Small-Molecule-Mediated Suppression of BMP Signaling by Selective Inhibition of BMP1-Dependent Chordin Cleavage.

BMP signaling is critical for many biological processes. Therefore, small molecules that modulate BMP signaling are useful for elucidating the function of BMP signaling and treating BMP signaling-related diseases. Here, we performed a phenotypic screening in zebrafish to examine the in vivo effects of N-substituted-2-amino-benzoic acid analogs NPL1010 and NPL3008 and found that they affect BMP signaling-dependent dorsal-ventral (D-V) patterning and bone formation in zebrafish embryos. Furthermore, NPL1010 and NPL3008 suppressed BMP signaling upstream of BMP receptors. BMP1 cleaves Chordin, an antagonist of BMP, and negatively regulates BMP signaling. Docking simulations demonstrated that NPL1010 and NPL3008 bind BMP1. We found that NPL1010 and NPL3008 partially rescued the disruptions in the D-V phenotype caused by bmp1 overexpression and selectively inhibited BMP1-dependent Chordin cleavage. Therefore, NPL1010 and NPL3008 are potentially valuable inhibitors of BMP signaling that act through selective inhibition of Chordin cleavage.

Artificial Notch Signaling Activation Method Using Immobilized Ligand Beads.

Activation of Notch signaling requires physical interaction between ligand- and receptor-expressing cells and pulling force to release the Notch intracellular domain. Therefore, the soluble recombinant ligand protein is not suitable for the activation of Notch signaling in a cell culture system. Here, we describe an efficient method for transient activation of Notch signaling using immobilized ligand beads. Using this method, the timing of Notch signaling can be efficiently controlled.

Transient activation of the Notch-her15.1 axis plays an important role in the maturation of V2b interneurons.

In the vertebrate ventral spinal cord, p2 progenitors give rise to two interneuron subtypes: excitatory V2a interneurons and inhibitory V2b interneurons. In the differentiation of V2a and V2b cells, Notch signaling promotes V2b fate at the expense of V2a fate. Later, V2b cells extend axons along the ipsilateral side of the spinal cord and express the inhibitory transmitter GABA. Notch signaling has been reported to inhibit the axonal outgrowth of mature neurons of the central nervous system; however, it remains unknown how Notch signaling modulates V2b neurite outgrowth and maturation into GABAergic neurons. Here, we have investigated neuron-specific Notch functions regarding V2b axon growth and maturation into zebrafish GABAergic neurons. We found that continuous neuron-specific Notch activation enhanced V2b fate determination but inhibited V2b axonal outgrowth and maturation into GABAergic neurons. These results suggest that Notch signaling activation is required for V2b fate determination, whereas its downregulation at a later stage is essential for V2b maturation. Accordingly, we found that a Notch signaling downstream gene, her15.1, showed biased expression in V2 linage cells and downregulated expression during the maturation of V2b cells, and continuous expression of her15.1 repressed V2b axogenesis. Our data suggest that spatiotemporal control of Notch signaling activity is required for V2b fate determination, maturation and axogenesis.