Global Transcriptomic Response Research Articles

FAILLA MEMORIAL LECTUREHow We Got Here: One Laboratory's Odyssey in the Field of Radiation-Inducible Genes.

This review focuses on early discoveries that contributed to our understanding and the scope of transcriptional responses after radiation damage. Before the development of modern approaches to assess overall global transcriptomic responses, the idea that mammalian cells could respond to DNA-damaging agents in a manner analogous to bacteria was not generally accepted. To investigate this possibility, the development of technology to identify differentially expressed low-abundance transcripts substantially facilitated our appreciation that DNA damaging agents like UV radiation and subsequently ionizing radiation did in fact produce robust transcriptional responses. Here we focus on our identification and characterization of radiation-inducible genes, and how even early studies on stress gene signaling highlighted the broad scope of transcriptional responses to radiation damage. Since then, the central role of transcriptional responses to radiation injury in maintaining genome integrity has been highlighted in many processes, including cell cycle checkpoint control, resistance to cancer by p53 and other key factors, cell senescence, and metabolism.

Brucella-driven host N-glycome remodeling controls infection

Many powerful methods have been employed to elucidate the global transcriptomic, proteomic, or metabolic responses to pathogen-infected host cells. However, the host glycome responses to bacterial infection remain largely unexplored, and hence, our understanding of the molecular mechanisms by which bacterial pathogens manipulate the host glycome to favor infection remains incomplete. Here, we address this gap by performing a systematic analysis of the host glycome during infection by the bacterial pathogen Brucella spp. that cause brucellosis. We discover, surprisingly, that a Brucella effector protein (EP) Rhg1 induces global reprogramming of the host cell N-glycome by interacting with components of the oligosaccharide transferase complex that controls N-linked protein glycosylation, and Rhg1 regulates Brucella replication and tissue colonization in a mouse model of brucellosis, demonstrating that Brucella exploits the EP Rhg1 to reprogram the host N-glycome and promote bacterial intracellular parasitism, thereby providing a paradigm for bacterial control of host cell infection.

Integrative transcriptomics reveals association of abscisic acid and lignin pathways with cassava whitefly resistance

BackgroundWhiteflies are a global threat to crop yields, including the African subsistence crop cassava (Manihot esculenta). Outbreaks of superabundant whitefly populations throughout Eastern and Central Africa in recent years have dramatically increased the pressures of whitefly feeding and virus transmission on cassava. Whitefly-transmitted viral diseases threaten the food security of hundreds of millions of African farmers, highlighting the need for developing and deploying whitefly-resistant cassava. However, plant resistance to whiteflies remains largely poorly characterized at the genetic and molecular levels. Knowledge of cassava-defense programs also remains incomplete, limiting characterization of whitefly-resistance mechanisms. To better understand the genetic basis of whitefly resistance in cassava, we define the defense hormone- and Aleurotrachelus socialis (whitefly)-responsive transcriptome of whitefly-susceptible (COL2246) and whitefly-resistant (ECU72) cassava using RNA-seq. For broader comparison, hormone-responsive transcriptomes of Arabidopsis thaliana were also generated.ResultsWhitefly infestation, salicylic acid (SA), jasmonic acid (JA), ethylene (ET), and abscisic acid (ABA) transcriptome responses of ECU72 and COL2246 were defined and analyzed. Strikingly, SA responses were largely reciprocal between the two cassava genotypes and we suggest candidate regulators. While susceptibility was associated with SA in COL2246, resistance to whitefly in ECU72 was associated with ABA, with SA-ABA antagonism observed. This was evidenced by expression of genes within the SA and ABA pathways and hormone levels during A. socialis infestation. Gene-enrichment analyses of whitefly- and hormone-responsive genes suggest the importance of fast-acting cell wall defenses (e.g., elicitor recognition, lignin biosynthesis) during early infestation stages in whitefly-resistant ECU72. A surge of ineffective immune and SA responses characterized the whitefly-susceptible COL2246’s response to late-stage nymphs. Lastly, in comparison with the model plant Arabidopsis, cassava’s hormone-responsive genes showed striking divergence in expression.ConclusionsThis study provides the first characterization of cassava’s global transcriptome responses to whitefly infestation and defense hormone treatment. Our analyses of ECU72 and COL2246 uncovered possible whitefly resistance/susceptibility mechanisms in cassava. Comparative analysis of cassava and Arabidopsis demonstrated that defense programs in Arabidopsis may not always mirror those in crop species. More broadly, our hormone-responsive transcriptomes will also provide a baseline for the cassava community to better understand global responses to other yield-limiting pests/pathogens.

