Comparative evolutionary studies have greatly aided our understanding of how the heart develops. Two recent articles from Benoit Bruneau’s laboratory reveal the power of the so-called evo-devo approach and provide yet further details of development, both of the heart itself and of the cells that build it.
Thanks to our many and varied primitive cousins, developmental biologists are gaining a good picture of the conserved genes and pathways controlling heart development.1 It appears that Natural Selection’s fiddly fingers have taken a basic set of genes and pathways—capable of giving rise to the simple peristaltic pumps of insects, for example—and then tweaked, tuned, and expanded them to build hearts that meet the needs of increasingly complex multicellular organisms.
Heart evolution has not just been about meeting needs, though. It has also been a driving force that has enabled increased complexity and new evolutionary adaptations, for example, the adaptation to warm-bloodedness.
Endotherms require more oxygen than their cold-blooded counterparts to supply their high metabolic activity. “One of the major evolutionary adaptations that allowed mammals to become warm-blooded was the formation of the four-chambered heart,” says Benoit Bruneau (Gladstone Institute of Cardiovascular Disease, San Francisco, Ca). Four chambers means no mixing of deoxygenated and oxygenated blood, and thus, more oxygen gets to the tissues.
This adaptation was not sufficient alone, however, Bruneau explains. “You need a certain lung structure. There’s certain changes to the breathing apparatus that are necessary, and other things as well that have to happen.” But the point is, he says, “None of that can happen if you don’t have a four-chambered heart.”
Cold-blooded fish and amphibians have three-chambered hearts, two atria and one ventricle. Reptiles are a mixed bag with some, like crocodiles and alligators, having four-chambered hearts and others, like lizards, having three-chambered hearts.
The crocodilian four-chambered heart has led people to speculate that crocodile ancestors may have been warm-blooded. But Bruneau thinks not. “My interpretation of the crocodile and alligator hearts,” he says, “is that they’ve sort of overshot their evolution: the heart part has moved ahead but the rest hasn’t caught up.”
The fourth chamber has arisen via the separation of one large ventricle into two. In animals in which this occurs, a transcription factor called Tbx5 concentrates in the prospective left ventricle during heart development. Bruneau’s team wondered what the pattern of Tbx5 mRNA expression would look like in the three-chambered hearts of non-crocodilian reptiles. They chose to compare a turtle with a lizard.2 Turtle hearts are interesting because, although they have just one large ventricle, there is evidence of a primitive interventricular septum (IVS)-like structure. Lizards, on the other hand, are considered evolutionarily less complex creatures and have no hint of an IVS.
At early stages of heart development in the turtle, Trachemys scripta elegans, and the lizard, Anolis carolinenis, Tbx5 was expressed broadly across the heart. At comparable stages in the chick and mouse, however, Tbx5 expression was tightly concentrated on the left side of the heart. Interestingly, later in the development of the turtle heart, Tbx5’s expression pattern changed. It diminished on the right side of the large single ventricle and increased on the left. Target genes of Tbx5 (Bmp10 and Nppa) were also restricted to the left side. The gradient of Tbx5 expression in the turtle heart was not as steep as that in chicken or mouse, and it occurred at a later stage of embryogenesis. This perhaps explains why only a partial IVS-like structure forms in the turtle heart. In the lizard, the distribution of Tbx5 remained broad.
Although this comparative study provided a pleasing correlation between Tbx5 expression patterns and the formation of the IVS, the question remained as to whether this gradient of Tbx5 expression was actually driving IVS patterning. The authors went some way to answering this by removing the Tbx5 gradient in developing mouse hearts. They conditionally deleted Tbx5 from the embryonic ventricles and also misexpressed Tbx5 across the entire early ventricle. Both of these experiments resulted in the loss of the IVS and the development of one large ventricle.
Christine “Kricket” Seidman (Harvard Medical School, Boston, Ma) describes Bruneau’s study as, “a creative integration of evolutionary and developmental biology, which provides new insights into cardiac septation.” Seidman explains, “Atrial and ventricular septal defects are among the most common of human congenital heart malformations.” Thus, she says, “Understanding this important process should help to elucidate why human patients often have defects in septation.” Seidman’s laboratory had first shown that mutations in TBX5 in humans could cause septal defects,3 and “now it seems likely that mutations in any factors that play a part in the level or localization of Tbx5 expression might also lead to malformation of the IVS,” Bruneau says.
Tbx5 functions in the development of other systems and organs besides the heart. Even within the heart, Tbx5 has a number of roles. Two articles published in Nature Genetics recently revealed that human TBX5 variants affect heart conduction in the general population,4,5 a role that had previously only been suspected.6 Tbx5 is also involved in forming fundamental building blocks of the heart—the cardiomyocytes that build the early heart tube.7,8 In fact, according to another recent study from Bruneau’s laboratory, Tbx5 is one of a minimal set of three factors that can switch mouse embryonic mesoderm cells from their assigned fate into becoming heart cells.9
Bruneau’s team was testing Tbx5 along with other cardiac gene regulatory factors to identify the minimal determinants of heart cell differentiation. They found that the factors Gata4 and Baf60c, when expressed together ectopically in mouse embryo mesoderm, could induce the expression of the early cardiac marker protein, Actc1. However, when Tbx5 was thrown into the mix, the cells differentiated beyond this early stage and showed signs of contractile function—they twitched.
