Spatial transcriptomics in embryonic mouse diaphragm muscle reveals regional gradients and subdomains of developmental gene expression.

The murine embryonic diaphragm is a primary model for studying myogenesis and neuro-muscular synaptogenesis, both representing processes regulated by spatially organized genetic programs of myonuclei located in distinct myodomains. However, a spatial gene expression pattern of embryonic mouse diaphragm has not been reported. Here, we provide spatially resolved gene expression data for horizontally sectioned embryonic mouse diaphragms at embryonic days E14.5 and E18.5. These data reveal gene signatures for specific muscle regions with distinct maturity and fiber type composition, as well as for a central neuromuscular junction (NMJ) and a peripheral myotendinous junction (MTJ) compartment. Comparing spatial expression patterns of wild-type mice with those of transgenic mice lacking either the skeletal muscle calcium channel CaV1.1 or ß-catenin, reveals curtailed muscle development and dysregulated expression of genes potentially involved in NMJ formation. Altogether, these datasets provide a powerful resource for further studies of muscle development and NMJ formation in the mouse.

Asymmetric contribution of a selectivity filter gate in triggering inactivation of CaV1.3 channels.

Voltage-dependent and Ca2+-dependent inactivation (VDI and CDI, respectively) of CaV channels are two biologically consequential feedback mechanisms that fine-tune Ca2+ entry into neurons and cardiomyocytes. Although known to be initiated by distinct molecular events, how these processes obstruct conduction through the channel pore remains poorly defined. Here, focusing on ultrahighly conserved tryptophan residues in the interdomain interfaces near the selectivity filter of CaV1.3, we demonstrate a critical role for asymmetric conformational changes in mediating VDI and CDI. Specifically, mutagenesis of the domain III-IV interface, but not others, enhanced VDI. Molecular dynamics simulations demonstrate that mutations in distinct selectivity filter interfaces differentially impact conformational flexibility. Furthermore, mutations in distinct domains preferentially disrupt CDI mediated by the N- versus C-lobes of CaM, thus uncovering a scheme of structural bifurcation of CaM signaling. These findings highlight the fundamental importance of the asymmetric arrangement of the pseudotetrameric CaV pore domain for feedback inhibition.