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Macas, J.

Publications and source records attributed to Macas, J..

18 recordsLinked to original sources

An atlas of eukaryotic centromere architecture reveals recurrent evolutionary dynamics

Centromeres evolved at the root of eukaryotes to segregate chromosomes during cell division. Despite their ancient origin, centromeric DNA sequences evolve rapidly and adopt diverse architectures, including point centromeres, satellite arrays, transposon clusters, and holocentrics. To analyse centromere evolution at a broad scale, we characterised architectures across 325 diverse Darwin Tree of Life genome assemblies. Centromere architecture is evolutionarily labile, and similar configurations arise independently across divergent lineages. In plants and animals, we modelled centromere evolution as a recurrent cycle, in which satellite- and transposon-based architectures interconvert, with independent origins of holocentricity. We curated >23 million satellite repeats comprising 263 families from 165 species. Despite sequence divergence between satellite families, higher order repeats are prevalent, indicating constraint on repeat architecture rather than primary sequence. Satellite arrays are heavily invaded by diverse transposon families, consistent with convergent adaptation to the centromeric niche. In 89 species, transposons themselves constitute the primary centromere structure. We observed centrophilic transposons forming tandem arrays, suggesting mechanisms for satellite regeneration. Our sample includes five independent origins of holocentricity in plants and animals, which vary in association with periodic satellite arrays. We propose that genetic instability, centrophilic transposition, and transmission distortion promote recurrent centromere architectural interconversions during evolution.

genomics↗

Benchmarking cell-type deconvolution in cross-platform transcriptomic data

BackgroundTranscriptomic data from diverse measurement technologies are widely used to study tissue heterogeneity. Cell-type deconvolution, which resolves mixed transcriptomic signals into cellular components, is a key analytical approach. However, achieving accurate deconvolution across platforms remains challenging due to platform-specific experimental and technological biases. ResultsWe systematically benchmarked deconvolution performance using real-world cross-platform datasets and simulated data modeling distinct technological features. Our analyses provide practical guidelines for experimental design and promote more robust and comparable cross-platform transcriptomic analyses. ConclusionLog-normal regression methods such as SpatialDecon demonstrated the most reliable and consistent performance across both simulated and experimental settings, establishing a robust framework for accurate cross-platform transcriptomic deconvolution. Moreover, our results highlight potential caveats in deconvolution predictions arising from specific experimental conditions, providing guidance for more informed experimental design and interpretation Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=60 SRC="FIGDIR/small/690141v1_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@585cbdorg.highwire.dtl.DTLVardef@13088c7org.highwire.dtl.DTLVardef@163ea75org.highwire.dtl.DTLVardef@b5ccf7_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗

Kinetochore mutations and histone phosphorylation pattern changes preceded holo- and macro-monocentromere evolution

Centromeres are essential for kinetochore assembly and spindle attachment. While chromosomes of most species are monocentric with a single centromere, a minority exhibit holocentricity, with a centromere along the chromatid length. Sporadic emergence of holocentricity suggests multiple independent transitions. To explore this, we compared the centromere and (epi)genome organization of two sister genera with contrasting centromere types: Chamaelirium luteum with large "macro-monocentromeres" and Chionographis japonica with holocentromeres. Both exhibit chromosome-wide histone phosphorylation patterns distinct from typical monocentric species. Kinetochore analysis revealed similar chimeric Borealin in both species, with additional KNL2 loss and NSL1 chimerism in Cha. luteum. The broad-scale synteny between both genomes supports de novo holocentromere formation in Chi. japonica. Despite sharing features with both centromere types, macro-monocentromeres do not represent a direct link between mono- and holocentromeres. We propose a model for the divergent evolution involving kinetochore gene mutations, altered histone phosphorylation patterns, and centromeric satellite DNA amplification.

evolutionary biology↗

A new genome assembly of the pea cultivar Cameor provides resources for functional genomics and genetics

