scholarly journals In Vivo Transcriptome of Lactobacillus acidophilus and Colonization Impact on Murine Host Intestinal Gene Expression

mBio ◽  
2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yong Jun Goh ◽  
Rodolphe Barrangou ◽  
Todd R. Klaenhammer
Keyword(s):  
Gene Expression ◽  
Lactobacillus Acidophilus ◽  
Expression Patterns ◽  
Transcriptional Response ◽  
Nutrient Acquisition ◽  
Content Type ◽  
Host Interactions ◽  
Core Genes

ABSTRACT Lactobacillus acidophilus NCFM is a probiotic strain commonly used in dairy products and dietary supplements. Postgenome in vitro studies of NCFM thus far have linked potential key genotypes to its probiotic-relevant attributes, including gut survival, prebiotic utilization, host interactions, and immunomodulatory activities. To corroborate and extend beyond previous in vivo and in vitro functional studies, we employed a dual RNA sequencing (RNA-seq) transcriptomic approach to identify genes potentially driving the gut fitness and activities of L. acidophilus NCFM in vivo, and in parallel, examine the ileal transcriptional response of its murine hosts during monocolonization. Spatial expression profiling of NCFM from the ileum through the colon revealed a set of 134 core genes that were consistently overexpressed during gut transit. These in vivo core genes are predominantly involved in the metabolism of carbohydrates, amino acids, and nucleotides, along with mucus-binding proteins and adhesion factors, confirming their functionally important roles in nutrient acquisition and gut retention. Functional characterization of the highly expressed major S-layer-encoding gene established its indispensable role as a cell shape determinant and maintenance of cell surface integrity, essential for viability and probiotic attributes. Host colonization by L. acidophilus resulted in significant downregulation of several proinflammatory cytokines and tight junction proteins. Genes related to redox signaling, mucin glycosylation, and circadian rhythm modulation were induced, suggesting impacts on intestinal development and immune functions. Metagenomic analysis of NCFM populations postcolonization demonstrated the genomic stability of L. acidophilus as a gut transient and further established its safety as a probiotic and biotherapeutic delivery platform. IMPORTANCE To date, our basis for comprehending the probiotic mechanisms of Lactobacillus acidophilus, one of the most widely consumed probiotic microbes, was largely limited to in vitro functional genomic studies. Using a germfree murine colonization model, in vivo-based transcriptional studies provided the first view of how L. acidophilus survives in the mammalian gut environment, including gene expression patterns linked to survival, efficient nutrient acquisition, stress adaptation, and host interactions. Examination of the host ileal transcriptional response, the primary effector site of L. acidophilus, has also shed light into the mechanistic roles of this probiotic microbe in promoting anti-inflammatory responses, maintaining intestinal epithelial homeostasis and modulation of the circadian-metabolic axis in its host.

scholarly journals DifferentialIn VivoGene Expression of Major Leptospira Proteins in Resistant or Susceptible Animal Models

2012 ◽  
Vol 78 (17) ◽  
pp. 6372-6376 ◽  
Author(s):  
Mariko Matsui ◽  
Marie-Estelle Soupé ◽  
Jérôme Becam ◽  
Cyrille Goarant
Keyword(s):  
Gene Expression ◽  
Animal Models ◽  
16S Rrna ◽  
Virulent Strain ◽  
Expression Patterns ◽  
Susceptible Animal ◽  
Content Type ◽  
Its Gene

ABSTRACTTranscripts ofLeptospira16S rRNA, FlaB, LigB, LipL21, LipL32, LipL36, LipL41, and OmpL37 were quantified in the blood of susceptible (hamsters) and resistant (mice) animal models of leptospirosis. We first validated adequate reference genes and then evaluated expression patternsin vivocompared toin vitrocultures. LipL32 expression was downregulatedin vivoand differentially regulated in resistant and susceptible animals. FlaB expression was also repressed in mice but not in hamsters. In contrast, LigB and OmpL37 were upregulatedin vivo. Thus, we demonstrated that a virulent strain ofLeptospiradifferentially adapts its gene expression in the blood of infected animals.

