Lysophosphatidic acid (LPA) is a potent lipid mediator with various biological functions mediated through six G protein–coupled receptors (GPCRs), LPA1–LPA6. Previous studies have demonstrated that LPA–Gα12/Gα13 signaling plays an important role in embryonic vascular development. However, the responsible LPA receptors and underlying mechanisms are poorly understood. Here, we show a critical role of LPA4 and LPA6 in developmental angiogenesis. In mice, Lpa4;Lpa6 double-knockout (DKO) embryos were lethal due to global vascular deficiencies, and endothelial cell–specific (EC-specific) Lpa4;Lpa6-DKO retinas had impaired sprouting angiogenesis. Mechanistically, LPA activated the transcriptional regulators YAP and TAZ through LPA4/LPA6–mediated Gα12/Gα13–Rho–ROCK signaling in ECs. YAP/TAZ knockdown increased endothelial expression of the Notch ligand delta-like ligand 4 (DLL4) that was mediated by β-catenin and Notch intracellular domain (NICD). Fibrin gel sprouting assay revealed that LPA4/LPA6, Gα12/Gα13, or YAP/TAZ knockdown consistently blocked EC sprouting, which was rescued by a Notch inhibitor. Notably, the inhibition of Notch signaling also ameliorated impaired retinal angiogenesis in EC-specific Lpa4;Lpa6-DKO mice. Overall, these results suggest that the Gα12/Gα13–coupled receptors LPA4 and LPA6 synergistically regulate endothelial Dll4 expression through YAP/TAZ activation. This could in part account for the mechanism of YAP/TAZ–mediated developmental angiogenesis. Our findings provide insight into the biology of GPCR-activated YAP/TAZ.

Molecular chaperones promote the folding and macromolecular assembly of a diverse set of ‘client’ proteins. How ubiquitous chaperone machineries direct their activities towards specific sets of substrates is unclear. Through the use of mouse genetics, imaging and quantitative proteomics we uncover that ZMYND10 is a novel co-chaperone that confers specificity for the FKBP8-HSP90 chaperone complex towards axonemal dynein clients required for cilia motility. Loss of ZMYND10 perturbs the chaperoning of axonemal dynein heavy chains, triggering broader degradation of dynein motor subunits. We show that pharmacological inhibition of FKBP8 phenocopies dynein motor instability associated with the loss of ZMYND10 in airway cells and that human disease-causing variants of ZMYND10 disrupt its ability to act as an FKBP8-HSP90 co-chaperone. Our study indicates that primary ciliary dyskinesia (PCD), caused by mutations in dynein assembly factors disrupting cytoplasmic pre-assembly of axonemal dynein motors, should be considered a cell-type specific protein-misfolding disease.

BACKGROUND: The heart comprises many types of cells such as cardiomyocytes, endothelial cells (ECs), fibroblasts, smooth muscle cells, pericytes, and blood cells. Every cell type responds to various stressors (eg, hemodynamic overload and ischemia) and changes its properties and interrelationships among cells. To date, heart failure research has focused mainly on cardiomyocytes; however, other types of cells and their cell-to-cell interactions might also be important in the pathogenesis of heart failure. METHODS: Pressure overload was imposed on mice by transverse aortic constriction and the vascular structure of the heart was examined using a tissue transparency technique. Functional and molecular analyses including single-cell RNA sequencing were performed on the hearts of wild-type mice and EC-specific gene knockout mice. Metabolites in heart tissue were measured by capillary electrophoresis-time of flight-mass spectrometry system. The vaccine was prepared by conjugating the synthesized epitope peptides with keyhole limpet hemocyanin and administered to mice with aluminum hydroxide as an adjuvant. Tissue samples from heart failure patients were used for single-nucleus RNA sequencing to examine gene expression in ECs and perform pathway analysis in cardiomyocytes. RESULTS: Pressure overload induced the development of intricately entwined blood vessels in murine hearts, leading to the accumulation of replication stress and DNA damage in cardiac ECs. Inhibition of cell proliferation by a cyclin-dependent kinase inhibitor reduced DNA damage in ECs and ameliorated transverse aortic constriction–induced cardiac dysfunction. Single-cell RNA sequencing analysis revealed upregulation of Igfbp7 (insulin-like growth factor-binding protein 7) expression in the senescent ECs and downregulation of insulin signaling and oxidative phosphorylation in cardiomyocytes of murine and human failing hearts. Overexpression of Igfbp7 in the murine heart using AAV9 (adeno-associated virus serotype 9) exacerbated cardiac dysfunction, while EC-specific deletion of Igfbp7 and the vaccine targeting Igfbp7 ameliorated cardiac dysfunction with increased oxidative phosphorylation in cardiomyocytes under pressure overload. CONCLUSIONS: Igfbp7 produced by senescent ECs causes cardiac dysfunction and vaccine therapy targeting Igfbp7 may be useful to prevent the development of heart failure.

Genetically modified mouse models are essential for in vivo investigation of gene function and human disease research. Targeted mutations can be introduced into mouse embryos using genome editing technology such as CRISPR-Cas. Although mice with small indel mutations can be produced, the production of mice carrying large deletions or gene fragment knock-in alleles remains inefficient. We introduced the nuclear localisation property of Cdt1 protein into the CRISPR-Cas system for efficient production of genetically engineered mice. Mouse Cdt1-connected Cas9 (Cas9-mC) was present in the nucleus of HEK293T cells and mouse embryos. Cas9-mC induced a bi-allelic full deletion of Dmd, GC-rich fragment knock-in, and floxed allele knock-in with high efficiency compared to standard Cas9. These results indicate that Cas9-mC is a useful tool for producing mouse models carrying targeted mutations.