MicroRNAs (miRNAs) are genomically encoded small RNAs that hybridize with messenger RNAs, resulting in degradation or translational inhibition of targeted transcripts. The potential for miRNAs to regulate cell-lineage determination or differentiation from pluripotent progenitor or stem cells is unknown. Here, we show that microRNA1 (miR-1) is an ancient muscle-specific gene conserved in sequence and expression in Drosophila. Drosophila miR-1 (dmiR-1) is regulated through a serum response factor-like binding site in cardiac progenitor cells. Loss-and gain-of-function studies demonstrated a role for dmiR-1 in modulating cardiogenesis and in maintenance of muscle-gene expression. We provide in vivo evidence that dmiR-1 targets transcripts encoding the Notch ligand Delta, providing a potential mechanism for the expansion of cardiac and muscle progenitor cells and failure of progenitor cell differentiation in some dmiR-1 mutants. These findings demonstrate that dmiR-1 may ''fine-tune'' critical steps involved in differentiation of cardiac and somatic muscle progenitors and targets a pathway required for progenitor cell specification and asymmetric cell division.Delta ͉ microRNA ͉ progenitor cells ͉ stem cells ͉ cardiogenesis

Cyclic guanosine monophosphate (cGMP) is a second messenger molecule that transduces nitric oxide (NO) and natriuretic peptide (NP) coupled signaling, stimulating phosphorylation changes by protein kinase G (PKG). Enhancing cGMP synthesis or blocking its degradation by phosphodiesterase type 5A (PDE5A) protects against cardiovascular disease1,2. However, cGMP stimulation alone is limited by counter-adaptions including PDE upregulation3. Furthermore, though PDE5A regulates NO-generated cGMP4,5, NO-signaling is often depressed by heart disease6. PDEs controlling NP-coupled cGMP remain uncertain. Here we show that cGMP-selective PDE9A7,8 is expressed in mammalian heart including humans, and is upregulated by hypertrophy and cardiac failure. PDE9A regulates NP rather than NO-stimulated cGMP in heart myocytes and muscle, and its genetic or selective pharmacological inhibition protects against pathological responses to neuro-hormones, and sustained pressure-overload stress. PDE9A inhibition reverses pre-established heart disease independent of NO-synthase (NOS) activity, whereas PDE5A inhibition requires active NOS. Transcription factor activation and phospho-proteome analyses of myocytes with each PDE selectively inhibited reveals substantial differential targeting, with phosphorylation changes from PDE5A inhibition being more sensitive to NOS activation. Thus, unlike PDE5A, PDE9A can regulate cGMP signaling independent of the NO-pathway, and its role in stress-induced heart disease suggests potential as a therapeutic target.

Guiding multipotent cells into distinct lineages and controlling their expansion remain fundamental challenges in developmental and stem cell biology. Members of the Wnt pathway control many pivotal embryonic events, often promoting self-renewal or expansion of progenitor cells. In contrast, canonical Wnt ligands are thought to negatively regulate cardiomyogenesis in several species. However, the cell-autonomous role of canonical Wnt signaling within precardiac mesoderm, through its obligatory transcriptional mediator, ␤-catenin, is unknown. Using tissue-specific in vivo genetic manipulation, we found that ␤-catenin is required for development of cardiac progenitors and is a positive regulator of proliferative expansion of such progenitor cells. At discrete windows of development in embryonic stem cells, activation of canonical Wnt signaling promoted expansion of cardiac progenitors after initial commitment and was required for cardiac differentiation. Together, these data provide in vivo and in vitro evidence that canonical Wnt signaling promotes the expansion of cardiac progenitors and differentiation of cardiomyocytes.cardiac development ͉ embryonic stem cells ͉ ␤-catenin ͉ second heart field

Cellular decisions of self-renewal or differentiation arise from integration and reciprocal titration of numerous regulatory networks. Notch and Wnt/β-Catenin signaling often intersect in stem and progenitor cells and regulate one another transcriptionally. The biological outcome of signaling through each pathway often depends on the context and timing as cells progress through stages of differentiation. Here, we show that membrane-bound Notch physically associates with unphosphorylated (active) β-Catenin in stem and colon cancer cells and negatively regulates post-translational accumulation of active β-Catenin protein. Notch-dependent regulation of β-Catenin protein did not require ligand-dependent membrane cleavage of Notch or the glycogen synthase kinase-3β-dependent activity of the β-catenin destruction complex. It did, however, require the endocytic adaptor protein, Numb, and lysosomal activity. This study reveals a previously unrecognized function of Notch in negatively titrating active β-Catenin protein levels in stem and progenitor cells.