44 I. 44 II. 45 III. 46 IV. 49 V. 57 VI. 58 58 References 58 SUMMARY: Conifers have been understudied at the genomic level despite their worldwide ecological and economic importance but the situation ...is rapidly changing with the development of next generation sequencing (NGS) technologies. With NGS, genomics research has simultaneously gained in speed, magnitude and scope. In just a few years, genomes of 20–24 gigabases have been sequenced for several conifers, with several others expected in the near future. Biological insights have resulted from recent sequencing initiatives as well as genetic mapping, gene expression profiling and gene discovery research over nearly two decades. We review the knowledge arising from conifer genomics research emphasizing genome evolution and the genomic basis of adaptation, and outline emerging questions and knowledge gaps. We discuss future directions in three areas with potential inputs from NGS technologies: the evolutionary impacts of adaptation in conifers based on the adaptation‐by‐speciation model; the contributions of genetic variability of gene expression in adaptation; and the development of a broader understanding of genetic diversity and its impacts on genome function. These research directions promise to sustain research aimed at addressing the emerging challenges of adaptation that face conifer trees.
Genetic and genomic studies are seeing an increase in sample sizes together with a wider range of species investigated in response to environmental change concerns. In turn, these changes may come ...with challenges including the time and difficulty to isolate nucleic acids (DNA or RNA), the sequencing cost and environmental impacts of the growing amount of plastic waste generated in the process. Pseudotsuga menziesii var. menziesii (Mirbel) Franco (PM), Tsuga heterophylla (Raf.) Sarg. (TH) and Thuja plicata Donn ex D.Don (TP) are conifer species found in diverse woodlands both as natives and naturalized exotics. Our study was carried out whilst investigating their genetics to understand their population structure and potential for adaptation. In the present study, we compared two different DNA isolation methods, i.e., spin-column DNeasy plant mini kit (QIAGEN), and temperature-driven enzymatic cocktail Plant DNA Extraction (MicroGEM). The quantity of recovered DNA and the quality of DNA were assessed along with the plastic footprint and time needed for three tree species. Both methods were optimised and proven to provide enough DNA for each studied species. The yield of DNA for each method depended on the species: QIAGEN showed higher yield in P. menziesii and T. heterophylla, while T. plicata recovered similar amount of DNA for both methods. The DNA quality was investigated using DNA barcoding techniques by confirming species identity and species discrimination. No difference was detected in the PCR amplification of the two barcoding loci, (rbcL and trnH-psbA), and the recovered sequences between DNA isolation methods. Measurement of the plastic use and the processing time per sample indicated that MicroGEM had a 52.64% lower plastic footprint and was 51.8% faster than QIAGEN. QIAGEN gave higher yields in two of the species although both methods showed similar quality results across all species. However, MicroGEM was clearly advantageous to decrease the plastic footprint and improve the time efficiency. Overall, MicroGEM recovers sufficient and reliable DNA to perform common downstream analyses such as PCR and sequencing. Our findings illustrate the benefits of research and efforts towards developing more sustainable methods and techniques to reduce the environmental footprint of molecular analyses.
The functional plasticity of T cell subsets Bluestone, Jeffrey A; Mackay, Charles R; O'Shea, John J ...
Nature reviews. Immunology,
11/2009, Letnik:
9, Številka:
11
Journal Article
Recenzirano
Odprti dostop
In 1986, Robert Coffman and Timothy Mossman first described the division of CD4(+) T cells into functional subsets, termed T helper 1 (T(H)1) and T(H)2, based on cytokine production, and in doing so ...unwittingly opened a Pandora's box of complexity and controversy. Although the mechanisms that regulate T(H)1 and T(H)2 cells are now well known, recent descriptions of other CD4(+) T cell subsets--such as regulatory T cells, T follicular helper cells, T(H)17, T(H)22 and most recently T(H)9 and T(H)22 cells--have questioned how we think of T cell subsets and what commitment to a functional T cell subset means. Here, Nature Reviews Immunology asks four leaders in the field their thoughts on the functional plasticity of T cell subsets.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Phytophthora ramorum severely affects both European larch (EL) and Japanese larch (JL) trees as indicated by high levels of mortality particularly in the UK. Field observations suggested that EL is ...less severely affected and so may be less susceptible to P. ramorum than JL; however, controlled inoculations have produced inconsistent or non-statistically significant differences. The present study aimed to compare RNA transcript accumulation profiles in EL and JL in response to inoculation with P. ramorum to improve our understanding of their defence responses.
