Transcriptome-wide Gene Research Articles

Examining the disconnect between prolactin and parental care in avian brood parasites.

Prolactin is often referred to as the "parental hormone" but there are examples in which prolactin and parental behavior are disconnected. One intriguing example is in avian obligate brood parasites; species exhibiting high circulating prolactin but no parental care. To understand this disconnect, we examined transcriptional and behavioral responses to prolactin in brown-headed (Molothrus ater) and bronzed (M aeneus) brood parasitic cowbirds. We first examine prolactin-dependent regulation of transcriptome wide gene expression in the preoptic area (POA), a brain region associated with parental care across vertebrates. We next examined prolactin-dependent abundance of seven parental care-related candidate genes in hypothalamic regions that are prolactin-responsive in other avian species. We found no evidence of prolactin sensitivity in cowbirds in either case. To understand this prolactin insensitivity, we compared prolactin receptor transcript abundance between parasitic and nonparasitic species and between prolactin treated and untreated cowbirds. We observed significantly lower prolactin receptor transcript abundance in brown-headed but not bronzed cowbird POA compared with a nonparasite and no prolactin-dependent changes in either parasitic species. Finally, estrogen-primed female brown-headed cowbirds with or without prolactin treatment exhibited significantly greater avoidance of nestling begging stimuli compared with untreated birds. Taken together, our results suggest that modified prolactin receptor distributions in the POA and surrounding hypothalamic regions disconnect prolactin from parental care in brood parasitic cowbirds.

Identification of Reproducible BCL11A Alterations in Schizophrenia Through Individual-Level Prediction of Coexpression.

Previous studies have provided evidence for an alteration of genetic coexpression in schizophrenia (SCZ). However, such analyses have thus far lacked biological specificity for individual genes, which may be critical for identifying illness-relevant effects. Therefore, we applied machine learning to identify gene-specific coexpression differences at the individual subject level and compared these between individuals with SCZ, bipolar disorder, major depressive disorder (MDD), autism spectrum disorder (ASD), and healthy controls. Utilizing transcriptome-wide gene expression data from 21 independent datasets, comprising a total of 9509 participants, we identified a reproducible decrease of BCL11A coexpression across 4 SCZ datasets that showed diagnostic specificity for SCZ when compared with ASD and MDD. We further demonstrate that individual-level coexpression differences can be combined in multivariate coexpression scores that show reproducible illness classification across independent datasets in SCZ and ASD. This study demonstrates that machine learning can capture gene-specific coexpression differences at the individual subject level for SCZ and identify novel biomarker candidates.

Comparative RNA sequencing based transcriptome profiling of regular bearing and alternate bearing mango (Mangifera indica L.) varieties reveals novel insights into the regulatory mechanisms underlying alternate bearing.

This study is to understand a comprehensive perspective on the molecular mechanisms underlying alternate bearing in mango (Mangifera indica L.) via transcriptome wide gene expression profiling of both regular and irregular mango varieties. Transcriptome data of regular (Neelam) and irregular (Dashehari) mango varieties revealed a total of 42,397 genes. Out of that 12,557 significantly differentially expressed genes were identified, of which 6453 were found to be up-regulated and 6104 were found to be down-regulated genes. Further, many of the common unigenes which were involved in hormonal regulation, metabolic processes, oxidative stress, ion homeostasis, alternate bearing etc. showed significant differences between these two different bearing habit varieties. Pathway analysis showed the highest numbers of differentially expressed genes were related with the metabolic processes (523). A total of 26 alternate bearing genes were identified and principally three genes viz; SPL-like gene (GBVX01015803.1), Rumani GA-20-oxidase-like gene (GBVX01019650.1) and LOC103420644 (GBVX01016070.1) were significantly differentially expressed (at log2FC and pval less than 0.05) while, only single gene (gbGBVW01004309.1) related with flowering was found to be differentially expressed. A total of 15 differentially expressed genes from three important pathways viz; alternate bearing, carbohydrate metabolism and hormone synthesis were validated using Real time PCR and results were at par with in silico analysis. Deciphering the differentially expressed genes (DEGs) and potential candidate genes associated with alternate bearing, hormone and carbohydrate metabolism pathways will help for illustrating the molecular mechanisms underlying the bearing tendencies in mango.

