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Spiders produce nature's toughest fiber using renewable components at ambient temperatures and with water as solvent, making it highly interesting to replicate for the materials industry. Despite this, much remains to be understood about the bioprocessing and composition of spider silk fibers. Here, we identify 18 proteins that make up the spiders' strongest silk type, the major ampullate fiber. Single-cell RNA sequencing and spatial transcriptomics revealed that the secretory epithelium of the gland harbors six cell types. These cell types are confined to three distinct glandular zones that produce specific combinations of silk proteins. Image analysis of histological sections showed that the secretions from the three zones do not mix, and proteomics analysis revealed that these secretions form layers in the final fiber. Using a multi-omics approach, we provide substantial advancements in the understanding of the structure and function of the major ampullate silk gland as well as of the architecture and composition of the fiber it produces.

The European sprat is a small plankton-feeding clupeid present in the northeastern Atlantic Ocean, the Mediterranean Sea as well as in the brackish Baltic Sea and Black Sea. This species is the target of a major fishery and therefore an accurate characterization of its genetic population structure is crucial to delineate proper stock assessments that aid ensuring the fishery's sustainability. Here we present (i) a draft genome assembly, (ii) pooled whole genome sequencing of 19 population samples covering most of the species' distribution range, and (iii) the design and test of a SNP-chip resource and use this to validate the population structure inferred from pooled sequencing. These approaches revealed, using the populations sampled here, three major groups of European sprat: Oceanic, Coastal, and Brackish with limited differentiation within groups even over wide geographical stretches. Genetic structure is largely driven by six large putative inversions that differentiate Oceanic and Brackish sprats, while Coastal populations display intermediate frequencies of haplotypes at each locus. Interestingly, populations from the Baltic and the Black Seas share similar frequencies of haplotypes at these putative inversions despite their distant geographic location. The closely related clupeids European sprat and Atlantic herring both show genetic adaptation to the brackish Baltic Sea, providing an opportunity to explore the extent of genetic parallelism. This analysis revealed limited parallelism because out of 125 independent loci detected in the Atlantic herring, three showed sharp signals of selection that overlapped between the two species and contained single genes such as PRLRA, which encodes the receptor for prolactin, a freshwater-adapting hormone in euryhaline species, and THRB, a receptor for thyroid hormones, important both for metabolic regulation and the development of red cone photoreceptors.

Distyly is an iconic floral polymorphism governed by a supergene, which promotes efficient pollen transfer and outcrossing through reciprocal differences in the position of sexual organs in flowers, often coupled with heteromorphic self-incompatibility. Distyly has evolved convergently in multiple flowering plant lineages, but has also broken down repeatedly, often resulting in homostylous, self-compatible populations with elevated rates of self-fertilization. Here, we aimed to study the genetic causes and genomic consequences of the shift to homostyly in Linum trigynum, which is closely related to distylous Linum tenue. Building on a high-quality genome assembly, we show that L. trigynum harbors a genomic region homologous to the dominant haplotype of the distyly supergene conferring long stamens and short styles in L. tenue, suggesting that loss of distyly first occurred in a short-styled individual. In contrast to homostylous Primula and Fagopyrum, L. trigynum harbors no fixed loss-of-function mutations in coding sequences of S-linked distyly candidate genes. Instead, floral gene expression analyses and controlled crosses suggest that mutations downregulating the S-linked LtWDR-44 candidate gene for male self-incompatibility and/or anther height could underlie homostyly and self-compatibility in L. trigynum. Population genomic analyses of 224 whole-genome sequences further demonstrate that L. trigynum is highly self-fertilizing, exhibits significantly lower genetic diversity genome-wide, and is experiencing relaxed purifying selection and less frequent positive selection on nonsynonymous mutations relative to L. tenue. Our analyses shed light on the loss of distyly in L. trigynum, and advance our understanding of a common evolutionary transition in flowering plants.

