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Genome divergence by repeat proliferation and/or loss is a process that plays a crucial role in species evolution. Nevertheless, knowledge of the variability related to repeat proliferation among species of the same family is still limited. Considering the importance of the Asteraceae family, here we present a first contribution towards the metarepeatome of five Asteraceae species. A comprehensive picture of the repetitive components of all genomes was obtained by genome skimming with Illumina sequence reads and by analyzing a pool of full-length long terminal repeat retrotransposons (LTR-REs). Genome skimming allowed us to estimate the abundance and variability of repetitive components. The structure of the metagenome of the selected species was composed of 67% repetitive sequences, of which LTR-REs represented the bulk of annotated clusters. The species essentially shared ribosomal DNA sequences, whereas the other classes of repetitive DNA were highly variable among species. The pool of full-length LTR-REs was retrieved from all the species and their age of insertion was established, showing several lineage-specific proliferation peaks over the last 15-million years. Overall, a large variability of repeat abundance at superfamily, lineage, and sublineage levels was observed, indicating that repeats within individual genomes followed different evolutionary and temporal dynamics, and that different events of amplification or loss of these sequences may have occurred after species differentiation.

Long terminal repeats (LTR) retrotransposons have a major role in determining genome size, structure and function, thanks to their ability to transpose. We performed a meta-analysis of LTR-retrotransposon expression in roots of sunflower plantlets treated with different plant hormones, chemicals and NaCl. By using Illumina cDNA libraries, available from public repositories, we measured the number of reads matching the retrotranscriptase domains isolated from a whole genome library of retrotransposons. LTR-retrotransposons resulted in general barely expressed, except for 4 elements, all belonging to the AleII lineage, which showed high transcription levels in roots of both control and treated plants. The expression of retrotransposons in treated plants was slightly higher than in the control. Transcribed elements belonged to specific chromosomal loci and were not abundant in the genome. A few elements resulted differentially expressed depending on the treatment. Results suggest that, although most retrotransposons are not expressed, the transcription of such elements is related to their abundance, to their position in the chromosome and to their lineage.

The sunflower (Helianthus annuus L.) is among the most widely cultivated crops in the world due to the oilseed production. Lipid transfer proteins (LTPs) are low molecular mass proteins encoded by a broad multigenic family in higher plants, showing a vast range of functions; these proteins have not been characterised in sunflower at the genomic level. In this work, we exploited the reliable genome sequence of sunflower to identify and characterise the LTP multigenic family in H. annuus. Overall, 101 sunflower putative LTP genes were identified using a homology search and the HMM algorithm. The selected sequences were characterised through phylogenetic analysis, exon–intron organisation, and protein structural motifs. Sunflower LTPs were subdivided into four clades, reflecting their genomic and structural organisation. This gene family was further investigated by analysing the possible duplication origin of genes, which showed the prevalence of tandem and whole genome duplication events, a result that is in line with polyploidisation events that occurred during sunflower genome evolution. Furthermore, LTP gene expression was evaluated on cDNA libraries constructed on six sunflower tissues (leaf, root, ligule, seed, stamen, and pistil) and from roots treated with stimuli mimicking biotic and abiotic stress. Genes encoding LTPs belonging to three out of four clades responded specifically to external stimuli, especially to abscisic acid, auxin, and the saline environment. Interestingly, genes encoding proteins belonging to one clade were expressed exclusively in sunflower seeds. This work is a first attempt of genome-wide identification and characterisation of the LTP multigenic family in a plant species.

The WUSCHEL-related homeobox (WOX) is a family of specific transcription factors involved in plant development and response to stress, characterized by the presence of a homeodomain. This study represents the first comprehensive characterization of the WOX family in a member of the Asteraceae family, the sunflower (H. annuus L.). Overall, we identified 18 putative HaWOX genes divided by phylogenetic analysis in three major clades (i.e., ancient, intermediate, and WUS). These genes showed conserved structural and functional motifs. Moreover, HaWOX has homogeneously distributed on H. annuus chromosomes. In particular, 10 genes originated after whole segment duplication events, underpinning a possible evolution of this family along with the sunflower genome. In addition, gene expression analysis evidenced a specific pattern of regulation of the putative 18 HaWOX during embryo growth and in ovule and inflorescence meristem differentiation, suggesting a pivotal role for this multigenic family in sunflower development. The results obtained in this work improved the understanding of the WOX multigenic family, providing a resource for future study on functional analysis in an economically valuable species such as sunflower.

