The study investigates the impact of targeted chromosome engineering on telomere dynamics, chromatin structure, gene expression, and phenotypic stability in Arabidopsis thaliana. Using precise CRISPR/Cas-based engineering, reciprocal translocations of chromosome arms were introduced between non-homologous chromosomes. The subsequent homozygous generations of plants were assessed for phenotype, transcriptomic changes and chromatin modifications near translocation breakpoints, and telomere length maintenance. Phenotypically, translocated lines were indistinguishable from wild-type plants, as confirmed through morphological assessments and principal component analysis. Gene expression profiling detected minimal differential expression, with affected genes dispersed across the genome, indicating negligible transcriptional impact. Similarly, ChIPseq analysis showed no substantial alterations in the enrichment of key histone marks (H3K27me3, H3K4me1, H3K56ac) near junction sites or across the genome. Finally, bulk and arm-specific telomere lengths remained stable across multiple generations, except for minor variations in one translocation line. These findings highlight the remarkable genomic and phenotypic robustness of A. thaliana despite large-scale chromosomal rearrangements. The study offers insights into the cis-acting mechanisms underlying chromosome arm-specific telomere length setting and establishes the feasibility of chromosome engineering for studies of plant genome evolution and crop improvement strategies.

• Keywords
• Arabidopsis thaliana, chromatin structure, chromosome engineering, chromosome translocation, gene expression, phenotype, telomeres,
• MeSH
• Arabidopsis * genetics   MeSH
• Chromatin * genetics   metabolism   MeSH
• Chromosomes, Plant * genetics   MeSH
• CRISPR-Cas Systems MeSH
• Phenotype MeSH
• Genetic Engineering MeSH
• Gene Expression Regulation, Plant MeSH
• Telomere * genetics   MeSH
• Translocation, Genetic MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Chromatin * MeSH

The current repertoire of methods available for studying RNA-protein interactions in plants is somewhat limited. Employing an RNA-centric approach, particularly with less abundant RNAs, presents various challenges. Many of the existing methods were initially designed for different model systems, with their application in plants receiving limited attention thus far. The Comprehensive Identification of RNA-Binding Proteins by Mass Spectrometry (ChIRP-MS) technique, initially developed for mammalian cells, has been adapted in this study for application in Arabidopsis thaliana. The procedures have been meticulously modified and optimized for telomerase RNA, a notable example of a low-abundance RNA recently identified. Following these optimization steps, ChIRP-MS can serve as an effective screening method for identifying candidate proteins interacting with any target RNA of interest.

• Keywords
• Arabidopsis thaliana, ChIRP-MS, RNA, RNA-centric methods, RNA-protein interactions, telomerase, telomerase RNA,
• Publication type
• Journal Article MeSH

Telomeres are essential nucleoprotein structures at the very ends of linear eukaryote chromosomes. They shelter the terminal genome territories against degradation and prevent the natural chromosome ends from being recognized by repair mechanisms as double-strand DNA breaks.There are two basic characteristics of telomeric DNA, its sequence and its length. The telomere sequence is important as a "landing area" for specific telomere-binding proteins, which function as signals and moderate the interactions required for correct telomere function. While the sequence forms the proper "landing surface" of telomeric DNA, its length is similarly important. Too short or exceptionally long telomere DNA cannot perform its function properly. In this chapter, methods for the investigation of these two basic telomere DNA characteristics are described, namely, telomere motif identification and telomere length measurement.

• Keywords
• BAL31-NGS, Pulsed-field gel electrophoresis (PFGE), Tandem repeats finder, Telomere, Terminal restriction fragments,
• MeSH
• DNA * genetics   MeSH
• DNA Breaks, Double-Stranded MeSH
• Telomere-Binding Proteins genetics   MeSH
• Telomere * genetics   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• DNA * MeSH
• Telomere-Binding Proteins MeSH

Telomere repeat binding proteins (TRBs) belong to a family of proteins possessing a Myb-like domain which binds to telomeric repeats. Three members of this family (TRB1, TRB2, TRB3) from Arabidopsis thaliana have already been described as associated with terminal telomeric repeats (telomeres) or short interstitial telomeric repeats in gene promoters (telo-boxes). They are also known to interact with several protein complexes: telomerase, Polycomb repressive complex 2 (PRC2) E(z) subunits and the PEAT complex (PWOs-EPCRs-ARIDs-TRBs). Here we characterize two novel members of the TRB family (TRB4 and TRB5). Our wide phylogenetic analyses have shown that TRB proteins evolved in the plant kingdom after the transition to a terrestrial habitat in Streptophyta, and consequently TRBs diversified in seed plants. TRB4-5 share common TRB motifs while differing in several others and seem to have an earlier phylogenetic origin than TRB1-3. Their common Myb-like domains bind long arrays of telomeric repeats in vitro, and we have determined the minimal recognition motif of all TRBs as one telo-box. Our data indicate that despite the distinct localization patterns of TRB1-3 and TRB4-5 in situ, all members of TRB family mutually interact and also bind to telomerase/PRC2/PEAT complexes. Additionally, we have detected novel interactions between TRB4-5 and EMF2 and VRN2, which are Su(z)12 subunits of PRC2.

