Setting the Tempo in Development: An Investigation of the Zebrafish Somite Clock Mechanism

Abstract

The somites of the vertebrate embryo are clocked out sequentially from the presomitic mesoderm (PSM) at the tail end of the embryo. Formation of each somite corresponds to one cycle of oscillation of the somite segmentation clock—a system of genes whose expression switches on and off periodically in the cells of the PSM. We have previously proposed a simple mathematical model explaining how the oscillations, in zebrafish at least, may be generated by a delayed negative feedback loop in which the products of two Notch target genes, her1 and her7, directly inhibit their own transcription, as well as that of the gene for the Notch ligand DeltaC; Notch signalling via DeltaC keeps the oscillations of neighbouring cells in synchrony. Here we subject the model to quantitative tests. We show how to read temporal information from the spatial pattern of stripes of gene expression in the anterior PSM and in this way obtain values for the biosynthetic delays and molecular lifetimes on which the model critically depends. Using transgenic lines of zebrafish expressing her1 or her7 under heat-shock control, we confirm the regulatory relationships postulated by the model. From the timing of somite segmentation disturbances following a pulse of her7 misexpression, we deduce that although her7 continues to oscillate in the anterior half of the PSM, it governs the future somite segmentation behaviour of the cells only while they are in the posterior half. In general, the findings strongly support the mathematical model of how the somite clock works, but they do not exclude the possibility that other oscillator mechanisms may operate upstream from the her7/her1 oscillator or in parallel with it. Somites—the embryonic segments of the vertebrate body—are formed sequentially, with a spacing determined by a gene expression oscillator, the segmentation clock, operating in the cells at the tail end of the embryo. This system provides a rare opportunity to analyse how the timing of at least one set of developmental events is controlled. We previously proposed a mathematical model, showing how the oscillations could be generated by a delayed negative feedback loop, in which the products of two genes, her1 and her7, act as inhibitors of their own expression, and how Notch signalling between adjacent cells keeps their individual oscillations synchronised. Here we test and find support for this model in two ways. First, we show how to use the spatial pattern of gene expression to measure some of the temporal delays and molecular lifetimes that are critical for the occurrence of synchronised oscillations. Second, we use transgenic fish in which expression of her1 or her7 can be switched on at will by heat shock to probe the dynamics of the system and to analyse the logic of the control circuitry.