Background Long Terminal Repeat retrotransposons (LTR-REs) are repetitive DNA sequences that constitute a large part of the genome. The improvement of sequencing technologies and sequence assembling strategies has achieved genome sequences with much greater reliability than those of the past, especially in relation to repetitive DNA sequences. Results In this study, we analysed the genome of Ficus carica L., obtained using third generation sequencing technologies and recently released, to characterise the complete complement of full-length LTR-REs to study their dynamics during fig genome evolution. A total of 1867 full-length elements were identified. Those belonging to the Gypsy superfamily were the most abundant; among these, the Chromovirus/Tekay lineage was the most represented. For the Copia superfamily, Ale was the most abundant lineage. Measuring the estimated insertion time of each element showed that, on average, Ivana and Chromovirus/Tekay were the youngest lineages of Copia and Gypsy superfamilies, respectively. Most elements were inactive in transcription, both constitutively and in leaves of plants exposed to an abiotic stress, except for some elements, mostly belonging to the Copia/Ale lineage. A relationship between the inactivity of an element and inactivity of genes lying in close proximity to it was established. Conclusions The data reported in this study provide one of the first sets of information on the genomic dynamics related to LTR-REs in a plant species with highly reliable genome sequence. Fig LTR-REs are highly heterogeneous in abundance and estimated insertion time, and only a few elements are transcriptionally active. In general, the data suggested a direct relationship between estimated insertion time and abundance of an element and an inverse relationship between insertion time (or abundance) and transcription, at least for Copia LTR-REs.

Transposable elements (TEs) are DNA sequences that can change their position within genomes. TEs are present in most organisms and can be an important genomic component. Their activities are manifold: restructuring of genome size, chromosomal rearrangements, induction of gene mutations, and alteration of gene activity by insertion near or within promoters, intronic regions, or enhancer. There are several examples of mutations and other genetic variations determined by the activity of TEs, associated with the evolution of prokaryotic and eukaryotic organisms and the domestication of plants. Generally, TE mobilization occurs when the organism is subjected to stress, which can include both biotic and abiotic stresses, polyploidy conditions, and interspecific hybridizations, very common events in plants. TEs are widely distributed among organisms. TEs also play essential roles in evolution, but most of them are either dormant or inactive. This is mainly determined by epigenetic silencing mechanisms, regulatory systems, and control systems that aim to limit its proliferation. Furthermore, the host has recruited many genes originated from TEs as transcriptional regulators, especially in defense against pathogens and invasive genetic elements; this phenomenon is called molecular domestication. Therefore, TEs are responsible for horizontal gene transfer and the movement of genetic material between organisms, even phylogenetically distant, with a consequent remixing of their gene pools.

Seagrasses as Posidonia oceanica reproduce mostly by vegetative propagation, which can reduce genetic variability within populations. Since, in clonally propagated species, insurgence of genetic variability can be determined by the activity of transposable elements, we have estimated the activity of such repeat elements by measuring their expression level in the leaves of plants from a Mediterranean site, for which Illumina complementary DNA (cDNA) sequence reads (produced from RNAs isolated by leaves of plants from deep and shallow meadows) were publicly available. Firstly, we produced a collection of retrotransposon-related sequences and then mapped Illumina cDNA reads onto these sequences. With this approach, it was evident that Posidonia retrotransposons are, in general, barely expressed; only nine elements resulted transcribed at levels comparable with those of reference genes encoding tubulins and actins. Differences in transcript abundance were observed according to the superfamily and the lineage to which the retrotransposons belonged. Only small differences were observed between retrotransposon expression levels in leaves of shallow and deep Posidonia meadow stands, whereas one TAR/Tork element resulted differentially expressed in deep plants exposed to heat. It can be concluded that, in P. oceanica, the contribution of retrotransposon activity to genetic variability is reduced, although the nine specific active elements could actually produce new structural variations.

Arbuscular mycorrhizal fungi (AMF) play a fundamental role in plant growth and nutrition in natural and agricultural ecosystems. Despite the importance of such symbionts, the different developmental changes occurring during the AMF life cycle have not been fully elucidated at the molecular level. Here, the RNA-seq approach was used to investigate Rhizoglomus irregulare specific and common transcripts at two different time points of mycorrhizal establishment in Helianthus annuus in vivo. Four days after inoculation, transcripts related to cellular remodeling (actin and tubulin), cellular signaling (calmodulin, serine/threonine protein kinase, 14-3-3 protein, and calcium transporting ATPase), lipid metabolism (fatty acid desaturation, steroid hormone, and glycerophospholipid biosynthesis), and biosynthetic processes were detected. In addition to such transcripts, 16 days after inoculation, expressed genes linked to binding and catalytic activities; ion (K+, Ca2+, Fe2+, Zn2+, Mn2+, Pi, ammonia), sugar, and lipid transport; and those involved in vacuolar polyphosphate accumulation were found. Knowledge of transcriptomic changes required for symbiosis establishment and performance is of great importance to understand the functional role of AMF symbionts in food crop nutrition and health, and in plant diversity in natural ecosystems.

