PREreview of On the role of mechanical feedback in synchronous to asynchronous transition during embryogenesis

Summary and Overall Assessment

The 2023 paper by A. Malmi-Kakkada et al. explores the physical mechanism underling transitions from synchronous to asynchronous (SAT) cell divisions during embryogenesis. The authors propose that mechanical feedback plays a critical role in regulating this transition, contributing to the robustness and variability observed during embryonic development. Using a combination of computational modeling and experimental validation, the study provides an analysis of how mechanical interactions influence cell cycle dynamics and tissue morphogenesis.

Overall, this study makes a valuable contribution to understanding the role of mechanical feedback in embryogenesis. The paper is well-structured and presents a compelling argument for the importance for the role of mechanical feedback in SAT. By combining computational models with experimental validation, the authors provide robust evidence for the significance of mechanical forces in regulating cell cycle dynamics, and the agreement between numerical simulations and experimental data strengthens the validity of the findings. However, there are several areas where the paper could benefit from further clarification and detail, particularly in the methodology and interpretation of results, a deeper discussion of the biological implications, and clearer justification of the experimental approaches used.

Major Points

1.     Methodological Clarity and Detail

1. The modelling description, including parameter selection and sensitivity analyses, is somewhat lacking e.g. boundary conditions should be specified. Although the supplement provides a more detailed explanation of the model setup, the abstract opens with mention of an agent-based framework without further elaboration. Perhaps the connection between equations of motion driving active cell division in the model should be clarified sooner and contextualized within the theory derived in the body of the paper.

2. Reference to the workflow from experimental methods to data analysis, with particular emphasis on techniques used to measure cell cycle progression would help facilitate better understanding of statistical calculations.

3. In the supplement, further explanation as to the versatility of the pressure term across different collectively growing systems is necessary to establish the relationship between multicellular spheroids and zebrafish embryogenesis. For instance, are there also shared assumptions about confinement, cell sorting, directional order etc.?

4. A death rate is defined in TABLE I but there is no mention of turnover or apoptosis affecting cellular dynamics or SAT. Given that the only reference is this paper, more motivation for this term is necessary. Is death a stochastic variable or do the same number of cells die in each cycle? Are cells removed from the aggregate and then immediately replaced by two daughter cells? The section “Cell birth, apoptosis and dormancy” is ambiguous. In the statement “Depending on the local forces, a cell can either be in the dormant (D) or in the growth (G) phase,” should apoptosis be its own phase, or does dormancy lead to death in which case these terms being used to describe timed effects?

2.     Interpretation of Results

1. The link between mechanical pressure and rapid cell division is not clear early in the paper. It would be beneficial to explicitly connect mechanical pressure with contact inhibition when discussing feedback signals regulating cell division.

2. The fit in Fig. 1C is only shown over the last four data points. If the fit includes more points, this should be clarified. If not, the rationale for focusing on these four points should be explained.

3. The statement "Variability in cell cycle duration has been observed long ago, but its origin is unclear. Our theory suggests that it is driven by fluctuations in cell division times." appears tautological. Clarifying the distinction between cell division and cell cycle, and how fluctuations in division times drive variability in the cycle, would improve understanding.

3.     Support for Conclusions

1. Discuss potential alternative explanations and limitations of the study, providing a balanced view of the findings. This would add nuance to the conclusions drawn about the influence of mechanical feedback on cell division synchronization.

2. While the stated objective is to provide a predictive theory for the number of divisions it takes Zebrafish to exhibit SAT, it would strengthen the conclusion to discuss generalizability of the model to other systems where N(t) data is available, such as the worm embryo, and demonstrate broader applicability beyond the case study from Keller’s 2008 paper.

Minor Points

1.     Presentation and Readability

1. Ensure figures and tables have descriptive captions. Including brief summaries of key findings within the figure captions would help readers quickly grasp the main points. For example, if Fig. 3 and Fig. 4a are based on simulations, this should be clearly stated in the captions.

2. Fig 3 axes labels should be enlarged.

2.     Literature Context

1. Include more recent studies that have investigated similar phenomena to place the current work in a broader context and highlight its novelty and significance.

2. It would be helpful to add a citation that justifies the fit of cell division times to a normal distribution.

Reviewer Expertise Statement

This review is informed by expertise in optical imaging and numerical modelling of embryogenesis.

Competing interests

The authors declare that they have no competing interests.