The B-cell acute lymphoblastic leukemia (ALL) cell line REH, with the t(12;21) ETV6::RUNX1 translocation, is known to have a complex karyotype defined by a series of large-scale chromosomal rearrangements. Taken from a 15-yr-old at relapse, the cell line offers a practical model for the study of pediatric B-ALL. In recent years, short- and long-read DNA and RNA sequencing have emerged as a complement to karyotyping techniques in the resolution of structural variants in an oncological context. Here, we explore the integration of long-read PacBio and Oxford Nanopore whole-genome sequencing, IsoSeq RNA sequencing, and short-read Illumina sequencing to create a detailed genomic and transcriptomic characterization of the REH cell line. Whole-genome sequencing clarified the molecular traits of disrupted ALL-associated genes including CDKN2A, PAX5, BTG1, VPREB1, and TBL1XR1, as well as the glucocorticoid receptor NR3C1. Meanwhile, transcriptome sequencing identified seven fusion genes within the genomic breakpoints. Together, our extensive whole-genome investigation makes high-quality open-source data available to the leukemia genomics community.

The European sprat is a small plankton-feeding clupeid present in the northeastern Atlantic Ocean, in the Mediterranean Sea, and in the brackish Baltic Sea and Black Sea. This species is the target of a major fishery and, therefore, an accurate characterization of its genetic population structure is crucial to delineate proper stock assessments that aid ensuring the fishery's sustainability. Here, we present (i) a draft genome assembly, (ii) pooled whole genome sequencing of 19 population samples covering most of the species' distribution range, and (iii) the design and test of a single nucleotide polymorphism (SNP)-chip resource and use this to validate the population structure inferred from pooled sequencing. These approaches revealed, using the populations sampled here, three major groups of European sprat: Oceanic, Coastal, and Brackish with limited differentiation within groups even over wide geographical stretches. Genetic structure is largely driven by six large putative inversions that differentiate Oceanic and Brackish sprats, while Coastal populations display intermediate frequencies of haplotypes at each locus. Interestingly, populations from the Baltic and the Black Seas share similar frequencies of haplotypes at these putative inversions despite their distant geographic location. The closely related clupeids European sprat and Atlantic herring both show genetic adaptation to the brackish Baltic Sea, providing an opportunity to explore the extent of genetic parallelism. This analysis revealed limited parallelism because out of 125 independent loci detected in the Atlantic herring, three showed sharp signals of selection that overlapped between the two species and contained single genes such as PRLRA, which encodes the receptor for prolactin, a freshwater-adapting hormone in euryhaline species, and THRB, a receptor for thyroid hormones, important both for metabolic regulation and the development of red cone photoreceptors.

The clouded apollo (Parnassius mnemosyne) is a palearctic butterfly distributed over a large part of western Eurasia, but population declines and fragmentation have been observed in many parts of the range. The development of genomic tools can help to shed light on the genetic consequences of the decline and to make informed decisions about direct conservation actions. Here, we present a high-contiguity, chromosome-level genome assembly of a female clouded apollo butterfly and provide detailed annotations of genes and transposable elements. We find that the large genome (1.5 Gb) of the clouded apollo is extraordinarily repeat rich (73%). Despite that, the combination of sequencing techniques allowed us to assemble all chromosomes (nc = 29) to a high degree of completeness. The annotation resulted in a relatively high number of protein-coding genes (22,854) compared with other Lepidoptera, of which a large proportion (21,635) could be assigned functions based on homology with other species. A comparative analysis indicates that overall genome structure has been largely conserved, both within the genus and compared with the ancestral lepidopteran karyotype. The high-quality genome assembly and detailed annotation presented here will constitute an important tool for forthcoming efforts aimed at understanding the genetic consequences of fragmentation and decline, as well as for assessments of genetic diversity, population structure, inbreeding, and genetic load in the clouded apollo butterfly.