Predicting gene expression changes upon epigenomic drug treatment

Background Tumors are characterized by global changes in epigenetic modifications such as DNA methylation and histone modifications that are functionally linked to tumor progression. Accordingly, several drugs targeting the epigenome have been proposed for cancer therapy, notably, histone deacetylase inhibitors (HDACi) such as vorinostat and DNA methyltransferase inhibitors (DNMTi) such as zebularine. However, a fundamental challenge with such approaches is the lack of genomic specificity, i.e., the transcriptional changes at different genomic loci can be highly variable, thus making it difficult to predict the consequences on the global transcriptome and drug response. For instance, treatment with DNMTi may upregulate the expression of not only a tumor suppressor but also an oncogene, leading to unintended adverse effect. Methods Given the pre-treatment transcriptome and epigenomic profile of a sample, we assessed the extent of predictability of locus-specific changes in gene expression upon treatment with HDACi using machine learning. Results We found that in two cell lines (HCT116 treated with Largazole at eight doses and RH4 treated with Entinostat at 1µM) where the appropriate data (pre-treatment transcriptome and epigenome as well as post-treatment transcriptome) is available, our model distinguished the post-treatment up versus downregulated genes with high accuracy (up to ROC of 0.89). Furthermore, a model trained on one cell line is applicable to another cell line suggesting generalizability of the model. Conclusions Here we present a first assessment of the predictability of genome-wide transcriptomic changes upon treatment with HDACi. Lack of appropriate omics data from clinical trials of epigenetic drugs currently hampers the assessment of applicability of our approach in clinical setting.

Enhanced Ferroptosis Induction Susceptibility in Diffuse Large B-Cell Lymphoma through Rocaglate-Mediated Translation Inhibition

Introduction: Diffuse Large B-cell Lymphoma (DLBCL) is a genomically complex hematologic malignancy characterized by heterogeneous pathogenesis across multiple molecular subtypes. Despite recent advances, standard frontline treatment options leave a third of patients with relapsed or refractory disease due to tumor populations resistant to therapy by a wide variety of mechanisms. Ferroptosis, an iron-dependent non-programmed cell death mechanism driven by lipid peroxidation, has emerged as a promising therapeutic opportunity in DLBCL. DLBCL demonstrates distinct sensitivity to ferroptosis activators compared to other cancer types, yet the implications in DLBCL prognosis, immune infiltration, and drug resistance remain elusive. Notably, the loss of histone deubiquitinase MYSM1 in hematopoietic stem cells significantly reduces protein synthesis, leading to reduced ferroptosis suppressors, ultimately rendering the cells more susceptible to ferroptosis. Moreover, NRF2, encoded by NFE2L, a known modulator of ferroptosis, has been linked to resistance to rocaglates (cap-dependent translation inhibitors). Intriguingly, our preliminary data indicate that rocaglates induce translation of key proteins involved in oxidative stress (i.e. SDHB) and transcription factors related to the oxidative stress response (NRF1, ATF2, NRF2). Additionally, our published findings in glioblastoma suggest the upregulation of the ferroptosis suppressor 4F2 ( SLC3A2) under translation inhibition conditions. Furthermore, our investigations uncover another potential mechanism, wherein rocaglates induce NF-κB, which was previously shown to be suppressed by dimethyl fumarate (DMF) upon ferroptosis induction in DLBCL. However, the intricate interplay between ferroptosis and translation inhibition in DLBCL remains uninvestigated, prompting our hypothesis that protein synthesis inhibitors might enhance therapeutic ferroptosis induction. Method: To test our hypothesis, we conducted a comprehensive analysis, employing TMT-pSILAC to assess the global transcriptome response to rocaglates treatment, and subsequently verified significant findings via western blot analysis. Additionally, we quantified reduced glutathione (GSH) levels, reactive oxygen species (ROS) levels, and synergy with ferroptosis inducers (e.g., erastin, RSL3) to elucidate the mechanism of rocaglates in enhancing susceptibility to the ferroptosis pathway. Moreover, we explored NF-κB, previously identified as a ferroptosis regulator in our RNAseq analysis, using ELISA and NF-κB reporter assays. Results: Our TMT-pSILAC analysis unveiled upregulation of key ferroptosis suppressors, specifically NFS1, CBS, and 4F2. Concurrent treatment with rocaglates resulted in a notable increase in GSH levels, which was attenuated when administered in combination with ferroptosis inducers (p-value < 0.005) concomitant with the reduction of Glutathione Peroxidase 4 (GPX4), modulator of lipid peroxidation. Although rocaglates alone did not elicit significant levels of reactive oxygen species (ROS), their combination with ferroptosis inducers resulted in dramatic elevation of ROS levels (p-value < 0.0005). Correspondingly, there was strong synergy between the clinical rocaglate zotatifin and different ferroptosis inducers (Bliss δ synergy score > 10) and antagonism with ferroptosis inhibitors (Bliss δ synergy score < -10). Furthermore, our investigation revealed a noteworthy upregulation of NFKB1 at the translatome level, accompanied by a significant increase in TNF-α expression at 48 hours. The NF-κB-EGFP reporter assay confirmed the enhanced expression of NF-κB in response to the treatment. Conclusion: In conclusion, rocaglate-induced stress upregulates multiple ferroptosis suppressors, suggesting that cap-dependent translation disruption triggers a protective response against ferroptosis. Strikingly, these factors did not protect DLBCL tumors from ferroptosis inducers, which synergized remarkably with rocaglates, yielding promising new combination drug strategies. The proteomic basis for ferroptosis sensitization by rocaglate therapy remains under active investigation. Understanding the intricate interplay between these pathways may unveil potential key mechanisms for DLBCL treatment, potentially offering novel therapeutic targets to improve patient outcomes.

