Periodic Reporting for period 1 - RiboHeterogeneity (Functional analysis of ribosome heterogeneity during zebrafish embryogenesis)
Reporting period: 2021-12-01 to 2023-11-30
Early embryonic development requires the coordination of many cellular processes that enable the formation of a multicellular organism from a single cell. Many of these changes occur at the molecular level and involve the production of specific proteins at the right place and time. To make this possible, the machinery responsible for the synthesis of proteins, the ribosome, must be stored in the oocyte for extended periods of time. Most of the time, ribosomes must be inactive in the oocyte because most proteins are not needed until later in embryogenesis. How ribosomes are stored in the oocyte has been a long-standing question in the field.
The study of ribosomes is important not only in the context of early embryonic development. Ribosomes are abundant in the cells of all living organisms, where they are responsible for the energy-intensive task of making proteins. To regulate energy consumption and control protein levels in cells, ribosomes are subject to regulation. For example, during starvation, ribosomes associate with proteins that block their function to conserve energy. In certain diseases, such as cancer, the number of ribosomes increases to sustain cell proliferation. Therefore, understanding the mechanisms that control ribosome function may have important therapeutic applications.
The embryo is an ideal system to study ribosome regulation because early embryogenesis occurs in the absence of ribosome synthesis. Using zebrafish as a model organism, the goals of this project were (1) to understand how oocytes store ribosomes, and (2) to investigate whether ribosomes play an active role in regulating protein synthesis during early embryogenesis. In addition to studying ribosomes, we also focused on the molecules that carry the instructions for making specific proteins, the messenger RNAs (mRNAs). Like ribosomes, mRNAs must also be stored for long periods of time and activated only at the right time during embryogenesis.
My research has led to the discovery of novel protein factors that associate with egg ribosomes and mRNAs to repress their function while extending their lifespan. These factors transiently bind to ribosomes and mRNAs, making them available for protein synthesis later during embryogenesis. By studying embryogenesis, we have gained insights into novel ways to regulate protein synthesis that can be applied to other systems and cells.
The study of ribosomes is important not only in the context of early embryonic development. Ribosomes are abundant in the cells of all living organisms, where they are responsible for the energy-intensive task of making proteins. To regulate energy consumption and control protein levels in cells, ribosomes are subject to regulation. For example, during starvation, ribosomes associate with proteins that block their function to conserve energy. In certain diseases, such as cancer, the number of ribosomes increases to sustain cell proliferation. Therefore, understanding the mechanisms that control ribosome function may have important therapeutic applications.
The embryo is an ideal system to study ribosome regulation because early embryogenesis occurs in the absence of ribosome synthesis. Using zebrafish as a model organism, the goals of this project were (1) to understand how oocytes store ribosomes, and (2) to investigate whether ribosomes play an active role in regulating protein synthesis during early embryogenesis. In addition to studying ribosomes, we also focused on the molecules that carry the instructions for making specific proteins, the messenger RNAs (mRNAs). Like ribosomes, mRNAs must also be stored for long periods of time and activated only at the right time during embryogenesis.
My research has led to the discovery of novel protein factors that associate with egg ribosomes and mRNAs to repress their function while extending their lifespan. These factors transiently bind to ribosomes and mRNAs, making them available for protein synthesis later during embryogenesis. By studying embryogenesis, we have gained insights into novel ways to regulate protein synthesis that can be applied to other systems and cells.
The results of our egg ribosome study were published this year in Nature. This work was done in collaboration with the Haselbach lab, experts in cryo-electron microscopy (cryo-EM). Friederike Leesch (PhD student) and I were co-first authors and the main driving forces behind this work. My contribution to this work was mainly the modeling and analysis of cryo-EM datasets, which allowed us to visualize how ribosomes of maternal origin “look” at near-atomic resolution. Using this technique, we were able to identify a set of four factors that bind to functionally important sites on the ribosome. Furthermore, using genetics, we were able to conclude that ribosome dormancy is key for embryonic survival. In this way, oocytes and early embryos ensure that the molecular machinery responsible for making proteins is repressed, reducing protein synthesis and saving energy for when it is needed.
In a second independent project, I was able to identify a protein factor, eIF4E1b, that binds to mRNAs that are stored at a specific time during oogenesis and/or early embryogenesis. Normally, mRNAs can be degraded by removing a specific structure at one of their ends, known as the mRNA cap. We report that eIF4E1b is present only in oocytes and early embryos, where it protects mRNAs by binding to their cap, preventing its removal and at the same time repressing mRNA translation. Using zebrafish genetics, cell biology, and biochemistry, we were able to identify an essential role for eIF4E1b in mRNA storage and repression. This work has been recently published in the journal EMBO Reports, with me being first and co-corresponding author.
In a second independent project, I was able to identify a protein factor, eIF4E1b, that binds to mRNAs that are stored at a specific time during oogenesis and/or early embryogenesis. Normally, mRNAs can be degraded by removing a specific structure at one of their ends, known as the mRNA cap. We report that eIF4E1b is present only in oocytes and early embryos, where it protects mRNAs by binding to their cap, preventing its removal and at the same time repressing mRNA translation. Using zebrafish genetics, cell biology, and biochemistry, we were able to identify an essential role for eIF4E1b in mRNA storage and repression. This work has been recently published in the journal EMBO Reports, with me being first and co-corresponding author.
My work has opened up new avenues of research related to the study of ribosome and mRNA dormancy. Our work on the ribosome has led to the discovery of a factor, Dap1b, that had never before been reported to associate with the ribosome. Our laboratory is currently investigating the potential role of Dap1b in other contexts, as we have observed that the presence of Dap1b is sufficient to induce ribosome dormancy in mammalian systems. Furthermore, the small size of Dap1b and its ability to repress protein synthesis make it a good candidate for therapeutics. For example, drugs that reduce protein synthesis have been successfully used to treat cancer.
Similarly, eIF4E1b can not only protect mRNAs, but also prevent them from being used as templates for protein synthesis. Thus, like Dap1b, eIF4E1b may be a potential candidate for regulating protein synthesis in other contexts. In addition, our work has raised new questions in the field, such as how mRNAs are selected for repression and how they are activated at a particular time during development. By providing data sets that identify many of the factors involved in these processes, our work on mRNA dormancy will serve as a starting point for addressing these questions in the future.
Similarly, eIF4E1b can not only protect mRNAs, but also prevent them from being used as templates for protein synthesis. Thus, like Dap1b, eIF4E1b may be a potential candidate for regulating protein synthesis in other contexts. In addition, our work has raised new questions in the field, such as how mRNAs are selected for repression and how they are activated at a particular time during development. By providing data sets that identify many of the factors involved in these processes, our work on mRNA dormancy will serve as a starting point for addressing these questions in the future.