Unveiling inter-embryo variability in spindle length over time: Towards quantitative phenotype analysis

How can inter-individual variability be quantified? Measuring many features per experiment raises the question of choosing them to recapitulate high-dimensional data. Tackling this challenge on spindle elongation phenotypes, we showed that only three typical elongation patterns describe spindle elongation in C. elegans one-cell embryo. These archetypes, automatically extracted from the experimental data using principal component analysis (PCA), accounted for more than 95% of inter-individual variability of more than 1600 experiments across more than 100 different conditions. The two first archetypes were related to spindle average length and anaphasic elongation rate. The third archetype, accounting for 6% of the variability, was novel and corresponded to a transient spindle shortening in late metaphase, reminiscent of kinetochore function-defect phenotypes. Importantly, these three archetypes were robust to the choice of the dataset and were found even considering only non-treated conditions. Thus, the inter-individual differences between genetically perturbed embryos have the same underlying nature as natural inter-individual differences between wild-type embryos, independently of the temperatures. We thus propose that beyond the apparent complexity of the spindle, only three independent mechanisms account for spindle elongation, weighted differently in the various conditions. Interestingly, the spindle-length archetypes covered both metaphase and anaphase, suggesting that spindle elongation in late metaphase is sufficient to predict the late anaphase length. We validated this idea using a machine-learning approach. Finally, given amounts of these three archetypes could represent a quantitative phenotype. To take advantage of this, we set out to predict interacting genes from a seed based on the PCA coefficients. We exemplified this firstly on the role of tpxl-1 whose homolog tpx2 is involved in spindle microtubule branching, secondly the mechanism regulating metaphase length, and thirdly the central spindle players which set the length at anaphase. We found novel interactors not in public databases but supported by recent experimental publications.Author summary: When quantifying the cell, scientists need to accurately measure cell-to-cell variability as it carries unique information about the underlying mechanisms. In this article, we focused on the spindle length as a proxy for correct mitosis. We used the nematode one-cell embryo, an established model organism, to investigate (stem-like) cell divisions. Through a data-only approach of spindle dynamics over time, we automatically extracted the most informative variability descriptors. We recalled two known ones: spindle length and anaphase elongation rate. We uncovered a new one –late-metaphase shortening –present in all conditions. Such a phenotype was previously confined to cells with defective chromosome attachments. These three descriptors account for 95% of variability, suggesting that the complex spindle choreography relies only on a few core mechanisms. Furthermore, we showed that the final spindle length at anaphase, important to set the daughter cell fates, is already determined in late metaphase, despite a complete spindle reassembly between both phases. Interestingly, the same descriptors explain variability in genetically perturbed and non-treated conditions. This suggests that no novel mechanism appears in defective cells. Only mechanism contributions are changing. Finally, we propose a tool to predict genes co-involved in a mechanism from a known gene to support candidate approaches.