Predicting gene expression changes upon epigenomic drug treatment

Background: Tumors are characterized by global changes in epigenetic modifications such as DNA methylation and histone modifications that are functionally linked to tumor progression. Accordingly, several drugs targeting the epigenome have been proposed for cancer therapy, notably, histone deacetylase inhibitors (HDACi) such as vorinostat and DNA methyltransferase inhibitors (DNMTi) such as zebularine. However, a fundamental challenge with such approaches is the lack of genomic specificity, i.e., the transcriptional changes at different genomic loci can be highly variable, thus making it difficult to predict the consequences on the global transcriptome and drug response. For instance, treatment with DNMTi may upregulate the expression of not only a tumor suppressor but also an oncogene, leading to unintended adverse effect. Methods: Given the pre-treatment transcriptome and epigenomic profile of a sample, we assessed the extent of predictability of locus-specific changes in gene expression upon treatment with HDACi using machine learning. Results: We found that in two cell lines (HCT116 treated with Largazole at eight doses and RH4 treated with Entinostat at 1µM) where the appropriate data (pre-treatment transcriptome and epigenome as well as post-treatment transcriptome) is available, our model distinguished the post-treatment up versus downregulated genes with high accuracy (up to ROC of 0.89). Furthermore, a model trained on one cell line is applicable to another cell line suggesting generalizability of the model. Conclusions: Here we present a first assessment of the predictability of genome-wide transcriptomic changes upon treatment with HDACi. Lack of appropriate omics data from clinical trials of epigenetic drugs currently hampers the assessment of applicability of our approach in clinical setting.

Temporal transcriptomic profiling elucidates sorghum defense mechanisms against sugarcane aphids

BackgroundThe sugarcane aphid (SCA; Melanaphis sacchari) has emerged as a key pest on sorghum in the United States that feeds from the phloem tissue, drains nutrients, and inflicts physical damage to plants. Previously, it has been shown that SCA reproduction was low and high on sorghum SC265 and SC1345 plants, respectively, compared to RTx430, an elite sorghum male parental line (reference line). In this study, we focused on identifying the defense-related genes that confer resistance to SCA at early and late time points in sorghum plants with varied levels of SCA resistance.ResultsWe used RNA-sequencing approach to identify the global transcriptomic responses to aphid infestation on RTx430, SC265, and SC1345 plants at early time points 6, 24, and 48 h post infestation (hpi) and after extended period of SCA feeding for 7 days. Aphid feeding on the SCA-resistant line upregulated the expression of 3827 and 2076 genes at early and late time points, respectively, which was relatively higher compared to RTx430 and SC1345 plants. Co-expression network analysis revealed that aphid infestation modulates sorghum defenses by regulating genes corresponding to phenylpropanoid metabolic pathways, secondary metabolic process, oxidoreductase activity, phytohormones, sugar metabolism and cell wall-related genes. There were 187 genes that were highly expressed during the early time of aphid infestation in the SCA-resistant line, including genes encoding leucine-rich repeat (LRR) proteins, ethylene response factors, cell wall-related, pathogenesis-related proteins, and disease resistance-responsive dirigent-like proteins. At 7 days post infestation (dpi), 173 genes had elevated expression levels in the SCA-resistant line and were involved in sucrose metabolism, callose formation, phospholipid metabolism, and proteinase inhibitors.ConclusionsIn summary, our results indicate that the SCA-resistant line is better adapted to activate early defense signaling mechanisms in response to SCA infestation because of the rapid activation of the defense mechanisms by regulating genes involved in monolignol biosynthesis pathway, oxidoreductase activity, biosynthesis of phytohormones, and cell wall composition. This study offers further insights to better understand sorghum defenses against aphid herbivory.

