Biology subjects

Nowrousian, M.

Publications and source records attributed to Nowrousian, M..

11 recordsLinked to original sources

bioRxiv2025

Spore type-specific gene expression profiles underlying development and leaf infection processes of Colletotrichum graminicola

Colletotrichum graminicola causes significant losses of the staple crop maize worldwide. The fungus produces two distinct asexual spore types, oval and falcate conidia, which show unique processes in development and plant interaction. Based on genome resequencing of our laboratory strain (CgM2/ M1.001), we investigated the gene expression profiles of oval and falcate conidia during development and early leaf infection using RNA-seq. Our results reveal specific gene expression profiles between the two spore types, indicating fundamental differences in their developmental programs that reflect different modes of infection. We identified expression patterns discriminating both conidia types from mycelium and spore type-specific ones for genes encoding transcription factors, conserved fungal developmental genes, transporters, genes of secondary metabolite clusters, and pathogenicity-related functions, including effectors and carbohydrate-active enzymes (CAZymes). Our study shows that despite the identical genomic basis, oval and falcate conidia show unique transcriptomes across vegetative development and early plant interaction. Taking together, these results provide new insights into the molecular mechanisms determining the biology of C. graminicola and its interaction with the plant host.

microbiology↗

bioRxiv2024

Essential genes encoded by the mating-type locus of the human fungal pathogen Cryptococcus neoformans

Fungal sexual reproduction is controlled by the mating-type (MAT) locus. In contrast to a majority of species in the phylum Basidiomycota that have tetrapolar mating-type systems, the opportunistic human pathogen Cryptococcus neoformans employs a bipolar mating-type system, with two mating types (a and ) determined by a single MAT locus that is unusually large ([~]120 kb) and contains more than 20 genes. While several MAT genes are associated with mating and sexual development, others control conserved cellular processes (e.g. cargo transport and protein synthesis), of which five (MYO2, PRT1, RPL22, RPL39, and RPO41) have been hypothesized to be essential. In this study, through genetic analysis involving sporulation of heterozygous diploid deletion mutants, as well as in some cases construction and analyses of conditional expression alleles of these genes, we confirmed that with the exception of MYO2, both alleles of the other four MAT genes are indeed essential for cell viability. We further showed that while MYO2 is not essential, its function is critical for infectious spore production, faithful cytokinesis, adaptation for growth at high temperature, and pathogenicity in vivo. Our results demonstrate the presence of essential genes in the MAT locus that are divergent between cells of opposite mating types. We discuss possible mechanisms to maintain functional alleles of these essential genes in a rapidly-evolving genomic region in the context of fungal sexual reproduction and mating-type evolution. IMPORTANCESexual reproduction is essential for long-term evolutionary success. Fungal cell type identity is governed by the MAT locus, which is typically rapidly evolving and highly divergent between different mating types. In this study, we show that the a and alleles of four genes encoded in the MAT locus of the opportunistic human fungal pathogen C. neoformans are essential. We demonstrate that a fifth gene, MYO2, which had been predicted to be essential, is in fact dispensable for cell viability. However, a functional MYO2 allele is important for cytokinesis and fungal pathogenicity. Our study highlights the need for careful genetic analyses in determining essential genes, which is complementary to high-throughput approaches. Additionally, the presence of essential genes in the MAT locus of C. neoformans provides insights into the function, maintenance, and evolution of these fast-evolving genomic regions.

genetics↗

bioRxiv2024

Innovations in Alginate Catabolism Leading to Heterotrophy and Adaptive Evolution of Diatoms