Significant improvements in sequencing technologies have allowed the development of more contiguous genome assemblies in many plant species. The pea genome is characterized by its richness in repeated elements and its long and complex centromeres. This makes its assembly challenging. In this paper, we present an improved version of the genome sequence of the French cultivar Cameor. This sequence was obtained by combining Nanopore and PacBio long-read sequencing, Hi-C contact maps and Bionano maps. The assembly of centromeres was refined using a combination of FISH and ultra-long Nanopore read analyses. Overall, Cameor_v2 genome assembly is a highly continuous pea genome assembly with small total gap size and a large contig N50. In this version, the orientation of chromosomes was revised according to internationally accepted karyotype rules. Gene annotation statistics indicated a high completeness of gene sequences, with most gene sequences with 3 and 5 UTR. This genome assembly with its associated data constitute a useful resource for pea genetics, comparative mapping and functional genomics.

genomics↗

Modulation of the ATP-adenosine signaling axis combined with radiotherapy facilitates anti-cancer immunity in brain metastasis

The immunosuppressive microenvironment in the brain poses a major limitation to successful therapy for brain metastases. Here we report that blockade of the ATP-to-adenosine-converting enzymes CD39 and CD73 and the adenosine receptor A2AR in combination with radiotherapy attenuates tumor progression in a breast-to-brain metastasis model by facilitating anti-cancer immunity. Immunophenotyping revealed loss of exhausted T cells and higher abundance of anti-cancer effector T cell populations. This effect was accompanied by a decrease of immunosuppressive lipid-laden macrophages and an expansion of CD14CD33high macrophages associated with antigen presentation. Analyses of human brain metastases samples supports a role of the ATP-adenosine signaling axis in modulating tumor inflammation and identified expression of CD39 and adenosine deaminase as predictive markers for patient survival and/or immune infiltration. Our findings demonstrate that the adenosine axis represents a druggable pathway to achieve local immunomodulation and treatment response, opening a new therapeutic avenue for brain metastases patients.

cancer biology↗

Identification and characterization of tertiary lymphoid structures in brain metastases

Brain metastases (BrM) are the most common cancers in the brain. We performed transcriptome-wide gene expression profiling combined with spatial immune cell profiling to characterize the tumor immune microenvironment in BrM from different primary tumors. We found that BrM from lung carcinoma and malignant melanoma showed overall higher immune cell infiltration as compared to BrM from breast carcinoma. RNA sequencing-based immune cell deconvolution revealed gene expression signatures indicative of tertiary lymphoid structures (TLS) in subsets of BrM, mostly from lung cancer and melanoma. This finding was corroborated by multiplex immunofluorescence staining of immune cells in BrM tissue sections. Detection of TLS signatures was more common in treatment-naive BrM and associated with prolonged survival after BrM diagnosis in lung cancer patients. Our findings highlight the cellular diversity of the tumor immune microenvironment in BrM of different cancer types and suggest a role of TLS formation for BrM patient outcome.

cancer biology↗

Glioma-associated tertiary lymphoid structures are sites of lymphocyte clonal expansion and plasma cell formation

Adult-type diffuse gliomas, the most common primary brain tumors, pose significant clinical challenges due to limited treatment options, restricted anti-tumor immune response and dismal patient prognosis. In this study, we elucidate the immunological function and clinical relevance of intra-tumoral tertiary lymphoid structures (TLS) in adaptive anti-glioma immunity. We conducted a comprehensive, unbiased analysis of lymphoid aggregation in 642 gliomas using a multi-modal approach that combines RNA sequencing with spatial transcriptome and proteome profiling. Our findings reveal that TLS are present in 15% of tumors and correlate with improved overall survival. Gliomas with TLS exhibit a remodeled perivascular space, marked by transcriptional upregulation and spatial redistribution of collagens associated with barrier functions. Furthermore, we demonstrate that TLS maturation into sites of dynamic adaptive immune responses, characterized by clonal T and B cell expansion and IgA+ and IgG+ plasma-cell formation, is driven by efficient early T cell recruitment to the perivascular space.