RNA-sequencing was carried out on bark tissues following the inoculation with P. ramorum of potted saplings in both EL and JL carried out under controlled environment conditions, with sampling at 1, 3, 10, and 25 days post inoculation in infected and control plants.
All of the inoculated trees rapidly developed lesions but no statistically significant differences were found in lesion lengths between EL and JL. RNA-Sequencing comparing control and inoculate saplings identified key differences in differentially expressed genes (DEGs) between the two larch species. European larch had rapid induction of defence genes within 24 hours of infection followed by sustained expression until 25 days after inoculation. Results in JL were more varied; upregulation was stronger but more transient and represented fewer defence pathways. Gene enrichment analyses highlighted differences in jasmonate signalling and regulation including NPR1 upregulation in EL only, and specific aspects of secondary metabolism. Some DEGs were represented by multiple responsive copies including lipoxygenase, chalcone synthase and nucleotide-binding, leucine-rich-repeat genes.
The variations between EL and JL in responsive DEGs of interest as potentially related to differences seen in the field and should be considered in the selection of trees for planting and future breeding.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Phytophthora ramorum is an invasive oomycete pathogen that has been causing significant mortality of larch trees (Larix spp.) in the United Kingdom since 2009. This is the first multiyear study of ...the natural infection processes of P. ramorum on Japanese larch (Larix kaempferi) and the factors influencing disease progression. Field surveys in south‐west Scotland, which suffered an extensive epidemic in 2012 from a highly virulent new lineage (EU2), allowed detailed examination of how P. ramorum infects individual trees and spreads across a site over an extended time period. A marked expansion of the disease in spring 2018 allowed us to consider how environmental conditions influence outbreaks, with summer precipitation and spring storms found to be significant factors. Field observations revealed that buds on fine shoots appeared to be the primary infection sites with infection occurring between late autumn and spring, and rapid lesion extension during flushing. Infection and spread of P. ramorum were not consistent within or between sites and it is likely that site‐specific factors influence the spread of infection and subsequent mortality. The climatic conditions identified in this study could help predict future disease expansions and inform the development of management strategies for larch in the UK.
Summer precipitation and spring storms are significant factors influencing the progression of Phytophthora ramorum on Japanese larch; new infections occur between late autumn and spring with rapid lesion extension during flushing.
The copper‐catalyzed azide–alkyne cycloaddition (CuAAC) reaction has proven to be a pivotal advance in chemical ligation strategies with applications ranging from polymer fabrication to ...bioconjugation. However, application in vivo has been limited by the inherent toxicity of the copper catalyst. Herein, we report the application of heterogeneous copper catalysts in azide–alkyne cycloaddition processes in biological systems ranging from cells to zebrafish, with reactions spanning from fluorophore activation to the first reported in situ generation of a triazole‐containing anticancer agent from two benign components, opening up many new avenues of exploration for CuAAC chemistry.
Localized drug synthesis: Biocompatible copper nanoparticle catalysts were synthesized and employed in the activation of a profluorophore in living cells and in zebrafish embryos. Furthermore, an anticancer drug was synthesized in situ from two benign components, leading to apoptosis in ovarian cancer cells.
