Chromatin remodeling in regulation of chromosomal Crossing-Over and seed production – ChromCO

The project explores multi-type approaches, including CRISPR-Cas9 gene editing, genetic mapping, mutant characterization, cytogenetics, immunostaining, microscopy, and genome-wide profiling, to functionally characterize INO but also to deeply increase our knowledge about euchromatin modifications and chromatin remodeling in regulation of CO events, meiosis and plant reproduction.

We contributed to the characterization of the Arabidopsis atino80 mutant and reported that AtINO80 has an important function in regulating nucleosome occupancy and incorporation of the histone variant H2A.Z at specific genes as well as at genome-wide level. Studies are also in good progress to characterize functions of active histone marks (e.g. H3K4me3, H3K36me3 and H2Bub1) during meiosis and Arabidopsis reproduction. To study functions of multiple copies of BnINO80, two different strategies are explored using the CRISPR-Cas9 gene editing technology. Target designs and constructs are made, and the first series of plant transformation give already some encouraging results. Phenotypic and molecular characterizations of transformants are in progress.

Our ChromCO project is expected to lead to: i) A mechanistic insight into the function of the INO gene in chromatin remodeling and CO regulation; ii) A genome-wide knowledge of CO distribution regulated by chromatin landscape and chromatin remodeling; iii) A comparative knowledge of similarities/specificities of chromatin regulation of COs between the diploid plant Arabidopsis thaliana (2n = 10) and the allotetraploid plant Brassica napus (AACC, 2n = 38).

Yang C, Yin L, Xie F, Ma M, Huang S, Zeng Y, Shen WH, Dong A, Li L. (2020) AtINO80 represses photomorphogenesis by modulating nucleosome density and H2A.Z incorporation in light-related genes. Proc Natl Acad Sci USA. doi: 10.1073/pnas.2001976117.

Rousseau-Gueutin M, Facon M, Taburel LM, Deniot G, Archambeau H, Dellero Y, Montes E, Veillet F, Chauvin L, Chèvre AM. Tuning a ménage à 4: how to deal with DUPLicated gene Expression. Colloque EFOR meeting, 10-11 mai 2021

The aim of the ChromCO project is to investigate function of chromatin landscape and chromatin remodeling in regulating Crossing-Over (CO) events during meiosis, a fundamental process ensuring sexual transmission of genetic material to next generation and meanwhile generating diversity within species by creating new chromosome/allele combinations. The project focuses on two species of the Brassicaceae family: Arabidopsis thaliana as the model plant and Brassica napus as being one of the most important oilseed crops in agriculture (ranked at the second place in the world and the first place in Europe). Recent studies in Arabidopsis start to uncover importance of chromatin landscape in CO distribution within the genome. Moreover, our unpublished data have identified an ATP-dependent chromatin-remodeling factor (INO) to play a crucial role in repressing CO frequency in Arabidopsis. INO is highly conserved in oilseed rape. Within the ChromCO project, the three Partners bring together their highly complementary scientific knowledge and expertise to investigate INO function and to address fundamental and emergent questions in chromatin-controlled CO formation.

The ChromCO project will combine multi-type approaches, including CRISPR-Cas9 gene editing, genetic mapping, mutant characterization, cytogenetics, immunostaining, microscopy, and genome-wide profiling, to functionally characterize INO but also to deeply increase our knowledge about euchromatin modifications and chromatin remodeling in regulation of CO events. More specifically, in addition to our already available INO-knockout mutants in Arabidopsis as well as histone-modification-defective mutants in Arabidopsis and in Brassica that will be characterized into detail, the Brassica INO genes (three copies) will also be knockout by using CRISPR-Cas9 gene editing technology. Thereafter, the impact of loss-of-function of one, two or all three genes of INO on CO frequency, distribution and meiotic stability will be deciphered genome-wide by genetic mapping. An increased knowledge about how INO functions and interplays with meiotic factors (RECQ4, HEI10/HEIP1) as well as with other chromatin regulators (SWR1, NRPs, SDG8, SDG2, HUB2) will be gained through genetic interaction studies. Immunostaining and microscopy imaging will be performed to gain knowledge about epigenetic marks (e.g. H2A.X, H2A.Z, and methylated H3) during meiosis in both Arabidopsis and Brassica. Furthermore, alteration of genome-wide distribution of DNA double-strand breaks (DSBs) and COs will be integrated with publicly accessible profiles of histone methylations (e.g. H3K4, H3K9, H3K27 and H3K36 methylations) and histone variants (H2A.Z, H3.3, H2A.W) to assess INO function associated with or not with a specific chromatin feature in Arabidopsis. The ChromCO project is expected to make following breakthroughs: i) A functional understanding of the importance of chromatin landscape and chromatin remodeling in CO formation and meiosis; ii) A mechanistic insight into the function of the INO gene in chromatin remodeling and CO regulation; iii) A genome-wide knowledge of CO distribution regulated by chromatin landscape and chromatin remodeling; iv) A comparative knowledge of similarities/specificities of chromatin regulation of COs between the diploid plant Arabidopsis thaliana (2n = 10) and the allotetraploid plant Brassica napus (AACC, 2n = 38).

The ChromCO project has implicating interests in agronomical applications as well as in evolutionary biology, speciation and plant environmental adaptation. Indeed, a deep understanding of the mechanisms that regulate CO frequency could provide efficient ways to increase recombination and will enable to create more rapidly and efficiently new elite cultivars via combining alleles of interests. An insight about CO events occurring between homologous (A and A or C and C) and homoleologous (A and C) chromosomes could help to understand evolution of hybrid speciation.