The Gata4/5/6 sub-family of zinc finger transcription factors regulate many aspects of cardiogenesis. However, critical roles in extra-embryonic endoderm also challenge comprehensive analysis during early mouse cardiogenesis, while zebrafish models have previously relied on knockdown assays. We generated targeted deletions to disrupt each gata4/5/6 gene in zebrafish and analyzed cardiac phenotypes in single, double and triple mutants. The analysis confirmed that loss of gata5 causes cardia bifida and validated functional redundancies for gata5/6 in cardiac precursor specification. Surprisingly, we discovered that gata4 is dispensable for early zebrafish development, while loss of one gata4 allele can suppress the bifid phenotype of the gata5 mutant. The gata4 mutants eventually develop an age-dependent cardiomyopathy. By combining combinations of mutant alleles, we show that cardiac specification depends primarily on an overall dosage of gata4/5/6 alleles rather than a specific gene. We also identify a specific role for gata6 in controlling ventricle morphogenesis through regulation of both the first and second heart field, while loss of both gata4/6 eliminates the ventricle. Thus, different developmental programs are dependent on total dosage, certain pairs, or specific gata4/5/6 genes during embryonic cardiogenesis.This article has an associated First Person interview with the first author of the paper.

Highlights d Over 47,000 small molecules were screened for activation of ALP in C2C12 cells d DIPQUO was identified, also promoting osteogenesis in human MSCs and zebrafish d Functional activity is associated specifically with activation of p38-b d DIPQUO is a promising lead candidate for development of bone therapeutics

SUMMARYAimed movements require that an animal accurately locates the target and correctly reaches that location. One such behavior is the defensive strike seen in Manduca sexta larva. These caterpillars respond to noxious mechanical stimuli applied to their abdomen with a strike of the mandibles towards the location of the stimulus. The accuracy with which the first strike movement reaches the stimulus site depends on the location of the stimulus. Reponses to dorsal stimuli are less accurate than those to ventral stimuli and the mandibles generally land ventral to the stimulus site. Responses to stimuli applied to anterior abdominal segments are less accurate than responses to stimuli applied to more posterior segments and the mandibles generally land posterior to the stimulus site. A trade-off between duration of the strike and radial accuracy is only seen in the anterior stimulus location (body segment A4). The lower accuracy of the responses to anterior and dorsal stimuli can be explained by the morphology of the animal; to reach these areas the caterpillar needs to move its body into a tight curve. Nevertheless, the accuracy is not exact in locations that the animal has shown it can reach, which suggests that consistently aiming more ventral and posterior of the stimulation site might be a defense strategy. Supplementary material available online at

SUMMARY Ten-eleven translocation (Tet) enzymes (Tet1/2/3) mediate 5-methylcytosine (5mC) hydroxylation, which can facilitate DNA demethylation and thereby impact gene expression. Studied mostly for how mutant isoforms impact cancer, the normal roles for Tet enzymes during organogenesis are largely unknown. By analyzing compound mutant zebrafish, we discovered a requirement for Tet2/3 activity in the embryonic heart for recruitment of epicardial progenitors, associated with development of the atrial-ventricular canal (AVC). Through a combination of methylation, hydroxymethylation, and transcript profiling, the genes encoding the activin A subunit Inhbaa (in endocardium) and Sox9b (in myocardium) were implicated as demethylation targets of Tet2/3 and critical for organization of AVC-localized extracellular matrix (ECM), facilitating migration of epicardial progenitors onto the developing heart tube. This study elucidates essential DNA demethylation modifications that govern gene expression changes during cardiac development with striking temporal and lineage specificities, highlighting complex interactions in multiple cell populations during development of the vertebrate heart.

Highlights d Zebrafish with compound tet2/3 mutations show defects in neutrophil maturation d Defects are associated with altered cytokine signaling and increased socs3b mRNA d The socs3b mRNA rather than the gene itself is hypermethylated d The data further support Tets as epitranscriptomic regulatory proteins