scholarly journals Regulation of SpeB in Streptococcus pyogenes by pH and NaCl: a Model for In Vivo Gene Expression

2006 ◽  
Vol 188 (2) ◽  
pp. 399-408 ◽  
Author(s):  
Jennifer A. Loughman ◽  
Michael Caparon
Keyword(s):  
Gene Expression ◽  
Streptococcus Pyogenes ◽  
Growth Phase ◽  
Soft Tissues ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Transcriptional Response ◽  
Culture Conditions

ABSTRACT For a pathogen such as Streptococcus pyogenes, ecological success is determined by its ability to sense the environment and mount an appropriate adaptive transcriptional response. Thus, determining conditions for analyses of gene expression in vitro that are representative of the in vivo environment is critical for understanding the contributions of transcriptional response pathways to pathogenesis. In this study, we determined that the gene encoding the SpeB cysteine protease is up-regulated over the course of infection in a murine soft-tissue model. Conditions were identified, including growth phase, acidic pH, and an NaCl concentration of <0.1 M, that were required for expression of speB in vitro. Analysis of global expression profiles in response to these conditions in vitro identified a set of coregulated genes whose expression patterns showed a significant correlation with that of speB when examined during infection of murine soft tissues. This analysis revealed that a culture medium that promotes high levels of SpeB expression in vitro produced an expression profile that showed significant correlation to the profile observed in vivo. Taken together, these studies establish culture conditions that mimic in vivo expression patterns; that growth phase, pH, and NaCl may mimic relevant cues sensed by S. pyogenes during infection; and that identification of other environmental cues that alter expression of speB in vitro may provide insight into the signals that direct global patterns of gene expression in vivo.

scholarly journals Arsenic Directly Binds to and Activates the Yeast AP-1-Like Transcription Factor Yap8

2015 ◽  
Vol 36 (6) ◽  
pp. 913-922 ◽  
Author(s):  
Nallani Vijay Kumar ◽  
Jianbo Yang ◽  
Jitesh K. Pillai ◽  
Swati Rawat ◽  
Carlos Solano ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Transcriptional Activation ◽  
Transcriptional Response ◽  
Transcriptional Regulator ◽  
Yeast Protein ◽  
Content Type ◽  
Arsenic Tolerance ◽  
Arsenic Sensing

The AP-1-like transcription factor Yap8 is critical for arsenic tolerance in the yeastSaccharomyces cerevisiae. However, the mechanism by which Yap8 senses the presence of arsenic and activates transcription of detoxification genes is unknown. Here we demonstrate that Yap8 directly binds to trivalent arsenite [As(III)]in vitroandin vivoand that approximately one As(III) molecule is bound per molecule of Yap8. As(III) is coordinated by three sulfur atoms in purified Yap8, and our genetic and biochemical data identify the cysteine residues that form the binding site as Cys132, Cys137, and Cys274. As(III) binding by Yap8 does not require an additional yeast protein, and Yap8 is regulated neither at the level of localization nor at the level of DNA binding. Instead, our data are consistent with a model in which a DNA-bound form of Yap8 acts directly as an As(III) sensor. Binding of As(III) to Yap8 triggers a conformational change that in turn brings about a transcriptional response. Thus, As(III) binding to Yap8 acts as a molecular switch that converts inactive Yap8 into an active transcriptional regulator. This is the first report to demonstrate how a eukaryotic protein couples arsenic sensing to transcriptional activation.

scholarly journals Synthetic gene regulatory networks in the opportunistic human pathogen Streptococcus pneumoniae