A major goal of evolutionary biology is to identify the genetic basis for the emergence of adaptive traits. Diatoms are ancestrally photosynthetic microalgae. However, in the genus Nitzschia, loss of photosynthesis led to a group of free-living secondary heterotrophs whose manner of energy acquisition is unclear. Here, we sequence the genome of the non-photosynthetic diatom Nitzschia sing1 and identify the genetic basis for its catabolism of the brown seaweed cell wall polysaccharide alginate. N. sing1 obtained an endolytic alginate lyase enzyme by horizontal gene transfer (HGT) from a marine bacterium. Subsequent gene duplication and transposition led to 91 genes in three distinct gene families. One family retains the ancestral endolytic enzyme function. By contrast, the two others underwent domain duplication, gain, loss, rearrangement, and mutation to encode novel functions that can account for oligosaccharide import through the endomembrane system and the exolytic production of alginate monosaccharides. Together, our results show how a single HGT event followed by substantial gene duplication and neofunctionalization led to alginate catabolism and access to a new ecological niche. HighlightsO_LIN. sing1 acquired an alginate lyase (ALY) gene by horizontal gene transfer from a marine bacterium C_LIO_LIThis founding gene expanded and diversified to comprise 3 major families across 30 loci C_LIO_LIDerived functions account for alginate import and processing into monomers C_LIO_LIDomain duplication, gain, loss, mutation, and de novo sequence evolution underlie ALY gene neofunctionalization C_LI O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=169 SRC="FIGDIR/small/610029v1_ufig1.gif" ALT="Figure 1000"> View larger version (44K): org.highwire.dtl.DTLVardef@7a47c7org.highwire.dtl.DTLVardef@1fe958forg.highwire.dtl.DTLVardef@10ce362org.highwire.dtl.DTLVardef@12465a1_HPS_FORMAT_FIGEXP M_FIG Graphical Abstract C_FIG

evolutionary biology↗

bioRxiv2023

Comparative genomics of Cryptococcus and Kwoniella reveals pathogenesis evolution and contrasting modes of karyotype evolution via chromosome fusion or intercentromeric recombination

A large-scale comparative genomic analysis was conducted for the global human fungal pathogens within the Cryptococcus genus, compared to non-pathogenic Cryptococcus species, and related species from the sister genus Kwoniella. Chromosome-level genome assemblies were generated for multiple species of both genera, resulting in a dataset encompassing virtually all of their known diversity. Although Cryptococcus and Kwoniella have comparable genome sizes (about 19.2 and 22.9 Mb) and similar gene content, hinting at pre-adaptive pathogenic potential, our analysis found evidence in pathogenic Cryptococcus species of specific examples of gene gain (via horizontal gene transfer) and gene loss, which might represent evolutionary signatures of pathogenic development. Genome analysis also revealed a significant variation in chromosome number and structure between the two genera. By combining synteny analysis and experimental centromere validation, we found that most Cryptococcus species have 14 chromosomes, whereas most Kwoniella species have fewer (11, 8, 5 or even as few as 3). Reduced chromosome number in Kwoniella is associated with formation of giant chromosomes (up to 18 Mb) through repeated chromosome fusion events, each marked by a pericentric inversion and centromere loss. While similar chromosome inversion-fusion patterns were observed in all Kwoniella species with fewer than 14 chromosomes, no such pattern was detected in Cryptococcus. Instead, Cryptococcus species with less than 14 chromosomes, underwent chromosome reductions primarily through rearrangements associated with the loss of repeat-rich centromeres. Additionally, Cryptococcus genomes exhibited frequent interchromosomal translocations, including intercentromeric recombination facilitated by transposons shared between centromeres. Taken together, our findings advance our understanding of genomic changes possibly associated with pathogenicity in Cryptococcus and provide a foundation to elucidate mechanisms of centromere loss and chromosome fusion driving distinct karyotypes in closely related fungal species, including prominent global human pathogens.

genomics↗

bioRxiv2023

Functional analysis of chromatin-associated proteins in Sordaria macrospora reveals similar roles for RTT109 and ASF1 in development and DNA damage response