immunology↗

DANTE and DANTE_LTR: computational pipelines implementing lineage-centered annotation of LTR-retrotransposons in plant genomes

Long terminal repeat (LTR) retrotransposons constitute a predominant class of repetitive DNA elements in most plant genomes. With the increasing number of sequenced plant genomes, there is an ongoing demand for computational tools facilitating efficient annotation and classification of LTR retrotransposons in plant genome assemblies. Herein, we introduce DANTE, a computational pipeline for Domain-based ANnotation of Transposable Elements, designed for sensitive detection of these elements via their conserved protein domain sequences. The identified protein domains are subsequently inputted into the DANTE_LTR pipeline to annotate complete element sequences by detecting their structural features, such as LTRs, in adjacent genomic regions. Leveraging domain sequences allows for precise classification of elements into phylogenetic lineages, offering a more granular annotation compared with coarser conventional superfamily-based classification methods. The efficiency and accuracy of this approach were evidenced via annotation of LTR retrotransposons in 93 plant genomes. Results were benchmarked against several established pipelines, showing that DANTE_LTR is capable of identifying significantly more intact LTR retrotransposons. DANTE and DANTE_LTR are provided as user-friendly Galaxy tools accessible via a public server (https://repeatexplorer-elixir.cerit-sc.cz), installable on local Galaxy instances from the Galaxy tool shed or executable from the command line.

bioinformatics↗

Repeat-based holocentromeres of the woodrush Luzula sylvatica reveal new insights into the evolutionary transition from mono- to holocentricity

Although the centromere is restricted to a single region of the chromosome in most studied eukaryotes, members of the rush family (Juncaceae) harbor either monocentric (Juncus) or holocentric (Luzula) chromosomes. This provides an opportunity to study the evolutionary mechanisms involved in the transition to holocentricity. Here by combining chromosome-scale genome assembly, epigenetic analyses, immuno-FISH, and super-resolution microscopy, we report the occurrence of repeat-based holocentromeres in L. sylvatica. We found an irregular distribution of genes, centromeric units, and most repeats along the chromosomes. We determined the centromere function predominantly associated with two satellite DNA repeats, Lusy1 and Lusy2 of 124- and 174-bp monomer length, respectively, while CENH3 also binds satellite-free gene-poor regions. Comparative repeat analysis revealed that Lusy1 is present in most Luzula species, suggesting a conserved centromere role of this repeat. Synteny between L. sylvatica (n = 6) and J. effusus (n = 21) genomes further evidenced a chromosome number reduction in Luzula derived from multiple chromosome fusions of ancestral J. effusus-like chromosomes. We propose that the transition to holocentricity in Luzula involves: (i) fusion of small chromosomes resembling Juncus-like centromeres; (ii) expansion of atypical centromeric units; and (iii) colonization of satellite DNA for centromere stabilization.

genomics↗

KNL1 and NDC80 represent new universal markers for the detection of functional centromeres in plants

Centromere is the chromosomal site of kinetochore assembly and microtubule attachment for chromosome segregation. Given its importance, markers that allow specific labeling of centromeric chromatin throughout the cell cycle and across all chromosome types are sought for facilitating various centromere studies. Antibodies against the N-terminal region of CENH3 are commonly used for this purpose, since CENH3 is the near-universal marker of functional centromeres. However, because the N-terminal region of CENH3 is highly variable among plant species, antibodies directed against this region usually function only in a small group of closely related species. As a more versatile alternative, we present here antibodies targeted to the conserved domains of two outer kinetochore proteins, KNL1 and NDC80. Sequence comparison of these domains across more than 350 plant species revealed a high degree of conservation, particularly within a six amino acid motif, FFGPVS in KNL1, suggesting that both antibodies would function in a wide range of plant species. This assumption was confirmed by immunolabeling experiments in angiosperm (monocot and dicot) and gymnosperm species, including those with mono-, holo-, and meta-polycentric chromosomes. In addition to centromere labeling on condensed chromosomes during cell division, both antibodies detected the corresponding regions in the interphase nuclei of most species tested. These results demonstrated that KNL1 and NDC80 are better suited for immunolabeling centromeres than CENH3, because antibodies against these proteins offer incomparably greater versatility across different plant species which is particularly convenient for studying the organization and function of the centromere in non-model species.