Background Similar to other urban trees, holm oaks (Quercus ilex L.) provide a physiological, ecological and social service in the urban environment, since they remove atmospheric pollution. However, the urban environment has several abiotic factors that negatively influence plant life, which are further exacerbated due to climate change, especially in the Mediterranean area. Among these abiotic factors, increased uptake of Na + and Cl − usually occurs in trees in the urban ecosystem; moreover, an excess of the tropospheric ozone concentration in Mediterranean cities further affects plant growth and survival. Here, we produced and annotated a de novo leaf transcriptome of Q. ilex as well as transcripts over- or under-expressed after a single episode of O3 (80 nl l-1, 5 h), a salt treatment (150 mM for 15 days) or a combination of these treatments, mimicking a situation that plants commonly face, especially in urban environments. Results Salinity dramatically changed the profile of expressed transcripts, while the short O3 pulse had less effect on the transcript profile. However, the short O3 pulse had a very strong effect in inducing over- or under-expression of some genes in plants coping with soil salinity. Many differentially regulated genes were related to stress sensing and signalling, cell wall remodelling, ROS sensing and scavenging, photosynthesis and to sugar and lipid metabolism. Most differentially expressed transcripts revealed here are in accordance with a previous report on Q. ilex at the physiological and biochemical levels, even though the expression profiles were overall more striking than those found at the biochemical and physiological levels. Conclusions We produced for the first time a reference transcriptome for Q. ilex, and performed gene expression analysis for this species when subjected to salt, ozone and a combination of the two. The comparison of gene expression between the combined salt + ozone treatment and salt or ozone alone showed that even though many differentially expressed genes overlap all treatments, combined stress triggered a unique response in terms of gene expression modification. The obtained results represent a useful tool for studies aiming to investigate the effects of environmental stresses in urban-adapted tree species. Electronic supplementary material The online version of this article (10.1186/s12864-018-5260-2) contains supplementary material, which is available to authorized users.

Pdzrn3, a multidomain protein with E3-ubiquitin ligase activity, has been reported to play a role in myoblast and osteoblast differentiation and, more recently, in neuronal and endothelial cell development. The expression of the pdzrn3 gene is developmentally regulated in various vertebrate tissues, including muscular, neural and vascular system. Little is known about its expression during kidney development, although genetic polymorphisms and alterations around the human pdzrn3 chromosomal region have been found to be associated with renal cell carcinomas and other kidney diseases. We investigated the pdzrn3 spatio-temporal expression pattern in Xenopus laevis embryos by in situ hybridization. We focused our study on the development of the pronephros, which is the embryonic amphibian kidney, functionally similar to the most primitive nephric structures of human kidney. To explore the role of pdzrn3 during renal morphogenesis, we performed loss-of-function experiments, through antisense morpholino injections and analysed the morphants using specific pronephric markers. Dynamic pdzrn3 expression was observed in embryonic tissues, such as somites, brain, eye, blood islands, heart, liver and pronephros. Loss of function experiments resulted in specific alterations of pronephros development. In particular, at early stages, pdzrn3 depletion was associated with a reduction of the pronephros anlagen and later, with perturbations of the tubulogenesis, including deformation of the proximal tubules. Rescue experiments, in which mRNA of the zebrafish pdzrn3 orthologue was injected together with the morpholino, allowed recovery of the kidney phenotypes. These results underline the importance of pdzrn3 expression for correct nephrogenesis.

Although Ficus carica L. (fig) is one of the most resistant fruit tree species to salinity, no comprehensive studies are currently available on its molecular responses to salinity. Here we report a transcriptome analysis of F. carica cv. Dottato exposed to 100 mM sodium chloride for 7 weeks, where RNA-seq analysis was performed on leaf samples at 24 and 48 days after the beginning of salinization; a genome-derived fig transcriptome was used as a reference. At day 24, 224 transcripts were significantly up-regulated and 585 were down-regulated, while at day 48, 409 genes were activated and 285 genes were repressed. Relatively small transcriptome changes were observed after 24 days of salt treatment, showing that fig plants initially tolerate salt stress. However, after an early down-regulation of some cell functions, major transcriptome changes were observed after 48 days of salinity. Seven weeks of 100 mM NaCl dramatically changed the repertoire of expressed genes, leading to activation or reactivation of many cell functions. We also identified salt-regulated genes, some of which had not been previously reported to be involved in plant salinity responses. These genes could be potential targets for the selection of favourable genotypes, through breeding or biotechnology, to improve salt tolerance in fig or other crops.

