Periodic Reporting for period 1 - MITGEST (Quality Control of the Mitochondrial Gene Expression System in Health and Disease)
Reporting period: 2022-10-01 to 2024-09-30
The MITGEST doctoral network aims to improve our understanding of mitochondrial function and dysfunction, with a particular focus on mitochondrial DNA (mtDNA) and its gene expression. The main goal of the project is to address the challenges associated with mitochondrial dysfunction underlying a broad spectrum of human diseases, including rare diseases, aging-related conditions and cancer. By filling the gaps in understanding the pathophysiology of mitochondrial diseases, MITGEST intends to foster innovation in diagnostic and therapeutic strategies while training the next generation of scientists through a multidisciplinary programme.
The main objectives of MITGEST are: 1) to explore the quality control mechanisms of mtDNA and its expression; 2) to develop innovative methods to study and treat mitochondrial abnormalities; 3) to identify biomarkers and treatment options for mitochondrial dysfunction. This comprehensive initiative combines scientific research with transferable skills training emphasizing collaboration between academia and industry to create a sustainable and impactful research environment.
MITGEST will unite world-class academic excellence in mtDNA maintenance and expression with private sector expertise in nucleic acid chemistry and the development of mitochondrial therapeutics to address this urgent need. As the first research training programme in this field to take a fully integrated approach, our 11 doctoral candidates (DCs) will elucidate how mtDNA is maintained and expressed and support the development of new methods to analyse RNA and DNA metabolism in mitochondria. These advances will complement and facilitate/enhance innovative treatment strategies and experimental medicine studies designed to mitigate mitochondrial dysfunction in a range of human diseases.
Our network builds on proven collaborations and establishes new partnerships to deliver intersectoral and multidisciplinary scientific and transferable skills training to our 11 PhD students to generate the next generation of highly-skilled scientists able to exploit the latest advances in mitochondrial biology and medicine.
The main objectives of MITGEST are: 1) to explore the quality control mechanisms of mtDNA and its expression; 2) to develop innovative methods to study and treat mitochondrial abnormalities; 3) to identify biomarkers and treatment options for mitochondrial dysfunction. This comprehensive initiative combines scientific research with transferable skills training emphasizing collaboration between academia and industry to create a sustainable and impactful research environment.
MITGEST will unite world-class academic excellence in mtDNA maintenance and expression with private sector expertise in nucleic acid chemistry and the development of mitochondrial therapeutics to address this urgent need. As the first research training programme in this field to take a fully integrated approach, our 11 doctoral candidates (DCs) will elucidate how mtDNA is maintained and expressed and support the development of new methods to analyse RNA and DNA metabolism in mitochondria. These advances will complement and facilitate/enhance innovative treatment strategies and experimental medicine studies designed to mitigate mitochondrial dysfunction in a range of human diseases.
Our network builds on proven collaborations and establishes new partnerships to deliver intersectoral and multidisciplinary scientific and transferable skills training to our 11 PhD students to generate the next generation of highly-skilled scientists able to exploit the latest advances in mitochondrial biology and medicine.
In the first reporting period, successfully recruited 11 Doctoral Candidates who have been well integrated in their respective research groups and into the network as a whole. The supervision arrangements were established and the DCs were enrolled in their respective PhD programmes. MITGEST already delivered a collection of scientific and transferable skills training at the network level through two in-person and three online meetings.
The four scientific work packages (WPs), focused on different aspect of mitochondrial biology, have already produced the first results. In WP1, Biology of Mitochondrial DNA and RNA, a protocol to label, isolate and analyse nascent mitochondrial transcripts has been established. In parallel, another protocol to identify proteins associated with mtRNA has been developed and successfully applied to identify a list of potential mtRNA interacting protein partners. In WP2, Mitochondrial Gene Expression Functionality, a complexome profiling of mitochondrial nucleic-acid protein complexes has been developed to facilitate the analysis of mitochondrial protein-nucleic acid interactions. To study the mitochondria-nucleus retrograde signalling under pathophysiological conditions, cell lines with different mtDNA repair capabilities were developed and transcriptome analyses will be performed in RP2 to determine metabolic pathways activated by mitochondria-nucleus feedback under physiological and stress conditions. The final part of this WP is related to the biochemical and structural characterization of mitoribosome rescue factors. On this regard, two factors have been identified that rescue trapped ribosomes during translation in mitochondria. All these methodological approaches developed in WPs 1 and 2 will be used in the RP2 for deeply investigate the mechanisms of mtDNA expression and the regulatory pathways activated in response to mitochondrial/nucleus crosstalk.
