Biology subjects

Grognet, P.

Publications and source records attributed to Grognet, P..

10 recordsLinked to original sources

bioRxiv2026

Hi-C data from the filamentous fungus Podospora anserina and associated 3D models to visualize the spatial organization of its chromosomes

ObjectivesModel species are essential for fundamental research in biology. While a complete genomic sequence is a prerequisite for genetic studies, it is not enough on its own. Understanding the three-dimensional organization of the genome is also important, allowing researchers to gain a more realistic understanding of the mechanisms governing genome function. In the fungal model Podospora anserina, although the genomic sequence has been established for a long time, the three-dimensional organization remained unknown. Here we obtained the first Hi-C datasets and present associated 3D models, providing the research community with a valuable resource for better multi-omics data integration. Data descriptionHi-C experiments were performed in duplicate, using nuclei purified from wild-type fungal mycelium. Four FASTQ files were obtained (two per replicate) and used as inputs for the 3DGB workflow with four different output resolutions, to observe the genome organization of P. anserina at different levels of detail (50 kb, 20 kb, 10 kb, and 5 kb). In a context where researchers already have, for this species, a large amount of traditional omics data (ChIP-seq, RNA-seq, etc.), these 3D models are helpful for complementing the linear representation of the genome, which is traditionally used in bioinformatic analyses.

genomics↗

bioRxiv2025

A recent shift in centromere size and DNA content in Podospora pseudocomata co-occurs with the loss of a fungal genome defense system

The centromere of the eukaryotic chromosome is necessary for the accurate segregation during cell division. Yet, centromeric DNA is highly variable and rapidly evolving. In fungi, centromeres range from point- to regional centromeres, some of which are hundreds of thousands of base pairs long and filled with transposable elements. As fungi have evolved several specialized defense mechanisms against transposable elements, these regional centromeres are intriguing sites for investigating the connection between genome defense and centromere evolution. Here, we investigated the structure of the centromeres of seven species of the Podospora anserina species complex, which is made up of closely related filamentous ascomycetes that diverged less than 1 MYA. We discovered that one species in the complex, P. pseudocomata, lacks the genomic signature of the specialized genome defense mechanism called Repeat Induced Point mutations (RIP). We identified the centromeric regions in P. anserina and P. pseudocomata using chromatin immunoprecipitation targeting the centromere-specific histone variant cenH3, and using comparative genomics we inferred the size of centromeric regions in the other species. We found that while the centromere structure in the complex is generally well conserved, the centromeric regions of P. pseudocomata has gone through a rapid change. Specifically, the size of the centromeres in P. pseudocomata are 35-46 kb, which is significantly smaller than those of the other species (44-90 kb), and the DNA-transposon discoglosse is the most abundant TE family instead of the typical LTR-retrotransposon crapaud. Taken together, our data strongly indicates a link between genome defense and centromere evolution in fungi.

genomics↗

bioRxiv2025

The fastest movements and the most extreme deformations of the cell nucleus are driven by microtrubules in the model fungus Podospora anserina

A growing number of studies shows that nuclear movement, positioning, and the mechanical constraints applied to the nucleus play important roles in cellular physiology and gene regulation. This process, known as nuclear mechanotransduction, is an emerging field of research, and its deregulation can lead to severe human pathologies. In this study, we examine the behavior of female and male nuclei during sexual reproduction in the model fungus Podospora anserina. After plasmogamy, the two types of nuclei do not immediately undergo karyogamy. Instead, once inside the trichogyne (the female-specific hyphae extending from the ascogonium) the male nuclei migrate over distances of up to several hundred micrometers from the trichogyne tip, where plasmogamy occurs, to the ascogonium at the center of the protoperithecium. Our study highlights the contrasting behaviors of the two nuclear types: female nuclei remain immobile and spherical, while male nuclei move rapidly and exhibit striking deformations and stretching. Trichogynes are branched and compartmentalized, consisting of articles separated by septa that communicate through the septal pores. We show that during their migration across these compartments, male nuclei undergo the most extreme movements, deformations, and speeds reported to date for nuclei in motion. We also investigated the role of the actin and microtubule (MT) cytoskeletons in male nuclei behavior. Both cytoskeletal networks are highly dynamic and abundant in trichogynes, particularly at septa. We demonstrate that while actin depolymerization does not impair male nuclei migration, MT disruption causes a complete arrest of both nuclear movement and stretching, indicating a critical role for MTs. Finally, we reveal that the MT network in trichogynes is highly polarized, offering new insights into how male nuclei navigate within branched trichogynes.