2019 ◽  
Author(s):  
Robin A. Sorg ◽  
Clement Gallay ◽  
Jan-Willem Veening
Keyword(s):  
Gene Expression ◽  
Streptococcus Pneumoniae ◽  
Regulatory Networks ◽  
Expression Patterns ◽  
Combinatorial Libraries ◽  
Regulatory Elements ◽  
Expression Control ◽  
Gene Expression Control

AbstractStreptococcus pneumoniae can cause disease in various human tissues and organs, including the ear, the brain, the blood and the lung, and thus in highly diverse and dynamic environments. It is challenging to study how pneumococci control virulence factor expression, because cues of natural environments and the presence of an immune system are difficult to simulate in vitro. Here, we apply synthetic biology methods to reverse-engineer gene expression control in S. pneumoniae. A selection platform is described that allows for straightforward identification of transcriptional regulatory elements out of combinatorial libraries. We present TetR- and LacI-regulated promoters that show expression ranges of four orders of magnitude. Based on these promoters, regulatory networks of higher complexity are assembled, such as logic AND and IMPLY gates. Finally, we demonstrate single-copy genome-integrated toggle switches that give rise to bimodal population distributions. The tools described here can be used to mimic complex expression patterns, such as the ones found for pneumococcal virulence factors, paving the way for in vivo investigations of the importance of gene expression control on the pathogenicity of S. pneumoniae.

68 GENE EXPRESSION COMPARISONS BETWEEN BOVINE IN VIVO AND CLONED EMBRYOS PRODUCED BY THREE DIFFERENT METHODS

2006 ◽  
Vol 18 (2) ◽  
pp. 142
Author(s):  
N. Ruddock ◽  
K. Wilson ◽  
M. Cooney ◽  
R. Tecirlioglu ◽  
V. Hall ◽  
...  
Keyword(s):  
Gene Expression ◽  
Chromatin Remodeling ◽  
Nuclear Transfer ◽  
Expression Patterns ◽  
Experimental Models ◽  
Gene Expression Patterns ◽  
Imprinted Genes ◽  
Mammalian Embryo

Developmental pathways in the mammalian embryo are profoundly influenced by the epigenetic interaction of the environment and the genome. Loss of epigenetic control has been implicated in aberrant gene expression and altered imprinting patterns with consequence to the physiology and viability of the conceptus. Bovine somatic cell nuclear transfer (SCNT) is contingent on in vitro culture, and both SCNT and culture conditions are known to induce changes in embryonic gene expression patterns. Using these experimental models, this study compared gene expression of Day 7 cloned blastocysts created from three different SCNT protocols using the same cell line, with Day 7 in vivo blastocysts to elucidate mechanisms responsible for variations in phenotypic outcomes. SCNT methods included: (1) traditional SCNT by subzonal injection (SI); (2) handmade cloning (HMC); and (3) modified serial nuclear transfer (SNT), developed within the group. Four imprinted genes (Grb10, Ndn, Nnat, and Ube3a), four chromatin remodeling genes (Cbx1, Cbx3, Smarca4, and Smarcb1) and two genes implicated in polycystic liver disease (Prkcsh and Sec63) were analyzed in single blastocysts from each treatment (n = 5). All blastocysts expressed Actin, Oct-4 and Ifn-tau. All genes were sequence verified. Several genes were expressed ubiquitously across all groups, including Ndn, Ube3a, Cbx1, Cbx3, and Smarcb1. Interestingly, Grb10 was not expressed in two HMCs and one SNT blastocyst. Nnat was weakly expressed in one in vivo blastocyst and in the majority of cloned blastocysts in all groups. Prkcsh and Sec63 were expressed in all but one HMC blastocyst. While gene expression patterns were mostly maintained following SCNT, the imprinted genes Nnat and Grb10 showed instances of differential or abnormal expression in SCNT embryos. The chromatin remodeling genes were maintained in all SCNT treatments. Prkcsh and Sec63 were both absent in one HMC blastocyst, with implications for liver dysfunction, a condition previously reported in abnormal cloned offspring. The variable mRNA expression following SCNT provides an insight into genetic and environmental factors controlling implantation, placentation, organ formation, and fetal growth.