Abstract A85: Prediagnostic 25-hydroxyvitamin D concentrations in relation to tumor molecular alterations and risk of breast cancer recurrence

Abstract Although vitamin D inhibits breast tumor growth and development in experimental settings, the findings from population-based studies remain inconclusive. Our goals were to investigate the association between prediagnostic plasma 25-hydroxyvitamin D [25(OH)D] concentration and breast cancer recurrence in prospective epidemiologic studies and to explore the molecular underpinnings linking 25(OH)D to slower progression of breast cancer. Our study included 659 women with invasive breast cancer from the Nurses’ Health Studies (NHS) diagnosed from 1989 to 2010. Plasma 25(OH)D was measured with a high-affinity protein binding assay or a radioimmunoassay. We profiled transcriptome-wide gene expression in breast tumors using microarrays. Hazard ratios (HRs) of breast cancer recurrence were estimated from covariate-adjusted Cox proportional hazards models. We examined differential gene expression regulation in association with 25(OH)D with covariate-adjusted linear regression and employed pathway analysis. We derived a gene expression score for 25(OH)D, and further assessed associations between the standardized score and breast cancer recurrence. Although 25(OH)D was not associated with breast cancer recurrence overall (hazard ratio (HR)=0.96; 95% confidence interval (CI): 0.86, 1.08) (median follow-up=12.8 years), the association varied by estrogen receptor (ER) status (p for interaction=0.005). Importantly, among ER-positive stage I to III cancers, every 5 ng/ml increase in prediagnostic plasma 25(OH)D was associated with a 13% lowered risk of recurrence (HR=0.87; 95% CI: 0.76; 0.99). No association was observed in women with stage I to III ER-negative cancers (HR=1.07; 95% CI: 0.91; 1.27). Pathway analysis identified multiple gene sets that were significantly (FDR<5%) downregulated in ER-positive breast tumors of women with high prediagnostic 25(OH)D (≥30 ng/ml), compared to those with low levels (<30 ng/ml). These gene sets are primarily involved in tumor proliferation, apoptosis, migration and invasion, inflammation, and cancer cell metabolism. The standardized score of 25(OH)D signatures derived from these gene sets was marginally significantly associated with reduced risk of recurrence in ER-positive diseases (HR=0.77; 95% CI: 0.59, 1.01). Using TIMER, a mathematical algorithm to deconvolute tumor cellular heterogeneity, we found more infiltrated CD8+ T cells in ER-negative tumors as compared to ER-positive tumors, and this higher immune infiltration in ER-negative tumors may contribute to the observed heterogeneous response. Our findings support an intriguing line of research for future studies to better understand the biologic mechanisms underlying the role of vitamin D in breast tumor progression, particularly for the ER-positive subtype. Citation Format: Yujing Heng, Donghao Lu, Natalie DuPre, Kimberly Glass, Peter Kraft, David Feldman, Susan Hankinson, Kathryn Rexrode, Heather Eliassen, Rulla Tamimi, Cheng Peng. Prediagnostic 25-hydroxyvitamin D concentrations in relation to tumor molecular alterations and risk of breast cancer recurrence [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2019 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(3 Suppl):Abstract nr A85.

Proteolysis and cartilage development are activated in the synovium after surgical induction of post traumatic osteoarthritis.

Anterior cruciate ligament (ACL) transection surgery in the minipig induces post-traumatic osteoarthritis (PTOA) in a pattern similar to that seen in human patients after ACL injury. Prior studies have reported the presence of cartilage matrix-degrading proteases, such as Matrix metalloproteinase-1 (MMP-1) and A disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS-4), in the synovial fluid of injured or arthritic joints; however, the tissue origin of these proteases is unknown. The objective of this study was to identify transcriptional processes activated in the synovium after surgical induction of PTOA with ACL transection, and to determine if processes associated with proteolysis were enriched in the synovium after ACL transection. Unilateral ACL transection was performed in adolescent Yucatan minipigs and synovium samples were collected at 1, 5, 9, and 14 days post-injury. Transcriptome-wide gene expression levels were determined using bulk RNA-Sequencing in the surgical animals and control animals with healthy knees. The greatest number of transcripts with significant changes was observed 1 day after injury. These changes were primarily associated with cellular proliferation, consistent with measurements of increased cellularity of the synovium at the two-week time point. At five to 14 days, the expression of transcripts relating to proteolysis and cartilage development was significantly enriched. While protease inhibitor-encoding transcripts (TIMP2, TIMP3) represented the largest fraction of protease-associated transcripts in the uninjured synovium, protease-encoding transcripts (including MMP1, MMP2, ADAMTS4) predominated after surgery. Cartilage development-associated transcripts that are typically not expressed by synovial cells, such as ACAN and COMP, were enriched in the synovium following ACL-transection. The upregulation in both catabolic processes (proteolysis) and anabolic processes (cartilage development) suggests that the synovium plays a complex, balancing role in the early response to PTOA induction.

