This study was aimed to gain new insights into the molecular mechanisms used by Lactobacillus pla... more This study was aimed to gain new insights into the molecular mechanisms used by Lactobacillus plantarum WCFS1 to respond to hydroxytyrosol (HXT), one of the main and health-relevant plant phenolics present in olive oil. To this goal, whole genome transcriptomic profiling was used to better understand the contribution of differential gene expression in the adaptation to HXT by this microorganism. The transcriptomic profile reveals an HXT-triggered antioxidant response involving genes from the ROS (reactive oxygen species) resistome of L. plantarum, genes coding for H2S-producing enzymes and genes involved in the response to thiol-specific oxidative stress. The expression of a set of genes involved in cell wall biogenesis was also upregulated, indicating that this subcellular compartment was a target of HXT. The expression of several MFS (major facilitator superfamily) efflux systems and ABC-transporters was differentially affected by HXT, probably to control its transport across the ...
Chronic hepatitis C virus (HCV) infection causes liver inflammation and fibrosis, which can lead ... more Chronic hepatitis C virus (HCV) infection causes liver inflammation and fibrosis, which can lead to development of cirrhosis and hepatocellular carcinoma (HCC). The recent approval of direct-acting antiviral (DAA) drug combinations has revolutionized antiviral therapy against HCV. These drugs enable virus eradication in virtually all treated patients regardless of their genotype and liver disease status. Based on clinical parameters, it has been proposed that elimination of infected cells by reactivated immune responses may be dispensable for virus eradication in contrast to previously used interferon-based therapies. It is thus formally possible that the patients, who are declared cured, do indeed carry formerly infected cells that display irreversible alterations due to prolonged chronic HCV infection. Although transcriptional profiles of biopsies from cured patients have been previously studied, it is difficult to determine the precise mechanisms by which permanent alterations ar...
Oleuropein (OLE) is a secoiridoid unique to Oleaceae known to play a role in the plant-herbivore ... more Oleuropein (OLE) is a secoiridoid unique to Oleaceae known to play a role in the plant-herbivore interaction. However, it is not clear how this molecule is induced to mediate plant responses to microbes and how microbes, in turn, withstand with OLE. To better understand how OLE affects the plant-microbe interaction, the contribution of differential gene expression in the adaptation to OLE was characterized by whole genome transcriptional profiling in Lactobacillus plantarum, a bacterium associated to the olive. OLE downregulated functions associated to rapid growth, remodeled membrane phospholipid biosynthesis pathways and markedly repressed the expression of several ABC transporters from L. plantarum. Genes encoding the plantaricin and lamABDCA quorum-sensing (QS) systems were down-regulated indicating the potential of OLE as a QS-antagonist. Notably, OLE diminished the expression of a set of genes encoding inmunomodulatory components and reoriented metabolic pathways to increase protein acetylation, probably to attenuate plant immunity. Responses were also triggered to repress the transport of acetoin and to buffer reactive oxygen species accumulation, two signals involved in plant development. The results suggest that OLE could act as a signaling molecule in the plant-microbe interaction and facilitate the accommodation of beneficial microbes such as L. plantarum by the plant host, via controlled expression of bacterial molecular players involved in this reciprocal interplay.
Background: Epigenetic phenomena are crucial for explaining the phenotypic plasticity seen in the... more Background: Epigenetic phenomena are crucial for explaining the phenotypic plasticity seen in the cells of different tissues, developmental stages and diseases, all holding the same DNA sequence. As technology is allowing to retrieve epigenetic information in a genome-wide fashion, massive epigenomic datasets are being accumulated in public repositories. New approaches are required to mine those data to extract useful knowledge. We present here an automatic approach for detecting genomic regions with epigenetic variation patterns across samples related to a grouping of these samples, as a way of detecting regions functionally associated to the phenomenon behind the classification. Results: We show that the regions automatically detected by the method in the whole human genome associated to three different classifications of a set of epigenomes (cancer vs. healthy, brain vs. other organs, and fetal vs. adult tissues) are enriched in genes associated to these processes. Conclusions: The method is fully automatic and can exhaustively scan the whole human genome at any resolution using large collections of epigenomes as input, although it also produces good results with small datasets. Consequently, it will be valuable for obtaining functional information from the incoming epigenomic information as it continues to accumulate.
