Drosophila may be small, but you can't accuse them of lacking heart. Unless they're missing the transcription factor Tinman, that is, in which case they fail to form a heart during embryogenesis (1, 2) or, if tinman is deleted later in development, they fail to maintain adult cardiac function (3). Qian et al. now reveal how Tinman works in conjunction with the small GTPase Cdc42 to regulate the heart in both flies and mice (4).
Tinman lies at the top of a complex hierarchical network that makes and maintains the heart. Much of this network—including Tinman itself—is conserved in mammals. “So we can use Drosophila to learn about the fundamental genetic mechanisms that are important for the development and function of the heart,” says Rolf Bodmer, from the Sanford-Burnham Medical Research Institute in La Jolla, CA.
To discover new genes that work with tinman to maintain adult heart function, Bodmer and colleagues performed a genetic screen for mutations that cause heart failure in flies heterozygous for a deletion of tinman (4). Using this approach, Qian et al. found a genetic interaction between tinman and Cdc42. Whereas flies lacking one copy of either tinman or Cdc42 were healthy, animals lacking one copy of both genes had multiple heart defects. “The flies’ hearts didn't beat as regularly, and the heart myofibrils weren't arranged in a parallel, organized fashion,” Bodmer recounts. Expressing a dominant-negative version of Cdc42 in cardiac tissue produced similar effects.
The heart arrhythmias in tinman/Cdc42 double heterozygotes prompted Qian et al. to investigate the expression in these flies of several ion channels that regulate heart beat patterns. The expression of two potassium channels—dSUR and slowpoke—was down-regulated in double-heterozygous flies, and in vitro experiments revealed that Tinman and Cdc42 act together genetically to promote dSUR expression by regulating its enhancer element. Furthermore, flies lacking one copy of Cdc42 and one copy of either dSUR or slowpoke also had irregular heartbeats and disorganized myofibrils.
Complete loss of either Cdc42 or Nkx2-5 (the mammalian homologue of tinman) causes heart defects in mice (5, 6). “So we wanted to see if the combination of Tinman and Cdc42 also affected mouse heart function,” Bodmer explains. Qian et al. found that the loss of one copy of each gene in the mouse, as in the fly, compromised cardiac contraction and rhythmicity, similar to double-heterozygous flies. “However, we didn't see any aberrations in myofibril organization in the mice,” Bodmer says.
“Drosophila genetics is a unique way of identifying polygenic interactions that might aggravate a disease.”
Therefore, although the precise phenotypes differ, Cdc42 and Tinman/Nkx2-5 genetically interact in both mouse and fly hearts. But what is the nature of this interaction? Qian et al. found that Nkx2-5 inhibited the expression of a microRNA, miR-1, that targets Cdc42 mRNA both in vitro and in mice. miR-1 appears to serve as an intermediate between Tinman and Cdc42 in flies as well. Drosophila overexpressing miR-1 in their hearts had decreased Cdc42 and slowpoke levels as well as increased arrhythmias.
Mutations in the human homologue of Nkx2-5 are associated with a variety of cardiomyopathies (7), suggesting that Cdc42 may also influence human heart disease. “Drosophila genetics is a unique way of identifying polygenic interactions that might aggravate a disease,” Bodmer says. Qian et al.’s collaborators performed a small-scale screen of human samples and found one congenital heart disease patient with a mutated version of Cdc42 not found in control samples. Bodmer now wants to examine families carrying mutations in Nkx2-5 to see whether they have different variants of Cdc42 that could explain differences in the severity of their cardiomyopathies. “We'd also like to look for more genes that interact with tinman or with other factors that we know are important for the heart,” Bodmer says.