Diatom flagellar genes and their expression during sexual reproduction in Leptocylindrus danicus

Background: Flagella have been lost in the vegetative phase of the diatom life cycle, but they are still present in male gametes of centric species, thereby representing a hallmark of sexual reproduction. This process, besides maintaining and creating new genetic diversity, in diatoms is also fundamental to restore the maximum cell size following its reduction during vegetative division. Nevertheless, sexual reproduction has been demonstrated in a limited number of diatom species, while our understanding of its different phases and of their genetic control is scarce. Results: In the transcriptome of Leptocylindrus danicus, a centric diatom widespread in the world's seas, we identified 22 transcripts related to the flagella development and confirmed synchronous overexpression of 6 flagellum-related genes during the male gamete formation process. These transcripts were mostly absent in the closely related species L. aporus, which does not have sexual reproduction. Among the 22 transcripts, L. danicus showed proteins that belong to the Intra Flagellar Transport (IFT) subcomplex B as well as IFT-A proteins, the latter previously thought to be absent in diatoms. The presence of flagellum-related proteins was also traced in the transcriptomes of several other centric species. Finally, phylogenetic reconstruction of the IFT172 and IFT88 proteins showed that their sequences are conserved across protist species and have evolved similarly to other phylogenetic marker genes. Conclusion: Our analysis describes for the first time the diatom flagellar gene set, which appears to be more complete and functional than previously reported based on the genome sequence of the model centric diatom, Thalassiosira pseudonana. This first recognition of the whole set of diatom flagellar genes and of their activation pattern paves the way to a wider recognition of the relevance of sexual reproduction in individual species and in the natural environment

TEspeX: consensus-specific quantification of transposable element expression preventing biases from exonized fragments

Summary: Transposable elements (TEs) play key roles in crucial biological pathways. Therefore, several tools enabling the quantification of their expression were recently developed. However, many of the existing tools lack the capability to distinguish between the transcription of autonomously expressed TEs and TE fragments embedded in canonical coding/non-coding non-TE transcripts. Consequently, an apparent change in the expression of a given TE may simply reflect the variation in the expression of the transcripts containing TE-derived sequences. To overcome this issue, we have developed TEspeX, a pipeline for the quantification of TE expression at the consensus level. TEspeX uses Illumina RNA-seq short reads to quantify TE expression avoiding counting reads deriving from inactive TE fragments embedded in canonical transcripts. Availability and implementation: The tool is implemented in python3, distributed under the GNU General Public License (GPL) and available on Github at https://github.com/fansalon/TEspeX (Zenodo URL: https://doi.org/10.5281/zenodo.6800331). Supplementary information: Supplementary data are available at Bioinformatics online

Shuffling of cis-regulatory elements is a pervasive feature of the vertebrate lineage

BACKGROUND: All vertebrates share a remarkable degree of similarity in their development as well as in the basic functions of their cells. Despite this, attempts at unearthing genome-wide regulatory elements conserved throughout the vertebrate lineage using BLAST-like approaches have thus far detected noncoding conservation in only a few hundred genes, mostly associated with regulation of transcription and development. RESULTS: We used a unique combination of tools to obtain regional global-local alignments of orthologous loci. This approach takes into account shuffling of regulatory regions that are likely to occur over evolutionary distances greater than those separating mammalian genomes. This approach revealed one order of magnitude more vertebrate conserved elements than was previously reported in over 2,000 genes, including a high number of genes found in the membrane and extracellular regions. Our analysis revealed that 72% of the elements identified have undergone shuffling. We tested the ability of the elements identified to enhance transcription in zebrafish embryos and compared their activity with a set of control fragments. We found that more than 80% of the elements tested were able to enhance transcription significantly, prevalently in a tissue-restricted manner corresponding to the expression domain of the neighboring gene. CONCLUSION: Our work elucidates the importance of shuffling in the detection of cis-regulatory elements. It also elucidates how similarities across the vertebrate lineage, which go well beyond development, can be explained not only within the realm of coding genes but also in that of the sequences that ultimately govern their expression

Meta-Analysis Suggests That Intron Retention Can Affect Quantification of Transposable Elements from RNA-Seq Data