scholarly journals 18β-Glycyrrhetinic Acid Inhibits Methicillin-Resistant Staphylococcus aureus Survival and Attenuates Virulence Gene Expression

2012 ◽  
Vol 57 (1) ◽  
pp. 241-247 ◽  
Author(s):  
Danyelle R. Long ◽  
Julia Mead ◽  
Jay M. Hendricks ◽  
Michele E. Hardy ◽  
Jovanka M. Voyich
Keyword(s):  
Gene Expression ◽  
Staphylococcus Aureus ◽  
Virulence Gene ◽  
Glycyrrhetinic Acid ◽  
Licorice Root ◽  
Methicillin Resistant ◽  
Content Type ◽  
Virulence Gene Expression

ABSTRACTMethicillin-resistantStaphylococcus aureus(MRSA) has become a major source of infection in hospitals and in the community. Increasing antibiotic resistance inS. aureusstrains has created a need for alternative therapies to treat disease. A component of the licorice rootGlycyrrhizaspp., 18β-glycyrrhetinic acid (GRA), has been shown to have antiviral, antitumor, and antibacterial activity. This investigation explores thein vitroandin vivoeffects of GRA on MRSA pulsed-field gel electrophoresis (PFGE) type USA300. GRA exhibited bactericidal activity at concentrations exceeding 0.223 μM. Upon exposure ofS. aureusto sublytic concentrations of GRA, we observed a reduction in expression of key virulence genes, includingsaeRandhla. In murine models of skin and soft tissue infection, topical GRA treatment significantly reduced skin lesion size and decreased the expression ofsaeRandhlagenes. Our investigation demonstrates that at high concentrations GRA is bactericidal to MRSA and at sublethal doses it reduces virulence gene expression inS. aureusbothin vitroandin vivo.

scholarly journals Adaptive Strategies and Pathogenesis of Clostridium difficile fromIn VivoTranscriptomics

2013 ◽  
Vol 81 (10) ◽  
pp. 3757-3769 ◽  
Author(s):  
Claire Janoir ◽  
Cécile Denève ◽  
Sylvie Bouttier ◽  
Frédéric Barbut ◽  
Sandra Hoys ◽  
...  
Keyword(s):  
Clostridium Difficile ◽  
Stress Responses ◽  
Differentially Expressed ◽  
Content Type ◽  
Host Interactions ◽  
Colonization Factor ◽  
Genes Encoding ◽  
Colonization Process

ABSTRACTClostridium difficileis currently the major cause of nosocomial intestinal diseases associated with antibiotic therapy in adults. In order to improve our knowledge ofC. difficile-host interactions, we analyzed the genome-wide temporal expression ofC. difficile630 genes during the first 38 h of mouse colonization to identify genes whose expression is modulatedin vivo, suggesting that they may play a role in facilitating the colonization process. In the ceca of theC. difficile-monoassociated mice, 549 genes of theC. difficilegenome were differentially expressed compared to their expression duringin vitrogrowth, and they were distributed in several functional categories. Overall, our results emphasize the roles of genes involved in host adaptation. Colonization results in a metabolic shift, with genes responsible for the fermentation as well as several other metabolic pathways being regulated inversely to those involved in carbon metabolism. In addition, several genes involved in stress responses, such as ferrous iron uptake or the response to oxidative stress, were regulatedin vivo. Interestingly, many genes encoding conserved hypothetical proteins (CHP) were highly and specifically upregulatedin vivo. Moreover, genes for all stages of sporulation were quickly inducedin vivo, highlighting the observation that sporulation is central to the persistence ofC. difficilein the gut and to its ability to spread in the environment. Finally, we inactivated two genes that were differentially expressedin vivoand evaluated the relative colonization fitness of the wild-type and mutant strains in coinfection experiments. We identified a CHP as a putative colonization factor, supporting the suggestion that thein vivotranscriptomic approach can unravel newC. difficilevirulence genes.

