Median facial clefts in Xenopus laevis: Roles of retinoic acid signaling and homeobox genes

Abstract

The upper lip and primary palate form an essential separation between the brain, nasal structures and the oral cavity. Surprisingly little is known about the development of these structures, despite the fact that abnormalities can result in various forms of orofacial clefts. We have uncovered that retinoic acid is a critical regulator of upper lip and primary palate development in Xenopus laevis. Retinoic acid synthesis enzyme, RALDH2, and retinoic acid receptor gamma (RARγ) are expressed in complementary and partially overlapping regions of the orofacial prominences that fate mapping revealed contribute to the upper lip and primary palate. Decreased RALDH2 and RARγ result in a median cleft in the upper lip and primary palate. To further understand how retinoic acid regulates upper lip and palate morphogenesis we searched for genes downregulated in response to RARγ inhibition in orofacial tissue, and uncovered homeobox genes lhx8 and msx2. These genes are both expressed in overlapping domains with RARγ, and together their loss of function also results in a median cleft in the upper lip and primary palate. Inhibition of RARγ and decreased Lhx8/Msx2 function result in decreased cell proliferation and failure of dorsal anterior cartilages to form. These results suggest a model whereby retinoic acid signaling regulates Lhx8 and Msx2, which together direct the tissue growth and differentiation necessary for the upper lip and primary palate morphogenesis. This work has the potential to better understand the complex nature of the upper lip and primary palate development which will lead to important insights into the etiology of human orofacial clefts.

Retinoic acid regulates Lhx8 expression via FGF-8b to the upper jaw development of chick embryo

“…Thus, the upregulation of Lhx8 by enhanced FGF signaling and its down-regulation by blocking of the FGF receptor, indicate that Lhx8 transcription is controlled by FGF signaling during growth of the maxillary prominence. It has been suggested that RA signaling regulates Msx2 and Lhx8 expressions directly or indirectly by activating intermediate pathway such as FGF (14). Our findings confirm that Lhx8 expression is primarily controlled by the FGF signaling pathway and depends on local RA concentration.…”

“…Insufficient levels of RA may induce the downregulation of those genes lead to specific defects on the maxilla because Msx gene products have a role on mediating the effect of RA on facial development (19). A study in Xenopus embryo suggests a model whereby RA signaling regulates Lhx8 and Msx2, which together direct the upper lip and palate morphogenesis (14). RA homeostasis appears to be vital for normal Lhx8 expression as both addition and inhibition of RA leads to aberrant expression of Lhx8.…”

“…Mutation of these genes has been associated with non syndromic cleft lip with or without cleft palate (13). A study at Xenopus suggests RA regulate Lhx8 and Msx2 directly or indirectly in the upper lip and primary palate development (14). We investigated whether RA signaling controls Lhx8 expression in craniofacial development.…”

A homozygous 237-kb deletion at 1p31 identified as the locus for midline cleft of the upper and lower lip in a consanguineous family

“…10 Loss of function of lhx8 (together with msx2) in frogs resulted in a median cleft in the upper lip and primary palate; both genes were suggested to be regulated by retinoic acid signalling. 11 Thus, animal studies indicate that the gene has a role in orofacial development and is a very good candidate for the trait observed in the presented family. Additional research could help determine whether or not the 273-kb deletion sequences contain any elements that regulate LHX8.…”

Facial Transplants in <em>Xenopus laevis</em> Embryos

“…Xenopus laevis is an ideal model for study of craniofacial development as embryos are large, develop externally, and the face is readily visible, allowing micromanipulation and imaging of development. Mechanisms underlying facial development appear conserved, indicating that findings made in the frog provide insight into human development 4,[15][16] .…”