Simple Summary Transposable elements (TEs) are repetitive sequences comprising more than one third of the human genome with the original ability to change their location within the genome. Owing to their repetitive nature, the quantification of TEs results often challenging. RNA-seq is a useful tool for genome-wide TEs quantification, nevertheless it also presents technical issues, including low reads mappability and erroneous quantification derived from the transcription of TEs fragments embedded in canonical transcripts. Fragments derived from TEs are found within the introns of most genes, which led to the hypothesis that intron retention (IR) can affect the unbiased quantification of TEs expression. Performing meta-analysis of public RNA-seq datasets, here we observe that IR can indeed impact the quantification of TEs by increasing the number of reads mapped on intronic TE copies. Our work highlights a correlation between IR and TEs expression measurement by RNA-seq that should be taken into account to achieve reliable TEs quantification, especially in samples characterized by extensive IR, because differential IR might be confused with differential TEs expression. Transposable elements (TEs), also known as "jumping genes", are repetitive sequences with the capability of changing their location within the genome. They are key players in many different biological processes in health and disease. Therefore, a reliable quantification of their expression as transcriptional units is crucial to distinguish between their independent expression and the transcription of their sequences as part of canonical transcripts. TEs quantification faces difficulties of different types, the most important one being low reads mappability due to their repetitive nature preventing an unambiguous mapping of reads originating from their sequences. A large fraction of TEs fragments localizes within introns, which led to the hypothesis that intron retention (IR) can be an additional source of bias, potentially affecting accurate TEs quantification. IR occurs when introns, normally removed from the mature transcript by the splicing machinery, are maintained in mature transcripts. IR is a widespread mechanism affecting many different genes with cell type-specific patterns. We hypothesized that, in an RNA-seq experiment, reads derived from retained introns can introduce a bias in the detection of overlapping, independent TEs RNA expression. In this study we performed meta-analysis using public RNA-seq data from lymphoblastoid cell lines and show that IR can impact TEs quantification using established tools with default parameters. Reads mapped on intronic TEs were indeed associated to the expression of TEs and influence their correct quantification as independent transcriptional units. We confirmed these results using additional independent datasets, demonstrating that this bias does not appear in samples where IR is not present and that differential TEs expression does not impact on IR quantification. We concluded that IR causes the over-quantification of intronic TEs and differential IR might be confused with differential TEs expression. Our results should be taken into account for a correct quantification of TEs expression from RNA-seq data, especially in samples in which IR is abundant

Transposons Acting as Competitive Endogenous RNAs: In-Silico Evidence from Datasets Characterised by L1 Overexpression

LINE L1 are transposable elements that can replicate within the genome by passing through RNA intermediates. The vast majority of these element copies in the human genome are inactive and just between 100 and 150 copies are still able to mobilize. During evolution, they could have been positively selected for beneficial cellular functions. Nonetheless, L1 deregulation can be detrimental to the cell, causing diseases such as cancer. The activity of miRNAs represents a fundamental mechanism for controlling transcript levels in somatic cells. These are a class of small non-coding RNAs that cause degradation or translational inhibition of their target transcripts. Beyond this, competitive endogenous RNAs (ceRNAs), mostly made by circular and non-coding RNAs, have been seen to compete for the binding of the same set of miRNAs targeting protein coding genes. In this study, we have investigated whether autonomously transcribed L1s may act as ceRNAs by analyzing public dataset in-silico. We observed that genes sharing miRNA target sites with L1 have a tendency to be upregulated when L1 are overexpressed, suggesting the possibility that L1 might act as ceRNAs. This finding will help in the interpretation of transcriptomic responses in contexts characterized by the specific activation of transposons

Promiscuity of enhancer, coding and non-coding transcription functions in ultraconserved elements