Gene Expression Profiling of Head and Neck Tumors Identifies FOXP1 and SOX10 Expression as Useful for Distinguishing Ameloblastoma From Basaloid Salivary Gland Tumors.

Odontogenic tumors show considerable morphologic heterogeneity and at times the diagnosis can be challenging. Ameloblastoma, the most common odontogenic tumor, can have morphologic similarity to some salivary gland tumors and therefore we sought to identify biomarkers that might aid in the diagnosis by performing transcriptome wide gene expression profiling of 80 odontogenic and salivary gland neoplasms. These data identified the FOXP1/SOX10 expression profile as characteristic of many odontogenic tumors including ameloblastoma but largely absent in salivary gland tumors. We then assessed 173 salivary gland tumors and 108 odontogenic tumors by immunohistochemistry for FOXP1 and SOX10 expression and found that 34/35 (97%) cases of ameloblastomas were diffusely positive for FOXP1 but completely negative for SOX10. None of the basaloid salivary neoplasms (basal cell adenoma, adenoid cystic carcinoma, polymorphous adenocarcinoma, and myoepitheloma) demonstrated FOXP1/SOX10 expression pattern. Taken together, the results of this study suggest that the FOXP1/SOX10 immunophenotype is common in odontogenic tumors including ameloblastoma and might be useful distinguishing these from similar appearing basaloid salivary gland tumors.

Longitudinal transcriptome-wide gene expression analysis of sleep deprivation treatment shows involvement of circadian genes and immune pathways

Therapeutic sleep deprivation (SD) rapidly induces robust, transient antidepressant effects in a large proportion of major mood disorder patients suffering from a depressive episode, but underlying biological factors remain poorly understood. Research suggests that these patients may have altered circadian molecular genetic ‘clocks’ and that SD functions through ‘resetting’ dysregulated genes; additional factors may be involved, warranting further investigation. Leveraging advances in microarray technology enabling the transcriptome-wide assessment of gene expression, this study aimed to examine gene expression changes accompanying SD and recovery sleep in patients suffering from an episode of depression. Patients (N = 78) and controls (N = 15) underwent SD, with blood taken at the same time of day before SD, after one night of SD and after recovery sleep. A transcriptome-wide gene-by-gene approach was used, with a targeted look also taken at circadian genes. Furthermore, gene set enrichment, and longitudinal gene set analyses including the time point after recovery sleep, were conducted. Circadian genes were significantly affected by SD, with patterns suggesting that molecular clocks of responders and non-responders, as well as patients and controls respond differently to chronobiologic stimuli. Notably, gene set analyses revealed a strong widespread effect of SD on pathways involved in immune function and inflammatory response, such as those involved in cytokine and especially in interleukin signalling. Longitudinal gene set analyses showed that in responders these pathways were upregulated after SD; in non-responders, little response was observed. Our findings emphasize the close relationship between circadian, immune and sleep systems and their link to etiology of depression at the transcriptomic level.

