Genotyping-by-sequencing resolves relationships in Polygonaceae tribe Eriogoneae

The resolution of cryptic diversity is essential for understanding the evolutionary diversification of lineages and establishing conservation priorities. We examine relationships in Eriogoneae (Polygonaceae), a diverse lineage in western North America. We ask whether Eriogonum umbellatum, a morphologically and ecologically diverse species, is monophyletic and whether its varieties represent evolutionary lineages. We use genotyping-by-sequencing to assemble a SNP dataset for 51 species in the genera Chorizanthe, Eriogonum and Sidotheca. We report a hierarchical phylogenetic analysis using maximum likelihood to estimate the evolutionary history of Eriogoneae. We illustrate admixture components for 21 populations of E. umbellatum, representing four varieties, and test for lineage structure using TreeMix. We identify strongly supported clades within Eriogoneae. Many relationships in the Eucycla + Oregonium and Latifolia clades are supported, while most relationships within the Eriogonum subg. Oligogonum clade and a clade with most Chorizanthe remain unresolved. Eriogonum congdonii resolves within the main E. umbellatum clade, while populations of three varieties of E. umbellatum are closely related to E. ursinum and are associated with serpentine soils. ADmixture and TreeMix analyses suggest E. umbellatum varieties represent evolutionary lineages. These results from SNP data are largely consistent with previous phylogenetic studies of Eriogoneae based on sequence variation. Structure within Oligogonum suggests consistent environmental association and radiation after initial colonization of serpentine. Morphology is unreliable for the infraspecific taxonomy of E. umbellatum. Additional molecular studies are needed to resolve the evolutionary relationships and ecological diversification within this species, in Oligogonum, and in Eriogoneae. © 2021 The Authors. TAXON published by John Wiley & Sons Ltd on behalf of International Association for Plant Taxonomy.We thank SGIker research support services at the University of the Basque Country, Leioa, Spain for DNA extraction and quality control, and acknowledge Centro Nacional de d-An?lisi Gen?mica in Barcelona, Spain for GBS sequencing. We thank J. Andre, N.J. Jensen and J. Steele for contributing samples and D. Marino for collaboration in nucleic acid extractions. We acknowledge the contributions of O. Lao Grueso to the?ADmixture and TreeMix analyses. We thank U.S. Forest Service employees D. Austin, J. Fedorchuk, M. Friend, J. Haas, D. Ikeda, L. Janeway, J. Nelson, D. Netz, A. Sanger, and S. Weis among others for facilitating permitting and collecting. This work was supported by funds from the Basque Government in support of the Terrestrial Plant Diversity group of the Department of Plant Biology and Ecology, University of the Basque Country, and an ERC Advanced Grant, FP7-IDEAS-ERC, ?ADAPT?, project 339941 awarded to T. Brown. We thank SGIker research support services at the University of the Basque Country, Leioa, Spain for DNA extraction and quality control, and acknowledge Centro Nacional de d‐Anàlisi Genòmica in Barcelona, Spain for GBS sequencing. We thank J. Andre, N.J. Jensen and J. Steele for contributing samples and D. Marino for collaboration in nucleic acid extractions. We acknowledge the contributions of O. Lao Grueso to the ADmixture and TreeMix analyses. We thank U.S. Forest Service employees D. Austin, J. Fedorchuk, M. Friend, J. Haas, D. Ikeda, L. Janeway, J. Nelson, D. Netz, A. Sanger, and S. Weis among others for facilitating permitting and collecting. This work was supported by funds from the Basque Government in support of the Terrestrial Plant Diversity group of the Department of Plant Biology and Ecology, University of the Basque Country, and an ERC Advanced Grant, FP7‐IDEAS‐ERC, ‘ADAPT’, project 339941 awarded to T. Brown

The origins, evolution, and functional potential of alternative splicing in vertebrates.