WP3, Mitochondrial Disease Models and Therapy, is a clinically oriented WP with a specific focus on the analysis of cellular response to mtDNA perturbation and their consequences on organ pathology and the impact of nutrients on mitochondrial DNA maintenance and expression. The activities are in close continuity with WP1 and 2 with the final goal to achieve a proof-of-concept for the treatments of mtDNA disorders. In this WP, key advances have come from the study of the Mpv17 deficient mouse that is a model of human mitochondrial DNA depletion syndrome. These studies have enabled investigate the cellular responses to mtDNA perturbation and the consequences for organ pathology, to develop small molecules therapies for mtDNA disorders, and to evaluate how energy production respiratory complexes are altered in disease states. We have also begun to create the first iPSC line from a mitochondrial disease patient.
The last scientific WP4, New Methods and Tools for Mitochondrial Studies, is a methodological WP focused on developing new methods for mitochondrial studies. With its technical nature, this WP is aimed at the development, improvement and implementation of protocols for the isolation of proteins on nascent RNA, cryo-EM studies of mitoribosome structure, delivery of active oligonucleotides within the mitochondria, and development of quantitative assay for measuring mitochondrial dynamics. During the RP1, click chemistry approaches for tagging nascent RNA and conjugation of nucleic acids with novel mitochondrial targeting agents have been implemented as well as purifications of human mitoribosomes from HEK293 cells and Jurkat cell lines followed by cryoEM. Overall, WP4 has made substantial progress in developing and optimizing methods for mitochondrial studies that will be applied in RP2.
The four scientific work packages (WPs), focused on different aspect of mitochondrial biology, have already produced the first results. In WP1, Biology of Mitochondrial DNA and RNA, a protocol to label, isolate and analyse nascent mitochondrial transcripts has been established. In parallel, another protocol to identify proteins associated with mtRNA has been developed and successfully applied to identify a list of potential mtRNA interacting protein partners. In WP2, Mitochondrial Gene Expression Functionality, a complexome profiling of mitochondrial nucleic-acid protein complexes has been developed to facilitate the analysis of mitochondrial protein-nucleic acid interactions. To study the mitochondria-nucleus retrograde signalling under pathophysiological conditions, cell lines with different mtDNA repair capabilities were developed and transcriptome analyses will be performed in RP2 to determine metabolic pathways activated by mitochondria-nucleus feedback under physiological and stress conditions. The final part of this WP is related to the biochemical and structural characterization of mitoribosome rescue factors. On this regard, two factors have been identified that rescue trapped ribosomes during translation in mitochondria. All these methodological approaches developed in WPs 1 and 2 will be used in the RP2 for deeply investigate the mechanisms of mtDNA expression and the regulatory pathways activated in response to mitochondrial/nucleus crosstalk.
WP3, Mitochondrial Disease Models and Therapy, is a clinically oriented WP with a specific focus on the analysis of cellular response to mtDNA perturbation and their consequences on organ pathology and the impact of nutrients on mitochondrial DNA maintenance and expression. The activities are in close continuity with WP1 and 2 with the final goal to achieve a proof-of-concept for the treatments of mtDNA disorders. In this WP, key advances have come from the study of the Mpv17 deficient mouse that is a model of human mitochondrial DNA depletion syndrome. These studies have enabled investigate the cellular responses to mtDNA perturbation and the consequences for organ pathology, to develop small molecules therapies for mtDNA disorders, and to evaluate how energy production respiratory complexes are altered in disease states. We have also begun to create the first iPSC line from a mitochondrial disease patient.
The last scientific WP4, New Methods and Tools for Mitochondrial Studies, is a methodological WP focused on developing new methods for mitochondrial studies. With its technical nature, this WP is aimed at the development, improvement and implementation of protocols for the isolation of proteins on nascent RNA, cryo-EM studies of mitoribosome structure, delivery of active oligonucleotides within the mitochondria, and development of quantitative assay for measuring mitochondrial dynamics. During the RP1, click chemistry approaches for tagging nascent RNA and conjugation of nucleic acids with novel mitochondrial targeting agents have been implemented as well as purifications of human mitoribosomes from HEK293 cells and Jurkat cell lines followed by cryoEM. Overall, WP4 has made substantial progress in developing and optimizing methods for mitochondrial studies that will be applied in RP2.
The main results obtained during the RP1 which can be considered advancement in the field of mitochondrial studies are: 1) the new/optimized methodological approaches for mitochondrial studies (WP1, 2, 3); 2) the identification and biochemical/structural characterization of two new factors involved in rescuing stalled mitochondrial translation processes (WP2); 3) the biochemical/metabolic data obtained in WP3 using the mitochondrial diseased mouse model Mpv17.