Gene Switches Wing To Leg, Salk Scientists Find
La Jolla, CA – A single gene can transform embryonic tissue destined to form a wing into a leg instead, Salk Institute investigators have found. Their studies were performed in chickens, and the gene, called Tbx4 , appears to be involved in limb formation in mammals as well, including humans.
Limb malformations are among the most common human birth defects, including the rare disorder Holt-Oram Syndrome, characterized by external shortened arms and hands and associated with defects in a gene closely related to Tbx4.
"We knew that these genes in the Tbx family are important determinants in whether a limb becomes a hind or fore limb," said Juan Carlos Izpisúa Belmonte, Salk associate professor and senior author on the study, which appears in the current issue of Nature. "But we didn't know if one of these genes by itself was sufficient to send a limb down one pathway or the other. The current study demonstrates that the Tbx4 gene, normally active in the leg, can convert tissue from the wing area into a complete and normal leg."
Previous studies had demonstrasted that switching on Tbx4 in the nascent wing or in tissue close to the wing could add some elements of "legness," often resulting in a limb of intermediate structure but not effecting a complete transformation.
"We think this is because, even though we turn on a 'leg' gene, the tissue is still receiving 'wing' commands due to its location," explained Concepción Rodriguez-Esteban, lead author and Salk postdoctoral fellow. "It's been very difficult to tease apart the contributions of individual genes and to show that any single gene carries the complete blueprint for a leg or a wing."
In the current study, the Salk team infected tissue in the wing area with a virus carrying the gene Tbx4. They began their experiment when the embryos were one day old, before limb identity is set. Then, a day later they transferred the nascent limbs to the gut areas of younger embryos.
"By moving the tissue, we could ensure that no additional limb cues would affect the course that had been set," said Tohru Tsukui, Salk postdoctoral fellow and co-author of the study.
The majority of the transferred limbs grew into legs, demonstrating "that Tbx4 contains all the information needed to make a leg," said Izpisúa Belmonte. Several became chimeric leg/wing structures, which he thought was "probably because the Tbx4-carrying virus did not penetrate 100 percent of the cells in the potential wing tissue."
In control experiments, wing tissue that did not receive any Tbx4 genes was transferred and always developed into wings.
"Now that we know Tbx4 contains the 'leg blueprint,' we need to learn more about which genes it turns on and off, and how the Tbx4 gene itself gets switched on. Much remains to be done, but we hope to eventually be in a position to address problems of limb truncations and malformations," said Izpisúa Belmonte.
Additional co-authors include Sayuri Yonei-Tamura, Jorge Magallon, and Koji Tamura, all members of Izpisúa Belmonte's laboratory. The study, titled "The T-box genes Tbx4 and Tbx5 regulate limb outgrowth and identity," was supported by the National Science Foundation, the G. Harold and Leila Y. Mathers Foundation, and the Pew Charitable Trust.
The Salk Institute for Biological Studies, located in La Jolla, Calif., is an independent nonprofit institution dedicated to fundamental discoveries in the life sciences, the improvement of human health and conditions, and the training of future generations of researchers. The Institute was founded in 1960 by Jonas Salk, M.D., with a gift of land from the City of San Diego and the financial support of the March of Dimes Birth Defects Foundation.