Longitudinal multi-omic changes in the transcriptome and proteome of peripheral blood cells after a 4 Gy total body radiation dose to Rhesus macaques

BackgroundNon-human primates, such as Rhesus macaques, are a powerful model for studies of the cellular and physiological effects of radiation, development of radiation biodosimetry, and for understanding the impact of radiation on human health. Here, we study the effects of 4 Gy total body irradiation (TBI) at the molecular level out to 28 days and at the cytogenetic level out to 56 days after exposure. We combine the global transcriptomic and proteomic responses in peripheral whole blood to assess the impact of acute TBI exposure at extended times post irradiation.ResultsThe overall mRNA response in the first week reflects a strong inflammatory reaction, infection response with neutrophil and platelet activation. At 1 week, cell cycle arrest and re-entry processes were enriched among mRNA changes, oncogene-induced senescence and MAPK signaling among the proteome changes. Influenza life cycle and infection pathways initiated earlier in mRNA and are reflected among the proteomic changes during the first week. Transcription factor proteins SRC, TGFβ and NFATC2 were immediately induced at 1 day after irradiation with increased transcriptional activity as predicted by mRNA changes persisting up to 1 week. Cell counts revealed a mild / moderate hematopoietic acute radiation syndrome (H-ARS) reaction to irradiation with expected lymphopenia, neutropenia and thrombocytopenia that resolved within 30 days. Measurements of micronuclei per binucleated cell levels in cytokinesis-blocked T-lymphocytes remained high in the range 0.27–0.33 up to 28 days and declined to 0.1 by day 56.ConclusionsOverall, we show that the TBI 4 Gy dose in NHPs induces many cellular changes that persist up to 1 month after exposure, consistent with damage, death, and repopulation of blood cells.

Insights into the Global Transcriptome Response of Lentinula edodes Mycelia during Aging.

The spawn of Lentinula edodes and other basidiomycete fungi tend to age with long-term culture. This causes heavy yield losses if aging spawn is used for propagation. In this study, we cultivated dikaryotic L. edodes mycelia in plates for 60 days to produce intrinsic aging phenotypes. We found that intracellular reactive oxygen species levels increased in contrast to mitochondrial depolarization and also observed greater DNA fragmentation with longer culture time. Transcriptome analysis of mycelia at different growth stages revealed pronounced expression differences between short- and long-term cultures. In particular, "phenylalanine, tyrosine, and tryptophan biosynthesis", "mitophagy and autophagy", "MAPK signaling pathway", and "ABC transporter" were among the enriched terms in the mycelial aging process. Weighted correlation network analysis identified LeAtg8, LeHog1, LePbs2, and LemTOR as key genes during aging. Western blotting confirmed that LeATG8 and phosphorylated LeHOG1 protein levels were significantly upregulated in aging mycelia. Our combined analytical approach provides insights into the mechanisms that regulate mycelial aging, indicating that autophagy/mitophagy plays a major role in counteracting the effects of age on mycelial growth development.

Global transcriptomic response of the AI-3 isomers 3,5-DPO and 3,6-DPO in Salmonella Typhimurium.

Bacterial intercellular signaling mediated by small molecules, also called autoinducers (AIs), enables synchronized behavior in response to environmental conditions, and in many bacterial pathogens, intercellular signaling controls virulence gene expression. However, in the intestinal pathogen Salmonella enterica subsp. enterica serovar Typhimurium (S. Typhimurium), although three signals, named AI-1, AI-2 and AI-3, have been described, their roles in virulence remain elusive. AI-3 is the 3,6- isomer of a previously described Vibrio cholerae signaling molecule; 3,5-dimethylpyrazin-2-ol (3,5-DPO). To elucidate the role of AI-3/DPO in S. Typhimurium, we have mapped the global transcriptomic responses to 3,5- and 3,6-DPO isomers in S. Typhimurium. Our studies showed that DPO affects expression of almost 8% of all genes. Specifically, expression of several genes involved in gut-colonization respond to DPO. Interestingly, most of the affected genes are similarly regulated by 3,5-DPO and 3,6-DPO, respectively, indicating that the two isomers have overlapping roles in S. Typhimurium.