Alternative splicing (AS) has the potential to greatly expand the functional repertoire of mammalian transcriptomes. However, few variant transcripts have been characterized functionally, making it difficult to assess the contribution of AS to the generation of phenotypic complexity and to study the evolution of splicing patterns. We have compared the AS of 309 protein-coding genes in the human ENCODE pilot regions against their mouse orthologs in unprecedented detail, utilizing traditional transcriptomic and RNAseq data. The conservation status of every transcript has been investigated, and each functionally categorized as coding (separated into coding sequence [CDS] or nonsense-mediated decay [NMD] linked) or noncoding. In total, 36.7% of human and 19.3% of mouse coding transcripts are species specific, and we observe a 3.6 times excess of human NMD transcripts compared with mouse; in contrast to previous studies, the majority of species-specific AS is unlinked to transposable elements. We observe one conserved CDS variant and one conserved NMD variant per 2.3 and 11.4 genes, respectively. Subsequently, we identify and characterize equivalent AS patterns for 22.9% of these CDS or NMD-linked events in nonmammalian vertebrate genomes, and our data indicate that functional NMD-linked AS is more widespread and ancient than previously thought. Furthermore, although we observe an association between conserved AS and elevated sequence conservation, as previously reported, we emphasize that 30% of conserved AS exons display sequence conservation below the average score for constitutive exons. In conclusion, we demonstrate the value of detailed comparative annotation in generating a comprehensive set of AS transcripts, increasing our understanding of AS evolution in vertebrates. Our data supports a model whereby the acquisition of functional AS has occurred throughout vertebrate evolution and is considered alongside amino acid change as a key mechanism in gene evolution

Close similarities between Cherry chlorotic rusty spot disease from Italy and Cherry leaf scorch from Spain

Cherry chlorotic rusty spot (CCRS), a disease affecting sweet and sour cherry in Southern Italy was regularly found associated with an unidentified fungus and with a complex pattern of viral-like double-stranded RNAs as well as with two small circular RNAs (cherry small circular RNAs, cscRNAs). Further studies revealed that i) the ds-RNAs correspond to the genome of different mycoviruses belonging to the genera Chrysovirus, Partitivirus and Totivirus and ii) the two viroid-like RNAs consist of two groups of variants with similar sequences but differing in size (394–415 and 372–377 nt for cscRNA1 and cscRNA2, respectively). Here we report that the dsRNAs of Chrysovirus and Partitivirus have been detected by RT-PCR analysis with CCRS specific primers in nucleic acid preparations from cherry leaves affected by cherry leaf scorch (CLS) in Spain, a disease whose etiological agent is the ascomycetes Apiognomonia erythrostoma, order Diaporthales. Moreover, Northern-blot hybridization assays showed that a viroid-like RNA comigrating and sharing high sequence similarity with the cscRNA1 previously reported in Italy, accumulate in leaves from CLS affected trees in Spain. These data, together with other evidence showing similar symptoms, disease cycle and fungal fructifications in CCRS and CLS affected trees, suggest a close relationship between the two cherry disorders.Keywords: dsRNAs, cscRNAs, Apiognomonia erythrostoma, Diaporthale

European sea bass brain DLB-1 cell line is susceptible to nodavirus: A transcriptomic study

Viral diseases are responsible for high rates of mortality and subsequent economic losses in modern aquaculture. The nervous necrosis virus (NNV) produces viral encephalopathy and retinopathy (VER), which affects the fish central nervous system. It is considered one of the most serious viral diseases in marine aquaculture, the European sea bass (Dicentrarchus labrax) being amongst the most susceptible. We have evaluated the European sea bass brain derived cell line (DLB-1) susceptibility to NNV genotypes and evaluated its transcriptomic profile. DLB-1 cells supported NNV gene transcription and replication since strains belonging to the four NNV genotypes produce cytopathic effects. Afterwards, DLB-1 cells were infected with an RGNNV strain, the one which showed the highest replication, for 12 and 72 h and an RNA-seq analysis was performed to identify potential genes involved in the host-NNV interactions. Differential expression analysis showed the up-regulation of many genes related to immunity, heat-shock proteins or apoptosis but not to proteasome or autophagy processes. These data suggest that the immune response, mainly the interferon (IFN) pathway, is not powerful enough to abrogate the infection, and cells finally suffer stress and die by apoptosis liberating infective particles. GO enrichment also revealed, for the first time, the down-regulation of terms related to brain/neuron biology indicating molecular mechanisms causing the pathogenic effect of NNV. This study opens the way to understand key elements in sea bass brain and NNV interactions.Versión del edito