Conifers are long-lived and slow-evolving, thus requiring effective defences against their fast-evolving insect natural enemies. The copy number variation (CNV) of two key acetophenone biosynthesis ...genes Ugt5/Ugt5b and βglu-1 may provide a plausible mechanism underlying the constitutively variable defence in white spruce (Picea glauca) against its primary defoliator, spruce budworm. This study develops a long-insert sequence capture probe set (Picea_hung_p1.0) for quantifying copy number of βglu-1-like, Ugt5-like genes and single-copy genes on 38 Norway spruce (Picea abies) and 40 P. glauca individuals from eight and nine provenances across Europe and North America respectively. We developed local assemblies (Piabi_c1.0 and Pigla_c.1.0), full-length transcriptomes (PIAB_v1 and PIGL_v1), and gene models to characterise the diversity of βglu-1 and Ugt5 genes. We observed very large copy numbers of βglu-1, with up to 381 copies in a single P. glauca individual. We observed among-provenance CNV of βglu-1 in P. glauca but not P. abies. Ugt5b was predominantly single-copy in both species. This study generates critical hypotheses for testing the emergence and mechanism of extreme CNV, the dosage effect on phenotype, and the varying copy number of genes with the same pathway. We demonstrate new approaches to overcome experimental challenges in genomic research in conifer defences.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Outbreaks of spruce budworm (SBW, Choristoneura fumiferana Clem.) cause major recurrent damage in boreal conifers such as white spruce (Picea glauca Moench Voss) and large losses of forest biomass in ...North America. Although defensive phenolic compounds have recently been linked to chemical resistance against SBW, their genetic basis remains poorly understood in forest trees, especially in conifers. Here, we used diverse association genetics approaches to discover genes and their variants that may control the accumulation of acetophenones, and dissect the genetic architecture of these defence compounds against SBW in white spruce mature trees.
Out of 4747 single nucleotide polymorphisms (SNPs) from 2312 genes genotyped in a population of 211 unrelated individuals, genetic association analyses identified 35 SNPs in 33 different genes that were significantly associated with the defence traits by using single-locus, multi-locus and multi-trait approaches. The multi-locus approach was particularly effective at detecting SNP-trait associations that explained a large fraction of the phenotypic variance (from 20 to 43%). Significant genes were regulatory including the NAC transcription factor, or they were involved in carbohydrate metabolism, falling into the binding, catalytic or transporter activity functional classes. Most of them were highly expressed in foliage. Weak positive phenotypic correlations were observed between defence and growth traits, indicating little or no evidence of defence-growth trade-offs.
This study provides new insights on the genetic architecture of tree defence traits, contributing to our understanding of the physiology of resistance mechanisms to biotic factors and providing a basis for the genetic improvement of the constitutive defence of white spruce against SBW.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Nucleotide-binding domain and Leucine-rich Repeat (NLR) immune receptor genes form a major line of defence in plants, acting in both pathogen recognition and resistance machinery activation. NLRs are ...reported to form large gene clusters in limber pine (Pinus flexilis) but it is unknown how widespread this genomic architecture may be among the extant species of conifers (Pinophyta). We used comparative genomic analyses to assess patterns in the abundance, diversity and genomic distribution of NLR genes. Chromosome-level whole genome assemblies and high-density linkage maps in the Pinaceae, Cupressaceae, Taxaceae and other gymnosperms were scanned for NLR genes using existing and customised pipelines. Discovered genes were mapped across chromosomes and linkage groups and analysed phylogenetically for evolutionary history. Conifer genomes are characterised by dense clusters of NLR genes, highly localised on one chromosome. These clusters are rich in TNL-encoding genes, which seem to have formed through multiple tandem duplication events. In contrast to angiosperms and non-coniferous gymnosperms, genomic clustering of NLR genes is ubiquitous in conifers. NLR-dense genomic regions are likely to influence a large part of the plant's resistance, informing our understanding of adaptation to biotic stress and the development of genetic resources through breeding.
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