We performed a functional analysis of two potential partners of ASF1, a highly conserved histone chaperone that plays a crucial role in the sexual development and DNA damage resistance in the ascomycete Sordaria macrospora. ASF1 is known to be involved in nucleosome assembly and disassembly, binding histones H3 and H4 during transcription, replication and DNA repair and has direct and indirect roles in histone recycling and modification as well as DNA methylation, acting as a chromatin modifier hub for a large network of chromatin-associated proteins. Here, we functionally characterized two of these proteins, RTT109 and CHK2. RTT109 is a fungal-specific histone acetyltransferase, while CHK2 is an ortholog to PRD-4, a checkpoint kinase of Neurospora crassa that performs similar cell cycle checkpoint functions as yeast RAD53. Through the generation and characterization of deletion mutants, we discovered striking similarities between RTT109 and ASF1 in terms of their contributions to sexual development, histone acetylation and protection against DNA damage. Phenotypic observations revealed a developmental arrest at the same stage in {Delta}rtt109 and {Delta}asf1 strains, accompanied by a loss of H3K56 acetylation, as detected by western blot analysis. Deletion mutants of rtt109 and asf1 are sensitive to the DNA damaging agent MMS (methylmethane sulfonate), but not HU (hydroxyurea). In contrast, chk2 mutants are fertile and resistant to MMS, but not HU. Our findings suggest a close functional association between ASF1 and RTT109 in the context of development, histone modification and DNA damage response, while indicating a role for CHK2 in separate pathways of the DNA damage response. Article summaryIn the filamentous fungus Sordaria macrospora, the conserved histone chaperone ASF1, which interacts with histones H3 and H4, was previously shown to be required for multicellular development and DNA damage response. Here, we have analyzed two additional chromatin-associated proteins. rtt109 encodes a histone acetyltransferase, and deletion of the gene in S. macrospora results in a phenotype similar to that of a {Delta}asf1 mutant, whereas chk2 is involved in different aspects of the DNA damage response, but not in development.

molecular biology↗

bioRxiv2023

Histone binding of ASF1 is required for fruiting body development, but not for genome stability in the filamentous fungus Sordaria macrospora

The highly conserved eukaryotic histone chaperone ASF1 is involved in the assembly and disassembly of nucleosomes during transcription, DNA replication and repair. It was the first chaperone discovered to be involved in all three of these processes. The filamentous fungus Sordaria macrospora is one of only two multicellular organisms where asf1 deletions are viable, which makes it useful for in vivo analysis of this central regulator of eukaryotic chromatin structure. Deletion of asf1 in S. macrospora leads to sterility, a reduction of DNA methylation, and upregulation of genes that are usually weakly expressed in the wild type. Here, we focused on the functions of the highly conserved core and the divergent C-terminal tail of ASF1, studied the effects of ASF1 on histone modifications and tested its relevance for genomic stability. By Co-IP and complementation analysis we showed that substitutions of amino acid V94 or truncations of the C-terminal tail abolish histone binding and do not complement the sterile mutant phenotype. {Delta}asf1 is sensitive to the DNA damaging agent MMS, while complementation strains, even those with non-histone-binding variants, regain wild type-like resistance. To aid in subsequent ChIP-seq and Hi-C analyses, we generated a chromosome-resolved genome assembly of S. macrospora. ChIP-Seq analysis revealed a global increase of H3K27me3 in {Delta}asf1, accompanied by a global decrease of H3K56ac. By using Hi-C we detected a tandem duplication of around 600 kb on chromosome 2 in the mutant. Crossing experiments indicated linkage between the viability of {Delta}asf1 strains and the presence of the duplication. ImportanceHistone chaperones are proteins that are involved in nucleosome assembly and disassembly and can therefore influence all DNA-dependent processes including transcription, DNA replication and repair. ASF1 is a histone chaperone that is conserved throughout eukaryotes. In contrast to most other multicellular organisms, a deletion mutant of asf1 in the fungus Sordaria macrospora is viable; however, the mutant is sterile. In this study, we could show that the histone binding ability of ASF1 is required for fertility in S. macrospora, whereas the function of ASF1 in maintenance of genome stability does not require histone binding. We also showed that the histone modifications H3K27me3 and H3K56ac are misregulated in the {Delta}asf1 mutant. Furthermore, we identified a large duplication on chromosome 2 of the mutant strain that is genetically linked to the {Delta}asf1 allele present on chromosome 6, suggesting that viability of the mutant might depend on the presence of the duplicated region.

molecular biology↗

bioRxiv2023

Lifestyle transitions in basidiomycetous fungi are reflected by tRNA composition and translation efficiency of metabolic genes

