Functional interplays between Arabidopsis linker proteins and impact on epigenome organization – EpiLinks

Uncovering how epigenome regulation influences gene expression has revolutionized former views in developmental and evolutionary biology. In particular, an ever-increasing number of studies are unveiling how modulation of chromatin status, compaction and accessibility, impact on the transcriptional regulation of genes and transposable elements. In most eukaryotes, chromatin compaction at specific domains involves linker histone H1 and Polycomb Repressive Complex 2 (PRC2). While PRC2 activity comprises an enzymatic modification of histones (H3K27me3 deposition), H1 displays intrinsic chromatin compaction properties involving its conserved 'globular H1' domain (GH1). Extensive studies in mammals recently established H1’s central function in instructing genome topology through PRC2 activity in vivo, yet, no molecular mechanism explaining how linker histones drive sequence-specific processes has been identified. More generally, we lag behind in understanding how, and to which extent, chromatin mechanisms operating at different genomic scales (from protein-coding genes to telomeres, centromeres or ribosomal gene arrays) interplay to allow for a concerted regulation of the epigenome landscape and chromosome architecture. Moreover, whether and how these large-scale regulations can impact organismic phenotypes and adaptive potential is completely unknown. Our groundwork in Arabidopsis thaliana shed light on telomeric sequences as an ideal system to tackle these emerging questions. These motifs occur in three genomic contexts: 1) telomeres protecting chromosome ends, 2) large internal Interstitial Telomeric Regions (ITRs) present in multiple organisms and impacting on genome stability and organization, and 3) single copy promoter elements acting in PRC2 recruitment for targeted gene regulation. We unveiled that H1 mediates a selective repression of PRC2 activity at A. thaliana telomeres (~20 kb) and ITRs (~500 kb), presumably acting in competition with two structurally-related GH1 protein families: High Mobility Group A (HMGA) and Telomere Repeat Binding (TRB) proteins. H1 prevents the formation of enormous H3K27me3 aggregates at ITRs while minimizing ITR-telomere and telomere-telomere distal interactions. Based on these findings we propose that interplays between GH1 proteins serve as a safeguarding mechanism that enables plants to cope with the deleterious consequences of telomeric motifs attractiveness for PRC2. The project design is composed of three complementary work packages (WP) using well-established experimental setups or pioneering approaches. WP1 addresses the physical and functional relationships between these three GH1 protein families and how their cooperative/competitive chromatin integration impacts on the epigenome landscape. WP2 is aimed at assessing how H1 contributes to plants' capability to accommodate abundant ITRs, and which role these genome domains play in epigenome and nuclear organization. This question pertains to evolutionary perspectives that will be explored using genome engineering and by exploiting ITR natural variation in A. thaliana ecotypes. In WP3, we propose to determine the biological meaning of GH1 protein interplays in PRC2-mediated transcription regulation and in plant development, considering both GH1-mediated regulation at genes and the potential role of ITR-encoded TERRA long non-coding RNAs (lncRNAs) in PRC2 recruitment in trans. Deciphering how chromatin machineries enable plants to restrict PRC2 activity at telomeric repeats while permitting it at protein-coding genes should help understanding how similar genetic elements are differentially controlled and drive distinct transcriptional and topological states. Our work plan is therefore aimed at reaching an integrated view of epigenome homeostasis from genes to telomeres and from genomes to populations, a much awaited perspective by the scientific community.