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October 1, 2002 - Scientists Identify Key Gene Involved in Cleft Lip and Palate

Scientists report in this month's Nature Genetics they have discovered the gene that causes Van der Woude syndrome, the most common of the syndromic forms of cleft lip and palate.  The term "syndromic" means babies are born with cleft lip and palate, in addition to other birth defects.

According to the scientists, the discovery could very possibly direct them to genes involved in "non-syndromic" cleft lip and palate, one of the most common birth defects in the world.  Among Caucasians, non-syndromic cleft lip and palate occurs in an estimated 1 in every 1,000 live births, and the frequency seems to be even higher in some Asian countries, such as China and The Philippines.

"Since there is so much clinical overlap between the two, we expect that similar genes and maybe even the same genes will be involved in the non-syndromic form," said Jeff Murray, M.D., a scientist at the University of Iowa and an author on the paper.

Murray noted that the gene, called IRF6, seems to play a key role in the normal formation of the lips, palate, skin, and genitalia.  He said further study of the gene should provide precise molecular clues into normal human development and suggest specific biological strategies to prevent birth defects, such as cleft lip and palate.

First described in the 1860s, Van der Woude syndrome (VWS) is involved in about 2 percent of all cases of cleft lip and palate, occurring in approximately 1 of every 33,000 live births.  Children with the syndrome are born with any of four characteristic birth defects:  Pits, or small indentations, in the lower lip, cleft lip, cleft palate, and undeveloped tooth buds.

What has puzzled scientists is those with the syndrome inherit different combinations of birth defects, not necessarily all of them.  For instance, half of the children born with VWS have cleft lip, 70 percent have pits in their lower lip, 20 percent have cleft palate, and only about 10 percent do not have a full set of teeth.  More puzzling, siblings are often born with different combinations of birth defects, though they inherit the same gene change.

Because the cleft lip and palate seen in people with VWS closely resembles the non-syndromic condition, scientists long have been intrigued with identifying its gene.  The hope was the gene not only would permit genetic testing of affected families, the gene might point them to the complex cascade of molecular signals during early gestation that prompt the formation of the lips and palate, critical information in developing viable strategies to prevent clefts.

In the mid 1980s, as scientists became more adept at identifying so-called "disease" genes, laboratories launched their initial attempts to find the one involved in VWS.  Based on data from a case report of an individual with VWS that was caused by a chromosomal abnormality on chromosome 1, Murray's group confirmed that other individuals affected with VWS shared an irregularity in the same region of chromosome one, the first step in finding the gene.

But, in trying to pinpoint the precise location of the irregularity, the scientists eventually ran into technical difficulties.  Part of the problem was making sense of the region's unusual terrain, which in some people includes a fairly large, but benign, deletion and long stretches of difficult-to-track repeat sequences of DNA. 

Another problem was simply wading through all of the genomic information that had become available to them during the late 1990s. According to Brian Schutte, Ph.D., a lead author on the paper and a scientist at the University of Iowa, one of the first genes the group evaluated was IRF6, the subject of this month's paper.  However, based on a preliminary analysis, they initially excluded it as the disease gene.

"We tested DNA samples from a subset of people with the syndrome, and we didn't find any mutations in IRF6," said Schutte.  "About this time, the full DNA sequence map of the region became available to us.  We suddenly had a long list of other genes to evaluate and decided to move on from IRF6."

Added Murray, "At the time, we understood that we were excluding candidate genes that could still potentially be the disease gene.  But you always have to choose the best place to focus your efforts, and our data at the time indicated that it was better to move on to other candidate genes."

A break came last year while a visiting graduate student in Murray's laboratory spoke on the telephone to a colleague in her native Brazil.  "Her colleague happened to mention that he had just seen so-called 'monozygotic twins,' in which one had van der Woude and the other didn't," remembered Murray.  "She told of us about it, and we got quite excited actually."

