RESUMEN
Tyrosine kinase inhibitors are widely used in the clinic, but limited information is available about their toxicity in developing organisms. Here, we tested the effect of tyrosine kinase inhibitors targeting the ErbB receptors for their effects on developing zebrafish ( Danio rerio) embryos. Embryos treated with wide-spectrum pan-ErbB inhibitors or erbb4a-targeting antisense oligonucleotides demonstrated reduced locomotion, reduced diameter of skeletal muscle fibers, and reduced expression of muscle-specific genes, as well as reduced motoneuron length. The phenotypes in the skeletal muscle, as well as the defect in motility, were rescued both by microinjection of human ERBB4 mRNA and by transposon-mediated muscle-specific ERBB4 overexpression. The role of ErbB4 in regulating motility was further controlled by targeted mutation of the endogenous erbb4a locus in the zebrafish genome by CRISPR/Cas9. These observations demonstrate a potential for the ErbB tyrosine kinase inhibitors to induce neuromuscular toxicity in a developing organism via a mechanism involving inhibition of ErbB4 function.
Asunto(s)
Embrión no Mamífero/metabolismo , Desarrollo de Músculos/efectos de los fármacos , Neurogénesis/efectos de los fármacos , Unión Neuromuscular/embriología , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Tirosina Quinasas Receptoras/antagonistas & inhibidores , Receptor ErbB-4/antagonistas & inhibidores , Proteínas de Pez Cebra/antagonistas & inhibidores , Pez Cebra/embriología , Animales , Secuencia de Bases , Embrión no Mamífero/efectos de los fármacos , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Morfolinos/farmacología , Neuronas Motoras/efectos de los fármacos , Neuronas Motoras/metabolismo , Células Musculares/efectos de los fármacos , Células Musculares/metabolismo , Desarrollo de Músculos/genética , Músculo Esquelético/metabolismo , Mutación/genética , Neurogénesis/genética , Unión Neuromuscular/efectos de los fármacos , Proteínas Tirosina Quinasas Receptoras/genética , Proteínas Tirosina Quinasas Receptoras/metabolismo , Receptor ErbB-4/genética , Receptor ErbB-4/metabolismo , Pez Cebra/genética , Proteínas de Pez Cebra/genética , Proteínas de Pez Cebra/metabolismoRESUMEN
Endothelial cells (ECs) line the inside of blood vessels and respond to mechanical cues generated by blood flow. Mechanical stimuli regulate the localization of YAP by reorganizing the actin cytoskeleton. Here we demonstrate blood-flow-mediated regulation of endothelial YAP in vivo. We indirectly monitored transcriptional activity of Yap1 (zebrafish YAP) and its spatiotemporal localization in living zebrafish and found that Yap1 entered the nucleus and promoted transcription in response to blood flow. In cultured human ECs, laminar shear stress induced nuclear import of YAP and its transcriptional activity in a manner independent of Hippo signaling. We uncovered a molecular mechanism by which flow induced the nuclear translocation of YAP through the regulation of filamentous actin and angiomotin. Yap1 mutant zebrafish showed a defect in vascular stability, indicating an essential role for Yap1 in blood vessels. Our data imply that endothelial Yap1 functions in response to flow to maintain blood vessels.