Early in the life of every vertebrate embryo, be it human or hamster, there is a moment when the heart comes together -- literally. Scientists at the University of California, San Francisco have discovered a molecule that directs the two halves of the primordial heart to join as one.
Under the molecule's influence, separate tubes of the would-be heart -- primordial heart buds, essentially -- migrate toward each other from opposite sides of the embryo, the researchers found. As the two halves join, the rudimentary heart begins to beat.
The discovery is based on studies of zebrafish, tiny creatures which are utterly transparent as embryos. Many pivotal early changes that take place in the human embryo can be witnessed in zebrafish as if they were unfolding in a test tube. The fish is also fairly easy to study genetically.
The research, published in the July 13 issue of the journal Nature, shows that the normal union of the two heart tubes in the zebrafish embryo requires the presence of S1P, a molecule already known to be involved with cell proliferation and survival. S1P, an abbreviation for sphingosine 1-phosphate, is also active during wound healing, and the researchers suspect that its capacity to draw cells together is crucial there as well.
The heart buds join to form the zebrafish heart 22 hours into the embryo's life, and the heart begins beating soon thereafter. In the mouse embryo, the crucial union that will produce the heart occurs on the eighth day of development, and in human embryos, the heart forms and begins beating at three weeks, says Didier Stainier, PhD, UCSF associate professor of biochemistry and biophysics and senior author of the Nature study.
"S1P is a very old molecule," Stainier says. "It is present in organisms as distant as yeast and humans, and it appears to have been used over and over again throughout evolution for different roles: for cell proliferation, for wound healing, and now as we have found, for bringing the two parts of the primordial heart together."
If it serves this role for the zebrafish heart, Stainier suggests, it probably performs the same function in the human embryo.
"And as we start to understand the molecular pathways involving S1P by studying heart formation in zebrafish, we should learn something about how it regulates other processes such as wound healing," he says.
In 1996, Stainier and colleagues published the identity of 58 different mutations affecting zebrafish heart development. These they had selected from a much larger number of mutations produced by exposing the zebrafish genome to a chemical agent. They zeroed in on the most interesting genes by examining the developmental problems their mutations caused. One mutation that prevented the primordial heart halves to join was named miles apart by a post-doctoral researcher pining for his lover across the sea. In the embryos affected by the miles apart mutation, the researchers found, the two primordial heart tubes failed to converge to the midline.
In the current research reported in Nature, Stainier's lab isolated the miles apart gene and went on to show that the gene codes for a receptor, or docking site, for S1P. They determined that the receptors are concentrated in the midline region toward which the two heart buds migrate. Presumably, when S1P molecules dock with their miles apart receptors, they create a field that attracts or allows the primordial heart buds to migrate to the midline. There they find each other and fuse.
The discovery that S1P is essential for the normal union of the zebrafish heart is surprising, Stainier says, because most molecules that guide developing cells to their destiny -- all discovered in the last five to ten years -- are proteins, the conventional cellular workhorses produced directly from genetic instructions. But S1P is a lipid - an oily substance produced not from a genetic code but as a product of chemical reactions triggered by enzymes. This is the first lipid molecule shown to be crucial in vertebrate development and greatly expands the roster of molecular candidates scientists must consider in efforts to tease apart the puzzles of how vertebrates develop.
"Proteins can be identified from newly sequenced genes, and they can be localized using antibodies. But lipids are the products of enzymatic pathways. No gene codes for them, and they are much harder to study."
Stainier has been studying zebrafish heart development for ten years. The fish undergo most of the early life-defining changes found in human embyros, yet their embryos are transparent and readily accessible. Because it is also amenable to genetic study, the zebrafish has become an important model organism for studying vertebrate development.
The blue-and-silver-striped fish, a native of the Ganges River, is less than an inch long when mature. At the critical time when S1P and its miles apart receptor join forces to urge the animal's heart to come together, the transparent embryo is no larger than the head of a pin. Still, the researchers can scrutinize the tiny embryos, focusing on the maladies created by different mutations.
"This study underscores the value of studying from mutations to genes, rather than the other way around, to understand the genetics of vertebrate development," Stainier says. "The vertebrate genome is full of nuggets, and this approach of random mutagenesis in zebrafish is the best way to get at those nuggets." Random mutagenesis is the practice of producing mutations throughout the genome and then studying the genes that underlie mutations of particular interest.
The July 13 issue of Nature containing the report on heart formation includes a perspective on the research by Wolfgang Driever, a prominent developmental biologist in Germany.
First author on the paper with Stainier is Erik Kupperman, PhD, post-doctoral researcher in Stainier's lab. Co-authors on the paper and collaborators on the research are Songzhu An, MD, PhD, UCSF assistant professor of medicine; Nick Osborne, a graduate student and Steven Waldron, a technician, both in Stainier's lab. All co-authors in Stainier's lab are in the UCSF biochemistry and biophysics department and UCSF programs in developmental biology, genetics and human genetics.
The research is funded by the American Heart Association, the Packard Foundation and the UCSF Program in Human Genetics.
REPORTERS AND EDITORS: A color photo of the zebrafish can be accessed at http://pubaffr.ucsf.edu/imagedb/imsearch.php3?keywords=zebrafish . Click on the words "300 dip tiff". Good quality images of the heart buds coming together - which appear as figures "d" and "e"in the Nature article - can be accessed at http://pubaffr.ucsf.edu/imagedb/imsearch.php3?iname=071020002 Image "d" shows the normal merging of the two heart halves at the midline of the developing zebrafish embryo. Image "e" shows the mutant form in which the heart buds fail to merge.
Journal
Nature