Decoding protein phosphorylation during oocyte meiotic divisions using phosphoproteomics

Reviewer #1 (Public review):

Summary:

The study aims to create a comprehensive repository about the changes in protein abundance and their modification during oocyte maturation in Xenopus laevis.

Strengths:

The results contribute meaningfully to the field.

Weaknesses:

The manuscript could have benefitted from more comprehensive analyses and clearer writing. Nonetheless, the key findings are robust and offer a valuable resource for the scientific community.

Reviewer #2 (Public review):

Summary:

The authors analyzed Xenopus oocytes at different stages of meiosis using quantitative phosphoproteomics. Their advanced methods and analyses revealed changes in protein abundances and phosphorylation states to an unprecedented depth and quantitative detail. In the manuscript they provide an excellent interpretation of these findings putting them in the context of past literature in Xenopus as well as in other model systems.

Strengths:

High quality data, careful and detailed analysis, outstanding interpretation in the context of the large body of the literature.

Weaknesses:

Merely a resource, none of the findings are tested in functional experiments.

I am very impressed by the quality of the data and the careful and detailed interpretation of the findings. In this form the manuscript will be an excellent resource to the cell division community in general, and it presents a very large number of hypotheses that can be tested in future experiments.

Xenopus has been and still is a popular and powerful model system that led to critical discoveries around countless cellular processes, including the spindle, nuclear envelope, translational regulation, just to name a few. This also includes a huge body of literature on the cell cycle describing its phosphoregulation. It is indeed somewhat frustrating to see that these earlier studies using phospho-mutants and phospho-antibodies were just scratching the surface. The phosphoproteomics analysis presented here reveals much more extensive and much more dynamic changes in phosphorylation states. Thereby, in my opinion, this manuscript opens a completely new chapter in this line of research, setting the stage for more systematic future studies.

Reviewer #3 (Public review):

Summary:

The authors performed time-resolved proteomics and phospho-proteomics in Xenopus oocytes from prophase I through the MII arrest of the unfertilized egg. The data contains protein abundance and phosphorylation sites of a large number set of proteins at different stages of oocyte maturation. The large sets of the data are of high quality. In addition, the authors discussed several key pathways critical for the maturation. The data is very useful for the researchers not only researchers in Xenopus oocytes but also those in oocyte biology in other organisms.

Strengths:

The data of proteomics and phospho-proteomics in Xenopus oocyte maturation is very useful for future studies to understand molecular networks in oocyte maturation.

Weaknesses:

Although the authors offered molecular pathways of the phosphorylation in the translation, protein degradation, cell cycle regulation, and chromosome segregation. The author did not check the validity of the molecular pathways based ontheir proteomic data by the experimentation.

Author response:

We are both honored and humbled by the high praise our work received from all three reviewers. Below, we address the common comments made by the reviewers:

(1) Value and Impact of the Resource: We are grateful for the recognition of our dataset as a valuable and high-quality resource. Our primary goal was to generate a comprehensive dataset on protein abundance and phosphorylation dynamics during Xenopus oocyte maturation. We are pleased that this work has been seen as a solid foundation for future studies in Xenopus research and beyond, with broader implications for oocyte and cell cycle biology.

(2) Focus on Functional Validation and Contextualization with Prior Studies: The manuscript was submitted as a Tools and Resources article, a format that emphasizes the creation and presentation of datasets, tools, and methodological advances to facilitate future discoveries. In alignment with this format, we ensured that the information is accessible and deployable for the broader scientific community. While we did not include functional validation of specific pathways, the dataset provides a robust framework for generating numerous testable hypotheses. We plan to pursue some of these follow-up studies in our labs and encourage the community to explore these further.

(3) Contextualization with Prior Studies: We appreciate the recognition of our efforts to integrate our findings with the existing body of literature. In conclusion, we would like to thank the reviewers for their evaluation and thoughtful suggestions. We look forward to seeing how this dataset contributes to future discoveries in the field.