TISSUE-SPECIFIC STEM CELLS Anterior Hox Genes Interact with Components of the Neural Crest Speciﬁcation Network to Induce Neural Crest Fates MINA GOUTI,a JAMES BRISCOE,b ANTHONY GAVALASa aDevelopmental Biology Laboratory, Biomedical Research Foundation of the Academy of Athens (BRFAA), Athens, Greece; bDivision of Developmental Neurobiology, MRC National Institute for Medical Research (NIMR), The Ridgeway, Mill Hill, London, United Kingdom Key Words. Hox genes • Neural crest speciﬁcation • Epithelial mesenchymal transition • Snail genes • Bmp and Notch signaling ABSTRACT Hox genes play a central role in neural crest (NC) pattern- ing particularly in the cranial region of the body. Despite evidence that simultaneous loss of Hoxa1 and Hoxb1 func- tion resulted in NC speciﬁcation defects, the role of Hox genes in NC speciﬁcation has remained unclear due to extended genetic redundancy among Hox genes. To circumvent this problem, we expressed anterior Hox genes in the trunk neural tube of the developing chick embryo. This demonstrated that anterior Hox genes play a central role in NC cell speciﬁcation by rapidly inducing the key transcription factors Snail2 and Msx1/2 and a neural progenitor to NC cell fate switch characterized by cell ad- Disclosure of potential conﬂicts of interest is found at the end of this article. INTRODUCTION Neural crest (NC) cells originate at the interface between the neural plate and the non-neural ectoderm and are character- ized by their multipotency and migratory capacity. They migrate extensively into the developing embryo differentiating into a wide array of cell types including neurons and glia of the peripheral nervous system, pigment cells, and facial carti- lage and bones in the cranial region of the body [1]. NC stem cells appear to be maintained in different tissues of the adult and can generate neurons, Schwann cells, myoﬁbroblasts, chondrocytes, and melanocytes. Their wide developmental potential and regenerative capacity have stimulated interest in using them in stem cell-based therapies but their isolation, expansion, and generation remain challenging [2]. Elucidating the mechanisms underlying their generation could bring their use in regenerative medicine closer. NC cells are speciﬁed in response to extrinsic signals and intrinsic factors. Bone morphogenetic protein (Bmp), Wnt, and Notch signaling have been implicated in NC cell induc- tion through the activation of key transcription factors [3, 4]. Fibroblast growth factor (Fgf) and retinoic acid (RA) signal- ing have been implicated in NC induction concomitantly with inducing posterior fates in the neural plate [5] and their hesion changes and an epithelial-to-mesenchymal transi- tion (EMT). Cells delaminated from dorsal and medial neural tube levels and generated ectopic neurons, glia progenitors, and melanocytes. The mobilization of the NC genetic cascade was dependent upon bone morphogenetic protein signaling and optimal levels of Notch signaling. Therefore, anterior Hox patterning genes participate in NC speciﬁcation and EMT by interacting with NC- inducing signaling pathways and regulating the expression of key genes involved in these processes. STEM CELLS 2011;29:858–870 expression in paraxial mesoderm [6]. Targets of their action in NC induction have not been identiﬁed, but both Fgf and RA signaling regulate expression of Hox genes either directly or through Cdx transcription factors [7–10]. Hox genes are expressed at the time of NC induction [11] but extensive func- tional redundancy among members of this family might have masked their role in this process. A battery of transcription factors such as Msx1/2, Gbx2, Pax3, and Pax7, are induced in the neural plate border in response to signaling from non-neural ectoderm and paraxial mesoderm and establish a region of competence for NC cell speciﬁcation. In turn, a second set of transcription factors such as Snail, Sox9, Sox10, and FoxD3 [12] are induced and imple- ment the genetic program deﬁning NC cells. This includes an epithelial-to-mesenchymal transition (EMT) mediated by reor- ganization of the actin cytoskeleton, loss of epithelial polarity, and alterations in cell adhesion properties [13]. Despite similarities in the molecular mechanisms underly- ing NC speciﬁcation in cranial and trunk regions there are im- portant differences including expression patterns of key genes and the order of their activation during NC speciﬁcation [14– 16]. Several gain and loss-of-function experiments result in distinct phenotypes in the cranial and the trunk region [14, 17–19]. Hox patterning genes are expressed from early neuru- lation stages in the developing neuroepithelium as well as the Author contributions: M.G.: conception and design, collection of data, data analysis and interpretation; J.B.: ﬁnancial support, provision of study material, data analysis and interpretation, manuscript writing; A.G.: conception and design, ﬁnancial support, collection of data, data analysis and interpretation, manuscript writing, ﬁnal approval of manuscript. Correspondence: Anthony Gavalas, Ph.D., Developmental Biology Laboratory, Biomedical Research Foundation of the Academy of Athens (BRFAA), Soranou Ephessiou 4, Athens 11527, Greece. Telephone: 003-210-6597209; Fax: 0030-210-6597545; e-mail: agavalas@bioacademy.gr Received January 11, 2011; accepted for publication March 2, 2011; ﬁrst published online in STEM CELLS EXPRESS March 23, 2011. VC AlphaMed Press 1066-5099/2009/$30.00/0 doi: 10.1002/stem.630 STEM CELLS 2011;29:858–870 www.StemCells.com