 |
The Molecular and Genetic Basis of Organ Development
Summary: Brigid Hogan is interested in the molecular mechanisms regulating the growth, differentiation, and patterning of the mouse embryo. Of particular interest are genes involved in the elaboration of complex organs from small, undifferentiated rudiments, and the development of primordial germ cells.
Our long-term goal is to understand the molecular, cellular, and genetic basis of organogenesis—the process by which complex organs such as the mammalian lung, eye, kidney, and axial skeleton develop from small embryonic rudiments of undifferentiated cells. Classical embryological experiments have shown that organogenesis is driven by a series of reciprocal interactions between different cell populations. Initially, small groups of cells, known as organizing centers, secrete signaling factors that influence the social behavior and cell fate decisions of their neighbors. These target cells, in turn, produce signals that affect adjacent populations, and so on.
Understanding how embryonic cells cooperate to generate a three-dimensional, functional organ is crucial to our understanding of birth defects. The information will also be important for future efforts to stimulate the repair and regeneration of damaged tissues in vivo. Our attention has focused on the role of members of two large gene families—one encoding secreted factors known as bone morphogenetic proteins and the other encoding transcription factors known as forkhead or winged-helix proteins.
Foxc1 and Foxc2—Essential Components of the Molecular Circuitry Controlling Somite Formation
One hallmark of the development of the vertebrate embryo is the sequential formation on either side of the midline of paired blocks of mesodermal cells known as somites. These are the precursors of the vertebrae, body muscles, and dermis of the skin. In the mouse, new somites bud every 90 minutes from the anterior ends of two unsegmented strips of cells, known as the presomitic mesoderm (PSM), that extend back to a pool of stem cells at the posterior of the embryo. The generation of somites is a highly orchestrated process; repetitive waves of gene expression sweep anteriorly through the PSM and then slow down and come to a halt in response to a hypothetical "wavefront" maturation signal propagated posteriorly from the most recently formed somite. A prepattern is thus established in the anterior PSM corresponding to alternating stripes about half a somite wide of cells expressing genes corresponding to either anterior or posterior somite fates.
Several years ago we identified two closely related forkhead genes, now known as Foxc1 and Foxc2, which are expressed at high levels in the PSM and also in other embryonic mesodermal tissues, including the developing cardiovascular system, kidney rudiments, and mesenchyme around the eyes. Based on their strong expression in the PSM, we predicted that the genes play an important role in somite formation. However, inactivation of either Foxc1 or Foxc2 alone resulted in relatively minor defects in the development of the vertebrae and body muscles. By contrast, there were serious abnormalities in the early development of blood vessels, eyes, and kidneys, and the etiology of these defects provided insights into certain congenital birth defects in humans. We have now succeeded in generating compound null Foxc1;Foxc2 mutant embryos and have finally uncovered an essential role for both genes in somite formation. This role was previously masked because one copy of either gene could initially compensate for the absence of all three others. In the complete absence of both proteins, no somites are formed at all, and the prepatterning of the anterior PSM is completely disrupted. Analysis of the compound mutants suggests that the two forkhead genes are previously unrecognized components of the molecular circuitry mediating Notch-dependent intercellular signaling during somite formation. This hypothesis is supported by experiments in zebrafish embryos in which Foxc1 protein expression is specifically blocked by injection of morpholino antisense oligonucleotides.
The Role of Bone Morphogenetic Proteins in Organogenesis and Primordial Germ Cell Formation
We continue to investigate the role in early mouse development of the secreted signaling factors known as bone morphogenetic proteins (BMPs). We previously found that at least some embryos lacking a functional Bmp4 gene develop as far as the 25-somite stage but then die due to the complete absence of the allantois, which is essential for establishing the vascular connection between the embryo and the placenta. All Bmp4-null embryos also lack primordial germ cells (PGCs). These findings led us to propose a model in which Bmp4 made in the extraembryonic trophoblast layer induces precursors of the allantois and PGCs in the adjacent pluripotent embryonic cell population (the epiblast). We have now obtained further evidence for the early role of Bmp4 in embryonic patterning by making chimeras between Bmp4-null embryonic stem cells and tetraploid wild-type embryos in which the wild-type cells contribute only to extraembryonic tissues. We have thus uncovered a requirement for Bmp4 in the proliferation and differentiation of posterior mesoderm cells, and for PGC survival. We have also uncovered a role for Bmp4 in establishing the early left/right asymmetry of the mouse embryo.
Branching Morphogenesis of the Embryonic Lung and Development of Hair Follicles
The embryonic mouse lung provides an excellent system in which to define the rules governing the stereotypic formation of branched organs from simple buds, and the specification of different cell types along the proximal-distal axis of the branches. In studies funded by the National Institutes of Health, we have used mutant and transgenic mice, and in vitro culture, to establish that branching and proximodistal patterning are cooperatively regulated by a number of signaling factors and their antagonists. These include Bmp4, fibroblast growth factor 10 (Fgf10), Sonic hedgehog (Shh), and Noggin. Experiments are now under way using subtractive hybridization and microarray analysis to identify additional genes differentially expressed in the epithelium at the tips of lung buds versus more proximal regions. We have also developed in vitro culture conditions for growing lung epithelial stem cells in the absence of surrounding mesenchyme and identifying factors that drive them along different lineage pathways in culture.
Many of the same signaling factor gene families that are involved in lung morphogenesis are also used in the development of organs such as the eye, limb, and hair follicles. We are using Cre-lox technology to inactivate specific genes in small groups of cells in developing tissues. In addition we are exploring the role of BMPs by misexpressing antagonists that bind the ligands and prevent their interaction with receptors. This approach has been successful in identifying a role for BMPs in the growth and differentiation of stem cells within the postnatal hair follicle.
Last updated April 23, 2001
|
|
|
|