Although these three factors were enough to induce mesoderm to transdifferentiate to a cardiomyocyte state, it is perhaps less likely that they would be sufficient to convert any cell type. The factors did not work when the team tried to convert endoderm cells, for example. Ultimately, if the three factors were able to convert, say, blood cells or skin cells into heart cells, it would be great news for heart injury patients: the clinicians would have an accessible source of the patient’s own cells with which to perform repairs. “That is the ideal situation,” says Bruneau, adding, “we may need additional factors or it might just be that that’s not a possibility, but we are certainly going to give it a try.”
If only making heart muscle cells were as easy as making skeletal muscle cells. According to Margaret Buckingham (Institut Pasteur, Paris, France), “If you overexpress MyoD, then you can get most cells to form skeletal muscle, even cells differentiated down a different pathway.”
MyoD is a transcription factor that activates skeletal muscle genes, but it has also been shown to have chromatin remodeling capability.10 Baf60c, one of the three required factors for heart cell differentiation, is itself a chromatin remodeler. “This is an exciting finding,” says Buckingham, “because it confirms the idea that was already suggested by the MyoD story, that actually you need to remodel chromatin in order to get a cell to change its destiny.”
The need for chromatin remodeling in cardiomyocyte transdifferentiation appears to be restricted to amniotes, because in other vertebrates, such as Xenopus and zebrafish, transcription factors alone (Mesp1 or Gata5) are enough. “That intrigues me,” says Buckingham.
Indeed, why would amniotes require what Bruneau describes as an additional layer of tissue-specific gene regulation? “Probably because of the additional complexity of the heart,” says Bruneau, “One could imagine that since the heart [of non-amniotes] is simpler, the regulation of gene expression doesn’t need to be as refined or complex. By providing an extra layer of regulation, you have more flexibility and versatility to control the same DNA binding factors.”
Seidman adds, “This work shows the considerable complexity of regulating gene expression during development.” The additional regulatory layer that chromatin provides, she goes on is, “Quite neat!”
Us amniotes might think our chromatin-level control of heart development is neat, but other vertebrates, arguably have the coup de grâce. As Buckingham points out, “You can chop off a bit of the zebrafish heart and it will regenerate perfectly, whereas if you have an infarction in your heart you have a terrible problem.” Perhaps evo-devo studies might one day reveal how fish and frogs achieve this amazing regeneration feat and allow us to artificially achieve the same. As Bruneau puts it, “The way I see this is that by looking backwards, we are able to better look forward.”
The opinions expressed in News and Views are not necessarily those of the editors or of the American Heart Association.
Olson EN. Gene regulatory networks in the evolution and development of the heart. Science. 2006; 313: 1922–1927.
Koshiba-Takeuchi K, Mori AD, Kaynak BL, Cebra-Thomas J, Sukonnik T, Georges RO, Latham S, Beck L, Henkelman RM, Black BL, Olson EN, Wade J, Takeuchi JK, Nemer M, Gilbert SF, Bruneau BG. Reptilian heart development and the molecular basis of cardiac chamber evolution. Nature. 2009; 461: 95–98.
Basson CT, Bachinsky DR, Lin RC, Levi T, Elkins JA, Soults J, Grayzel D, Kroumpouzou E, Traill TA, Leblanc-Straceski J, Renault B, Kucherlapati R, Seidman JG, Seidman CE. Mutations in human TBX5 [corrected] cause limb and cardiac malformation in Holt-Oram syndrome. Nat Genet. 1997; 15: 30–35.
Holm H, Gudbjartsson DF, Arnar DO, Thorleifsson G, Thorgeirsson G, Stefansdottir H, Gudjonsson SA, Jonasdottir A, Mathiesen EB, Njølstad I, Nyrnes A, Wilsgaard T, Hald EM, Hveem K, Stoltenberg C, Løchen ML, Kong A, Thorsteinsdottir U, Stefansson K. Several common variants modulate heart rate, PR interval and QRS duration. Nat Genet. 2010; Jan 10 [Epub ahead of print].
Pfeufer A, van Noord C, Marciante KD, Arking DE, Larson MG, Smith AV, Tarasov KV, Müller M, Sotoodehnia N, Sinner MF, Verwoert GC, Li M, Kao WH, Köttgen A, Coresh J, Bis JC, Psaty BM, Rice K, Rotter JI, Rivadeneira F, Hofman A, Kors JA, Stricker BH, Uitterlinden AG, van Duijn CM, Beckmann BM, Sauter W, Gieger C, Lubitz SA, Newton-Cheh C, Wang TJ, Magnani JW, Schnabel RB, Chung MK, Barnard J, Smith JD, Van Wagoner DR, Vasan RS, Aspelund T, Eiriksdottir G, Harris TB, Launer LJ, Najjar SS, Lakatta E, Schlessinger D, Uda M, Abecasis GR, Müller-Myhsok B, Ehret GB, Boerwinkle E, Chakravarti A, Soliman EZ, Lunetta KL, Perz S, Wichmann HE, Meitinger T, Levy D, Gudnason V, Ellinor PT, Sanna S, Kääb S, Witteman JC, Alonso A, Benjamin EJ, Heckbert SR. Genome-wide association study of PR interval. Nat Genet. 2010; Jan 10 [Epub ahead of print].
Postma AV, van de Meerakker JB, Mathijssen IB, Barnett P, Christoffels VM, Ilgun A, Lam J, Wilde AA, Lekanne Deprez RH, Moorman AF. A gain-of-function TBX5 mutation is associated with atypical Holt-Oram syndrome and paroxysmal atrial fibrillation. Circ Res. 2008; 102: 1433–1442.
Gerber AN, Klesert TR, Bergstrom DA, Tapscott SJ. Two domains of MyoD mediate transcriptional activation of genes in repressive chromatin: a mechanism for lineage determination in myogenesis. Genes Dev. 1997; 11: 436–450.