cell biology↗

bioRxiv2025

Stepwise recombination suppression around the mating-type locus associated with a diploid-like life cycle in Schizothecium fungi

Recombination suppression often evolves around sex-determining loci and extends stepwise, resulting in adjacent regions with different levels of divergence between sex chromosomes, called evolutionary strata. In Ascomycota fungi, evolutionary strata have been found around the mating-type (MAT) locus only in pseudo-homothallic species, i.e., with a diploid-like lifecycle and mycelia carrying nuclei of both mating types. In contrast, no recombination suppression has been observed in fungi with a haploid-like lifecycle, such as heterothallic fungi (with mycelial colonies of a single mating type each). Here, we investigated the evolution of recombination suppression in a clade of dung fungi encompassing 16 pseudo-homothallic and three heterothallic sibling species from the Schizothecium genus (Ascomycota, Sordariales). The analysis of genetic divergence based on genome sequencing indicated recombination suppression around the MAT locus in all investigated 13 pseudo-homothallic species. The non-recombining region ranged from 600 kb to 1.6 Mb and harbored multiple evolutionary strata, varying in size and number among species. The separation of alleles associated with alternative mating types in gene genealogies across strains within species, the high linkage disequilibrium and an inversion in one species supported the lack of recombination in the MAT-proximal region in pseudo-homothallic species. The overall lack of trans-specific polymorphism suggested multiple independent events of recombination suppression or the occurrence of rare events of recombination or genic conversion. Progeny analyses showed the occurrence of recombination close to the MAT locus in heterothallic strains. We thus revealed here multiple and likely independent evolutionary strata, associated with an extended diploid-like stage in Schizothecium fungi, which provides a good model for research on sex-related chromosome evolution.

evolutionary biology↗

bioRxiv2024

Sexual reproduction is controlled by successive transcriptomic waves in Podospora anserina

Despite the inherent challenge of finding suitable mating partners, most eukaryotes use sexual reproduction to produce offspring endowed with increase genetic diversity and fitness. The persistence of this mode of reproduction is a key question in evolutionary biology. Fungi offer valuable insights into this question, due to their diversity and short lifecycle. This study focuses on Podospora anserina, a pseudohomothallic ascomycete fungus that bypasses self-sterility by maintaining two compatible nuclei in one mycelium. We performed genome-wide gene expression profiling during ten stages of P. anserina sexual reproduction and identified five major expression patterns. Our expert annotation approaches identified differentially expressed genes related to secondary metabolite production, fungal vegetative incompatibility, programmed cell death, and epigenetic regulation. In addition, master transcriptional regulators and their target networks were uncovered. This study provides a comprehensive database for future functional genomics experiments and novel pathway characterization during sexual reproduction.

genomics↗

bioRxiv2024

Genetic differentiation in the MAT-proximal region is not sufficient for suppressing recombination in Podospora anserina