The joint expression of particular colors, morphologies, and behaviors is a common feature of adaptation, but the genetic basis for such "phenotypic syndromes" remains poorly understood. Here, we identified a complex genetic architecture associated with a sexually selected syndrome in common wall lizards, by capitalizing on the adaptive introgression of coloration and morphology into a distantly related lineage. Consistent with the hypothesis that the evolution of phenotypic syndromes in vertebrates is facilitated by developmental linkage through neural crest cells, most of the genes associated with the syndrome are involved in neural crest cell regulation. A major locus was a similar to 400-kb region, characterized by standing structural genetic variation and previously implied in the evolutionary innovation of coloration and beak size in birds. We conclude that features of the developmental and genetic architecture contribute to maintaining trait integration, facilitating the extensive and rapid introgressive spread of suites of sexually selected characters.

Background Malignant gliomas, the most common malignant brain tumors in adults, represent a heterogeneous group of diseases with poor prognosis. Retroviruses can cause permanent genetic alterations that modify genes close to the viral integration site. Methods Here we describe the use of a high-throughput pipeline coupled to the commonly used tissue-specific retroviral RCAS-TVA mouse tumor model system. Utilizing next-generation sequencing, we show that retroviral integration sites can be reproducibly detected in malignant stem cell lines generated from RCAS-PDGFB-driven glioma biopsies. Results A large fraction of common integration sites contained genes that have been dysregulated or misexpressed in glioma. Others overlapped with loci identified in previous glioma-related forward genetic screens, but several novel putative cancer-causing genes were also found. Integrating retroviral tagging and clinical data, Ppfibp1 was highlighted as a frequently tagged novel glioma-causing gene. Retroviral integrations into the locus resulted in Ppfibp1 upregulation, and Ppfibp1-tagged cells generated tumors with shorter latency on orthotopic transplantation. In human gliomas, increased PPFIBP1 expression was significantly linked to poor prognosis and PDGF treatment resistance. Conclusions Altogether, the current study has demonstrated a novel approach to tagging glioma genes via forward genetics, validating previous results, and identifying PPFIBP1 as a putative oncogene in gliomagenesis.

The major histocompatibility complex (MHC) is of central importance to the immune system, and an optimal MHC diversity is believed to maximize pathogen elimination. Birds show substantial variation in MHC diversity, ranging from few genes in most bird orders to very many genes in passerines. Our understanding of the evolutionary trajectories of the MHC in passerines is hampered by lack of data on genomic organization. Therefore, we assembled and annotated the MHC genomic region of the great reed warbler (Acrocephalus arundinaceus), using long-read sequencing and optical mapping. The MHC region is large (>5.5 Mb), characterized by structural changes compared to hitherto investigated bird orders and shows higher repeat content than the genome average. These features were supported by analyses in three additional passerines. MHC genes in passerines are found in two different chromosomal arrangements, either as single copy MHC genes located among non-MHC genes, or as tandemly duplicated tightly linked MHC genes. Some single copy MHC genes are old and putative orthologues among species. In contrast tandemly duplicated MHC genes are monophyletic within species and have evolved by simultaneous gene duplication of several MHC genes. Structural differences in the MHC genomic region among bird orders seem substantial compared to mammals and have possibly been fuelled by clade-specific immune system adaptations. Our study provides methodological guidance in characterizing complex genomic regions, constitutes a resource for MHC research in birds, and calls for a revision of the general belief that avian MHC has a conserved gene order and small size compared to mammals.

The complete mitochondrial genome of the Carniolan honeybee (Apis mellifera carnica) from Slovenia, a homeland of this subspecies, was acquired in two contigs from WGS data and annotated. The newly obtained mitochondrial genome is a circular closed loop of 16,447 bp. It comprises 37 genes (13 protein coding genes, 22 tRNA genes, and 2 rRNA genes) and an AT-rich control region. The order of the tRNA genes resembles the order characteristic of A. mellifera. The mitogenomic sequence of A. m. carnica from Slovenia contains 44 uniquely coded sites in comparison to the closely related subspecies A. m. ligustica and to A. m. carnica from Austria. Furthermore, 24 differences were recognised in comparison between A. m. carnica and A. m. ligustica subspecies. Among them, there are three SNPs that affect translation in the nd2, nd4, and cox2 genes, respectively. The phylogenetic placement of A. m. carnica from Slovenia within C lineage deviates from the expected position and changes the perspective on relationship between C and O lineages. The results of this study represent a valuable addition to the information available in the phylogenomic studies of A. mellifera-a pollinator species of worldwide importance. Such genomic information is essential for this local subspecies' conservation and preservation as well as its breeding and selection.