Efficient depletion of ribosomal RNA for RNA sequencing in planarians

BackgroundThe astounding regenerative abilities of planarian flatworms prompt steadily growing interest in examining their molecular foundation. Planarian regeneration was found to require hundreds of genes and is hence a complex process. Thus, RNA interference followed by transcriptome-wide gene expression analysis by RNA-seq is a popular technique to study the impact of any particular planarian gene on regeneration. Typically, the removal of ribosomal RNA (rRNA) is the first step of all RNA-seq library preparation protocols. To date, rRNA removal in planarians was primarily achieved by the enrichment of polyadenylated (poly(A)) transcripts. However, to better reflect transcriptome dynamics and to cover also non-poly(A) transcripts, a procedure for the targeted removal of rRNA in planarians is needed.ResultsIn this study, we describe a workflow for the efficient depletion of rRNA in the planarian model species S. mediterranea. Our protocol is based on subtractive hybridization using organism-specific probes. Importantly, the designed probes also deplete rRNA of other freshwater triclad families, a fact that considerably broadens the applicability of our protocol. We tested our approach on total RNA isolated from stem cells (termed neoblasts) of S. mediterranea and compared ribodepleted libraries with publicly available poly(A)-enriched ones. Overall, mRNA levels after ribodepletion were consistent with poly(A) libraries. However, ribodepleted libraries revealed higher transcript levels for transposable elements and histone mRNAs that remained underrepresented in poly(A) libraries. As neoblasts experience high transposon activity this suggests that ribodepleted libraries better reflect the transcriptional dynamics of planarian stem cells. Furthermore, the presented ribodepletion procedure was successfully expanded to the removal of ribosomal RNA from the gram-negative bacterium Salmonella typhimurium.ConclusionsThe ribodepletion protocol presented here ensures the efficient rRNA removal from low input total planarian RNA, which can be further processed for RNA-seq applications. Resulting libraries contain less than 2% rRNA. Moreover, for a cost-effective and efficient removal of rRNA prior to sequencing applications our procedure might be adapted to any prokaryotic or eukaryotic species of choice.

Exposure to Dimethyl Selenide (DMSe)-Derived Secondary Organic Aerosol Alters Transcriptomic Profiles in Human Airway Epithelial Cells.

Dimethyl selenide (DMSe) is one of the major volatile organoselenium compounds released from aquatic and terrestrial environments through microbial transformation and plant metabolism. The detailed processes of DMSe leading to secondary organic aerosol (SOA) formation and the pulmonary health effects induced by inhalation of DMSe-derived SOA remain largely unknown. In this study, we characterized the chemical composition and formation yields of SOA produced from the oxidation of DMSe with OH radicals and O3 in controlled chamber experiments. Further, we profiled the transcriptome-wide gene expression changes in human airway epithelial cells (BEAS-2B) after exposure to DMSe-derived SOA. Our analyses indicated a significantly higher SOA yield resulting from the OH-initiated oxidation of DMSe. The oxidative potential of DMSe-derived SOA, as measured by the dithiothreitol (DTT) assay, suggested the presence of oxidizing moieties in DMSe-derived SOA at levels higher than typical ambient aerosols. Utilizing RNA sequencing (RNA-Seq) techniques, gene expression profiling followed by pathway enrichment analysis revealed several major biological pathways perturbed by DMSe-derived SOA, including elevated genotoxicity, DNA damage, and p53-mediated stress responses, as well as downregulated cholesterol biosynthesis, glycolysis, and interleukin IL-4/IL-13 signaling. This study highlights the significance of DMSe-derived SOA as a stressor in human airway epithelial cells.

ZNF518B as a transcriptional factor involved in colorectal cancer progression through the epithelial to mesenchymal transition

