Time-lapse observation of stepwise regression of Erk activity in zebrafish presomitic mesoderm

Abstract

During somite segmentation, clock genes oscillate within the posterior presomitic mesoderm (PSM). The temporal information ties up with the posteriorly moving FGF gradient, leading to the formation of a presumptive somite within the PSM. We previously investigated Erk activity downstream of FGF signaling by collecting stained zebrafish embryos, and discovered that the steep gradient of Erk activity was generated in the PSM, and the Erk activity border regularly shifted in a stepwise manner. However, since these interpretations come from static analyses, we needed to firmly confirm them by applying an analysis that has higher spatiotemporal resolutions. Here we developed a live imaging system for Erk activity in zebrafish embryos, using a Förster resonance energy transfer (FRET)-based Erk biosensor. With this system, we firmly showed that Erk activity exhibits stepwise regression within the PSM. Although our static analyses could not detect the stepwise pattern of Erk activity in clock-deficient embryos, our system revealed that, in clock-deficient embryos, the stepwise regression of Erk activity occurs at an irregular timing, eventually leading to formation of irregularly-sized somites. Therefore, our system overcame the limitation of static analyses and revealed that clock-dependent spatiotemporal regulation of Erk is required for proper somitogenesis in zebrafish.

Size-reduced embryos reveal a gradient scaling based mechanism for zebrafish somite formation

“…Without gradient scaling, the clock and wavefront model predicts a 31 single smaller somite following the induced larger somite, but the size of the following somites 1 immediately returns to normal (Fig. 6L), consistent with previous theoretical work (Baker et al, 2 2006). Interestingly, for a class of mechanisms that assumes that the "size" of a somite is what is 3 determined, rather than the "position" of the next somitic furrow (e.g.…”

“…3P). Recent work using the Erk live reporter showed that this L50 is a good17 approximation of the future somite boundary(Sari et al, 2018). Time course analysis of L50 in 18 both control and chopped embryos confirmed that the Fgf activity gradient scales with PSM size19 (Fig.…”

“…While the scaling of morphogen-based patterning has received significant 31 al., 2006), understanding has been limited for scaling of other patterning processes, such as 1 somite segmentation. 2 During embryogenesis, somites provide the first body segments in vertebrates, eventually 3 giving rise to tissues such as the vertebrae and axial skeletal muscles. Somite segmentation 4 occurs sequentially in an anterior to posterior progression along the presomitic mesoderm (PSM), 5 with temporal and spatial periodicity.…”

“…29 Key aspects of the clock and scaled gradient model are the 4-cycle delay between somite 30 specification and formation, and the feedback between newly formed somites and gradient length. 31 We thus reasoned that if we experimentally created one larger somite, it would shorten the PSM 1 and rescale the gradient in a jump, which would then result in another larger somite four cycles 2 later, and this process would repeat creating "echoes" of larger somites with a ~4-cycle 3 periodicity ( Fig. 6A).…”

Quantitative study of the somitogenetic wavefront in zebrafish

“…The MapK activity boundary then jumps to its rightmost stability boundary. These coordinated jumps of the MapK activity domain have been reported recently (Sari et al, 2018).…”