scholarly journals Developmental and molecular correlates of bovine preimplantation embryos

Reproduction ◽  
2006 ◽  
Vol 131 (5) ◽  
pp. 895-904 ◽  
Author(s):  
Hakan Sagirkaya ◽  
Muge Misirlioglu ◽  
Abdullah Kaya ◽  
Neal L First ◽  
John J Parrish ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methyltransferase ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Blastocyst Stage ◽  
Culture Conditions ◽  
Preimplantation Embryos ◽  
Developmental Potential

Expression of embryonic genes is altered in different culture conditions, which influence developmental potential both during preimplantation and fetal development. The objective of this study was to define the effects of culture conditions on: bovine embryonic development to blastocyst stage, blastocyst cell number, apoptosis and expression patterns of a panel of developmentally important genes. Bovine embryos were culturedin vitroin three culture media containing amino acids, namely potassium simplex optimization medium (KSOMaa), Charles Rosenkrans 1 (CR1aa) and synthetic oviductal fluid (SOFaa). Apoptosis in blastocysts was determined by TUNEL assay and expression profiles of developmentally important genes were assayed by real-time PCR.In vivo-produced bovine blastocysts were used as controls for experiments determining gene expression patterns. While the cleavage rates did not differ, embryos cultured in SOFaa had higher rates of development to blastocyst stage (P< 0.05). Mean cell numbers and percentages of apoptotic cells per blastocyst did not differ among the groups. Expression of the heat shock protein 70 (Hsp70) gene was significantly up-regulated in both CR1aa and KSOMaa when compared with SOFaa (P< 0.001). DNA methyltransferase 3a (Dnmt3a) expression was higher in embryos cultured in CR1aa than in those cultured in SOFaa (P< 0.001). Expression of interferon tau (IF-τ) and insulin-like growth factor II receptor (Igf-2r) genes was significantly up-regulated in KSOMaa when compared with CR1aa (P< 0.001). Gene expression did not differ betweenin vivo-derived blastocysts and theirin vitro-derived counterparts. In conclusion, SOFaa supports higher development to blastocyst stage than KSOMaa and CR1aa, and the culture conditions influence gene expression.

scholarly journals Recapitulation of Transcriptomic Characteristics of Primary Breast Tumours in Patient-Derived 3D Cultures in Vitro

2021 ◽  
Vol 75 (1) ◽  
pp. 20-24
Author(s):  
Dina Nitiša ◽  
Nityanand Jain ◽  
Arvīds Irmejs ◽  
Valdis Pirsko ◽  
Inese Čakstiņa
Keyword(s):  
Gene Expression ◽  
Quantitative Polymerase Chain Reaction ◽  
Expression Patterns ◽  
3D Cell Culture ◽  
Model System ◽  
Gene Expression Patterns ◽  
3D Cultures ◽  
Culture Development

AbstractBreast cancer (BC) is the most common cause of cancer-related deaths among women in Europe and worldwide. Adherent (2D) cell cultures have been the routine in vitro model system employed in preclinical BC research for the last half-century. Over the past decade, new protocols have been developed allowing patient-derived three-dimensional organoid (3D) cell culture development from a range of solid tumours, including BC. These 3D models offer a promise of closer resemblance to the native tumour than the 2D cultures. To test the assumption that an in vitro 3D BC model system provides increased faithfulness to the molecular processes happening in vivo, as compared to 2D BC cultures, post-operational material from three BC patients was used to simultaneously develop 2D and 3D cultures in vitro. When analysed by quantitative polymerase chain reaction (PCR), the gene expression patterns of the cells from 3D cultures resembled the original tissues, while the gene expression patterns of the conventional 2D cultures were more distant.

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