Periodic Reporting for period 1 - PAMFGAL (Ancestral molecular function of Polycomb Repressive Complex 2 (PRC2) in green algae lineage)
Okres sprawozdawczy: 2023-01-01 do 2024-12-31
Epigenetic modifications of histones and DNA ensure the stable heritability of gene expression, maintain the repression of transposable elements, and facilitate transcriptional responses to environmental changes. The landscape of different epigenetic modifications determines chromatin structure, DNA accessibility, and gene transcription. One of the better-known and described factors in this process is the Polycomb repressive system, consisting of Polycomb group (PcG) proteins that assemble into complexes. These complexes maintain cellular identity in multicellular organisms and regulate developmental phase transitions. Polycomb Repressive Complex 2 (PRC2) is an evolutionarily conserved complex that catalyses the trimethylation of histone H3 at lysine 27 (H3K27me3) to create facultative heterochromatin. PRC2 is extensively studied in flowering plants and determines cell fate during development. Putative homologs of PRC2 subunits have been reported to exist in the genomes of unicellular green algae. Still, there was no molecular data on the composition, genomic targeting, and function of the PRC2 complex. How chromatin structure and histone modifications contribute to the cellular identities in unicellular green algae was not known. The project's main aim was to characterize the molecular functions of PRC2 in the unicellular green algae to elucidate its potential ancestral role and its interplay with other epigenetic mechanisms.
To obtain an overview of the presence of different post-transcriptional modifications of the N-terminal H3 tail in green microalgae, we performed a mass spectrometry-based proteomic analysis of histone modifications in the two species Chlamydomonas reinhardtii and Chlorella sorokiniana, which led us to the conclusion that only the latter has all canonical histone modifications at the analysed sites. In addition, we investigated the landscape of epigenetic modifications, including histone post-translational modifications, DNA methylation, and gene expression under different trophic conditions. This allowed us to define chromatin states and reveal unique overlaps of epigenetic mechanisms in unicellular green algae. Furthermore, using the data obtained in this project, we have identified genes involved in the development, metabolism, and basic cellular processes as orthologues with conserved H3K27me3 marks from unicellular green algae to land plants. We also performed experiments to understand the influence of specific amino acids neighboring the K27 residue on the activity of PRC2 and other methyltransferases in the green lineage. Furthermore, we investigated the composition of PRC2 core subunits in the green lineage using in silico and in vivo methods, which revealed common patterns but also the diversity of PRC2 systems in the green lineage.
Part of the results obtained have been summarised in a collaborative manuscript that is available as a preprint (DOI: 10.1101/2024.09.22.614159 recently accepted in Molecular Biology and Evolution). This manuscript describes the relative distribution of H3K9me2 and H3K27me3 in Chlorella sorokiniana and shows a preference for targeting of young genes by H3K9 methylation in the green lineage. Another manuscript is currently in preparation and we expect publication of the manuscript in the first half of 2025. This manuscript will comprehensively describe chromatin states and associated gene expression patterns during the trophic transitions in Chlorella sorokiniana.
Throughout the project, I have presented our results as poster presentations at six conferences and practical courses: YAS 2023 - Young Algaeneers Symposium, 9-11 May 2023; EMBO Practical Course: Characterization of post-translational modifications in cellular signaling, 14-22 June 2023, Odense, Denmark; 8th European Phycological Congress, Brest, France, 20-26 August 2023, Brest, France; Methods in Plant Sciences, 24-27 September 2023, Srni, Czechia; EMBO Workshop Plant genome stability and change, 16-19 June 2024, Olomouc, Czechia; SEB Epiplant Conference, 10-12 July 2024, Clermond-Ferrand, France. I also gave two talks: SEB Annual Meeting, SEB Conference, 2-5 July 2024, Prague, CZ; Plant Science Seminars, 5 December 2024, České Budějovice, CZ While working on this project, I also presented my work 9 times at regular lab meetings of Iva Mozgová's group at the Biology Centre CAS.
Part of the results obtained have been summarised in a collaborative manuscript that is available as a preprint (DOI: 10.1101/2024.09.22.614159 recently accepted in Molecular Biology and Evolution). This manuscript describes the relative distribution of H3K9me2 and H3K27me3 in Chlorella sorokiniana and shows a preference for targeting of young genes by H3K9 methylation in the green lineage. Another manuscript is currently in preparation and we expect publication of the manuscript in the first half of 2025. This manuscript will comprehensively describe chromatin states and associated gene expression patterns during the trophic transitions in Chlorella sorokiniana.
Throughout the project, I have presented our results as poster presentations at six conferences and practical courses: YAS 2023 - Young Algaeneers Symposium, 9-11 May 2023; EMBO Practical Course: Characterization of post-translational modifications in cellular signaling, 14-22 June 2023, Odense, Denmark; 8th European Phycological Congress, Brest, France, 20-26 August 2023, Brest, France; Methods in Plant Sciences, 24-27 September 2023, Srni, Czechia; EMBO Workshop Plant genome stability and change, 16-19 June 2024, Olomouc, Czechia; SEB Epiplant Conference, 10-12 July 2024, Clermond-Ferrand, France. I also gave two talks: SEB Annual Meeting, SEB Conference, 2-5 July 2024, Prague, CZ; Plant Science Seminars, 5 December 2024, České Budějovice, CZ While working on this project, I also presented my work 9 times at regular lab meetings of Iva Mozgová's group at the Biology Centre CAS.
The project has led to new insights into the evolution of PRC2 in the green lineage. We have uncovered the distribution of histone PTMs in C. sorokiniana, their interplay with DNA methylation and their association with gene expression levels, providing the first insights into the epigenomic landscape in unicellular green algae. Furthermore, we have defined several distinct chromatin states, their connection to gene expression, and their dynamics during trophic transition. Our results contribute to a better understanding of the role of chromatin dynamics in phase transitions of unicellular organisms. We have identified the role of PRC2 in defining both facultative and constitutive heterochromatin. Our established and improved methods for studying epigenetics in C. sorokiniana provide a good basis for establishing this species as a green algal model. Together with our collaborators, we have provided insights into the conservation of PRC2 silencing. We have identified genes that evolved during plant terrestrialization, and that are preferentially silenced by the epigenetic mark H3K27me3 introduced by PRC2. We have uncovered common and ancient features of epigenetic gene regulation in the green lineage. Our results provide insights into the evolutionary dynamics driving the adaptation of epigenetic regulation. We have shown that the recruitment of PRC2 and the deposition of H3K27me3 in genes occurred before the emergence of land plants. In addition, the results of the project have made a significant contribution to the scientific community by being presented at conferences and seminars, facilitating the dissemination of knowledge and fostering scientific collaboration. By sharing our results and engaging in meaningful discussions, we have helped to emphasize the broader societal impact of an improved understanding of plant biology. It is anticipated that our results will attract interest from both the general public and the algal industry, providing valuable insights and practical applications in the future.