Abstract Background Colorectal cancer (CRC) represents a relevant public health problem. The identification of new markers involved in the mechanisms of invasiveness represents a priority in order to better understand cancer development and generate new therapeutic targets. Recently, our group demonstrated overexpression of ZNF518B gene, which encodes an unknown zinc finger transcription factor, in CRC. A transcriptome-wide gene expression profile revealed its implication in different biological processes related to the progression of CRC, especially in the epithelial to mesenchymal transition (EMT). Methods To study the biological processes regulated by ZNF518B, we performed a ClariomS Array (Affimetrix) by silencing this gene with a specific siRNA in DLD1 and HCT116 CRC cell lines. All data were analysed using the TAC (Transcriptomic Analysis ConsoleTM) software. The array validation was performed by RT-qPCR. To explore the in vitro mechanisms related to EMT, an overexpression of ZNF518B was performed in RKO, cell line that does not express the gene. Invasion and migration assays were done using transwells and we also test adhesion capacity to type I collagen plates. Results The analysis of ClariomS showed, after the silencing of the transcription factor ZNF518B, an alteration of 64 genes commonly modified in the DLD1 and HCT116 cell lines. These differentially expressed genes are involved in different biological processes involved in cellular focal adhesion, in the EMT process and in cell cycle. On the other hand, some proliferation pathways are also altered like MAPK, RAS, WNT and VEGFR2 signalling pathways. Through in vitro analysis, we showed that the increased expression of ZNF518B enhanced cell migration, invasion and adhesion to type I collagen. These results in vitro support the altered gene expression profile after ZNF518B silencing observed in the ClariomS array, suggesting that this gene has an important role in the progression of CRC throughout EMT. Conclusions The transcription factor ZNF518B could be involved in the progression of tumorigenesis by inducing EMT in CRC.The transcription factor ZNF518B could be involved in the progression of tumorigenesis by inducing EMT in CRC. Legal entity responsible for the study INCLIVA, Biomedical Research Institute. Funding Instituto de Salud Carlos III, Rio Hortega Contract, ESMO Translational Research Fellowship Programme. Disclosure A. Cervantes: Advisory / Consultancy, Speaker Bureau / Expert testimony, Research grant / Funding (institution): Merck Serono; Advisory / Consultancy, Speaker Bureau / Expert testimony, Research grant / Funding (institution): Roche; Advisory / Consultancy, Research grant / Funding (institution): Beigene; Advisory / Consultancy, Research grant / Funding (institution): Bayer; Advisory / Consultancy, Speaker Bureau / Expert testimony, Research grant / Funding (institution): Servier; Advisory / Consultancy, Research grant / Funding (institution): Lilly; Advisory / Consultancy, Research grant / Funding (institution): Novartis; Advisory / Consultancy, Research grant / Funding (institution): Takeda; Advisory / Consultancy, Research grant / Funding (institution): Astellas; Advisory / Consultancy: Pierre Fabre; Research grant / Funding (self): Genentech; Research grant / Funding (institution): Fibrogene; Research grant / Funding (institution): Amcure; Research grant / Funding (institution): Sierra Oncology; Research grant / Funding (institution): AstraZeneca; Research grant / Funding (institution): MedImmune; Research grant / Funding (institution): BMS; Research grant / Funding (institution): MSD; Speaker Bureau / Expert testimony: Amgen; Speaker Bureau / Expert testimony: Fundation Medicine. All other authors have declared no conflicts of interest.

Two Distinct Subtypes Revealed in Blood Transcriptome of Breast Cancer Patients With an Unsupervised Analysis.

Background: Breast cancer (BC) is a highly heterogeneous cancer. The interaction between immune system and BC is complex, widespread yet unclear. In this study, we aimed to reveal the heterogeneity of host systemic immune response to BC and understand the possible mechanisms that may drive the heterogeneity using transcriptomic data from peripheral blood mononuclear cells (PBMCs).Methods: Transcriptome-wide gene expressions of PBMCs in 33 BC patients were generated by RNA sequencing. An unsupervised clustering algorithm was employed to discover PBMC transcriptome subtypes among BC patients. Association analysis between PBMC subtypes and age, clinical stage, abundance of immune cells, and other clinical factors was performed to understand the underlying biological processes that may drive this heterogeneity. Immune gene signature identification and in silico survival analysis were performed to investigate the potential clinical implications of these PBMC subtypes. The findings were validated using the whole blood transcriptomes of an independent cohort.Results: We observed that established BC subtypes were not associated with PBMC gene expression profiles. Instead, we discovered and validated two new BC subtypes using PBMC transcriptome, which have distinct immune cell proportions, especially for lymphocytes (P = 5.22 × 10−12) and neutrophils (P = 1.13 × 10−14). Enrichment analysis of differentially expressed genes revealed that these two subtypes had distinct patterns of immune responses, including osteoclast differentiation and interleukin-10 signaling pathway. We developed two immune gene signatures that can differentiate these two BC PBMC subtypes. Further analysis suggested they had the ability to predict the clinical outcome of BC patients.Conclusions: PBMC transcriptome profiles can classify BC patients into two distinct subtypes. These two subtypes are mainly shaped by different immune cell abundance, which may have implications on clinical outcomes.

