules that are enriched for alternative transcript usage in different cell types and species, including ribosome biogenesis and mRNA splicing. Analysis at the transcript level allows data integration with scATAC-seq on individual promoters, improved correlation with protein expression data, and linked mutations known to confer drug resistance to transcriptome heterogeneity.

Due to its essential role in movement, insulating the internal organs, generating heat to maintain core body temperature, and acting as a major energy storage depot, any impairment to skeletal muscle structure and function may lead to an increase in both morbidity and mortality. In the context of skeletal muscle, altered metabolism is directly associated with numerous pathologies and disorders, including diabetes, and obesity, while many skeletal muscle pathologies have secondary changes in metabolism, including cancer cachexia, sarcopenia and the muscular dystrophies. Furthermore, the importance of cellular metabolism in the regulation of skeletal muscle stem cells is beginning to receive significant attention. Thus, it is clear that skeletal muscle metabolism is intricately linked to the regulation of skeletal muscle mass and regeneration. The aim of this review is to discuss some of the recent findings linking a change in metabolism to changes in skeletal muscle mass, as well as describing some of the recent studies in developmental, cancer and stem-cell biology that have identified a role for cellular metabolism in the regulation of stem cell function, a process termed “metabolic reprogramming.”

While metabolism was initially thought to play a passive role in cell biology by generating ATP to meet bioenergetic demands, recent studies have identified critical roles for metabolism in the generation of new biomass and provision of obligate substrates for the epigenetic modification of histones and DNA. This review details how metabolites generated through glycolysis and the tricarboxylic acid cycle are utilized by somatic stem cells to support cell proliferation and lineage commitment. Importantly, we also discuss the evolving hypothesis that histones can act as an energy reservoir during times of energy stress. Finally, we discuss how cells integrate both extrinsic metabolic cues and intrinsic metabolic machinery to regulate cell fate.

Myostatin (MSTN) is a transforming growth factor-β (TGF-β) family member that normally acts to limit muscle growth. The function of MSTN is partially redundant with that of another TGF-β family member, activin A. MSTN and activin A are capable of signaling through a complex of type II and type I receptors. Here, we investigated the roles of two type II receptors (ACVR2 and ACVR2B) and two type I receptors (ALK4 and ALK5) in the regulation of muscle mass by these ligands by genetically targeting these receptors either alone or in combination specifically in myofibers in mice. We show that targeting signaling in myofibers is sufficient to cause significant increases in muscle mass, showing that myofibers are the direct target for signaling by these ligands in the regulation of muscle growth. Moreover, we show that there is functional redundancy between the two type II receptors as well as between the two type I receptors and that all four type II/type I receptor combinations are utilized in vivo. Targeting signaling specifically in myofibers also led to reductions in overall body fat content and improved glucose metabolism in mice fed either regular chow or a high-fat diet, demonstrating that these metabolic effects are the result of enhanced muscling. We observed no effect, however, on either bone density or muscle regeneration in mice in which signaling was targeted in myofibers. The latter finding implies that MSTN likely signals to other cells, such as satellite cells, in addition to myofibers to regulate muscle homeostasis.