The overlooked manipulation of nucleolar functions by plant pathogen effectors.

Pathogens need to manipulate plant functions to facilitate the invasion of their hosts. They do this by secreting a cocktail of molecules called effectors. Studies of these molecules have mostly focused on the mechanisms underlying their recognition and the subsequent transcriptional reprogramming of cells, particularly in the case of R gene-dependent resistance.

Extrachromosomal circular DNA and structural variants highlight genome instability in Arabidopsis epigenetic mutants.

Abundant extrachromosomal circular DNA (eccDNA) is associated with transposable element (TE) activity. However, how the eccDNA compartment is controlled by epigenetic regulations and what is its impact on the genome is understudied. Here, using long reads, we sequence both the eccDNA compartment and the genome of Arabidopsis thaliana mutant plants affected in DNA methylation and post-transcriptional gene silencing.

NICOTIANAMINE SYNTHASE activity affects nucleolar iron accumulation and impacts rDNA silencing and RNA methylation in Arabidopsis.

In plant cells, a large pool of iron (Fe) is contained in the nucleolus, as well as in chloroplasts and mitochondria. A central determinant for intracellular distribution of Fe is nicotianamine (NA) generated by NICOTIANAMINE SYNTHASE (NAS). Here, we used Arabidopsis thaliana plants with disrupted NAS genes to study the accumulation of nucleolar iron and understand its role in nucleolar functions and more specifically in rRNA gene expression.

Glutathione-mediated thermomorphogenesis and heat stress responses in Arabidopsis thaliana.

In the context of climate change, the global rise of temperature and intense heat waves affect plant development and productivity. Among the molecular perturbations that high temperature induces in living cells is the accumulation of reactive oxygen species (ROS), which perturbs the cellular redox state. In plants, the dynamics of the cellular and subcellular redox state have been poorly investigated under high temperature.

The plant-specific DDR factor SOG1 increases chromatin mobility in response to DNA damage.

Homologous recombination (HR) is a conservative DNA repair pathway in which intact homologous sequences are used as a template for repair. How the homology search happens in the crowded space of the cell nucleus is, however, still poorly understood. Here, we measure chromosome and double-strand break (DSB) site mobility in Arabidopsis thaliana, using lacO/LacI lines and two GFP-tagged HR reporters.

ANCHOR: A Technical Approach to Monitor Single-Copy Locus Localization .

Together with local chromatin structure, gene accessibility, and the presence of transcription factors, gene positioning is implicated in gene expression regulation. Although the basic mechanisms are expected to be conserved in eukaryotes, less is known about the role of gene positioning in plant cells, mainly due to the lack of a highly resolutive approach. In this study, we adapted the use of the ANCHOR system to perform real-time single locus detection .

Large tandem duplications affect gene expression, 3D organization, and plant-pathogen response.

Rapid plant genome evolution is crucial to adapt to environmental changes. Chromosomal rearrangements and gene copy number variation (CNV) are two important tools for genome evolution and sources for the creation of new genes. However, their emergence takes many generations.

Tidying-up the plant nuclear space: domains, functions, and dynamics.

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.

Nucleolus-associated chromatin domains are maintained under heat stress, despite nucleolar reorganization in Arabidopsis thaliana.

Several layers of mechanisms participate in plant adaptation to heat-stress. For example, the plant metabolism switches from cell growth mode to stress adaptation mode. Ribosome biogenesis is one of the most energy costly pathways.

Three-dimensional nuclear organization in Arabidopsis thaliana.

In recent years, the study of plant three-dimensional nuclear architecture received increasing attention. Enabled by technological advances, our knowledge on nuclear architecture has greatly increased and we can now access large data sets describing its manifold aspects. The principles of nuclear organization in plants do not significantly differ from those in animals.

Chromatin domains in space and their functional implications.

Genome organization displays functional compartmentalization. Many factors, including epigenetic modifications, transcription factors, chromatin remodelers, and RNAs, shape chromatin domains and the three-dimensional genome organization. Various types of chromatin domains with distinct epigenetic and spatial features exhibit different transcriptional activities.

Ribosomal RNA genes shape chromatin domains associating with the nucleolus.

Genomic interactions can occur in addition to those within chromosome territories and can be organized around nuclear bodies. Several studies revealed how the nucleolus anchors higher order chromatin structures of specific chromosome regions displaying heterochromatic features. In this review, we comment on advances in this emerging field, with a particular focus on a recent study published by Quinodoz et al.

An Effector from the Cyst Nematode Derepresses Host rRNA Genes by Altering Histone Acetylation.

Cyst nematodes are plant-pathogenic animals that secrete effector proteins into plant root cells to alter host gene expression and reprogram these cells to form specialized feeding sites, known as syncytia. The molecular mechanisms of these effectors are mostly unknown. We determined that the sugar beet cyst nematode () 32E03 effector protein strongly inhibits the activities of histone deacetylases including the HDT1 enzyme, which has a known function in the regulation of rRNA gene expression through chromatin modifications.

Nucleolar Proteome Analysis and Proteasomal Activity Assays Reveal a Link between Nucleolus and 26S Proteasome in .

