Absence of heartbeat in the Xenopus tropicalis mutation muzak is caused by a nonsense mutation in cardiac myosin myh6

Abstract

Mechanisms coupling heart function and cardiac morphogenesis can be accessed in lower vertebrate embryos that can survive to swimming tadpole stages on diffused oxygen. Forward genetic screens in Xenopus tropicalis have identified more than 80 mutations affecting diverse developmental processes, including cardiac morphogenesis and function. In the first positional cloning of a mutation in X. tropicalis, we show that non-contractile hearts in muzak (muz) embryos are caused by a premature stop codon in the cardiac myosin heavy chain gene myh6. The mutation deletes the coiled-coil domain responsible for polymerization into thick filaments, severely disrupting the cardiomyocyte cytoskeleton. Despite the lack of contractile activity and absence of a major structural protein, early stages of cardiac morphogenesis including looping and chamber formation are grossly normal. Muz hearts subsequently develop dilated chambers with compressed endocardium and fail to form identifiable cardiac valves and trabeculae.

The Impact of Mechanical Forces in Heart Morphogenesis

“…17 Similarly, disruption of atrial myosin heavy chain causes defective atrial septal formation in the chick 18 and failure to develop valves or trabeculae in Xenopus tropicalis. 19 Mice lacking atrial myosin regulatory light chain (MLC2a) display enlarged and amorphous heart tubes, defects in looping architecture and abnormalities of cardiac chamber morphology. 20 Mouse embryos with targeted mutations of cardiac troponin T and the Na 2 -Ca 2ϩ exchanger, which prevent the onset of heart beating, display looping and endocardial cushion defects 12 as well as thin ventricular walls with underdeveloped trabeculae.…”

The Hitchhiker's guide to Xenopus genetics

“…ENU creates point mutations forming both nulls (Abu-Daya et al , 2009) and milder hypomorphic alleles (Geach and Zimmerman, 2010) more typical of human disease variants. Since dosage of the toxic and labile ENU is difficult to control, in vitro mutagenesis of sperm offers a practical opportunity to select a dose producing both high rates of dominant lethal abnormalities visible in the days after in vitro fertilization as well as viable F1 carriers to raise.…”

“…Higher resolution mapping to specific genes can be accomplished using conventional positional cloning methods as well as candidate gene approaches (Abu-Daya et al , 2009; Geach and Zimmerman, 2010). Efficiency of these strategies has greatly increased with improved genome contiguity and annotation and meiotic map marker density, and they remain useful for evaluating candidate genes.…”

“…The muzak ( muz ) mutation completely lacks heartbeat, and is caused by a truncation of the cardiac specific myosin heavy chain gene myh6 (Figure 4E–H); skeletal muscle is unaffected (Abu-Daya et al , 2009). Muzak permits study of the roles of heartbeat, bloodflow, and myofibrils in cardiac morphogenesis.…”

“…The last five years have seen dramatic advances in Xenopus genetics (Harland and Grainger, 2011), including the first screens for induced mutations in amphibians (Goda et al , 2006), cloning of novel mutations (Abu-Daya et al , 2010; Abu-Daya et al , 2009; Geach and Zimmerman, 2010), generation of a meiotic map (Wells et al , 2010) and a genomic sequence assembly (Hellsten et al , 2010), reverse genetics (Goda et al , 2006; Young et al , 2011), and stock centers on either side of the Atlantic (see Pearl et al in this issue). Recent breakthroughs in high throughput sequencing, together with improved genomic resources, are revolutionizing both forward and reverse genetic strategies.…”