Biology subjects

Ziolkowski, P. A.

Publications and source records attributed to Ziolkowski, P. A..

7 recordsLinked to original sources

bioRxiv2026

Chromatin crosstalk between HDA19 and NuA4 sets thresholds for stress gene activation in Arabidopsis

Histone acetylation by the plant NuA4 complex promotes high expression of growth-related genes while preventing gene body H2A.Z depletion and spurious activation of stress-responsive loci. How this NuA4-dependent chromatin state is modulated by histone deacetylases (HDACs) in plants remains unclear. Here, we investigate the interplay between NuA4 and HDACs in Arabidopsis by generating a collection of HDAC loss-of-function mutants in NuA4-proficient and NuA4-deficient backgrounds. Loss of individual HDACs did not rescue the severe growth defects of the NuA4(-) mutant; instead, most combinations further aggravated the phenotype, indicating that HDACs predominantly support rather than counteract NuA4 function. Focusing on the hda19-5Atepl1ballele, we show that loss of HDA19 causes strong developmental defects and constitutive activation of biotic stress-responsive genes, but not canonical heat-response genes. 3' RNA-seq and ChIP-seq reveal that upregulated genes in hda19-5Atepl1bdisplay increased H3K9ac and H2A.Zac with largely unchanged gene-body H2A.Z, and substantially overlap with genes induced in the NuA4-null Atepl1-2 mutant and in wild-type plants exposed to elevated temperature. Integration of our datasets with published HDA19 ChIP-seq maps shows that H2A.Zac is selectively increased at HDA19-bound H2A.Z peaks in hda19-5Atepl1b, consistent with a direct role for HDA19 in H2A.Z deacetylation. However, elevated H2A.Zac is neither necessary nor sufficient for transcriptional activation, whereas changes in H3K9ac correlate strongly with gene induction. We propose that NuA4 and HDA19 cooperate to tune chromatin at stress-related loci, with H3K9 acetylation as the primary driver of transcription and H2A.Z acetylation acting as a modulatory mark that shapes stress gene responsiveness.

plant biology↗

bioRxiv2025

Modifying meiotic recombination by targeting chromatin modifiers to crossover hotspots in Arabidopsis

The impact of specific chromatin modifications on meiotic crossover frequency is usually inferred from correlative studies, leaving open the question of causality. To address this, we used a dCas9-based system to target histone H3 methylation modifiers to defined genomic loci. Targeting methyltransferases responsible for H3K9me3 and H3K27me3 had little effect on recombination at selected hotspots, whereas targeting H3K4me3- and H3K36me3-associated enzymes often silenced the endogenous genes. The strongest effect was observed with the demethylase JMJ14, which reduced local H3K4me3 levels and decreased crossover frequency within the targeted interval. This was accompanied by reduced transcription of a long non-coding RNA (lncRNA) located at the hotspot and altered crossover topology. Suppressed recombination was also seen at neighbouring, untargeted hotspots. Conversely, directing the transcriptional activator VP64 to the same region elevated lncRNA expression, increased crossover frequency, and raised H3K4me3 levels. Our results reveal a causal relationship between H3K4me3, transcription, and local crossover activity, demonstrating that H3K4me3 levels are tightly associated with both transcriptional output and recombination frequency at specific genomic sites.

genetics↗

bioRxiv2024

COmapper: High-resolution mapping of meiotic crossovers by long-read sequencing in Arabidopsis

O_LIMeiotic crossovers rearrange existing genetic variation between homologous chromosomes, profoundly affecting genomic diversity. Crossovers are typically constrained to 1-3 events per chromosome pair, and their distribution is shaped by chromatin accessibility and DNA polymorphisms. Genome-wide crossover maps can be generated in plants by high-throughput short-read sequencing or linked-read sequencing. C_LIO_LIHere, we use long-read nanopore sequencing technology to develop a crossover mapping pipeline, COmapper, for high-resolution mapping of genome-wide crossovers from pooled DNA of F1 hybrid pollen and F2 recombinant seedlings derived from a cross between Arabidopsis thaliana accessions Col and Ler. We validate the high accuracy of COmapper by applying nanopore long-read sequencing to pooled DNA of Arabidopsis F2 individuals with crossovers mapped by short-read sequencing. C_LIO_LIUsing the COmapper, we constructed high-resolution genomic maps of crossovers using F1 hybrid pollen and F2 seedlings in wild type and crossover-elevated recq4a recq4b mutant, showing results comparable to short-read sequencing. Crossovers were enriched at gene-proximal promoters in wild type and increased but reshaped by high polymorphism density in recq4a recq4b. C_LIO_LIWe propose that COmapper will be widely applicable for exploring the effects of genetic, epigenetic and environmental changes on the crossover patterns across diverse plant species. C_LI

plant biology↗

bioRxiv2024

Genetic dissection of MutL complexes in Arabidopsis meiosis

During meiosis, homologous chromosomes exchange genetic material through crossing-over. The main crossover pathway relies on ZMM proteins, including ZIP4 and HEI10, and is typically resolved by the MLH1/MLH3 heterodimer, MutL{gamma}. Our analysis of plant fertility and bivalent formation revealed that the MUS81 endonuclease can partially compensate for the MutL{gamma} loss. Comparing genome-wide crossover maps of the mlh1 mutant with ZMM-deficient mutants and lines with varying HEI10 levels reveals that while crossover interference persists in mlh1, it is weakened. Additionally, mlh1 show reduced crossover assurance, leading to a higher incidence of aneuploidy in offspring. This is likely due to MUS81 resolving intermediates without the crossover bias seen in MutL{gamma}. Comparing mlh1 mlh3 mus81 and zip4 mus81 mutants suggests that additional crossover pathways emerge in the absence of both MutL{gamma} and MUS81. The loss of MutL{gamma} can also be suppressed by eliminating the FANCM helicase. Elevated expression of MLH1 or MLH3 increases crossover frequency, while their overexpression significantly reduces crossover numbers and plant fertility, highlighting the importance for tight control of MLH1/MLH3 levels. By contrast, PMS1, a component of the MutL endonuclease, appears not to be involved in crossing-over. Together, these findings demonstrate the unique role of MutL{gamma} in ZMM-dependent crossover regulation.