Influenza A virus (IAV) infection can be severe or even lethal in toddlers, the elderly and patie... more Influenza A virus (IAV) infection can be severe or even lethal in toddlers, the elderly and patients with certain medical conditions. Infection of apparently healthy individuals nonetheless accounts for many severe disease cases and deaths, suggesting that viruses with increased pathogenicity co-circulate with pandemic or epidemic viruses. Looking for potential virulence factors, we have identified a polymerase PA D529N mutation detected in a fatal IAV case, whose introduction into two different recombinant virus backbones, led to reduced defective viral genomes (DVGs) production. This mutation conferred low induction of antiviral response in infected cells and increased pathogenesis in mice. To analyze the association between low DVGs production and pathogenesis in humans, we performed a genomic analysis of viruses isolated from a cohort of previously healthy individuals who suffered highly severe IAV infection requiring admission to Intensive Care Unit and patients with fatal outc...
Severe acute respiratory syndrome coronavirus (SARS-CoV) causes lethal disease in humans, which i... more Severe acute respiratory syndrome coronavirus (SARS-CoV) causes lethal disease in humans, which is characterized by exacerbated inflammatory response and extensive lung pathology. To address the relevance of small non-coding RNAs in SARS-CoV pathology, we deep sequenced RNAs from the lungs of infected mice and discovered three 18-22 nt small viral RNAs (svRNAs). The three svRNAs were derived from the nsp3 (svRNA-nsp3.1 and -nsp3.2) and N (svRNA-N) genomic regions of SARS-CoV. Biogenesis of CoV svRNAs was RNase III, cell type, and host species independent, but it was dependent on the extent of viral replication. Antagomir-mediated inhibition of svRNA-N significantly reduced in vivo lung pathology and pro-inflammatory cytokine expression. Taken together, these data indicate that svRNAs contribute to SARS-CoV pathogenesis and highlight the potential of svRNA-N antagomirs as antivirals.
Lower respiratory tract infections are among the top five leading causes of human death. Fighting... more Lower respiratory tract infections are among the top five leading causes of human death. Fighting these infections is therefore a world health priority. Searching for induced alterations in host gene expression shared by several relevant respiratory pathogens represents an alternative to identify new targets for wide-range host-oriented therapeutics. With this aim, alveolar macrophages were independently infected with three unrelated bacterial (Streptococcus pneumoniae, Klebsiella pneumoniae, and Staphylococcus aureus) and two dissimilar viral (respiratory syncytial virus and influenza A virus) respiratory pathogens, all of them highly relevant for human health. Cells were also activated with bacterial lipopolysaccharide (LPS) as a prototypical pathogen-associated molecular pattern. Patterns of differentially expressed cellular genes shared by the indicated pathogens were searched by microarray analysis. Most of the commonly up-regulated host genes were related to the innate immune ...
The CRISPR/Cas technology is enabling targeted genome editing in multiple organisms with unpreced... more The CRISPR/Cas technology is enabling targeted genome editing in multiple organisms with unprecedented accuracy and specificity by using RNAguided nucleases. A critical point when planning a CRISPR/Cas experiment is the design of the guide RNA (gRNA), which directs the nuclease and associated machinery to the desired genomic location. This gRNA has to fulfil the requirements of the nuclease and lack homology with other genome sites that could lead to off-target effects. Here we introduce the Breaking-Cas system for the design of gR-NAs for CRISPR/Cas experiments, including those based in the Cas9 nuclease as well as others recently introduced. The server has unique features not available in other tools, including the possibility of using all eukaryotic genomes available in ENSEMBL (currently around 700), placing variable PAM sequences at 5 or 3 and setting the guide RNA length and the scores per nucleotides. It can be freely accessed at: http://bioinfogp.cnb.csic.es/ tools/breakingcas, and the code is available upon request.