genetics↗

Disruption of the standard kinetochore in holocentric Cuscuta species

Segregation of chromosomes depends on the centromere. Most species are monocentric, with the centromere restricted to a single region per chromosome. In some organisms, monocentric organization changed to holocentric, in which the centromere activity is distributed over the entire chromosome length. However, the causes and consequences of this transition are poorly understood. Here, we show that the transition in the genus Cuscuta was associated with dramatic changes in the kinetochore, a protein complex that mediates the attachment of chromosomes to microtubules. We found that in holocentric Cuscuta species the KNL2 genes were lost; the CENP-C, KNL1, and ZWINT1 genes were truncated; the centromeric localization of CENH3, CENP-C, KNL1, MIS12, and NDC80 proteins was disrupted; and the spindle assembly checkpoint (SAC) was degenerated. Our results demonstrate that holocentric Cuscuta species lost the ability to form a standard kinetochore and do not employ SAC to control the attachment of microtubules to chromosomes.

genetics↗

Plasticity in centromere organization: Holocentromeres can consist of merely a few megabase-sized satellite arrays

The centromere is the chromosome region where the microtubules attach during cell division. In contrast to monocentric chromosomes with one centromere location, holocentric species usually distribute hundreds of centromere units along the entire chromatid. We assembled the chromosome-scale reference genome and analyzed the holocentromere and (epi)genome organization of the lilioid Chionographis japonica. Remarkably, each of its holocentric chromatids consists of only 7 to 11 evenly-spaced megabase-sized centromere-specific histone H3-positive units. These units contain satellite arrays of 23 and 28 bp-long monomers capable of forming palindromic structures. Like monocentric species, C. japonica forms distinctly clustered centromeres in chromocenters at interphase. Additionally, the large-scale eu- and heterochromatin arrangement differs between C. japonica and other known holocentric species. Using polymer simulations, we modeled the formation of prometaphase line-like holocentromeres from interphase centromere clusters. Our findings broaden the knowledge about the diversity of centromere organization, showing that holocentricity is not restricted to species with numerous and small centromere units.

plant biology↗

Assembly of the 81.6 Mb centromere of pea chromosome 6 elucidates the structure and evolution of metapolycentric chromosomes

Centromeres in the legume genera Pisum and Lathyrus exhibit unique morphological characteristics, including extended primary constrictions and multiple separate domains of centromeric chromatin. These so-called metapolycentromeres resemble an intermediate form between monocentric and holocentric types, and therefore provide a great opportunity for studying the transitions between different types of centromere organizations. However, because of the exceedingly large and highly repetitive nature of metapolycentromeres, highly contiguous assemblies needed for these studies are lacking. Here, we report on the assembly and analysis of a 177.6 Mb region of pea (Pisum sativum) chromosome 6, including the 81.6 Mb centromere region (CEN6) and adjacent chromosome arms. Genes, DNA methylation profiles, and most of the repeats were uniformly distributed within the centromere, and their densities in CEN6 and chromosome arms were similar. The exception was an accumulation of satellite DNA in CEN6, where it formed multiple arrays up to 2 Mb in length. Centromeric chromatin, characterized by the presence of the CENH3 protein, was predominantly associated with arrays of three different satellite repeats; however, five other satellites present in CEN6 lacked CENH3. The presence of CENH3 chromatin was found to determine the spatial distribution of the respective satellites during the cell cycle. Finally, oligo-FISH painting experiments, performed using probes specifically designed to label the genomic regions corresponding to CEN6 in Pisum, Lathyrus, and Vicia species, revealed that metapolycentromeres evolved via the expansion of centromeric chromatin into neighboring chromosomal regions and the accumulation of novel satellite repeats. However, in some of these species, centromere evolution also involved chromosomal translocations and centromere repositioning. SignificanceDespite their conserved function, plant centromeres exhibit considerable variation in their morphology and sequence composition. For example, centromere activity is restricted to a single region in monocentric chromosomes, but is distributed along the entire chromosome length in holocentric chromosomes. The principles of centromere evolution that led to this variation are largely unknown, partly due to the lack of high-quality centromere assemblies. Here, we present an assembly of the pea metapolycentromere, a unique type of centromere that represents an intermediate stage between monocentric and holocentric organizations. This study not only provides a detailed insight into sequence organization, but also reveals possible mechanisms for the formation of the metapolycentromere through the spread of centromeric chromatin and the accumulation of satellite DNA.

