Features
Cancer Research Takes Flight:
Wnt Signaling in Development and Disease
It turns out that Wnt signaling controls this process of self-renewal, operating as a kind of master switch between proliferation of stem cells and differentiation into epithelial cells. Several studies have shown that Wnt-target gene expression occurs in a gradient that is strongest at the base of the intestinal crypt and weakens further away. And loss of β-catenin, the key transducer of Wnt signaling, dramatically reduces intestinal cell proliferation.
"The connection of the Wnt pathway to human cancer is very strong— mutations in this pathway are associated with 85-90 percent of human colorectal cancers."
Yamaguchi and his colleagues have compared the gene expression patterns they observed in the embryonic mesoderm with that of the adult intestinal crypt and found a remarkable 60 percent of the Wnt-target genes that they identified in mesodermal stem cells are also found in the adult intestinal stem cells. Identifying these genes is the first step in establishing the critical molecular network that is responsible for stem cell maintenance, whether in the embryo or in the adult. Functional follow-up studies will be necessary to establish their roles in cellular renewal (see "The Power of Embryonic Stem Cells").
Development Gone Awry
The precise regulation of development, once tampered with, can quickly give rise to abnormal growth that is the hallmark of cancer. "From my developmental perspective, cancer is developmental signaling gone awry," explained Yamaguchi. Already in 1989, mutations of the gene adenomatous polyposis coli (APC) were found in patients with familial adenomatous polyposis (FAP) and in sporadic colorectal cancers before it was understood that APC was a critical component of the Wnt signaling pathway. Since then, several other mutations in the canonical Wnt signaling cascade have been associated with cancers.
Terry Yamaguchi, Ph.D. (left), with Postdoctoral Fellow Ravi Chalamalasetty, Ph.D. (right). (Photo: R. Baer)
Hans Clevers, M.D., Ph.D., and colleagues at the Utrecht University Medical Centre, The Netherlands, showed in a paper published in Nature this year that deleting APC in long-lived intestinal crypt stem cells—but not in differentiated cells migrating away from the intestinal crypt—leads to intestinal adenomas. The transformed stem cells appear to remain in the crypts, steadily fueling growth of the adenomas, and they may represent one of the best examples of a true cancer stem cell.
Studying the same Wnt3a-target genes that he originally identified in embryonic mesoderm, Yamaguchi has shown that 40 percent of these genes are expressed in intestinal adenomas. "So, we’re asking whether any of these genes are required downstream of Wnt signaling for tumor formation." Specifically, in a mouse model of intestinal adenomas in which the β-catenin pathways are constitutively active, Yamaguchi and his colleagues are asking whether they can reduce the tumor burden by knocking out some of the Wnt-target genes. Conversely, they are also trying to make transgenic mice that overexpress individual target genes specifically in the intestinal epithelium to ask whether they alone are sufficient to form tumors.
"The connection of the Wnt pathway to human cancer is very strong— mutations in this pathway are associated with 85–90 percent of human colorectal cancers. And this case is one where the animal model, although not perfect, is quite good for human cancer. The molecular mutations in both cases are essentially the same. Thus, we have a great opportunity to apply what we know from normal biology to an animal model of cancer."
The Power of Embryonic Stem Cells
Embryonic stem cells are not only part of the biology fueling Terry Yamaguchi, Ph.D.’s, intellectual curiosity, but they are also a means to satisfy that curiosity. As a graduate student, Yamaguchi first exploited the ability of embryonic stem cells to differentiate in vitro to identify the growth factors important for the specification of mesodermal fates. His work led to the discovery of a family of receptors that regulates the stem cells that give rise to blood and blood vessels. Now Yamaguchi is an enthusiastic adopter of a powerful embryonic stem cell system generated by Michael Kyba, Ph.D.’s, group at the University of Minnesota.
While in vivo studies are critical to developmental biology, testing biochemical mechanisms is difficult to do in an embryo of only a few thousand cells. "We wanted to move away from an embryo to a stem cell population we could manipulate in vitro," explained Yamaguchi. Dr. Kyba had recently developed an inducible system for expressing genes of interest in embryonic stem cells in a consistent and reliable way. The gene is always integrated at the same genetic locus, and it is induced by the application of doxycycline.
"These [embryonic stem] cells are a great compromise—we can do so much manipulation genetically and biochemically in vitro—and the sky is the limit because the efficiency of targeting is so good," enthused Yamaguchi. In their laboratory, and with these cells, the probability of a successfully engineered cell is almost 95 percent for any gene of interest. They are, therefore, able to screen all of the Wnt3a target genes they identify by transcriptional profiling for functional activity and biochemical interactions. Postdoctoral Fellow Ravi Chalamalasetty, Ph.D., is using this approach to identify the targets of Mesogenin 1 (see main text).
Once stable embryonic stem cell lines are engineered with the inducible gene of interest, these cells can be taken back to the embryo to produce a transgenic mouse. Additionally, once the culture conditions are solved, these cell lines can be differentiated in vitro into any cell type (e.g., a heart cell or a neuron or, maybe one day, an intestinal stem cell). "For me," said Yamaguchi, "the possibilities are endless with these embryonic stem cells."