ABSTRACTInfluenza A viruses generate annual epidemics and occasional pandemics of respiratory dis... more ABSTRACTInfluenza A viruses generate annual epidemics and occasional pandemics of respiratory disease with important consequences for human health and the economy. Therefore, a large effort has been devoted to the development of new anti-influenza virus drugs directed to viral targets, as well as to the identification of cellular targets amenable to anti-influenza virus therapy. Here we have addressed the identification of such potential cellular targets by screening collections of drugs approved for human use. We reasoned that screening with a green fluorescent protein-based recombinant replicon system would identify cellular targets involved in virus transcription/replication and/or gene expression and hence address an early stage of virus infection. By using such a strategy, we identified Montelukast (MK) as an inhibitor of virus multiplication. MK inhibited virus gene expression but did not alter viral RNA synthesisin vitroor viral RNA accumulationin vivo. The low selectivity in...
Expression of the downstream regulatory element antagonist modulator (DREAM) protein in dorsal ro... more Expression of the downstream regulatory element antagonist modulator (DREAM) protein in dorsal root ganglia and spinal cord is related to endogenous control mechanisms of acute and chronic pain. In primary sensory trigeminal neurons high levels of endogenous DREAM protein are preferentially localized in the nucleus, suggesting a major transcriptional role. Here, we show that transgenic mice expressing a dominant active mutant of DREAM in trigeminal neurons show increased responses following orofacial sensory stimulation, which correlates with a decreased expression of prodynorphin and brain-derived neurotrophic factor (BDNF) in trigeminal ganglia. Genome-wide analysis of trigeminal neurons in daDREAM transgenic mice identified cathepsin L (CTSL) and the monoglyceride lipase (MGLL) as two new DREAM transcriptional targets related to pain. Our results suggest a role for DREAM in the regulation of trigeminal nociception. This article is protected by copyright. All rights reserved.
This study was aimed to gain new insights into the molecular mechanisms used by Lactobacillus pla... more This study was aimed to gain new insights into the molecular mechanisms used by Lactobacillus plantarum WCFS1 to respond to hydroxytyrosol (HXT), one of the main and health-relevant plant phenolics present in olive oil. To this goal, whole genome transcriptomic profiling was used to better understand the contribution of differential gene expression in the adaptation to HXT by this microorganism. The transcriptomic profile reveals an HXT-triggered antioxidant response involving genes from the ROS (reactive oxygen species) resistome of L. plantarum, genes coding for H2S-producing enzymes and genes involved in the response to thiol-specific oxidative stress. The expression of a set of genes involved in cell wall biogenesis was also upregulated, indicating that this subcellular compartment was a target of HXT. The expression of several MFS (major facilitator superfamily) efflux systems and ABC-transporters was differentially affected by HXT, probably to control its transport across the ...
Chronic hepatitis C virus (HCV) infection causes liver inflammation and fibrosis, which can lead ... more Chronic hepatitis C virus (HCV) infection causes liver inflammation and fibrosis, which can lead to development of cirrhosis and hepatocellular carcinoma (HCC). The recent approval of direct-acting antiviral (DAA) drug combinations has revolutionized antiviral therapy against HCV. These drugs enable virus eradication in virtually all treated patients regardless of their genotype and liver disease status. Based on clinical parameters, it has been proposed that elimination of infected cells by reactivated immune responses may be dispensable for virus eradication in contrast to previously used interferon-based therapies. It is thus formally possible that the patients, who are declared cured, do indeed carry formerly infected cells that display irreversible alterations due to prolonged chronic HCV infection. Although transcriptional profiles of biopsies from cured patients have been previously studied, it is difficult to determine the precise mechanisms by which permanent alterations ar...