In all eukaryotic cells, the nucleolus is functionally and structurally linked to rRNA synthesis and ribosome biogenesis. This compartment contains as well factors involved in other cellular activities, but the functional interconnection between non-ribosomal activities and the nucleolus (structure and function) still remains an open question. Here, we report a novel mass spectrometry analysis of isolated nucleoli from plants using the FANoS (Fluorescence Assisted Nucleolus Sorting) strategy.

A Method to Identify Nucleolus-Associated Chromatin Domains (NADs).

The nuclear context needs to be taken into consideration to better understand the mechanisms shaping the epigenome and its organization, and therefore its impact on gene expression. For example, in Arabidopsis, heterochromatin is preferentially localized at the nuclear and the nucleolar periphery. Although chromatin domains associating with the nuclear periphery remain to be identified in plant cells, Nucleolus Associated chromatin Domains (NADs) can be identified thanks to a protocol allowing the isolation of pure nucleoli.

Hybrid incompatibility caused by an epiallele.

Hybrid incompatibility resulting from deleterious gene combinations is thought to be an important step toward reproductive isolation and speciation. Here, we demonstrate involvement of a silent epiallele in hybrid incompatibility. In accession Cvi-0, one of the two copies of a duplicated histidine biosynthesis gene, , is mutated, making essential.

Selective nucleolus organizer inactivation in Arabidopsis is a chromosome position-effect phenomenon.

Nucleolus organizer regions (NORs) are chromosomal loci where hundreds of rRNA genes are clustered. Despite being nearly identical in sequence, specific rRNA genes are selected for silencing during development via choice mechanism(s) that remain unclear. In Arabidopsis thaliana, rRNA gene subtypes that are silenced during development were recently mapped to the NOR on chromosome 2, NOR2, whereas active rRNA genes map to NOR4, on chromosome 4.

Plant nucleolar DNA: Green light shed on the role of Nucleolin in genome organization.

The nucleolus forms as a consequence of ribosome biogenesis, but it is also implicated in other cell functions. The identification of nucleolus-associated chromatin domains (NADs) in animal and plant cells revealed the presence of DNA sequences other than rRNA genes in and around the nucleolus. NADs display repressive chromatin signatures and harbour repetitive DNA, but also tRNA genes and RNA polymerase II-transcribed genes.

Fluorescence-Activated Nucleolus Sorting in Arabidopsis.

Nucleolar isolation allows exhaustive characterization of the nucleolar content. Centrifugation-based protocols are not adapted to isolation of nucleoli directly from a plant tissue because of copurification of cellular debris. We describe here a method that allows the purification of nucleoli using fluorescent-activated cell sorting from Arabidopsis thaliana leaves.

Identification of Nucleolus-Associated Chromatin Domains Reveals a Role for the Nucleolus in 3D Organization of the A. thaliana Genome.

The nucleolus is the site of rRNA gene transcription, rRNA processing, and ribosome biogenesis. However, the nucleolus also plays additional roles in the cell. We isolated nucleoli using fluorescence-activated cell sorting (FACS) and identified nucleolus-associated chromatin domains (NADs) by deep sequencing, comparing wild-type plants and null mutants for the nucleolar protein NUCLEOLIN 1 (NUC1).

Chromosome-specific NOR inactivation explains selective rRNA gene silencing and dosage control in Arabidopsis.

In eukaryotes, scores of excess ribosomal RNA (rRNA) genes are silenced by repressive chromatin modifications. Given the near sequence identity of rRNA genes within a species, it is unclear how specific rRNA genes are reproducibly chosen for silencing. Using Arabidopsis thaliana ecotype (strain) Col-0, a systematic search identified sequence polymorphisms that differ between active and developmentally silenced rRNA gene subtypes.

Subunit compositions of Arabidopsis RNA polymerases I and III reveal Pol I- and Pol III-specific forms of the AC40 subunit and alternative forms of the C53 subunit.

Using affinity purification and mass spectrometry, we identified the subunits of Arabidopsis thaliana multisubunit RNA polymerases I and III (abbreviated as Pol I and Pol III), the first analysis of their physical compositions in plants. In all eukaryotes examined to date, AC40 and AC19 subunits are common to Pol I (a.k.

A duplicated NUCLEOLIN gene with antagonistic activity is required for chromatin organization of silent 45S rDNA in Arabidopsis.

In plants as well as in animals, hundreds to thousands of 45S rRNA gene copies localize in Nucleolus Organizer Regions (NORs), and the activation or repression of specific sets of rDNA depends on epigenetic mechanisms. Previously, we reported that the Arabidopsis thaliana nucleolin protein NUC1, an abundant and evolutionarily conserved nucleolar protein in eukaryotic organisms, is required for maintaining DNA methylation levels and for controlling the expression of specific rDNA variants in Arabidopsis. Interestingly, in contrast with animal or yeast cells, plants contain a second nucleolin gene.

A two-step process for epigenetic inheritance in Arabidopsis.

In Arabidopsis, multisubunit RNA polymerases IV and V orchestrate RNA-directed DNA methylation (RdDM) and transcriptional silencing, but what identifies the loci to be silenced is unclear. We show that heritable silent locus identity at a specific subset of RdDM targets requires HISTONE DEACETYLASE 6 (HDA6) acting upstream of Pol IV recruitment and siRNA biogenesis. At these loci, epigenetic memory conferring silent locus identity is erased in hda6 mutants such that restoration of HDA6 activity cannot restore siRNA biogenesis or silencing.