<p>Abstract</p> <p>Background</p> <p>Ultraconserved elements (UCEs) are highly constrained elements of mammalian genomes, whose functional role has not been completely elucidated yet. Previous studies have shown that some of them act as enhancers in mouse, while some others are expressed in both normal and cancer-derived human tissues. Only one UCE element so far was shown to present these two functions concomitantly, as had been observed in other isolated instances of single, non ultraconserved enhancer elements.</p> <p>Results</p> <p>We used a custom microarray to assess the levels of UCE transcription during mouse development and integrated these data with published microarray and next-generation sequencing datasets as well as with newly produced PCR validation experiments. We show that a large fraction of non-exonic UCEs is transcribed across all developmental stages examined from only one DNA strand. Although the nature of these transcripts remains a mistery, our meta-analysis of RNA-Seq datasets indicates that they are unlikely to be short RNAs and that some of them might encode nuclear transcripts. In the majority of cases this function overlaps with the already established enhancer function of these elements during mouse development. Utilizing several next-generation sequencing datasets, we were further able to show that the level of expression observed in non-exonic UCEs is significantly higher than in random regions of the genome and that this is also seen in other regions which act as enhancers.</p> <p>Conclusion</p> <p>Our data shows that the concurrent presence of enhancer and transcript function in non-exonic UCE elements is more widespread than previously shown. Moreover through our own experiments as well as the use of next-generation sequencing datasets, we were able to show that the RNAs encoded by non-exonic UCEs are likely to be long RNAs transcribed from only one DNA strand.</p

Intraspecific diversity in the cold stress response of transposable elements in the diatom leptocylindrus aporus

Transposable elements (TEs), activated as a response to unfavorable conditions, have been proposed to contribute to the generation of genetic and phenotypic diversity in diatoms. Here we explore the transcriptome of three warm water strains of the diatom Leptocylindrus aporus, and the possible involvement of TEs in their response to changing temperature conditions. At low temperature (13 \ub0C) several stress response proteins were overexpressed, confirming low temperature to be unfavorable for L. aporus, while TE-related transcripts of the LTR retrotransposon superfamily were the most enriched transcripts. Their expression levels, as well as most of the stress-related proteins, were found to vary significantly among strains, and even within the same strains analysed at different times. The lack of overexpression after many months of culturing suggests a possible role of physiological plasticity in response to growth under controlled laboratory conditions. While further investigation on the possible central role of TEs in the diatom stress response is warranted, the strain-specific responses and possible role of in-culture evolution draw attention to the interplay between the high intraspecific variability and the physiological plasticity of diatoms, which can both contribute to the adaptation of a species to a wide range of conditions in the marine environment

Genomewide transcriptional reprogramming in the seagrass Cymodocea nodosa under experimental ocean acidification

Here, we report the first use of massive-scale RNA-sequencing to explore seagrass response to CO2-driven ocean acidification (OA). Large-scale gene expression changes in the seagrass Cymodocea nodosa occurred at CO2 levels projected by the end of the century. C. nodosa transcriptome was obtained using Illumina RNA-Seq technology and de novo assembly, and differential gene expression was explored in plants exposed to short-term high CO2/low pH conditions. At high pCO(2), there was a significant increased expression of transcripts associated with photosynthesis, including light reaction functions and CO2 fixation, and also to respiratory pathways, specifically for enzymes involved in glycolysis, in the tricarboxylic acid cycle and in the energy metabolism of the mitochondrial electron transport. The upregulation of respiratory metabolism is probably supported by the increased availability of photo-synthates and increased energy demand for biosynthesis and stress-related processes under elevated CO2 and low pH. The upregulation of several chaperones resembling heat stress-induced changes in gene expression highlighted the positive role these proteins play in tolerance to intracellular acid stress in seagrasses. OA further modifies C. nodosa secondary metabolism inducing the transcription of enzymes related to biosynthesis of carbon-based secondary compounds, in particular the synthesis of polyphenols and isoprenoid compounds that have a variety of biological functions including plant defence. By demonstrating which physiological processes are most sensitive to OA, this research provides a major advance in the understanding of seagrass metabolism in the context of altered seawater chemistry from global climate change.Portuguese FCT project HighGrass [PTDC/MAR-EST/3687/2012

Identification of sex determination genes and their evolution in Phlebotominae sand flies (Diptera, Nematocera)

Phlebotomine sand flies (Diptera, Nematocera) are important vectors of several pathogens, including Leishmania parasites, causing serious diseases of humans and dogs. Despite their importance as disease vectors, most aspects of sand fly biology remain unknown including the molecular basis of their reproduction and sex determination, aspects also relevant for the development of novel vector control strategies

Identification of the meiotic toolkit in diatoms and exploration of meiosis-specific SPO11 and RAD51 homologs in the sexual species Pseudo-nitzschia multistriata and Seminavis robusta (vol 16, 930, 2015)

Following the publication of this article [1], the authors reported that the link to Additionalfile11 linked to the wrong set of data. The correct supplementary data is provided in this Correction article (Additionalfile11)