ß 2006 Wiley-Liss, Inc. American Journal of Medical Genetics Part A 140A:1366–1374 (2006) Research Review Dlx Homeobox Gene Control of Mammalian Limb and Craniofacial Development Petra Kraus and Thomas Lufkin* Stem Cell and Developmental Biology, Genome Institute of Singapore, Singapore Received 21 September 2005; Accepted 3 October 2005 The Dlx homeobox gene family is of ancient origin and crucial for embryonic development in invertebrates and vertebrates. The Dlx proteins are thought to function as DNA-binding transcriptional regulators, likely controlling large numbers of downstream effector genes. In mammals gene expression analysis of the six members of the Dlx gene family has been demonstrated in the nervous system, neural crest derivatives, branchial arches, and developing appen- dages. Through genetic approaches the Dlx genes have been implicated in patterning and development of the brain, craniofacial structures, and the axial and appendicular skeleton. Substantial functional redundancy within the Dlx gene family has prevented the analysis of single gene mutations from demonstrating the full developmental con- trol exerted by the Dlx proteins. Here, we will discuss data from recent combined loss and gain-of-function genetic mutations, which have given greater insight into the role of Dlx homeobox genes in craniofacial, limb, and bone development. ß 2006 Wiley-Liss, Inc. Key words: distal-less; homeodomain; limb; AER; SHFM; bone; embryo; development; patterning; ectrodactyly How to cite this article: Kraus P, Lufkin T. 2006. Dlx homeobox gene control of mammalian limb and craniofacial development. Am J Med Genet Part A 140A:1366 – 1374. INTRODUCTION An evolutionary conserved group of homeobox containing transcription factors, the Dlx gene family, is represented by three bigene clusters in vertebrates: Dlx1/Dlx2, Dlx3/Dlx7 and Dlx5/Dlx6, all homologs of the Drosophila distal-less (dll) gene. Gene expression analysis of members of the Dlx gene family has demonstrated expression in the developing nervous system, neural crest deriva- tives, branchial arches (BAs), and developing appendages. Recently, in support of data acquired in Drosophila demonstrating distal-less as crucial for patterning of distal appendages (limb and anten- nae) [Cohen et al., 1989], the Dlx gene family has also been implicated as important players in the development of distal limb structures and apical ectodermal ridge (AER) maintenance in mouse. A simultaneous targeted interruption of Dlx5 and Dlx6 genes resulted in a phenocopy of the autosomal dominant form of the split-hand/split- foot malformation (SHFM), a heterogeneous limb malformation, characterized by the absence of central digits and the claw-like appearance of the autopod (hands/feet). Dlx HOMEOBOX GENES PLAY AN ACTIVE ROLE IN LIMB PATTERNING During mid-gestation development of the verte- brate embryo, the successful formation of limbs requires outgrowth and patterning along three axes: anterior–posterior (A–P), dorsal–ventral (D–V), and proximal–distal (Pr–Di). This is achieved by the well-orchestrated interaction of signaling centers for each axis: The zone of polarizing activity (ZPA) via Shh signaling directs A–P patterning; the non- ridge limb ectoderm via the transcription factors En1 and Wnt7a determines D – V patterning; and the AER with its long known key players the Fgf family members plays a major role in Pr–Di patterning (Fig. 1). The AER is a transient structure during limb development, a stratified epithelium running along the D – V interface on the distal tip of the mouse limb bud. The AER is the signaling center directing Pr – Di *Correspondence to: Thomas Lufkin, Ph.D., Genome Institute of Singapore, 60 Biopolis Street, 138672 Singapore. E-mail: lufkin@gis.a-star.edu.sg DOI 10.1002/ajmg.a.31252