Recombination is advantageous over the long-term, as it allows efficient selection and purging deleterious mutations. Nevertheless, recombination suppression has repeatedly evolved in sex chromosomes and mating-type chromosomes. The evolutionary causes for recombination suppression and the proximal mechanisms preventing crossing overs are still poorly understood. Several hypotheses have recently been suggested based on theoretical models, and in particular, that divergence could accumulate neutrally around a sex-determining region and reduce recombination rates, a self-reinforcing process that could foster progressive extension of recombination suppression. The ascomycete fungus Podospora anserina is an excellent model for investigating these questions. A 0.8 Mb region around the mating-type locus is non-recombining, despite being collinear between the two mating types. This fungus is mostly selfing, so that strains are highly homozygous, except in the non-recombining region around the mating-type locus that displays differentiation between mating types. Here, we generated a mutant to test the hypothesis that sequence divergence alone is responsible for recombination cessation. We replaced the mat-idiomorph by the sequence of the mat+ idiomorph, to obtain a strain that is sexually compatible with the mat-reference strain and isogenic to this strain in the MAT-proximal region. Crosses showed that recombination was still suppressed in the MAT-proximal region in the mutant strains, indicating that other proximal mechanisms than inversions or mere sequence divergence are responsible for recombination suppression in this fungus. This finding suggests that selective mechanisms likely acted for suppressing recombination, as the neutral model does not seem to hold, at least in this fungus. ARTICLE SUMMARYIn many organisms, a non-recombining region is observed around the sex-determining locus. In the fungus Podospora anserina, recombination is suppressed within an 800kb region around the mating-type locus. In natural strains, the genome is isogenic between two mating-types, except in this region displaying heterozygosity. To determine if this heterozygosity can be responsible for the lack of recombination, compatible strains were engineered to be isogenic, including in the non-recombining region. Recombination inhibition persisted, indicating that mere sequence divergence does not cause recombination suppression. Our study provides an interesting insight on the molecular and evolutionary causes of recombination inhibition.

genetics↗

bioRxiv2023

Visual integration of omics data to improve 3D models of fungal chromosomes

The functions of eukaryotic chromosomes and their spatial architecture in the nucleus are reciprocally dependent. Hi-C experiments are routinely used to study chromosome 3D organization by probing chromatin interactions. Standard representation of the data has relied on contact maps that show the frequency of interactions between parts of the genome. In parallel, it has become easier to build 3D models of the entire genome based on the same Hi-C data, and thus benefit from the methodology and visualization tools developed for structural biology. 3D modeling of entire genomes leverages the understanding of their spatial organization. However, this opportunity for original and insightful modeling is under exploited. In this paper, we show how seeing the spatial organization of chromosomes can bring new perspectives to Hi-C data analysis. We assembled state-of-the-art tools into a workflow that goes from Hi-C raw data to fully annotated 3D models and we re-analysed public Hi-C datasets available for three fungal species. Besides the well-described properties of the spatial organization of their chromosomes (Rabl conformation, hypercoiling and chromosome territories), our 3D models highlighted i) in Saccharomyces cerevisiae, the backbones of the cohesin anchor regions, which were aligned all along the chromosomes, ii) in Schizosaccharomyces pombe, the oscillations of the coiling of chromosome arms throughout the cell cycle and iii) in Neurospora crassa, the massive relocalization of histone marks in mutants of heterochromatin regulators. 3D modeling of the chromosomes brings new opportunities for visual integration. This holistic perspective supports intuition and lays the foundation for building new concepts.

genomics↗

bioRxiv2022

Re-exploiting multiple RNA-seq data to identify transcript variations in Podospora anserina

BackgroundPublicly available RNA-seq datasets are often underused although being helpful to improve functional annotation of eukaryotic genomes. This is especially true for filamentous fungi genomes which structure differs from most well annotated yeast genomes. Podospora anserina is a filamentous fungal model, which genome has been sequenced and annotated in 2008. Still, the current annotation lacks information about cis-regulatory elements, including promoters, transcription starting sites and terminators, which are instrumental to integrate epigenomic features into global gene regulation strategies. ResultsHere we took advantage of 37 RNA-seq experiments that were obtained in contrasted developmental and physiological conditions, to complete the functional annotation of P. anserina genome. Out of the 10,800 previously annotated genes, 5UTR and 3UTR were defined for 7,554, among which, 3,328 showed differential transcriptional signal starts and/or transcriptional end sites. In addition, alternative splicing events were detected for 2350 genes, mostly due alternative 3splice site and 1,958 novel transcriptionally active regions (nTARs) in unannotated regions were identified. ConclusionsOur study provides a comprehensive genome-wide functional annotation release of P. anserina genome, including chromatin features, cis-acting elements such as UTRs, alternative splicing events and transcription of non-coding regions. These new findings will likely improve our understanding of gene regulation strategies in compact genomes, such as those of filamentous fungi. Characterization of alternative transcripts and nTARs paves the way to the discovery of putative new genes, alternative peptides or regulatory non-coding RNAs.