Transcriptional reprogramming during Garcinia-type recalcitrant seed germination of Garcinia mangostana

Mangosteen (Garcinia mangostana L.) is an apomictic tropical fruit tree species which asexually produces recalcitrant seeds that are sensitive to desiccation and not viable for long-term storage. It is important to understand seed germination to inform strategy for the propagation and conservation of important plants, especially for recalcitrant species. However, little is known about the molecular physiology of tropical seed germination. Garcinia-type seed germination is unique such that the primary root and shoot emerge from the opposite ends, which is akin to in vivo direct organogenesis. For the first time, time-course transcriptome-wide gene expression during mangosteen seed germination from day 0 to day 7 was performed through RNA-sequencing with RT-qPCR validation using independent samples in triplicates. The analyses of differentially expressed genes (DEGs) and expression profile identified D3 as a key turning point for transcriptional reprogramming with an indication of a transient shift to anaerobic respiration. Particularly, the activation of ethylene signalling through increased expression of genes involved in its biosynthesis and the upregulation of ethylene-responsive transcription factors lead to the induction of direct organogenesis concurrently with seed germination. Abscisic acid (ABA) signalling appeared to play a role in stress response while gibberellin (GA) provides growth potential during mangosteen seed germination. The antagonistic relationship between ABA and GA is conserved in mangosteen as manifested by global gene expression changes based on the Arabidopsis co-expression network analysis. The current transcriptomic study comprehensively describes various molecular aspects of Garcinia-type recalcitrant seed germination and highlights similarities to in vitro somatic embryogenesis but without a differentiated embryo. This study provides insights to improve seed germination for the mass propagation of mangosteen.

Transcriptomic abnormalities in peripheral blood in bipolar disorder, and discrimination of the major psychoses

We performed a transcriptome-wide meta-analysis and gene co-expression network analysis to identify genes and gene networks dysregulated in the peripheral blood of bipolar disorder (BD) cases relative to unaffected comparison subjects, and determined the specificity of the transcriptomic signatures of BD and schizophrenia (SZ). Nineteen genes and 4 gene modules were significantly differentially expressed in BD cases. Thirteen gene modules were shown to be differentially expressed in a combined case-group of BD and SZ subjects called “major psychosis”, including genes biologically linked to apoptosis, reactive oxygen, chromatin remodeling, and immune signaling. No modules were differentially expressed between BD and SZ cases. Machine-learning classifiers trained to separate diagnostic classes based solely on gene expression profiles could distinguish BD cases from unaffected comparison subjects with an area under the curve (AUC) of 0.724, as well as BD cases from SZ cases with AUC = 0.677 in withheld test samples. We introduced a novel and straightforward method called “polytranscript risk scoring” that could distinguish BD cases from unaffected subjects (AUC = 0.672) and SZ cases (AUC = 0.607) significantly better than expected by chance. Taken together, our results highlighted gene expression alterations common to BD and SZ that involve biological processes of inflammation, oxidative stress, apoptosis, and chromatin regulation, and highlight disorder-specific changes in gene expression that discriminate the major psychoses.

Integrative analysis of transcriptome-wide association study and gene expression profiling identifies candidate genes associated with stroke.

BackgroundStroke is a major public health burden worldwide. Although genetic variation is known to play a role in the pathogenesis of stroke, the specific pathogenic mechanisms are still unclear. Transcriptome-wide association studies (TWAS) is a powerful approach to prioritize candidate risk genes underlying complex traits. However, this approach has not been applied in stroke.MethodsWe conducted an integrative analysis of TWAS using data from the MEGASTROKE Consortium and gene expression profiling to identify candidate genes for the pathogenesis of stroke. Gene ontology (GO) enrichment analysis was also conducted to detect functional gene sets.ResultsThe TWAS identified 515 transcriptome-wide significant tissue-specific genes, among which SLC25A44 (P = 5.46E−10) and LRCH1 (P = 1.54E−6) were significant by Bonferroni test for stroke. After validation with gene expression profiling, 19 unique genes were recognized. GO enrichment analysis identified eight significant GO functional gene sets, including regulation of cell shape (P = 0.0059), face morphogenesis (P = 0.0247), and positive regulation of ATPase activity (P = 0.0256).ConclusionsOur study identified multiple stroke-associated genes and gene sets, and this analysis provided novel insights into the genetic mechanisms underlying stroke.

