The primary cilium is required for Hedgehog (Hh) signaling in vertebrates. Hh leads to ciliary accumulation and activation of the transmembrane protein Smoothened (Smo) and affects the localization of several pathway components, including the Gli family of transcriptional regulators, within different regions of primary cilia. Genetic analysis indicates that the kinesin protein Kif7 both promotes and inhibits mouse Hh signaling. Using mass spectrometry, we identified liprin-α1 (PPFIA1) and the protein phosphatase PP2A as Kif7-interacting proteins, and we showed that they were important for the trafficking of Kif7 and Gli proteins to the tips of cilia and for the transcriptional output of Hh signaling. Our results suggested that PPFIA1 functioned with PP2A to promote the dephosphorylation of Kif7, triggering Kif7 localization to the tips of primary cilia and promoting Gli transcriptional activity.

SUMMARYDevelopment of the heart requires recruitment of cardiovascular progenitor cells (CPCs) to the future heart-forming region. CPCs are the building blocks of the heart, and have the potential to form all the major cardiac lineages. However, little is known regarding what regulates CPC fate and behavior. Activity of GATA4, SMARCD3 and TBX5 -the 'cardiac BAF' (cBAF) complex, can promote myocardial differentiation in embryonic mouse mesoderm. Here, we exploit the advantages of the zebrafish embryo to gain mechanistic understanding of cBAF activity. Overexpression of smarcd3b and gata5 in zebrafish results in an enlarged heart, whereas combinatorial loss of cBAF components inhibits cardiac differentiation. In transplantation experiments, cBAF acts cell autonomously to promote cardiac fate. Remarkably, cells overexpressing cBAF migrate to the developing heart and differentiate as cardiomyocytes, endocardium and smooth muscle. This is observed even in host embryos that lack endoderm or cardiac mesoderm. Our results reveal an evolutionarily conserved role for cBAF activity in cardiac differentiation. Importantly, they demonstrate that Smarcd3b and Gata5 can induce a primitive, CPC-like state. KEY WORDS: Cardiovascular progenitor, Cell fate, ZebrafishSmarcd3b and Gata5 promote a cardiac progenitor fate in the zebrafish embryo Mounting evidence indicates that the initiation of cell differentiation is highly regulated at the epigenetic level. In vertebrates, the Swi/Snf-like multi-subunit BAF chromatin remodeling complex plays a key role in modifying DNA-histone contacts (Kwon et al., 1994). Work in ES cells and neurons has shown that the BAF complex can engage in a number of cellspecific events via differential incorporation of subunit variants, providing a 'combinatorial code' of activity (Ho et al., 2009;Yoo et al., 2009). Smarcd3/Baf60c, which encodes a variant Smarcd/Baf60 subunit, plays an essential role in early murine heart development (Lickert et al., 2004). Recently, lipofection of mouse embryos with Smarcd3/Gata4/Tbx5 was shown to promote myocardial differentiation of non-cardiogenic mesoderm (Takeuchi and Bruneau, 2009). The authors demonstrated that these three proteins bind to each other and form part of a cardiac BAF (termed cBAF) complex that synergistically upregulate the expression of myocardial-specific genes. Smarcd3 was further shown to recruit Gata4 to target genes, possibly acting as a 'pioneer' factor to effect myocardial differentiation. Although these recent cardiac reprogramming experiments illustrate that myocardial 'reprogramming' of cells is feasible, they do not address the in vivo mechanisms through which the earliest events of cardiac differentiation are initiated.Here, we exploit the advantages of the zebrafish embryo to study cBAF activity in vivo. We find that Gata5/Smarcd3b activity promotes myocardial development, and that combined loss of gata5, smarcd3b and tbx5 results in pronounced defects in cardiogenesis. Via transplantation, we find that cBAF plays a cell autonomous role i...

The Apelin receptor (Aplnr) is essential for heart development, controlling the early migration of cardiac progenitors. Here we demonstrate that in zebrafish Aplnr modulates Nodal/TGFβ signaling, a key pathway essential for mesendoderm induction and migration. Loss of Aplnr function leads to a reduction in Nodal target gene expression whereas activation of Aplnr by a non-peptide agonist increases the expression of these same targets. Furthermore, loss of Aplnr results in a delay in the expression of the cardiogenic transcription factors mespaa/ab. Elevating Nodal levels in aplnra/b morphant and double mutant embryos is sufficient to rescue cardiac differentiation defects. We demonstrate that loss of Aplnr attenuates the activity of a point source of Nodal ligands Squint and Cyclops in a non-cell autonomous manner. Our results favour a model in which Aplnr is required to fine-tune Nodal output, acting as a specific rheostat for the Nodal/TGFβ pathway during the earliest stages of cardiogenesis.DOI: http://dx.doi.org/10.7554/eLife.13758.001

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