Fungi are ubiquitous and inhabit every known terrestrial habitat. Pathogenic fungi are highly diverse, and recent years have seen an uprise in the emergence of new pathogens on crops, animals and humans. The order Trichosporonales (Tremellomycetes, Agaricomycotina, Basidiomycota) harbours saprobic and a few opportunistic human pathogenic species. These emerging pathogens cause superficial skin irritations, as well as invasive life-threatening infections. Yet, little is known about their evolution, ecology, virulence mechanisms and transition to pathogenic lifestyles. In this study we aimed to determine genomic signatures associated with lifestyle transitions, virulence and host/substrate specialization among 30 Trichosporonales species from a total of 41 genome sequences. We used comparative analyses of genome content, including gene functional categories, repetitive element content and tRNA composition among saprotrophic and reported opportunistic human pathogens. A genome-scale phylogenetic reconstruction revealed that even though the different genera are monophyletic, opportunistic pathogenic species are present in distantly-related clades. Statistical analyses showed that differences in genome structure among species did not correlate with predicted lifestyles. Intriguingly, we found that tRNA content varied widely across species (from 51 to 1455 manually curated tRNA genes). The expansion was independent from the phylogenetic structure. Opportunistic pathogenic species showed an overall increased efficiency in the translation of genes associated with host colonization (i.e. lipid metabolism), while exclusively saprotrophic species showed an increase translation efficiency for genes associated with a saprotrophic lifestyle (i.e. carbohydrate metabolism). This pattern was consistent among distantly-related saprotrophic and pathogenic Cryptococcus species (order Tremellales). In conclusion, our analyses link genomic information with ecology and fungal lifestyles across an entire order. We find evidence for an evolutionary scenario where distinct habitats select for an optimized translation of genes involved in successful proliferation in the respective habitat. We predict that lifestyles are not strictly defined by gene repertoires, but also by expression profiles in fungal pathogens.

genomics↗

bioRxiv2022

Obligate sexual reproduction of a homothallic fungus closely related to the Cryptococcus pathogenic species complex

Sexual reproduction is a ubiquitous and ancient trait of eukaryotic life. While sexual organisms are usually faced with the challenge of finding a compatible mating partner, species as diverse as animals, plants, and fungi have repeatedly evolved the ability to reproduce sexually without an obligate requirement for another individual. Here, we uncovered the underlying mechanism of self-compatibility (homothallism) in Cryptococcus depauperatus, a fungal species sister to the clinically relevant human fungal pathogens Cryptococcus neoformans and Cryptococcus gattii species complexes. In contrast to C. neoformans or C. gattii, which grow as a yeast in the asexual stage, and produce hyphae, basidia, and infectious spores during the sexual stage, C. depauperatus grows exclusively as hyphae decorated with basidia and abundant spores and appears to be continuously engaged in sexual reproduction. By combining the insights from comparative genomics and genetic analyses of mutants defective in key mating and meiosis genes, we demonstrate the sexual cycle of C. depauperatus involves meiosis, and reveal that self-compatibility is orchestrated by the expression, in the same cell, of an unlinked mating receptor (Ste3a) and pheromone ligand (MF) pair seemingly derived from opposite mating types of a heterothallic (self-sterile) ancestor. We identified a putative mating-type (MAT) determining region containing genes phylogenetically aligned with MATa alleles of other species, and a few MAT gene alleles scattered and unlinked throughout the genome, but no homologs of the mating-type homeodomain genes SXI1 (HD1) and SXI2 (HD2). Comparative genomic analyses suggested a dramatic remodeling of the MAT locus possibly owing to reduced selective constraints to maintain mating-type genes in tight linkage, associated with a transition to self-fertility. Our findings support C. depauperatus as an obligately sexual, homothallic fungal species and provide additional insight into the repeated transitions between modes of sexual reproduction that have occurred throughout the fungal kingdom.

microbiology↗

bioRxiv2019

Long transposon-rich centromeres in an oomycete reveal divergence of centromere features in Stramenopila-Alveolata-Rhizaria lineages