Eye Degeneration and Loss of otx5b Expression in the Cavefish Sinocyclocheilus tileihornes

Cave animals possess remarkable phenotypes associated with existence in their dark environments. The Chinese cavefish Sinocyclocheilus tileihornes shows substantial eye degeneration, a trait shared by most cave species. The extent to which independent evolution of troglomorphic traits uses convergent molecular genetic mechanisms is as yet unknown. We performed transcriptome-wide gene expression profiling in S. tileihornes eyes and compared results with those from the closely related surface species S. angustiporus and an independently derived congeneric cavefish, S. anophthalmus. In total, 52.85 million 100 bp long paired-end clean reads were generated for S. tileihornes, and we identified differentially expressed genes between the three possible pairs of species. Functional analysis of genes differentially expressed between S. tileihornes and S. angustiporus revealed that phototransduction (KEGG id: dre04744) was the most significantly enriched pathway, indicating the obvious differences in response to captured photons between the cavefish S. tileihornes and the surface species S. angustiporus. Analysis of key genes regulating eye development showed complete absence of otx5b (orthodenticle homolog 5) expression in S. tileihornes eyes, probably related to degradation of rods, but normal expression of crx (cone-rod homeobox). The enriched pathways and Otx5 are involved in phototransduction, photoreceptor formation, and regulation of photoreceptor-related gene expression. Unlike the S. tileihornes reported here, S. anophthalmus has reduced crx and otx5 expression. These results show that different species of cavefish within the same genus that independently evolved troglodyte characteristics can have different genetic mechanisms of eye degeneration.

RNA Sequencing Data: Hitchhiker's Guide to Expression Analysis

Gene expression is the fundamental level at which the results of various genetic and regulatory programs are observable. The measurement of transcriptome-wide gene expression has convincingly switched from microarrays to sequencing in a matter of years. RNA sequencing (RNA-seq) provides a quantitative and open system for profiling transcriptional outcomes on a large scale and therefore facilitates a large diversity of applications, including basic science studies, but also agricultural or clinical situations. In the past 10 years or so, much has been learned about the characteristics of the RNA-seq data sets, as well as the performance of the myriad of methods developed. In this review, we give an overview of the developments in RNA-seq data analysis, including experimental design, with an explicit focus on the quantification of gene expression and statistical approachesfor differential expression. We also highlight emerging data types, such as single-cell RNA-seq and gene expression profiling using long-read technologies.

Transcriptomic resilience, symbiont shuffling, and vulnerability to recurrent bleaching in reef-building corals.

As climate change progresses and extreme temperature events increase in frequency, rates of disturbance may soon outpace the capacity of certain species of reef-building coral to recover from bleaching. This may lead to dramatic shifts in community composition and ecosystem function. Understanding variation in rates of bleaching recovery among species and how that translates to resilience to recurrent bleaching is fundamental to predicting the impacts of increasing disturbances on coral reefs globally. We tracked the response of two heat sensitive species in the genus Acropora to repeated bleaching events during the austral summers of 2015 and 2017. Despite a similar bleaching response, the species Acropora gemmifera recovered faster based on transcriptome-wide gene expression patterns and had a more dynamic algal symbiont community than Acropora hyacinthus growing on the same reef. Moreover, A.gemmifera had higher survival to repeated heat extremes, with six-fold lower mortality than A.hyacinthus. These patterns suggest that speed of recovery from a first round of bleaching, based on multiple mechanisms, contributes strongly to sensitivity to a second round of bleaching. Furthermore, our data uncovered intragenus variation in a group of corals thought generally to be heat-sensitive and therefore paint a more nuanced view of the future health of coral reef ecosystems against a backdrop of increasing thermal disturbances.

Myeloid Cell and Transcriptome Signatures Associated With Inflammation Resolution in a Model of Self-Limiting Acute Brain Inflammation.