Oleuropein (OLE) is a secoiridoid unique to Oleaceae known to play a role in the plant-herbivore ... more Oleuropein (OLE) is a secoiridoid unique to Oleaceae known to play a role in the plant-herbivore interaction. However, it is not clear how this molecule is induced to mediate plant responses to microbes and how microbes, in turn, withstand with OLE. To better understand how OLE affects the plant-microbe interaction, the contribution of differential gene expression in the adaptation to OLE was characterized by whole genome transcriptional profiling in Lactobacillus plantarum, a bacterium associated to the olive. OLE downregulated functions associated to rapid growth, remodeled membrane phospholipid biosynthesis pathways and markedly repressed the expression of several ABC transporters from L. plantarum. Genes encoding the plantaricin and lamABDCA quorum-sensing (QS) systems were down-regulated indicating the potential of OLE as a QS-antagonist. Notably, OLE diminished the expression of a set of genes encoding inmunomodulatory components and reoriented metabolic pathways to increase protein acetylation, probably to attenuate plant immunity. Responses were also triggered to repress the transport of acetoin and to buffer reactive oxygen species accumulation, two signals involved in plant development. The results suggest that OLE could act as a signaling molecule in the plant-microbe interaction and facilitate the accommodation of beneficial microbes such as L. plantarum by the plant host, via controlled expression of bacterial molecular players involved in this reciprocal interplay.
Background: Epigenetic phenomena are crucial for explaining the phenotypic plasticity seen in the... more Background: Epigenetic phenomena are crucial for explaining the phenotypic plasticity seen in the cells of different tissues, developmental stages and diseases, all holding the same DNA sequence. As technology is allowing to retrieve epigenetic information in a genome-wide fashion, massive epigenomic datasets are being accumulated in public repositories. New approaches are required to mine those data to extract useful knowledge. We present here an automatic approach for detecting genomic regions with epigenetic variation patterns across samples related to a grouping of these samples, as a way of detecting regions functionally associated to the phenomenon behind the classification. Results: We show that the regions automatically detected by the method in the whole human genome associated to three different classifications of a set of epigenomes (cancer vs. healthy, brain vs. other organs, and fetal vs. adult tissues) are enriched in genes associated to these processes. Conclusions: The method is fully automatic and can exhaustively scan the whole human genome at any resolution using large collections of epigenomes as input, although it also produces good results with small datasets. Consequently, it will be valuable for obtaining functional information from the incoming epigenomic information as it continues to accumulate.
Influenza A virus (IAV) infection can be severe or even lethal in toddlers, the elderly and patie... more Influenza A virus (IAV) infection can be severe or even lethal in toddlers, the elderly and patients with certain medical conditions. Infection of apparently healthy individuals nonetheless accounts for many severe disease cases and deaths, suggesting that viruses with increased pathogenicity co-circulate with pandemic or epidemic viruses. Looking for potential virulence factors, we have identified a polymerase PA D529N mutation detected in a fatal IAV case, whose introduction into two different recombinant virus backbones, led to reduced defective viral genomes (DVGs) production. This mutation conferred low induction of antiviral response in infected cells and increased pathogenesis in mice. To analyze the association between low DVGs production and pathogenesis in humans, we performed a genomic analysis of viruses isolated from a cohort of previously healthy individuals who suffered highly severe IAV infection requiring admission to Intensive Care Unit and patients with fatal outc...
Severe acute respiratory syndrome coronavirus (SARS-CoV) causes lethal disease in humans, which i... more Severe acute respiratory syndrome coronavirus (SARS-CoV) causes lethal disease in humans, which is characterized by exacerbated inflammatory response and extensive lung pathology. To address the relevance of small non-coding RNAs in SARS-CoV pathology, we deep sequenced RNAs from the lungs of infected mice and discovered three 18-22 nt small viral RNAs (svRNAs). The three svRNAs were derived from the nsp3 (svRNA-nsp3.1 and -nsp3.2) and N (svRNA-N) genomic regions of SARS-CoV. Biogenesis of CoV svRNAs was RNase III, cell type, and host species independent, but it was dependent on the extent of viral replication. Antagomir-mediated inhibition of svRNA-N significantly reduced in vivo lung pathology and pro-inflammatory cytokine expression. Taken together, these data indicate that svRNAs contribute to SARS-CoV pathogenesis and highlight the potential of svRNA-N antagomirs as antivirals.
