Periodic Reporting for period 4 - RiboInflam (Assessing the role of ribosomes and mRNA translation in shaping the inflammatory response)
Periodo di rendicontazione: 2023-07-01 al 2024-12-31
Host responses to infection rely on a complex network of cellular and molecular mechanisms that regulate gene expression both in infected cells and in cells of the immune system. A key but not yet fully understood aspect of these responses is how gene expression is regulated at the level of mRNA translation and how translation interfaces with other cellular processes. mRNA translation is a fundamental biological process in which ribosomes decode messenger RNAs (mRNAs) to produce functional proteins. This process is highly dynamic, allowing rapid cellular adaptation to environmental changes. Being in direct contact with both the mRNA and the nascent protein, ribosomes act as central hubs for diverse regulatory mechanisms. They can mediate post-translational modifications, influence mRNA stability, and decode specific mRNA sequences or chemical modifications to adjust protein synthesis accordingly. Understanding these regulatory layers is important for deciphering the fine-tuning of immune cell activation and host-pathogen interactions, with potential implications for developing new therapeutic strategies for infectious diseases, immune disorders, and cancer.
The RIBOINFLAM project aimed to investigate the role of ribosomes and translation in shaping immune responses in both virus-infected and immune cells. Its overall objectives were to 1: Characterize the precise composition of ribosomes in cells of the immune system and in virus-infected cells to identify new actors that regulate the translation process and potentially uncover new roles for ribosomes in regulating other cellular related processes; 2. Study the relationship between mRNA translation and the degradation of the mRNA substrate; 3. Study the cross-talk between the two ribosome machineries present within cells (one residing in the cytosol to produce the bulk of cellular proteins and the other residing in the mitochondria to produce a handful of proteins involved in cellular metabolism) during activation of immune cells, which undergo a strong metabolic change to mount their effector functions.
The results obtained during the RIBOINFLAM project led to the identification of hundreds of new ribosome-associated proteins. We also showed that the composition of ribosomes is strongly reshuffled during immune cell activation and following virus infection, notably identifying new ribosome-associated proteins acting as positive or negative regulators of virus replication. We also uncovered rules that govern the relationship between translation and mRNA degradation in immune cells as well as their dynamics during the course of cell activation. Finally, we discovered a specific role for mitochondrial translation in the survival of a specific subset of T lymphocytes. Overall, our findings contributed to expanding the roles of ribosomes and mRNA translation in regulating gene expression, particularly during host-pathogen interactions.
Several results from the project are still part of ongoing studies and will be published in future research articles.
The RIBOINFLAM project aimed to investigate the role of ribosomes and translation in shaping immune responses in both virus-infected and immune cells. Its overall objectives were to 1: Characterize the precise composition of ribosomes in cells of the immune system and in virus-infected cells to identify new actors that regulate the translation process and potentially uncover new roles for ribosomes in regulating other cellular related processes; 2. Study the relationship between mRNA translation and the degradation of the mRNA substrate; 3. Study the cross-talk between the two ribosome machineries present within cells (one residing in the cytosol to produce the bulk of cellular proteins and the other residing in the mitochondria to produce a handful of proteins involved in cellular metabolism) during activation of immune cells, which undergo a strong metabolic change to mount their effector functions.
The results obtained during the RIBOINFLAM project led to the identification of hundreds of new ribosome-associated proteins. We also showed that the composition of ribosomes is strongly reshuffled during immune cell activation and following virus infection, notably identifying new ribosome-associated proteins acting as positive or negative regulators of virus replication. We also uncovered rules that govern the relationship between translation and mRNA degradation in immune cells as well as their dynamics during the course of cell activation. Finally, we discovered a specific role for mitochondrial translation in the survival of a specific subset of T lymphocytes. Overall, our findings contributed to expanding the roles of ribosomes and mRNA translation in regulating gene expression, particularly during host-pathogen interactions.
Several results from the project are still part of ongoing studies and will be published in future research articles.
Throughout the RIBOINFLAM project, we explored how cells regulate protein production during immune responses and viral infections. Our research focused on three key questions, leading to the following results:
1. How do ribosomes change during viral infections and immune responses?
During the project, we developed protocols to purify ribosomes in cultured cells and in vivo, allowing us to analyze how their composition changes in response to infections and immune activation. Using these protocols, we identified host cell factors that help or hinder viral replication. In addition, we explored how immune cells adjust their ribosomes when they are activated to fight infections, discovering unexpected factors with potential new roles in immune defense.