Inflammation contributes to tissue repair and restoration of function after infection or injury. However, some forms of inflammation can cause tissue damage and disease, particularly if inappropriately activated, excessive, or not resolved adequately. The mechanisms that prevent excessive or chronic inflammation are therefore important to understand. This is particularly important in the central nervous system where some effects of inflammation can have particularly harmful consequences, including irreversible damage. An increasing number of neurological disorders, both acute and chronic, and their complications are associated with aberrant neuroinflammatory activity. Here we describe a model of self-limiting acute brain inflammation optimized to study mechanisms underlying inflammation resolution. Inflammation was induced by intracerebral injection of lipopolysaccharide (LPS) and the temporal profile of key cellular and molecular changes were defined during the progression of the inflammatory response. The kinetics of accumulation and loss of neutrophils in the brain enabled well-demarcated phases of inflammation to be operatively defined, including induction and resolution phases. Microglial reactivity and accumulation of monocyte-derived macrophages were maximal at the onset of and during the resolution phase. We profiled the transcriptome-wide gene expression changes at representative induction and resolution timepoints and used gene coexpression network analysis to identify gene clusters. This revealed a distinct cluster of genes associated with inflammation resolution that were induced selectively or maximally during this phase and indicated an active programming of gene expression that may drive resolution as has been described in other tissues. Induction of gene networks involved in lysosomal function, lipid metabolism, and a comparative switch to MHC-II antigen presentation (relative to MHC-I during induction) were prominent during the resolution phase. The restoration and/or further induction of microglial homeostatic signature genes was notable during the resolution phase. We propose the current model as a tractable reductionist system to complement more complex models for further understanding how inflammation resolution in the brain is regulated and as a platform for in vivo testing/screening of candidate resolution-modifying interventions. Our data highlight how resolution involves active cellular and transcriptome reprogramming and identify candidate gene networks associated with resolution-phase adaptations that warrant further study.

Increases in Genistein in Medicago sativa Confer Resistance against the Pisum Host Race of Acyrthosiphon pisum.

Interspecific interaction with host plants have important consequences for the host race formation of herbivorous insects. Plant secondary metabolites, particularly those that are involved in host races specializing on plants, warrant the theory of host specialization. Acyrthosiphon pisum comprises various host races that adapt to different Fabaceae plants, which provides an ideal system for determining the behavioral and physiological mechanisms underlying host-adaptive diversification. The current study evaluated the effects of host transfer on population fitness, feeding behavior and the transcriptome-wide gene expression of the two host races of A. pisum, one of which was originally from Medicago sativa and the other from Pisum sativum. The results showed that the Pisum host race of A. pisum had a lower population abundance and feeding efficiency than the Medicago host race in terms of a longer penetration time and shorter duration times of phloem ingestion when fed on M. sativa. In contrast, few differences were found in the population abundance and feeding behavior of A. pisum between the two host races when fed on P. sativum. Meanwhile, of the nine candidate phenolic compounds, only genistein was significantly affected by aphid infestation; higher levels of genistein were detected in M. sativa after feeding by the Pisum host race, but these levels were reduced relative to uninfested controls after feeding by the Medicago host race, which suggested that genistein may be involved in the specialization of the aphid host race on M. sativa. Further exogenous application of genistein in artificial diets showed that the increase in genistein reduced the survival rate of the Pisum host race but had little effect on that of the Medicago host race. The transcriptomic profiles indicated that the transcripts of six genes with functions related to detoxification were up-regulated in the Pisum host race relative to the Medicago host race of A. pisum. These results suggested that the inducible plant phenolics and associated metabolic process in aphids resulted in their differential adaptations to their Fabaceae host.

Caste- and pesticide-specific effects of neonicotinoid pesticide exposure on gene expression in bumblebees.

Social bees are important insect pollinators of wildflowers and agricultural crops, making their reported declines a global concern. A major factor implicated in these declines is the widespread use of neonicotinoid pesticides. Indeed, recent research has demonstrated that exposure to low doses of these neurotoxic pesticides impairs bee behaviours important for colony function and survival. However, our understanding of the molecular‐genetic pathways that lead to such effects is limited, as is our knowledge of how effects may differ between colony members. To understand what genes and pathways are affected by exposure of bumblebee workers and queens to neonicotinoid pesticides, we implemented a transcriptome‐wide gene expression study. We chronically exposed Bombus terrestriscolonies to either clothianidin or imidacloprid at field‐realistic concentrations while controlling for factors including colony social environment and worker age. We reveal that genes involved in important biological processes including mitochondrial function are differentially expressed in response to neonicotinoid exposure. Additionally, clothianidin exposure had stronger effects on gene expression amplitude and alternative splicing than imidacloprid. Finally, exposure affected workers more strongly than queens. Our work demonstrates how RNA‐Seq transcriptome profiling can provide detailed novel insight on the mechanisms mediating pesticide toxicity to a key insect pollinator.