A complex oscillating network of signaling genes underlies the mouse segmentation clock

The segmental pattern of the spine is established early in development, when the vertebral precursors, the somites, are rhythmically produced from the presomitic mesoderm. Microarray studies of the mouse presomitic mesoderm transcriptome reveal that the oscillator associated with this process, the segmentation clock, drives the periodic expression of a large network of cyclic genes involved in cell signaling. Mutually exclusive activation of the notch–fibroblast growth factor and Wnt pathways during each cycle suggests that coordinated regulation of these three pathways underlies the clock oscillator. The segmentation clock drives the expres-sion of a very limited number of geneswhose mRNA shows a dynamic expres-sion sequence that is repeated in the presomitic mesoderm (PSM) each time a new somite forms (1). Most of the known cyclic genes are com-ponents of the notch pathway oscillating in phase with each other. In chick and mouse, they include genes coding for transcription factors of the hairy and enhancer of split (Hes) family and the lunatic fringe (Lfng) glycosyltransferase (1). Additionally, in mouse, a single component of the Wnt pathway, Axin2, oscillates out of phase with the notch pathway cyclic genes (2). We have used gene expression arrays sys-tematically to explore the cyclic transcription program associated with the segmentation clock in the mouse PSM. During the formation of each somite, Lfng is expressed in the PSM as a wave that sweeps across the tissue in a posterior-to-anterior direction (1). Therefore, by visually comparing the anteroposterior position of the Lfng expression stripes in the PSM in stained embryos, it is possible to define an approximate chronological order of the embryos along the segmentation clock oscillation cycle (3, 4). We collected PSM samples from 40 mouse embryos ranging from 19 to 23 somites and used their Lfng expression patterns as a proxy to select 17 samples covering an entire oscillation cycle (fig. S1 and Figs. 1 and 2, A and B). Probes were produced from RNA extracted from the dis-sected PSMs by using a two-step amplification protocol and were hybridized to Affymetri