genomics↗

The giant diploid faba genome unlocks variation in a global protein crop

Increasing the proportion of locally produced plant protein in currently meat-rich diets could substantially reduce greenhouse gas emission and loss of biodiversity. However, plant protein production is hampered by the lack of a cool-season legume equivalent to soybean in agronomic value. Faba bean (Vicia faba L.) has a high yield potential and is well-suited for cultivation in temperate regions, but genomic resources are scarce. Here, we report a high-quality chromosome-scale assembly of the faba bean genome and show that it has grown to a massive 13 Gb in size through an imbalance between the rates of amplification and elimination of retrotransposons and satellite repeats. Genes and recombination events are evenly dispersed across chromosomes and the gene space is remarkably compact considering the genome size, though with significant copy number variation driven by tandem duplication. Demonstrating practical application of the genome sequence, we develop a targeted genotyping assay and use high-resolution genome-wide association (GWA) analysis to dissect the genetic basis of hilum colour. The resources presented constitute a genomics-based breeding platform for faba bean, enabling breeders and geneticists to accelerate improvement of sustainable protein production across Mediterranean, subtropical, and northern temperate agro-ecological zones.

plant biology↗

Anatomy and transcription dynamics of wheat ribosomal RNA loci revealed by optical mapping and RNA sequencing

Background and AimsThree out of four RNA components of ribosomes are encoded by 45S rDNA loci, whose transcripts are processed into 18S, 5.8S and 26S ribosomal RNAs. The loci are organized as long head-to-tail tandem arrays of nearly identical units spanning over several megabases of sequence. Due to this peculiar structure, the number of rRNA genes, their sequence composition and expression status remain unclear, especially in complex polyploid genomes harbouring multiple loci. Here we conducted a complex study to decipher structure and activity of both major and minor rRNA loci in hexaploid bread wheat (Triticum aestivum). MethodsWe employed an original, multi-omics approach, combining chromosome flow sorting and optical mapping with transcriptome and methylome sequencing. Key ResultsThe former two techniques enabled unbiased quantification of rDNA units in particular loci of the wheat genome. Total number of rRNA genes organized in tandem arrays was 4388, with 64.1, 31.4, 3.9 and 0.7% located in short arms of chromosomes 6B, 1B, 5D and 1A, respectively. At the expression level, only 1B and 6B loci contributed to transcription at roughly 2:1 ratio. The 1B:6B ratio varied among five analysed tissues (embryo, coleoptile, root tip, primary leaf, mature leaf), being the highest (2.64:1) in mature leaf and lowest (1.72:1) in coleoptile. Cytosine methylation was considerably higher in CHG contexts in the silenced 5D locus compared to the active 1B and 6B loci. ConclusionsA fine genomic organization and tissue-specific expression of rRNA loci were deciphered, for the first time, in a complex polyploid species. We documented various mechanisms of rRNA dosage control, including gene elimination and stable inactivation related to nucleolar subdominance of A and D-genome loci, and a subtle, developmentally regulated silencing of one of the major loci. The results are discussed in the context of wheat evolution and transcription regulation.