• Keywords
• PEAT, PRC2, TERT, TRB, Telomere repeat binding, Telomeric,
• MeSH
• Arabidopsis * genetics   metabolism   MeSH
• Phylogeny MeSH
• Arabidopsis Proteins * genetics   metabolism   MeSH
• Telomere-Binding Proteins genetics   metabolism   MeSH
• Soil MeSH
• Telomerase * genetics   metabolism   MeSH
• Telomere genetics   metabolism   MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Arabidopsis Proteins * MeSH
• Telomere-Binding Proteins MeSH
• Soil MeSH
• Telomerase * MeSH

Polycomb repressive complex 2 (PRC2) represents a group of evolutionarily conserved multi-subunit complexes that repress gene transcription by introducing trimethylation of lysine 27 on histone 3 (H3K27me3). PRC2 activity is of key importance for cell identity specification and developmental phase transitions in animals and plants. The composition, biochemistry, and developmental function of PRC2 in animal and flowering plant model species are relatively well described. Recent evidence demonstrates the presence of PRC2 complexes in various eukaryotic supergroups, suggesting conservation of the complex and its function. Here, we provide an overview of the current understanding of PRC2-mediated repression in different representatives of eukaryotic supergroups with a focus on the green lineage. By comparison of PRC2 in different eukaryotes, we highlight the possible common and diverged features suggesting evolutionary implications and outline emerging questions and directions for future research of polycomb repression and its evolution.

• Keywords
• H3K27me3, PRC2, SAR, algae, animal, evolution, fungi, green lineage, plant, polycomb,
• Publication type
• Journal Article MeSH
• Review MeSH

Involvement of epigenetic mechanisms in the regulation of telomeres and transposable elements (TEs), genomic regions with the protective and potentially detrimental function, respectively, has been frequently studied. Here, we analyzed telomere lengths in Arabidopsis thaliana plants of Columbia, Landsberg erecta and Wassilevskija ecotypes exposed repeatedly to the hypomethylation drug zebularine during germination. Shorter telomeres were detected in plants growing from seedlings germinated in the presence of zebularine with a progression in telomeric phenotype across generations, relatively high inter-individual variability, and diverse responses among ecotypes. Interestingly, the extent of telomere shortening in zebularine Columbia and Wassilevskija plants corresponded to the transcriptional activation of TEs, suggesting a correlated response of these genomic elements to the zebularine treatment. Changes in lengths of telomeres and levels of TE transcripts in leaves were not always correlated with a hypomethylation of cytosines located in these regions, indicating a cytosine methylation-independent level of their regulation. These observations, including differences among ecotypes together with distinct dynamics of the reversal of the disruption of telomere homeostasis and TEs transcriptional activation, reflect a complex involvement of epigenetic processes in the regulation of crucial genomic regions. Our results further demonstrate the ability of plant cells to cope with these changes without a critical loss of the genome stability.

• Keywords
• Arabidopsis thaliana, cytosine methylation, epigenetics, telomeres, transposable elements, zebularine,
• MeSH
• Transcriptional Activation genetics   MeSH
• Arabidopsis genetics   metabolism   MeSH
• Cytidine analogs & derivatives   genetics   MeSH
• Cytosine metabolism   MeSH
• Epigenesis, Genetic genetics   MeSH
• Telomere Homeostasis genetics   MeSH
• DNA Methylation genetics   MeSH
• Plant Cells metabolism   MeSH
• Telomere genetics   MeSH
• DNA Transposable Elements genetics   MeSH
• Telomere Shortening genetics   MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Cytidine MeSH
• Cytosine MeSH
• pyrimidin-2-one beta-ribofuranoside MeSH Browser
• DNA Transposable Elements MeSH

Understanding how the packaging of chromatin in the nucleus is regulated and organized to guide complex cellular and developmental programmes, as well as responses to environmental cues is a major question in biology. Technological advances have allowed remarkable progress within this field over the last years. However, we still know very little about how the 3D genome organization within the cell nucleus contributes to the regulation of gene expression. The nuclear space is compartmentalized in several domains such as the nucleolus, chromocentres, telomeres, protein bodies, and the nuclear periphery without the presence of a membrane around these domains. The role of these domains and their possible impact on nuclear activities is currently under intense investigation. In this review, we discuss new data from research in plants that clarify functional links between the organization of different nuclear domains and plant genome function with an emphasis on the potential of this organization for gene regulation.

• Keywords
• 3D Chromatin organization, chromocentres, gene expression, liquid–liquid phase separation (LLPS), nuclear bodies, nuclear domains, nuclear periphery, nucleolus, telomeres, topologically associated domains (TADs),
• MeSH
• Cell Nucleolus MeSH
• Cell Nucleus * MeSH
• Chromatin * MeSH
• Gene Expression Regulation MeSH
• Plants genetics   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Review MeSH
• Names of Substances
• Chromatin * MeSH