Mycobacterium abscessus DosRS two-component system controls a species-specific regulon required for adaptation to hypoxia.

Mycobacterium abscessus (Mab), an emerging opportunistic pathogen, predominantly infects individuals with underlying pulmonary diseases such as cystic fibrosis (CF). Current treatment outcomes for Mab infections are poor due to Mab's inherent antibiotic resistance and unique host interactions that promote phenotypic tolerance and hinder drug access. The hypoxic, mucus-laden airways in the CF lung and antimicrobial phagosome within macrophages represent hostile niches Mab must overcome via alterations in gene expression for survival. Regulatory mechanisms important for the adaptation and long-term persistence of Mab within the host are poorly understood, warranting further genetic and transcriptomics study of this emerging pathogen. DosRS Mab , a two-component signaling system (TCS), is one proposed mechanism utilized to subvert host defenses and counteract environmental stress such as hypoxia. The homologous TCS of Mycobacterium tuberculosis (Mtb), DosRS Mtb , is known to induce a ~50 gene regulon in response to hypoxia, carbon monoxide (CO) and nitric oxide (NO) in vitro and in vivo. Previously, a small DosR Mab regulon was predicted using bioinformatics based on DosR Mtb motifs however, the role and regulon of DosRS Mab in Mab pathogenesis have yet to be characterized in depth. To address this knowledge gap, our lab generated a Mab dosRS knockout strain (MabΔdosRS) to investigate differential gene expression, and phenotype in an in vitro hypoxia model of dormancy. qRT-PCR and lux reporter assays demonstrate Mab_dosR and 6 predicted downstream genes are induced in hypoxia. In addition, RNAseq revealed induction of a much larger hypoxia response comprised of >1000 genes, including 127 differentially expressed genes in a dosRS mutant strain. Deletion of DosRS Mab led to attenuated growth under low oxygen conditions, a shift in morphotype from smooth to rough, and down-regulation of 216 genes. This study provides the first look at the global transcriptomic response of Mab to low oxygen conditions encountered in the airways of CF patients and within macrophage phagosomes. Our data also demonstrate the importance of DosRS Mab for adaptation of Mab to hypoxia, highlighting a distinct regulon (compared to Mtb) that is significantly larger than previously described, including both genes conserved across mycobacteria as well as Mab-specific genes.

Distinct differences in immunological properties of equine orthobiologics revealed by functional and transcriptomic analysis using an activated macrophage readout system.

Multiple biological therapies for orthopedic injuries are marketed to veterinarians, despite a lack of rigorous comparative biological activity data to guide informed decisions in selecting a most effective compound. Therefore, the goal of this study was to use relevant bioassay systems to directly compare the anti-inflammatory and immunomodulatory activity of three commonly used orthobiological therapies (OTs): mesenchymal stromal cells (MSC), autologous conditioned serum (ACS), and platelet rich plasma (PRP). Equine monocyte-derived macrophages were used as the readout system to compare therapies, including cytokine production and transcriptomic responses. Macrophages were stimulated with IL-1ß and treated 24 h with OTs, washed and cultured an additional 24 h to generate supernatants. Secreted cytokines were measured by multiplex immunoassay and ELISA. To assess global transcriptomic responses to treatments, RNA was extracted from macrophages and subjected to full RNA sequencing, using an Illumina-based platform. Data analysis included comparison of differentially expressed genes and pathway analysis in treated vs. untreated macrophages. All treatments reduced production of IL-1ß by macrophages. Secretion of IL-10 was highest in MSC-CM treated macrophages, while PRP lysate and ACS resulted in greater downregulation of IL-6 and IP-10. Transcriptomic analysis revealed that ACS triggered multiple inflammatory response pathways in macrophages based on GSEA, while MSC generated significant downregulation of inflammatory pathways, and PRP lysate induced a mixed immune response profile. Key downregulated genes in MSC-treated cultures included type 1 and type 2 interferon response, TNF-α and IL-6. PRP lysate cultures demonstrated downregulation of inflammation-related genes IL-1RA, SLAMF9, ENSECAG00000022247 but concurrent upregulation of TNF-α, IL-2 signaling, and Myc targets. ACS induced upregulation of inflammatory IL-2 signaling, TNFα and KRAS signaling and hypoxia, but downregulation of MTOR signaling and type 1 interferon signaling. These findings, representing the first comprehensive look at immune response pathways for popular equine OTs, reveal distinct differences between therapies. These studies address a critical gap in our understanding of the relative immunomodulatory properties of regenerative therapies commonly used in equine practice to treat musculoskeletal disease and will serve as a platform from which further in vivo comparisons may build.