genomics↗

Impact of parasitic lifestyle and different types of centromere organization on chromosome and genome evolution in the plant genus Cuscuta

SummaryThe parasitic genus Cuscuta (Convolvulaceae) is exceptional among plants with respect to centromere organization, including both monocentric and holocentric chromosomes, and substantial variation in genome size and chromosome number. We investigated 12 species representing the diversity of the genus in a phylogenetic context to reveal the molecular and evolutionary processes leading to diversification of their genomes.We measured genome sizes and investigated karyotypes and centromere organization using molecular cytogenetic techniques. We also performed low-pass whole genome sequencing and comparative analysis of repetitive DNA composition.A remarkable 102-fold variation in genome sizes (342–34,734 Mbp/1C) was detected for monocentric Cuscuta species, while genomes of holocentric species were of moderate sizes (533–1,545 Mbp/1C). The genome size variation was primarily driven by the differential accumulation of repetitive sequences. The transition to holocentric chromosomes in the subgenus Cuscuta was associated with loss of histone H2A phosphorylation and elimination of centromeric retrotransposons. In addition, the basic chromosome number (x) decreased from 15 to 7, presumably due to chromosome fusions.We demonstrated that the transition to holocentricity in Cuscuta was accompanied by significant changes in epigenetic marks, chromosome number and the repetitive DNA sequence composition.Competing Interest StatementThe authors have declared no competing interest.View Full Text

genomics↗

Genome-wide characterization of satellite DNA arrays in a complex plant genome using nanopore reads

BackgroundAmplification of monomer sequences into long contiguous arrays is the main feature distinguishing satellite DNA from other tandem repeats, yet it is also the main obstacle in its investigation because these arrays are in principle difficult to assemble. Here we explore an alternative, assembly-free approach that utilizes ultra-long Oxford Nanopore reads to infer the length distribution of satellite repeat arrays, their association with other repeats and the prevailing sequence periodicities.\n\nResultsWe have developed a computational workflow for similarity-based detection and downstream analysis of satellite repeats in individual nanopore reads that led to genome-wide characterization of their properties. Using the satellite DNA-rich legume plant Lathyrus sativus as a model, we demonstrated this approach by analyzing eleven major satellite repeats using a set of nanopore reads ranging from 30 to over 200 kb in length and representing 0.73x genome coverage. We found surprising differences between the analyzed repeats because only two of them were predominantly organized in long arrays typical for satellite DNA. The remaining nine satellites were found to be derived from short tandem arrays located within LTR-retrotransposons that occasionally expanded in length. While the corresponding LTR-retrotransposons were dispersed across the genome, this array expansion occurred mainly in the primary constrictions of the L. sativus chromosomes, which suggests that these genome regions are favorable for satellite DNA accumulation.\n\nConclusionsThe presented approach proved to be efficient in revealing differences in long-range organization of satellite repeats that can be used to investigate their origin and evolution in the genome.

genomics↗

Nondisjunction and unequal spindle organization accompany the drive of Aegilops speltoides B chromosomes

Supernumerary B chromosomes (Bs), which are often preferentially inherited, deviating from usual Mendelian segregation. This chromosome drive is one of the most important features of Bs. Here we analyzed the drive mechanism of Aegilops speltoides Bs and provide direct insight into its cellular mechanism. Comparative genomics resulted in the identification of the tandem repeat AesTR-183 of Ae. speltoides Bs, which also can be found on the Bs of Ae. mutica and rye, was used to track Bs during microgametogenesis. Nondisjunction of CENH3-positive, tubulin interacting B sister chromatids and an asymmetric spindle during first pollen grain mitosis are likely components of the accumulation process. A quantitative flow cytometric approach revealed, that independent on the number of Bs present in the mother plant Bs accumulate in the generative nuclei with more than 93%. Nine of eleven tested (peri)centromeric repeats were shared by A and B chromosomes. A common origin of the drive process in Poaceae is likely.

genetics↗