2. How does protein production influence the lifespan of genetic messages?
Cells do not uniquely synthesize mRNAs and proteins, they also break them down to maintain balance. Our research aimed to better understand how ribosomes influence the process of mRNA degradation. Scientists knew that translating an mRNA into a protein can also mark it for degradation, but the precise rules governing this were unclear. Using computational and molecular techniques to track how long different mRNAs survive we discovered that certain genetic features—such as how ribosomes interact with an mRNA, the length of regulatory regions, and the composition of genetic "words" (codons), determine how quickly an mRNA is degraded. We also found that translation regulates mRNA stability differently when T lymphocytes are activated, a discovery with potential implications for mRNA vaccine design.
3. How do two different protein-making systems work together in immune cells?
Our cells have two types of ribosomes: cytosolic ribosomes, which produce most cellular proteins, and mitochondrial ribosomes, which produce a small number of proteins essential for energy production. Although these two systems are separated, we explored whether they communicate and influence each other during immune responses. To investigate this, we created genetically modified mice in which mitochondrial ribosome function could be selectively turned off in specific immune cells as well as genetically modified mice in which mitochondrial ribosomes have been modified to include a “handle” that allows their easy purification to study their composition. Our experiments revealed that mitochondrial protein production is crucial for maintaining a special type of immune cell called "Virtual Memory" CD8+ T lymphocytes. We are now working to understand exactly how mitochondrial ribosomes support these cells.
Some of the above mentioned results have been published in peer-reviewed journals or patented, while others are currently being prepared for publications or are still in a preliminary phase, requiring additional research work before being submitted for publication. Finally, during the project, we continuously presented our results at conferences and invited seminars.
1. How do ribosomes change during viral infections and immune responses?
During the project, we developed protocols to purify ribosomes in cultured cells and in vivo, allowing us to analyze how their composition changes in response to infections and immune activation. Using these protocols, we identified host cell factors that help or hinder viral replication. In addition, we explored how immune cells adjust their ribosomes when they are activated to fight infections, discovering unexpected factors with potential new roles in immune defense.
2. How does protein production influence the lifespan of genetic messages?
Cells do not uniquely synthesize mRNAs and proteins, they also break them down to maintain balance. Our research aimed to better understand how ribosomes influence the process of mRNA degradation. Scientists knew that translating an mRNA into a protein can also mark it for degradation, but the precise rules governing this were unclear. Using computational and molecular techniques to track how long different mRNAs survive we discovered that certain genetic features—such as how ribosomes interact with an mRNA, the length of regulatory regions, and the composition of genetic "words" (codons), determine how quickly an mRNA is degraded. We also found that translation regulates mRNA stability differently when T lymphocytes are activated, a discovery with potential implications for mRNA vaccine design.
3. How do two different protein-making systems work together in immune cells?
Our cells have two types of ribosomes: cytosolic ribosomes, which produce most cellular proteins, and mitochondrial ribosomes, which produce a small number of proteins essential for energy production. Although these two systems are separated, we explored whether they communicate and influence each other during immune responses. To investigate this, we created genetically modified mice in which mitochondrial ribosome function could be selectively turned off in specific immune cells as well as genetically modified mice in which mitochondrial ribosomes have been modified to include a “handle” that allows their easy purification to study their composition. Our experiments revealed that mitochondrial protein production is crucial for maintaining a special type of immune cell called "Virtual Memory" CD8+ T lymphocytes. We are now working to understand exactly how mitochondrial ribosomes support these cells.
Some of the above mentioned results have been published in peer-reviewed journals or patented, while others are currently being prepared for publications or are still in a preliminary phase, requiring additional research work before being submitted for publication. Finally, during the project, we continuously presented our results at conferences and invited seminars.
The aim of the RIBOINFLAM project was to characterize the role of ribosomes and mRNA translation during viral infections and in cells of the immune system. Results obtained during the project have advanced our understanding of how cells regulate protein production in response to infections and immune activation. By uncovering new roles for ribosomes and their associated factors, our work revealed unexpected mechanisms that influence viral replication and immune cell function. We also demonstrated how cells rapidly adjust protein production by coupling it to the degradation of messenger RNAs, results with potential implications for improving mRNA-based vaccines and therapies. Furthermore, RIBOINFLAM opened new research avenues on the critical role of mitochondrial protein synthesis in supporting specific immune cells that protect the body against infections and cancer. Finally, RIBOINFLAM allowed us to develop innovative "virus inspired" tools for the delivery of proteins into cell that enable precise genetic modification of primary cells and tissues.
Although the RIBOINFLAM project has officially ended, our research group will continue to investigate the aspects that could not be fully explored within the project's timeframe, as well as the many new questions that emerged from our work.
Although the RIBOINFLAM project has officially ended, our research group will continue to investigate the aspects that could not be fully explored within the project's timeframe, as well as the many new questions that emerged from our work.