Centromeres are chromosomal regions that serve as platforms for kinetochore assembly and spindle attachments, ensuring accurate chromosome segregation during cell division. Despite functional conservation, centromere DNA sequences are diverse and often repetitive, making them challenging to assemble and identify. Here, we describe centromeres in an oomycete Phytophthora sojae by combining long-read sequencing-based genome assembly and chromatin immunoprecipitation for the centromeric histone CENP-A followed by high-throughput sequencing (ChIP-seq). P. sojae centromeres cluster at a single focus at different life stages and during nuclear division. We report an improved genome assembly of the P. sojae reference strain, which enabled identification of 15 enriched CENP-A binding regions as putative centromeres. By focusing on a subset of these regions, we demonstrate that centromeres in P. sojae are regional, spanning 211 to 356 kb. Most of these regions are transposon-rich, poorly transcribed, and lack the histone modification H3K4me2 but are embedded within regions with the heterochromatin marks H3K9me3 and H3K27me3. Strikingly, we discovered a Copia-like transposon (CoLT) that is highly enriched in the CENP-A chromatin. Similar clustered elements are also found in oomycete relatives of P. sojae, and may be applied as a criterion for prediction of oomycete centromeres. This work reveals a divergence of centromere features in oomycetes as compared to other organisms in the Stramenopila-Alveolata-Rhizaria (SAR) supergroup including diatoms and Plasmodium falciparum that have relatively short and simple regional centromeres. Identification of P. sojae centromeres in turn also advances the genome assembly. Author summaryOomycetes are fungal-like microorganisms that belong to the stramenopiles within the Stramenopila-Alveolata-Rhizaria (SAR) supergroup. The Phytophthora oomycetes are infamous as plant killers, threatening crop production worldwide. Because of the highly repetitive nature of their genomes, assembly of oomycete genomes presents challenges that impede identification of centromeres, which are chromosomal sites mediating faithful chromosome segregation. We report long-read sequencing-based genome assembly of the Phytophthora sojae reference strain, which facilitated the discovery of centromeres. P. sojae harbors large regional centromeres fully embedded in heterochromatin, and enriched for a Copia-like transposon that is also found in discrete clusters in other oomycetes. This study provides insight into the oomycete genome organization, broadens our knowledge of the centromere structure, function and evolution in eukaryotes, and may help elucidate the high frequency of aneuploidy during oomycete reproduction.

microbiology↗

bioRxiv2019

Convergent evolution of linked mating-type loci in basidiomycetes: an ancient fusion event that has stood the test of time

Sexual development is a key evolutionary innovation of eukaryotes. In many species, mating involves interaction between compatible mating partners that can undergo cell and nuclear fusion and subsequent steps of development including meiosis. Mating compatibility in fungi is governed by mating type determinants, which are localized at mating type (MAT) loci. In basidiomycetes, the ancestral state is hypothesized to be tetrapolar (bifactorial), with two genetically unlinked MAT loci containing homeodomain transcription factor genes (HD locus) and pheromone and pheromone receptor genes (P/R locus), respectively. Alleles at both loci must differ between mating partners for completion of sexual development. However, there are also basidiomycete species with bipolar (unifactorial) mating systems, which can arise through genomic linkage of the HD and P/R loci. In the order Tremellales, which is comprised of mostly yeast-like species, bipolarity is found only in the human pathogenic Cryptococcus species. Here, we describe the analysis of MAT loci from the Trichosporonales, a sister order to the Tremellales. We analyzed genome sequences from 29 strains that belong to 24 species, including two new genome sequences generated in this study. Interestingly, in all of the species analyzed, the MAT loci are fused and a single HD gene is present in each mating type. This is similar to the organization in the pathogenic Cryptococci, which also have linked MAT loci and carry only one HD gene per MAT locus instead of the usual two HD genes found in the vast majority of basidiomycetes. However, the HD and P/R allele combinations in the Trichosporonales are different from those in the pathogenic Cryptococcus species. The differences in allele combinations compared to the bipolar Cryptococci as well as the existence of tetrapolar Tremellales sister species suggest that fusion of the HD and P/R loci and differential loss of one of the two HD genes per MAT allele occurred independently in the Trichosporonales and pathogenic Cryptococci. This finding supports the hypothesis of convergent evolution at the molecular level towards fused mating-type regions in fungi, similar to previous findings in other fungal groups. Unlike the fused MAT loci in several other basidiomycete lineages though, the gene content and gene order within the fused MAT loci are highly conserved in the Trichosporonales, and there is no apparent suppression of recombination extending from the MAT loci to adjacent chromosomal regions, suggesting different mechanisms for the evolution of physically linked MAT loci in these groups.\n\nAuthor summarySexual development in fungi is governed by genes located within a single mating type (MAT) locus or at two unlinked MAT loci. While the latter is thought to be the ancestral state in basidiomycetes, physical linkage of the two MAT loci has occurred multiple times during basidiomycete evolution. Here, we show that physically linked MAT loci are present in all analyzed species of the basidiomycete order Trichosporonales. In contrast to previously studied basidiomycetes, the fused MAT loci in the Trichosporonales have highly conserved gene order, suggesting that this fusion might date back to the common ancestor of this lineage.