Arbuscular mycorrhizal (AM) fungi are essential elements of soil fertility, plant nutrition and productivity, facilitating soil mineral nutrient uptake. Helianthus annuus is a non-model, widely cultivated species. Here we used an RNA-seq approach for evaluating gene expression variation at early and late stages of mycorrhizal establishment in sunflower roots colonized by the arbuscular fungus Rhizoglomus irregulare. mRNA was isolated from roots of plantlets at 4 and 16 days after inoculation with the fungus. cDNA libraries were built and sequenced with Illumina technology. Differential expression analysis was performed between control and inoculated plants. Overall 726 differentially expressed genes (DEGs) between inoculated and control plants were retrieved. The number of up-regulated DEGs greatly exceeded the number of down-regulated DEGs and this difference increased in later stages of colonization. Several DEGs were specifically involved in known mycorrhizal processes, such as membrane transport, cell wall shaping, and other. We also found previously unidentified mycorrhizal-induced transcripts. The most important DEGs were carefully described in order to hypothesize their roles in AM symbiosis. Our data add a valuable contribution for deciphering biological processes related to beneficial fungi and plant symbiosis, adding an Asteraceae, non-model species for future comparative functional genomics studies.

The availability of transcriptomic data sequence is a key step for functional genomics studies. Recently, a repertoire of predicted genes of a Japanese cultivar of fig (Ficus carica L.) was released. Because of the great phenotypic variability that can be found in this species, we decided to study another fig genotype, the Italian cv. Dottato, in order to perform comparative studies between the two cultivars and extend the pan genome of this species. We isolated, sequenced and assembled fig genomic DNA from young fruits of cv. Dottato. Then, putative gene sequences were predicted and annotated. Finally, a comparison was performed between cvs. Dottato and Horaishi predicted transcriptomes. Our data provide a resource (available at the Sequence Read Archive database under SRP109082) to be used for functional genomics of fig, in order to fill the gap of knowledge still existing in this species concerning plant development, defense and adaptation to the environment.

Retrotransposon expression during arbuscular mycorrhizal (AM) fungal colonisation of sunflower roots (Helianthus annuus) was analysed using Illumina RNA-Seq, in order to verify whether mycorrhizal symbiosis can activate retrotransposable elements. Illumina cDNA libraries were produced from RNAs isolated from the roots of sunflower plants at 4 and 16 days after inoculation with the AM fungus Rhizoglomus irregulare and from their respective control plants. Illumina reads were mapped to a library of reverse transcriptase-encoding sequences, putatively belonging to long terminal repeat retrotransposons of Gypsy and Copia superfamilies. Forty-six different reverse transcriptase sequences were transcribed, although at a low rate, in mycorrhizal or control roots and only four were significantly over-expressed at day 16, compared with control roots. Almost all expressed or over-expressed sequences belonged to low-copy elements, mostly, of the Copia superfamily. A meta-analysis, using publicly available Illumina cDNA libraries obtained from sunflower plants treated with different hormones and chemicals, mimicking stimuli produced by abiotic and biotic stresses, was also conducted. Such analyses indicated that the four reverse transcriptase sequences over-expressed in mycorrhizal roots were explicitly induced only by AM symbiosis, showing the specificity of AM stimuli compared to that of other fungal/plant interactions.

Ozone (O3) and salinity are usually tested as combined factors on plant performance. However, the response to a single episode of O3 in plants already stressed by an excess of NaCl as occurs in the natural environment has never been investigated, but is important given that it is commonly experienced in Mediterranean areas. Three-year-old Quercus ilex L. (holm oak) saplings were exposed to salinity (150 mM NaCl, 15 days), and the effect on photosynthesis, hydric relations and ion partitioning was evaluated (Experiment I). In Experiment II, salt-treated saplings were exposed to 80 nl l-1 of O3 for 5 h, which is a realistic dose in a Mediterranean environment. Gas exchanges, chlorophyll fluorescence and antioxidant systems were characterized to test whether the salt-induced stomatal closure limited O3 uptake and stress or whether the pollutant represents an additional stressor for plants. Salt-dependent stomatal closure depressed the photosynthetic process (-71.6% of light-saturated rate of photosynthesis (A380)) and strongly enhanced the dissipation of energy via the xanthophyll cycle. However, salt-treated plants had higher values of net assimilation rate/stomatal conductance (A/gs) than the controls, which was attributable to a greater mesophyll conductance gm/gs and carboxylation efficiency (higher gm/maximal rate of Rubisco carboxylation (Vcmax)), thus suggesting no damage to chloroplasts. O3 did not exacerbate the effect of salinity on photosynthesis, however a general enhancement of the Halliwell-Asada cycle was necessary to counteract the O3-triggered oxidative stress. Despite the 79.4% gs reduction in salt-stressed plants, which strongly limited the O3 uptake, a single peak in the air pollutant led to an additional burden for the antioxidant system when plants had been previously subjected to salinity.