C OMMUNICATION A Molecular Clock Operates During Chick Autopod Proximal-distal Outgrowth Susana Pascoal 1 , Cláudia R. Carvalho 2 , Joaquín Rodriguez-León 2 Marie-Claire Delfini 3 , Delphine Duprez 3 , Sólveig Thorsteinsdóttir 2,4 and Isabel Palmeirim 1 ⁎ 1 Life and Health Sciences Research Institute (ICVS), School of Health Sciences, University of Minho, 4710-057 Braga, Portugal 2 Instituto Gulbenkian de Ciência, R. Quinta Grande 6, 2780-156 Oeiras, Portugal 3 CNRS, UMR 7622, Biologie du développement, Université Pierre et Marie Curie, Campus de Jussieu, 9 quai Saint Bernard, Bâtiment C, 6ème Etage, Case 24, 75252 Paris Cedex 05 - France 4 Departamento de Biologia Animal, Centro de Biologia Ambiental, Faculdade de Ciências, Universidade de Lisboa, 1740-016 Lisboa, Portugal Temporal control can be considered the fourth dimension in embryonic development. The identification of the somitogenesis molecular clock provided new insight into how embryonic cells measure time. We provide the first evidence of a molecular clock operating during chick fore-limb autopod outgrowth and patterning, by showing that the expression of the somitogenesis clock component hairy2 cycles in autopod chondrogenic precursor cells with a 6 h periodicity. We determined the length of time required to form an autopod skeletal limb element, and established a correlation between the latter and the periodicity of cyclic hairy2 gene expression. We suggest that temporal control exerted by cyclic gene expression can be a widespread mechanism providing cellular temporal information during vertebrate embryonic development. © 2007 Elsevier Ltd. All rights reserved. *Corresponding author Keywords: hairy2; molecular clock; limb development; cycling genes; vertebrate embryo Embryo limb development requires precise orchestration of cell proliferation and differentiation in time and space. 1 Limb skeletal elements emerge as cartilaginous primordia in a proximal-distal (p-d) sequence. Two models seek to explain cell fate specification along the p-d limb axis. Although fundamentally different, both models imply the existence of a limb bud distal zone where cells reside until they reach the time to differentiate, the progress zone (PZ) model, 2 or to expand, the early specification model. 3,4 However, how these cells measure time is not known. Time control during embryo development is particularly evident during somitogenesis. Chick presomitic cells were shown to undergo several cycles of hairy1 gene expression, providing evi- dence for the existence of a molecular clock underlying the rhythm of somitogenesis. 5 An increasing number of genes have been implicated in the molecular clock machinery; 6,7 namely, hairy2, Present addresses: C. R. Carvalho, Departamento de Morfologia, ICB, Universidade Federal de Minas Gerais, 31270-901, Belo Horizonte, MG, Brazil; J. Rodriguez-León, Center of Regenerative Medicine in Barcelona, Dr. Aiguader 80, 08003 Barcelona, Spain; M.-C. Delfini, Developmental Biology Institute of Marseille, Laboratoire de Génétique et Physiologie du Développement, Campus de Luminy, Case 907, University of Aix-Marseille II, 13288 Marseille Cedex 09, France. Abbreviations used: p-d, proximal-distal; AER, apical ectodermal ridge. E-mail address of the corresponding author: ipalmeirim@ecsaude.uminho.pt doi:10.1016/j.jmb.2007.01.089 J. Mol. Biol. (2007) 368, 303–309 0022-2836/$ - see front matter © 2007 Elsevier Ltd. All rights reserved.