evolutionary biology↗

bioRxiv2019

Genetic and genomic analyses reveal boundaries between species closely related to Cryptococcus pathogens

Speciation is a central mechanism of biological diversification. While speciation is well studied in plants and animals, in comparison, relatively little is known about speciation in fungi. One fungal model is the Cryptococcus genus, which is best known for the pathogenic Cryptococcus neoformans/Cryptococcus gattii species complex that causes over 200,000 new infections in humans annually. The closest non-human pathogenic relatives are the sibling species, Cryptococcus amylolentus and Tsuchiyaea wingfieldii. However, because relatively few isolates of each species are available, it is unclear whether they represent divergent lineages of the same species or different biological species. The recent isolation of an additional strain, preliminarily identified as T. wingfieldii, prompted us to reexamine this group as it may inform about the evolutionary processes underlying the diversification of both non-pathogenic and pathogenic Cryptococcus lineages. Using genomic data, we reappraised the phylogenetic relationship of the four available strains and confirmed the genetic separation of C. amylolentus and T. wingfieldii (now Cryptococcus wingfieldii), and revealed an additional cryptic species, for which the name Cryptococcus floricola is proposed. Comparison of full-length chromosome assemblies revealed approximately 6% pairwise sequence divergence between the three species, and identified significant genomic changes, including inversions as well as a reciprocal translocation that involved inter-centromeric ectopic recombination, which together likely impose significant barriers to genetic exchange. Using genetic crosses, we show that while C. wingfieldii cannot interbreed with any of the other strains, C. floricola can undergo sexual reproduction with C. amylolentus. However, most of the spores resulting from this cross were inviable, and many were sterile, indicating that the two species are genetically isolated through intrinsic post-zygotic barriers and possibly due to niche differentiation. Genome sequencing and analysis of the progeny demonstrated decreased recombination frequency during meiosis in heterospecific crosses compared to C. amylolentus conspecific crosses. This study advances our understanding of speciation in fungi and highlights the power of genomics in assisting our ability to correctly identify and discriminate fungal species. Author SummaryThe idea of species as discrete natural units seems rather intuitive for most people, just as cells are the basic units of life. However, when observing variation across a species range, boundaries can become blurred making it less than obvious when different populations evolve into separate species. Additionally, separate species can still interbreed, such as lions breeding with tigers to produce a liger or a tigon (depending on the paternal and maternal species of origin), but the resulting offspring is usually inviable or sterile, which in turn is evidence that the parents involved are distinct species. Therefore, what species are and how they originate is still an open question in evolutionary biology. While recent advances have been made in the fields of animal and plant speciation, many other important components of biological diversity, such as fungi, are still understudied. Genome sequencing is now providing new tools to address the genetic mechanisms that drive divergence and reproductive isolation between populations, including genetic incompatibilities, sequence divergence, and chromosomal rearrangements. Here we focus on the Cryptococcus amylolentus species complex, a non-pathogenic fungal lineage closely related to the human pathogenic Cryptococcus neoformans/Cryptococcus gattii complex. Using genetic and genomic analysis we reexamined the species boundaries of four available isolates within the C. amylolentus complex and revealed three genetically isolated species. The genomes of these species are ~6% divergent and exhibit chromosome rearrangements, including translocations and small-scale inversions. Although two of the species (C. amylolentus and newly described C. floricola) are still able to interbreed, the resulting hybrid progeny were mostly inviable, and many were sterile, indicating that barriers to reproduction have already been established. Our results will foster additional studies addressing the transitions between non-pathogenic and pathogenic Cryptococcus lineages.

evolutionary biology↗