Disruption of ECE-1 and ECE-2 reveals a role for endothelin-converting enzyme-2 in murine cardiac development
Hiromi Yanagisawa, Robert E. Hammer, James A. Richardson, Noriaki Emoto, S. Clay Williams, Shin-ichi Takeda, David E. Clouthier, Masashi Yanagisawa
Hiromi Yanagisawa, Robert E. Hammer, James A. Richardson, Noriaki Emoto, S. Clay Williams, Shin-ichi Takeda, David E. Clouthier, Masashi Yanagisawa
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Disruption of ECE-1 and ECE-2 reveals a role for endothelin-converting enzyme-2 in murine cardiac development

Abstract

Endothelin-converting enzyme-1 and -2 (ECE-1 and -2) are membrane-bound metalloproteases that can cleave biologically the inactive endothelin-1 (ET-1) precursor to form active ET-1 in vitro. We previously reported developmental defects in specific subsets of neural crest–derived tissues, including branchial arch–derived craniofacial structures, aortic arch arteries, and the cardiac outflow tract in ECE-1 knockout mice. To examine the role of ECE-2 in cardiovascular development, we have now generated a null mutation in ECE-2 by homologous recombination. ECE-2 null mice develop normally, are healthy into adulthood, are fertile in both sexes, and live a normal life span. However, when they are bred into an ECE-1–null background, defects in cardiac outflow structures become more severe than those in ECE-1 single knockout embryos. In addition, ECE-1–/–; ECE-2–/– double null embryos exhibited abnormal atrioventricular valve formation, a phenotype never seen in ECE-1 single knockout embryos. In the developing mouse heart, ECE-2 mRNA is expressed in the endocardial cushion mesenchyme from embyronic day (E) 12.5, in contrast to the endocardial expression of ECE-1. Levels of mature ET-1 and ET-2 in whole ECE-1–/–; ECE-2–/– embryos at E12.5 do not differ appreciably from those of ECE-1–/– embryos. The significant residual ET-1/ET-2 in the ECE-1–/–; ECE-2–/– embryos indicates that proteases distinct from ECE-1 and ECE-2 can carry out ET-1 activation in vivo.

Authors

Hiromi Yanagisawa, Robert E. Hammer, James A. Richardson, Noriaki Emoto, S. Clay Williams, Shin-ichi Takeda, David E. Clouthier, Masashi Yanagisawa

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Parasagittal sections of the heart of term wild-type (a), ECE-1–/– (b), ...
Parasagittal sections of the heart of term wild-type (a), ECE-1–/– (b), and ECE-1–/–; ECE-2–/– (c and d) embryos, and frontal sections of ECE-1–/–; ECE-2–/– embryos (e and f) stained with hematoxylin and eosin. Original magnification is ×11. (a) The interventricular septum (ivs) and aortic outflow (Ao) are seen. (b) Overriding of aorta with a small VSD (arrowhead) is seen in an ECE-1–/– embryo. (c) Persistent truncus arteriosus in an ECE-1–/–; ECE-2–/– embryo. One outflow (arrow) is separated into aortic outflow (Ao) and pulmonary (P) outflows. A large VSD (arrowhead) is shown. Displacement of the left AV valve (asterisk) is seen, opening toward the VSD. (d) Aorticopulmonary window in an ECE-1–/–; ECE-2–/– embryo. Septation of the aortic and pulmonary outflow tracts is incomplete at the base of the outflow tract (arrow). Note that the pulmonary outflow becomes the dorsal aorta (da) via the ductus arteriosus (dc). Pa, pulmonary artery. (e) Pulmonary and aortic outflow tracts originate from the right ventricle (DORV) in an ECE-1–/–; ECE-2–/– embryo. A large conotruncal ridge defect is seen underneath the aortic and pulmonary valves (filled diamond). A large VSD (arrowhead) is also shown. Note that the muscular layers of the pulmonary trunk (P) and aorta (Ao) are hypoplastic, and thin mesenchyme is observed between the two vessels (arrow). (f) DORV in an ECE-1–/–; ECE-2–/– embryo. The conotruncal ridge (filled diamond) is well developed compared with that in e. The arrowhead indicates a VSD.