Lower respiratory tract infections are among the top five leading causes of human death. Fighting... more Lower respiratory tract infections are among the top five leading causes of human death. Fighting these infections is therefore a world health priority. Searching for induced alterations in host gene expression shared by several relevant respiratory pathogens represents an alternative to identify new targets for wide-range host-oriented therapeutics. With this aim, alveolar macrophages were independently infected with three unrelated bacterial (Streptococcus pneumoniae, Klebsiella pneumoniae, and Staphylococcus aureus) and two dissimilar viral (respiratory syncytial virus and influenza A virus) respiratory pathogens, all of them highly relevant for human health. Cells were also activated with bacterial lipopolysaccharide (LPS) as a prototypical pathogen-associated molecular pattern. Patterns of differentially expressed cellular genes shared by the indicated pathogens were searched by microarray analysis. Most of the commonly up-regulated host genes were related to the innate immune ...
The CRISPR/Cas technology is enabling targeted genome editing in multiple organisms with unpreced... more The CRISPR/Cas technology is enabling targeted genome editing in multiple organisms with unprecedented accuracy and specificity by using RNAguided nucleases. A critical point when planning a CRISPR/Cas experiment is the design of the guide RNA (gRNA), which directs the nuclease and associated machinery to the desired genomic location. This gRNA has to fulfil the requirements of the nuclease and lack homology with other genome sites that could lead to off-target effects. Here we introduce the Breaking-Cas system for the design of gR-NAs for CRISPR/Cas experiments, including those based in the Cas9 nuclease as well as others recently introduced. The server has unique features not available in other tools, including the possibility of using all eukaryotic genomes available in ENSEMBL (currently around 700), placing variable PAM sequences at 5 or 3 and setting the guide RNA length and the scores per nucleotides. It can be freely accessed at: http://bioinfogp.cnb.csic.es/ tools/breakingcas, and the code is available upon request.
ABSTRACTInfluenza A viruses generate annual epidemics and occasional pandemics of respiratory dis... more ABSTRACTInfluenza A viruses generate annual epidemics and occasional pandemics of respiratory disease with important consequences for human health and the economy. Therefore, a large effort has been devoted to the development of new anti-influenza virus drugs directed to viral targets, as well as to the identification of cellular targets amenable to anti-influenza virus therapy. Here we have addressed the identification of such potential cellular targets by screening collections of drugs approved for human use. We reasoned that screening with a green fluorescent protein-based recombinant replicon system would identify cellular targets involved in virus transcription/replication and/or gene expression and hence address an early stage of virus infection. By using such a strategy, we identified Montelukast (MK) as an inhibitor of virus multiplication. MK inhibited virus gene expression but did not alter viral RNA synthesisin vitroor viral RNA accumulationin vivo. The low selectivity in...
Expression of the downstream regulatory element antagonist modulator (DREAM) protein in dorsal ro... more Expression of the downstream regulatory element antagonist modulator (DREAM) protein in dorsal root ganglia and spinal cord is related to endogenous control mechanisms of acute and chronic pain. In primary sensory trigeminal neurons high levels of endogenous DREAM protein are preferentially localized in the nucleus, suggesting a major transcriptional role. Here, we show that transgenic mice expressing a dominant active mutant of DREAM in trigeminal neurons show increased responses following orofacial sensory stimulation, which correlates with a decreased expression of prodynorphin and brain-derived neurotrophic factor (BDNF) in trigeminal ganglia. Genome-wide analysis of trigeminal neurons in daDREAM transgenic mice identified cathepsin L (CTSL) and the monoglyceride lipase (MGLL) as two new DREAM transcriptional targets related to pain. Our results suggest a role for DREAM in the regulation of trigeminal nociception. This article is protected by copyright. All rights reserved.
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