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Abstract

Vertebrate embryos robustly pattern their anteroposterior axis with a relatively invariant species-specific number of somites despite conspecific variation in body size. The periodicity with which somites segment from the anterior end of the unsegmented presomitic mesoderm (PSM) has been linked to the activity of a molecular oscillator known as the segmentation clock, which involves tissue-level oscillatory signaling in the PSM. A striking feature of anteroposterior axis patterning is that somite segmentation occurs concomitantly with axis elongation. The central hypothesis of this thesis posits that axis elongation and somite segmentation are integrated in the PSM via segmentation clock dynamics to ensure the production of a correct number of size-adjusted somites, regardless of PSM size. The work presented here explores this hypothesis in medaka (Oryzias latipes) through two independent projects.

Since axis elongation has not been characterized in medaka, the first project focused on providing the first tissue-level description of elongation and volumetric growth dynamics in this model organism. I used real-time imaging of medaka tail explants during secondary body formation and described a method to segment individual tissues from the resulting time-series data. Morphometric analysis of the paraxial mesoderm, neural tube, and notochord tissues revealed similar elongation rates, but significant tissue-specific differences in volumetric growth, with the notochord exhibiting a significantly higher fold volume increase than other tissues. Additionally, the study revealed that the PSM elongated over time despite a net decrease in its length and, intriguingly, that its elongation was accompanied by volumetric shrinkage.

The second project delved into the integration of axis elongation and somite segmentation in medaka tail explants, with a specific focus on segmentation clock dynamics. I used aphidicolin to disrupt axis elongation within the her7-Venus segmentation clock reporter line and assessed its impact on somite segmentation, in terms of both morphology and tissue-level signaling dynamics. This perturbation resulted in an accelerated reduction in the net length of the PSM, indicating an imbalance between tissue elongation and segmentation. Additionally, real-time imaging and quantitative analysis of Her7-Venus expression in the PSM revealed a reduction in the oscillation period across the tissue, leading to an upward shift in the period gradient. Further examination of phase distribution in the PSM demonstrated that the phase gradient scaled with PSM length, such that a consistent phase amplitude was reached in the tissue irrespective of its size. This finding supports the notion that the phase gradient plays a functional role in the scaling mechanism, ensuring robust patterning of the anteroposterior axis.

To sum up, this thesis sheds light on previously unexplored aspects of medaka development and provides valuable insights into the intricate coordination of tissue-level processes essential for the robust patterning of the anteroposterior axis in vertebrates.