Murray hypothesized that the twins were genetically identical with one exception:  The affected twin would have a change in the VWS gene. Shinji Kondo, a post-doctoral fellow in Murray's lab, then sequenced the relevant portions of DNA from the twins and, soon thereafter, they discovered a syndrome-causing deletion in IRF6.

"I certainly knew it was possible in theory," said Murray.  "But, to be honest, if I knew as much then about twins as I do now, I might have been less excited. "

Murray explained that the theory hinged on the possibility the mutation occurred soon after the twins divided into separate embryos, meaning all cells in the body of the affected twin would carry the mutation. If it had occurred later in the developmental process, after various distinct cell lineages have emerged, only some of the affected twin's cells would have carried the mutation.  "If that were the case, we might have missed it," said Murray.

 "Monozygotic twins are actually quite common in other complex conditions, including diabetes and heart disease," added Murray.  "I think our work offers proof of principle that monozygotic twins are a powerful model to search for genes involved in complex conditions, one that may have been overlooked in the past."

To confirm the discovery, Schutte and colleagues sequenced and found mutations in IRF6 among 46 unrelated families with the syndrome. Interestingly, the group also detected changes in 13 additional families with a related condition called popliteal pterygium syndrome, or PPS.  This rare disorder involves the same birth defects as VWS, plus underdeveloped genitalia and webbed skin on the back of the legs.  Further confirmation was obtained by Mike Dixon's lab at the University of Manchester in England.  Dixon's lab showed that IRF6 is made in all the tissues affected by both VWS and PPS during the critical times in development.

Schutte said the gene's involvement in causing both conditions is not necessarily surprising.   Researchers have noted previously that some so-called PPS families include members who only have orofacial abnormalities reminiscent of VWS, not those in other parts of the body.   "I think that as we learn more about the biochemistry of IRF6, we'll be able to make better predictions about how these syndromes will manifest themselves," said Schutte.  "At the same time, there are also probably many other genetic and environmental factors involved, too." 

One intriguing possibility raised by this month's paper is cleft lip and palate conceivably could be associated with some type of viral infection during pregnancy.  Though poorly studied, IRF6 is similar to other genes that help to trigger the production of interferons, immune signaling proteins involved in responding to viral infections.

Stressing that the viral connection is still extremely speculative, Murray noted, "This would be a wonderful outcome of this paper because, in 2002 at least, attacking viruses is much easier than manipulating genes. "

Already, the group has determined that IRF6 is highly expressed in the tooth buds, hair follicles, genitalia, skin, and palate of the developing human embryo.  By further studying the gene and its protein, Schutte said it should be possible to tap into the molecular signals that it helps to relay during human development.  "We know that IRF6 is a transcription factor, meaning it binds to certain genes and allows their information to be copied, the first step in producing a protein," he said. "We can now ask which genes it activates and when?  All of these genes might be involved in causing cleft lip and palate.  Or they could contribute to other birth defects.  The point is we stand to learn a great deal from this gene discovery."

The article, which is published in the October 2002 issue of Nature Genetics, is titled, "Mutations in IRF6 cause Van der Woude and popliteal pterygium syndromes."  Its authors are: Shinji Kondo, Brian C. Schutte, Rebecca J. Richardson, Bryan C. Bjork, Alexandra S. Knight, Yoriko Watanabe, Emma Howard, Renata L. L. Ferreira de Lima, Sandra Daack-Hirsch, Achim Sander, Donna M. McDonald-McGinn, Elaine H. Zackai, Ed Lammer, Art Aylsworth, Holly Ardinger, Andrew C. Lidral, Barbara Pober, Lina Moreno, Mauricio Arcos-Burgos, Consuelo Valencia, Claude Houdayer, Michel Bahuau, Danilo Moretti-Ferreira, Antonio Richieri-Costa, Michael J. Dixon, Jeffrey C. Murray.  The research was supported by the NIDCR.

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This page last updated: February 26, 2014