Comparative Analysis of Transcriptomic Changes including mRNA and microRNA Expression Induced by the Xenoestrogens Zearalenone and Bisphenol A in Human Ovarian Cells.

Xenoestrogens are natural or synthetic compounds that mimic the effect of endogenous estrogens and might cause cancer. We aimed to compare the global transcriptomic response to zearalenone (ZEA; mycotoxin) and bisphenol A (BPA; plastic additive) with the effect of physiological estradiol (E2) in the PEO1 human ovarian cell line by mRNA and microRNA sequencing. Estrogen exposure induced remarkable transcriptomic changes: 308, 288 and 63 genes were upregulated (log2FC > 1); 292, 260 and 45 genes were downregulated (log2FC < -1) in response to E2 (10 nM), ZEA (10 nM) and BPA (100 nM), respectively. Furthermore, the expression of 13, 11 and 10 miRNAs changed significantly (log2FC > 1, or log2FC < -1) after exposure to E2, ZEA and BPA, respectively. Functional enrichment analysis of the significantly differentially expressed genes and miRNAs revealed several pathways related to the regulation of cell proliferation and migration. The effect of E2 and ZEA was highly comparable: 407 genes were coregulated by these molecules. We could identify 83 genes that were regulated by all three treatments that might have a significant role in the estrogen response of ovarian cells. Furthermore, the downregulation of several miRNAs (miR-501-5p, let-7a-2-3p, miR-26a-2-3p, miR-197-5p and miR-582-3p) was confirmed by qPCR, which might support the proliferative effect of estrogens in ovarian cells.

Impact of Pomegranate on Probiotic Growth, Viability, Transcriptome and Metabolism

Despite rising interest in understanding intestinal bacterial survival in situ, relatively little attention has been devoted to deciphering the interaction between bacteria and functional food ingredients. Here, we examined the interplay between diverse beneficial Lactobacillaceae species and a pomegranate (POM) extract and determined the impact of this functional ingredient on bacterial growth, cell survival, transcription and target metabolite genesis. Three commercially available probiotic strains (Lactobacillus acidophilus NCFM, Lacticaseibacillus rhamnosus GG and Lactiplantibacillus plantarum Lp-115) were used in growth assays and flow cytometry analysis, indicating differential responses to the presence of POM extract across the three strains. The inclusion of POM extract in the growth medium had the greatest impact on L. acidophilus cell counts. LIVE/DEAD staining determined significantly fewer dead cells when L. acidophilus was grown with POM extract compared to the control with no POM (1.23% versus 7.23%). Whole-transcriptome analysis following exposure to POM extract showed markedly different global transcriptome responses, with 15.88% of the L. acidophilus transcriptome, 19.32% of the L. rhamnosus transcriptome and only 2.37% of the L. plantarum transcriptome differentially expressed. We also noted strain-dependent metabolite concentrations in the medium with POM extract compared to the control medium for punicalagin, ellagic acid and gallic acid. Overall, the results show that POM extract triggers species-specific responses by probiotic strains and substantiates the rising interest in using POM as a prebiotic compound.

How to survive pig farming: Mechanism of SCCmec element deletion and metabolic stress adaptation in livestock-associated MRSA.