genetics↗

bioRxiv2023

MSH2 stimulates interfering and inhibits non-interfering crossovers in response to genetic polymorphism

In Arabidopsis, local sequence polymorphism between homologs can stimulate double-strand break (DSB) repair via crossover formation in a MSH2-dependent manner. To understand how MSH2 regulates crossovers formed by the independent interfering and non-interfering pathways, we combine mutants that elevate non-interfering crossovers with msh2. We demonstrate that MSH2 blocks non-interfering crossovers at polymorphic loci, which is the opposite effect to interfering crossovers. We also observe MSH2-independent crossover inhibition at highly polymorphic sites. We measure recombination along the chromosome arms, in lines differing in the heterozygosity pattern, and observe a dramatic crossover increase at the boundaries between heterozygous and homozygous regions, which is MSH2-dependent. Together, we show that MSH2 is a master regulator of meiotic DSB repair, with antagonistic effects on interfering and non-interfering crossovers, that shapes the crossover landscape in relation to interhomolog polymorphism.

genetics↗

bioRxiv2019

NuA4-dependent H4 acetylation is adopted to specifically control photosynthesis-related genes

NuA4, an essential histone acetyltransferase complex, is required for efficient transcription in eukaryotes. Using genome editing, genomic approaches and biochemical assays, we characterized plant homologues of two key components of this complex, EPL1 and EAF1 in Arabidopsis thaliana. Surprisingly, we found that loss of AtEPL1, which is necessary for enzymatic activity of NuA4, is not lethal. Contrary to yeast, mutants lacking AtEAF1, responsible for complex targeting, display severe pleiotropic phenotype which copies that of Atepl1. Atepl1 and Ateaf1 mutants grow slowly, contain reduced chlorophyll levels and small chloroplasts. We provide evidence that these alterations are not caused by disturbance of GLK transcription factors, the major regulators of chloroplast development. Using ChIP-seq we show that H4 acetylation levels are dramatically reduced in the chromatin of the Atepl1 mutant, while H3 acetylation remains mostly unchanged. We use our data to define NuA4-dependent genes and show that chloroplast-related genes are significantly overrepresented in this group, consistent with the pale-green phenotypes of the mutants. We propose that NuA4 was adopted in plants to control nuclear-encoded photosynthesis genes. SignificancePhotosynthesis depends on chloroplast proteins, most of which are nucleus-encoded and thus subject to control mechanisms common across eukaryotes. Here we show that NuA4, an evolutionary conserved transcriptional coactivator, is necessary for proper development of photosynthetic apparatus. Surprisingly, in contrast to yeast and metazoans, plants engineered to lack core NuA4 subunits are capable of vegetative development despite dramatic genome-wide loss of NuA4-dependent H4K5 acetylation. This chromatin perturbation seems to directly affect 350 genes which, in addition to reduced H4K5ac levels, display decreased transcript levels but no loss of transcription-related H3K9ac. A significant proportion of these genes are related to chloroplast function, particularly to translation, an intriguing parallel to the yeast NuA4s role in transcription of ribosome biogenesis-related genes.

molecular biology↗

bioRxiv2017

Nucleosomes and DNA methylation shape meiotic DSB frequency in Arabidopsis transposons and gene regulatory regions

Meiotic recombination initiates via DNA double strand breaks (DSBs) generated by SPO11 topoisomerase-like complexes. Recombination frequency varies extensively along eukaryotic chromosomes, with hotspots controlled by chromatin and DNA sequence. To map meiotic DSBs throughout a plant genome, we purified and sequenced Arabidopsis SPO11-1-oligonucleotides. DSB hotspots occurred in gene promoters, terminators and introns, driven by AT-sequence richness, which excludes nucleosomes and allows SPO11-1 access. A strong positive relationship was observed between SPO11-1 DSBs and final crossover levels. Euchromatic marks promote recombination in fungi and mammals, and consistently we observe H3K4me3 enrichment in proximity to DSB hotspots at gene 5-ends. Repetitive transposons are thought to be recombination-silenced during meiosis, in order to prevent non-allelic interactions and genome instability. Unexpectedly, we found strong DSB hotspots in nucleosome-depleted Helitron/Pogo/Tc1/Mariner DNA transposons, whereas retrotransposons were coldspots. Hotspot transposons are enriched within gene regulatory regions and in proximity to immunity genes, suggesting a role as recombination-enhancers. As transposon mobility in plant genomes is restricted by DNA methylation, we used the met1 DNA methyltransferase mutant to investigate the role of heterochromatin on the DSB landscape. Epigenetic activation of transposon meiotic DSBs occurred in met1 mutants, coincident with reduced nucleosome occupancy, gain of transcription and H3K4me3. Increased met1 SPO11-1 DSBs occurred most strongly within centromeres and Gypsy and CACTA/EnSpm coldspot transposons. Together, our work reveals complex interactions between chromatin and meiotic DSBs within genes and transposons, with significance for the diversity and evolution of plant genomes.

genomics↗