Rapid production of pure recombinant actin isoforms in Pichia pastoris

Actins are major eukaryotic cytoskeletal proteins, which perform many important cell functions, including cell division, cell polarity, wound healing, and muscle contraction. Despite obvious drawbacks, muscle actin, which is easily purified, is used extensively presently for biochemical studies of actin cytoskeleton from other organisms / cell types. Here we report a rapid and cost-effective method to purify heterologous actins expressed in the yeast Pichia pastoris. Actin is expressed as a fusion with the actin-binding protein thymosin β4 and purified using an affinity tag introduced in the fusion. Following cleavage of thymosin β4 and the affinity tag, highly purified functional full-length actin is liberated. We purify actins from S. cerevisiae, S. pombe, and the β- and γ- isoforms of human actin. We also report a modification of the method that facilitates expression and purification of arginylated actin, a form of actin thought to regulate actin dendritic networks in mammalian cells. The methods we describe can be performed in all laboratories equipped for molecular biology, and should greatly facilitate biochemical and cell biological studies of the actin cytoskeleton

The Origins, Evolution, and Functional Potential of Alternative Splicing in Vertebrates

Alternative splicing (AS) has the potential to greatly expand the functional repertoire of mammalian transcriptomes. However, few variant transcripts have been characterized functionally, making it difficult to assess the contribution of AS to the generation of phenotypic complexity and to study the evolution of splicing patterns. We have compared the AS of 309 protein-coding genes in the human ENCODE pilot regions against their mouse orthologs in unprecedented detail, utilizing traditional transcriptomic and RNAseq data. The conservation status of every transcript has been investigated, and each functionally categorized as coding (separated into coding sequence [CDS] or nonsense-mediated decay [NMD] linked) or noncoding. In total, 36.7% of human and 19.3% of mouse coding transcripts are species specific, and we observe a 3.6 times excess of human NMD transcripts compared with mouse; in contrast to previous studies, the majority of species-specific AS is unlinked to transposable elements. We observe one conserved CDS variant and one conserved NMD variant per 2.3 and 11.4 genes, respectively. Subsequently, we identify and characterize equivalent AS patterns for 22.9% of these CDS or NMD-linked events in nonmammalian vertebrate genomes, and our data indicate that functional NMD-linked AS is more widespread and ancient than previously thought. Furthermore, although we observe an association between conserved AS and elevated sequence conservation, as previously reported, we emphasize that 30% of conserved AS exons display sequence conservation below the average score for constitutive exons. In conclusion, we demonstrate the value of detailed comparative annotation in generating a comprehensive set of AS transcripts, increasing our understanding of AS evolution in vertebrates. Our data supports a model whereby the acquisition of functional AS has occurred throughout vertebrate evolution and is considered alongside amino acid change as a key mechanism in gene evolution

A chromosome-level genome assembly enables the identification of the follicule stimulating hormone receptor as the master sex determining gene in the flatfish Solea senegalensis

Sex determination (SD) shows huge variation among fish and a high evolutionary rate, as illustrated by the Pleuronectiformes (flatfishes). This order is characterized by its adaptation to demersal life, compact genomes and diversity of SD mechanisms. Here, we assembled the Solea senegalensis genome, a flatfish of great commercial value, into 82 contigs (614 Mb) combining long- and short-read sequencing, which were next scaffolded using a highly dense genetic map (28,838 markers, 21 linkage groups), representing 98.9% of the assembly. Further, we established the correspondence between the assembly and the 21 chromosomes by using BAC-FISH. Whole genome resequencing of six males and six females enabled the identification of 41 SNP variants in the follicle stimulating hormone receptor (fshr) consistent with an XX / XY SD system. The observed sex association was validated in a broader independent sample, providing a novel molecular sexing tool. Fshr displayed differential gene expression between male and female gonads from 86 days post-fertilization, when the gonad is still an undifferentiated primordium, concomitant with the activation of amh and cyp19a1a, testis and ovary marker genes, respectively, in males and females. The Y-linked fshr allele, which included 24 non-synonymous variants and showed a highly divergent 3D protein structure, was overexpressed in males compared to the X-linked allele at all stages of gonadal differentiation. We hypothesize a mechanism hampering the action of the follicle stimulating hormone driving the undifferentiated gonad toward testis.info:eu-repo/semantics/acceptedVersio