Previous research on methicillin susceptible Staphylococcus aureus (MSSA) belonging to livestock-associated (LA-) sequence type (ST) 398, isolated from pigs and their local surroundings, indicated that differences between these MSSA and their methicillin resistant predecessors (MRSA) are often limited to the absence of the staphylococcal cassette chromosome mec (SCCmec) and few single nucleotide polymorphisms. So far, our understanding on how LA-MRSA endure the environmental conditions associated with pig-farming as well as the putative impact of this particular environment on the mobilisation of SCCmec elements is limited. Thus, we performed in-depth genomic and transcriptomic analyses using the LA-MRSA ST398 strain IMT38951 and its methicillin susceptible descendant. We identified a mosaic-structured SCCmec region including a putative replicative SCCmecVc which is absent from the MSSA chromosome through homologous recombination. Based on our data, such events occur between short repetitive sequences identified within and adjacent to two distinct alleles of the large cassette recombinase genes C (ccrC). We further evaluated the global transcriptomic response of MRSA ST398 to particular pig-farm associated conditions, i.e., contact with host proteins (porcine serum) and a high ammonia concentration. Differential expression of global regulators involved in stress response control were identified, i.e., ammonia-induced alternative sigma factor B-depending activation of genes for the alkaline shock protein 23, the heat shock response and the accessory gene regulator (agr)-controlled transcription of virulence factors. Exposure to serum transiently induced the transcription of distinct virulence factor encoding genes. Transcription of genes reported for mediating the loss of methicillin resistance, especially ccrC, was not significantly different compared to the unchallenged controls. We concluded that, from an evolutionary perspective, bacteria may save energy by incidentally dismissing a fully replicative SCCmec element in contrast to the induction of ccr genes on a population scale. Since the genomic SCCmec integration site is a hot-spot of recombination, occasional losses of elements of 16 kb size may restore capacities for the uptake of foreign genetic material. Subsequent spread of resistance, on the other hand, might depend on the autonomous replication machinery of the deleted SCCmec elements that probably enhance chances for reintegration of SCCmec into susceptible genomes by mere multiplication.

Transcriptome profiling of Arabidopsis slac1-3 mutant reveals compensatory alterations in gene expression underlying defective stomatal closure.

Plants adjust their stomatal aperture for regulating CO2 uptake and transpiration. S-type anion channel SLAC1 (slow anion channel-associated 1) is required for stomatal closure in response to various stimuli such as abscisic acid, CO2, and light/dark transitions etc. Arabidopsis slac1 mutants exhibited defects in stimulus-induced stomatal closure, reduced sensitivity to darkness, and faster water loss from detached leaves. The global transcriptomic response of a plant with defective stimuli-induced stomatal closure (particularly because of defects in SLAC1) remains to be explored. In the current research we attempted to address the same biological question by comparing the global transcriptomic changes in Arabidopsis slac1-3 mutant and wild-type (WT) under dark, and dehydration stress, using RNA-sequencing. Abscisic acid (ABA)- and dark-induced stomatal closure was defective in Arabidopsis slac1-3 mutants, consequently the mutants had cooler leaf temperature than WT. Next, we determined the transcriptomic response of the slac1-3 mutant and WT under dark and dehydration stress. Under dehydration stress, the molecular response of slac1-3 mutant was clearly distinct from WT; the number of differentially expressed genes (DEGs) was significantly higher in mutant than WT. Dehydration induced DEGs in mutant were related to hormone signaling pathways, and biotic and abiotic stress response. Although, overall number of DEGs in both genotypes was not different under dark, however, the expression pattern was very much distinct; whereas majority of DEGs in WT were found to be downregulated, in slac1-3 majority were upregulated under dark. Further, a set 262 DEGs was identified with opposite expression pattern between WT and mutant under light–darkness transition. Amongst these, DEGs belonging to stress hormone pathways, and biotic and abiotic stress response were over-represented. To sum up, we have reported gene expression reprogramming underlying slac1-3 mutation and resultantly defective stomatal closure in Arabidopsis. Moreover, the induction of biotic and abiotic response in mutant under dehydration and darkness could be suggestive of the role of stomata as a switch in triggering these responses. To summarize, the data presented here provides useful insights into the gene expression reprogramming underlying slac1-3 mutation and resultant defects in stomatal closure.

Global transcriptomic response of Escherichia coli to p-coumaric acid

The aromatic compound p-coumaric acid (p-CA) is a secondary metabolite produced by plants. This aromatic acid and derived compounds have positive effects on human health, so there is interest in producing them in biotechnological processes with recombinant Escherichia coli strains. To determine the physiologic response of E. coli W3110 to p-CA, dynamic expression analysis of selected genes fused to a fluorescent protein reporter as well as RNA-seq and RT-qPCR were performed. The observed transcriptional profile revealed the induction of genes involved in functions related to p-CA active export, synthesis of cell wall and membrane components, synthesis of amino acids, detoxification of formaldehyde, phosphate limitation, acid stress, protein folding and degradation. Downregulation of genes encoding proteins involved in energy production, carbohydrate import and metabolism, as well as several outer and plasma membrane proteins was detected. This response is indicative of cell envelope damage causing the leakage of intracellular components including amino acids and phosphate-containing compounds. The cellular functions responding to p-CA that were identified in this study will help in defining targets for production strains improvement.

