Emergence of Xin Demarcates a Key Innovation in Heart Evolution

The mouse Xin repeat-containing proteins (mXinα and mXinβ) localize to the intercalated disc in the heart. mXinα is able to bundle actin filaments and to interact with β-catenin, suggesting a role in linking the actin cytoskeleton to N-cadherin/β-catenin adhesion. mXinα-null mouse hearts display progressively ultrastructural alterations at the intercalated discs, and develop cardiac hypertrophy and cardiomyopathy with conduction defects. The up-regulation of mXinβ in mXinα-deficient mice suggests a partial compensation for the loss of mXinα. To elucidate the evolutionary relationship between these proteins and to identify the origin of Xin, a phylogenetic analysis was done with 40 vertebrate Xins. Our results show that the ancestral Xin originated prior to the emergence of lamprey and subsequently underwent gene duplication early in the vertebrate lineage. A subsequent teleost-specific genome duplication resulted in most teleosts encoding at least three genes. All Xins contain a highly conserved β-catenin-binding domain within the Xin repeat region. Similar to mouse Xins, chicken, frog and zebrafish Xins also co-localized with β-catenin to structures that appear to be the intercalated disc. A putative DNA-binding domain in the N-terminus of all Xins is strongly conserved, whereas the previously characterized Mena/VASP-binding domain is a derived trait found only in Xinαs from placental mammals. In the C-terminus, Xinαs and Xinβs are more divergent relative to each other but each isoform from mammals shows a high degree of within-isoform sequence identity. This suggests different but conserved functions for mammalian Xinα and Xinβ. Interestingly, the origin of Xin ca. 550 million years ago coincides with the genesis of heart chambers with complete endothelial and myocardial layers. We postulate that the emergence of the Xin paralogs and their functional differentiation may have played a key role in the evolutionary development of the heart

Heavy and light roles: myosin in the morphogenesis of the heart

Myosin is an essential component of cardiac muscle, from the onset of cardiogenesis through to the adult heart. Although traditionally known for its role in energy transduction and force development, recent studies suggest that both myosin heavy-chain and myosin lightchain proteins are required for a correctly formed heart. Myosins are structural proteins that are not only expressed from early stages of heart development, but when mutated in humans they may give rise to congenital heart defects. This review will discuss the roles of myosin, specifically with regards to the developing heart. The expression of each myosin protein will be described, and the effects that altering expression has on the heart in embryogenesis in different animal models will be discussed. The human molecular genetics of the myosins will also be reviewed

A functional knock-out of titin results in defective myofibril assembly

van der Ven PFM, Bartsch JW, Gautel M, Jockusch H, Furst DO. A functional knock-out of titin results in defective myofibril assembly. JOURNAL OF CELL SCIENCE. 2000;113(8):1405-1414.Titin, also called connectin, is a giant muscle protein that spans the distance from the sarcomeric Z-disc to the M-band. Titin is thought to direct the assembly of sarcomeres and to maintain sarcomeric integrity by interacting with numerous sarcomeric proteins and providing a mechanical linkage. Since severe defects of such an important molecule are likely to result in embryonic lethality, a cell culture model should offer the best practicable tool to probe the cellular functions of titin. The myofibroblast cell line BHK-21/C13 was described to assemble myofibrils in culture. We have now characterized the sub-line BHK-21-Bi, which bears a small deletion within the titin gene. RNA analysis revealed that in this mutant cell line only a small internal portion of the titin mRNA is deleted. However, western blots, immunofluorescence microscopy and immunoprecipitation experiments showed that only the N-terminal, approx. 100 kDa central Z-disc portion of the 3 MDa titin protein is expressed, due to the homozygous deletion in the gene. Most importantly, in BHK-21-Bi cells the formation of thick myosin filaments and the assembly of myofibrils are impaired, although sarcomeric proteins are expressed. Lack of thick filament formation and of ordered actin-myosin arrays was confirmed by electron microscopy. Myogenisation induced by transfection with MyoD yielded myofibrils only in myotubes formed from wild type and not from mutant cells, ruling out that a principal failure in myogenic commitment of the BHK-21-Bi cells might cause the observed effects. These experiments provide the first direct evidence for the crucial role of titin in both thick filament formation as a molecular ruler and in the coordination of myofibrillogenesis