genomics↗

bioRxiv2020

Loss of Polycomb Protein EZH2 causes major depletion of H3K27 and H3K9 tri-methylation and developmental defects in the fungus Podospora anserina

Selective gene silencing is key to development. The H3K27me3 enriched heterochromatin maintains transcription repression established during early development and regulates cell fate. Conversely, H3K9me3 enriched heterochromatin prevents differentiation but constitutes a permanent protection against transposable element. We exploited the fungus Podospora anserina, a valuable alternative to higher eukaryote models to question the biological relevance and interplay of these two distinct heterochromatin conformations. We found that H3K27me3 and H3K9me3 modifications are mutually exclusive within gene-rich regions but not within repeats. Lack of PaKmt6 EZH2-like enzyme resulted in loss of H3K27me3 and in significant H3K9me3 reduction, whereas lack of PaKmt1 SU(VAR)3-9-like enzyme caused loss of H3K9me3 only. We established that P. anserina developmental programs require H3K27me3 mediated silencing unlike most fungi studied to date. Our findings provide new insight into roles of these histone marks and into the relationship between chromatin modifications and development.

genetics↗

bioRxiv2019

A RID-like cytosine methyltransferase homologue controls sexual development in the fungus Podospora anserina

DNA methyltransferases are ubiquitous enzymes conserved in bacteria, plants and opisthokonta. These enzymes, which methylate cytosines, are involved in numerous biological processes, notably development. In mammals and higher plants, methylation patterns established and maintained by the cytosine DNA methyltransferases (DMTs) are essential to zygotic development. In fungi, some members of an extensively conserved fungal-specific DNA methyltransferase class are both mediators of the Repeat Induced Point mutation (RIP) genome defense system and key players of sexual reproduction. Yet, no DNA methyltransferase activity of these purified RID (RIP deficient) proteins could be detected in vitro. These observations led us to explore how RID-like DNA methyltransferase encoding genes would play a role during sexual development of fungi showing very little genomic DNA methylation, if any.\n\nTo do so, we used the model ascomycete fungus P. anserina. We identified the PaRid gene, encoding a RID-like DNA methyltransferase and constructed knocked-out {Delta}PaRid defective mutants. Crosses involving P. anserina {Delta}PaRid mutants are sterile. Our results show that, although gametes are readily formed and fertilization occurs in a {Delta}PaRid background, sexual development is blocked just before the individualization of the dikaryotic cells leading to meiocytes. Complementation of {Delta}PaRid mutants with ectopic alleles of PaRid, including GFP-tagged, point-mutated, inter-specific and chimeric alleles, demonstrated that the catalytic motif of the putative PaRid methyltransferase is essential to ensure proper sexual development and that the expression of PaRid is spatially and temporally restricted. A transcriptomic analysis performed on mutant crosses revealed an overlap of the PaRid-controlled genetic network with the well-known mating-types gene developmental pathway common to an important group of fungi, the Pezizomycotina.\n\nAuthor SummarySexual reproduction is considered to be essential for long-term persistence of eukaryotic species. Sexual reproduction is controlled by strict mechanisms governing which haploids can fuse (mating) and which developmental paths the resulting zygote will follow. In mammals, differential genomic DNA methylation patterns of parental gametes, known as DNA methylation imprints are essential to zygotic development, while in plants, global genomic demethylation often results in female-sterility. Although animal and fungi are evolutionary related, little is known about epigenetic regulation of gene expression and development in multicellular fungi. Here, we report on a gene of the model fungus Podospora anserina, encoding a protein called PaRid that looks like a DNA methyltrasferase. We showed that expression of the catalytically functional version of the PaRid protein is required in the maternal parental strain to form zygotes. By establishing the transcriptional profile of PaRid mutant strain, we identified a set of PaRid direct and/or indirect target genes. Half of them are also targets of a mating-type transcription factor known to be a major regulator of sexual development. So far, there was no other example of identified RID targets shared with a well-known developmental pathway that is common to an important group of fungi, the Pezizomycotina

genetics↗