A Path-Based Analysis of Infected Cell Line and COVID-19 Patient Transcriptome Reveals Novel Potential Targets and Drugs Against SARS-CoV-2.

Most transcriptomic studies of SARS-CoV-2 infection have focused on differentially expressed genes, which do not necessarily reveal the genes mediating the transcriptomic changes. In contrast, exploiting curated biological network, our PathExt tool identifies central genes from the differentially active paths mediating global transcriptomic response. Here we apply PathExt to multiple cell line infection models of SARS-CoV-2 and other viruses, as well as to COVID-19 patient-derived PBMCs. The central genes mediating SARS-CoV-2 response in cell lines were uniquely enriched for ATP metabolic process, G1/S transition, leukocyte activation and migration. In contrast, PBMC response reveals dysregulated cell-cycle processes. In PBMC, the most frequently central genes are associated with COVID-19 severity. Importantly, relative to differential genes, PathExt-identified genes show greater concordance with several benchmark anti-COVID-19 target gene sets. We propose six novel anti-SARS-CoV-2 targets ADCY2, ADSL, OCRL, TIAM1, PBK, and BUB1, and potential drugs targeting these genes, such as Bemcentinib, Phthalocyanine, and Conivaptan.

Transcriptomic modulation in response to an intoxication with deltamethrin in a population of Triatoma infestans with low resistance to pyrethroids.

Triatoma infestans is the main vector of Chagas disease in the Southern Cone. The resistance to pyrethroid insecticides developed by populations of this species impairs the effectiveness of vector control campaigns in wide regions of Argentina. The study of the global transcriptomic response to pyrethroid insecticides is important to deepen the knowledge about detoxification in triatomines. We used RNA-Seq to explore the early transcriptomic response after intoxication with deltamethrin in a population of T. infestans which presents low resistance to pyrethroids. We were able to assemble a complete transcriptome of this vector and found evidence of differentially expressed genes belonging to diverse families such as chemosensory and odorant-binding proteins, ABC transporters and heat-shock proteins. Moreover, genes related to transcription and translation, energetic metabolism and cuticle rearrangements were also modulated. Finally, we characterized the repertoire of previously uncharacterized detoxification-related gene families in T. infestans and Rhodnius prolixus. Our work contributes to the understanding of the detoxification response in vectors of Chagas disease. Given the absence of an annotated genome from T. infestans, the analysis presented here constitutes a resource for molecular and physiological studies in this species. The results increase the knowledge on detoxification processes in vectors of Chagas disease, and provide relevant information to explore undescribed potential insecticide resistance mechanisms in populations of these insects.

Parental legacy versus regulatory innovation in salt stress responsiveness of allopolyploid cotton (Gossypium) species.

SUMMARYPolyploidy provides an opportunity for evolutionary innovation and species diversification, especially under stressful conditions. In allopolyploids, the conditional dynamics of homoeologous gene expression can be either inherited from ancestral states pre‐existing in the parental diploids or novel upon polyploidization, the latter potentially permitting a wider range of phenotypic responses to stresses. To gain insight into regulatory mechanisms underlying the diversity of salt resistance in Gossypium species, we compared global transcriptomic responses to modest salinity stress in two allotetraploid (AD‐genome) cotton species, Gossypium hirsutum and G. mustelinum, relative to their model diploid progenitors (A‐genome and D‐genome). Multivariate and pairwise analyses of salt‐responsive changes revealed a profound alteration of gene expression for about one third of the transcriptome. Transcriptional responses and associated functional implications of salt acclimation varied across species, as did species‐specific coexpression modules among species and ploidy levels. Salt responsiveness in both allopolyploids was strongly biased toward the D‐genome progenitor. A much lower level of transgressive downregulation was observed in the more salt‐tolerant G. mustelinum than in the less tolerant G. hirsutum. By disentangling inherited effects from evolved responses, we show that expression biases that are not conditional upon salt stress approximately equally reflect parental legacy and regulatory novelty upon allopolyploidization, whereas stress‐responsive biases are predominantly novel, or evolved, in allopolyploids. Overall, our work suggests that allopolyploid cottons acquired a wide range of stress response flexibility relative to their diploid ancestors, most likely mediated by complex suites of duplicated genes and regulatory factors.