Over the last six decades, populations of the bumblebees Bombus sylvicola and Bombus balteatus in Colorado have experienced decreases in tongue length, a trait important for plant-pollinator mutualisms. It has been hypothesized that this observation reflects selection resulting from shifts in floral composition under climate change. Here we used morphometrics and population genomics to determine whether morphological change is ongoing, investigate the genetic basis of morphological variation, and analyse population structure in these populations. We generated a genome assembly of B. balteatus. We then analysed whole-genome sequencing data and morphometric measurements of 580 samples of both species from seven high-altitude localities. Out of 281 samples originally identified as B. sylvicola, 67 formed a separate genetic cluster comprising a newly-discovered cryptic species ("incognitus"). However, an absence of genetic structure within species suggests that gene flow is common between mountains. We found a significant decrease in tongue length between bees collected between 2012-2014 and in 2017, indicating that morphological shifts are ongoing. We did not discover any genetic associations with tongue length, but a SNP related to production of a proteolytic digestive enzyme was implicated in body size variation. We identified evidence of covariance between kinship and both tongue length and body size, which is suggestive of a genetic component of these traits, although it is possible that shared environmental effects between colonies are responsible. Our results provide evidence for ongoing modification of a morphological trait important for pollination and indicate that this trait probably has a complex genetic and environmental basis.

Supergenes govern multi-trait-balanced polymorphisms in a wide range of systems; however, our understanding of their origins and evolution remains incomplete. The reciprocal placement of stigmas and anthers in pin and thrum floral morphs of distylous species constitutes an iconic example of a balanced polymorphism governed by a supergene, the distyly S-locus. Recent studies have shown that the Primula and Turnera distyly supergenes are both hemizygous in thrums, but it remains unknown whether hemizygosity is pervasive among distyly S-loci. As hemizygosity has major consequences for supergene evolution and loss, clarifying whether this genetic architecture is shared among distylous species is critical. Here, we have characterized the genetic architecture and evolution of the distyly supergene in Linum by generating a chromosome-level genome assembly of Linum tenue, followed by the identification of the S-locus using population genomic data. We show that hemizygosity and thrum-specific expression of S-linked genes, including a pistil-expressed candidate gene for style length, are major features of the Linum S-locus. Structural variation is likely instrumental for recombination suppression, and although the non-recombining dominant haplotype has accumulated transposable elements, S-linked genes are not under relaxed purifying selection. Our findings reveal remarkable convergence in the genetic architecture and evolution of independently derived distyly supergenes, provide a counterexample to classic inversion-based supergenes, and shed new light on the origin and maintenance of an iconic floral polymorphism.

Genome assemblies are currently being produced at an impressive rate by consortia and individual laboratories. The low costs and increasing efficiency of sequencing technologies now enable assembling genomes at unprecedented quality and contiguity. However, the difficulty in assembling repeat-rich and GC-rich regions (genomic "dark matter") limits insights into the evolution of genome structure and regulatory networks. Here, we compare the efficiency of currently available sequencing technologies (short/linked/long reads and proximity ligation maps) and combinations thereof in assembling genomic dark matter. By adopting different de novo assembly strategies, we compare individual draft assemblies to a curated multiplatform reference assembly and identify the genomic features that cause gaps within each assembly. We show that a multiplatform assembly implementing long-read, linked-read and proximity sequencing technologies performs best at recovering transposable elements, multicopy MHC genes, GC-rich microchromosomes and the repeat-rich W chromosome. Telomere-to-telomere assemblies are not a reality yet for most organisms, but by leveraging technology choice it is now possible to minimize genome assembly gaps for downstream analysis. We provide a roadmap to tailor sequencing projects for optimized completeness of both the coding and noncoding parts of nonmodel genomes.