scholarly journals In vivo transcriptome analysis provides insights into host-dependent expression of virulence factors by Yersinia entomophaga MH96, during infection of Galleria mellonella

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Amber R Paulson ◽  
Maureen O’Callaghan ◽  
Xue-Xian Zhang ◽  
Paul B Rainey ◽  
Mark R H Hurst
Keyword(s):  
Galleria Mellonella ◽  
Functional Enrichment ◽  
Natural Environments ◽  
Insecticidal Toxin ◽  
Heat Stable ◽  
Expression Of Genes ◽  
Genes Encoding ◽  
Cell Densities

Abstract The function of microbes can be inferred from knowledge of genes specifically expressed in natural environments. Here, we report the in vivo transcriptome of the entomopathogenic bacterium Yersinia entomophaga MH96, captured during initial, septicemic, and pre-cadaveric stages of intrahemocoelic infection in Galleria mellonella. A total of 1285 genes were significantly upregulated by MH96 during infection; 829 genes responded to in vivo conditions during at least one stage of infection, 289 responded during two stages of infection, and 167 transcripts responded throughout all three stages of infection compared to in vitro conditions at equivalent cell densities. Genes upregulated during the earliest infection stage included components of the insecticidal toxin complex Yen-TC (chi1, chi2, and yenC1), genes for rearrangement hotspot element containing protein yenC3, cytolethal distending toxin cdtAB, and vegetative insecticidal toxin vip2. Genes more highly expressed throughout the infection cycle included the putative heat-stable enterotoxin yenT and three adhesins (usher-chaperone fimbria, filamentous hemagglutinin, and an AidA-like secreted adhesin). Clustering and functional enrichment of gene expression data also revealed expression of genes encoding type III and VI secretion system-associated effectors. Together these data provide insight into the pathobiology of MH96 and serve as an important resource supporting efforts to identify novel insecticidal agents.

scholarly journals In vivo transcriptome analysis provides insights into host-dependent expression of virulence factors by Yersinia entomophaga MH96, during infection of Galleria mellonella

2020 ◽  
Author(s):  
Amber R. Paulson ◽  
Maureen O’Callaghan ◽  
Xue-Xian Zhang ◽  
Paul B. Rainey ◽  
Mark R.H. Hurst
Keyword(s):  
Galleria Mellonella ◽  
Functional Enrichment ◽  
Natural Environments ◽  
Insecticidal Toxin ◽  
Heat Stable ◽  
Expression Of Genes ◽  
Genes Encoding ◽  
Three Stages ◽  
Two Stages

AbstractThe function of microbes can be inferred from knowledge of genes specifically expressed in natural environments. Here we report the in vivo transcriptome of the entomopathogenic bacterium Yersinia entomophaga MH96, captured during three stages of intrahemocoelic infection in Galleria mellonella. A total of 1,285 genes were significantly upregulated by MH96 during infection; 829 genes responded to in vivo conditions during at least one stage of infection, 289 responded during two stages of infection and 167 transcripts responded throughout all three stages of infection. Genes upregulated during the earliest infection stage included components of the insecticidal toxin complex Yen-TC (chi1, chi2 and yenC1), genes for rearrangement hotspot element containing protein yenC3, cytolethal distending toxin cdtAB and vegetative insecticidal toxin vip2. Genes more highly expressed throughout the infection cycle included the putative heat-stable enterotoxin yenT and three adhesins (usher-chaperone fimbria, filamentous hemagglutinin and an AidA-like secreted adhesin). Clustering and functional enrichment of gene expression data also revealed expression of genes encoding type III and VI secretion system-associated effectors. Together these data provide insight into the pathobiology of MH96 and serve as an important resource supporting efforts to identify novel insecticidal agents.

scholarly journals Designing P. aeruginosa synthetic phages with reduced genomes

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Diana P. Pires ◽  
Rodrigo Monteiro ◽  
Dalila Mil-Homens ◽  
Arsénio Fialho ◽  
Timothy K. Lu ◽  
...  
Keyword(s):  
Galleria Mellonella ◽  
Antibacterial Properties ◽  
Genetically Engineered ◽  
In Vivo Studies ◽  
Infection Model ◽  
Promising Therapeutic Approach ◽  
Genes Encoding ◽  
First Time

AbstractIn the era where antibiotic resistance is considered one of the major worldwide concerns, bacteriophages have emerged as a promising therapeutic approach to deal with this problem. Genetically engineered bacteriophages can enable enhanced anti-bacterial functionalities, but require cloning additional genes into the phage genomes, which might be challenging due to the DNA encapsulation capacity of a phage. To tackle this issue, we designed and assembled for the first time synthetic phages with smaller genomes by knocking out up to 48% of the genes encoding hypothetical proteins from the genome of the newly isolated Pseudomonas aeruginosa phage vB_PaeP_PE3. The antibacterial efficacy of the wild-type and the synthetic phages was assessed in vitro as well as in vivo using a Galleria mellonella infection model. Overall, both in vitro and in vivo studies revealed that the knock-outs made in phage genome do not impair the antibacterial properties of the synthetic phages, indicating that this could be a good strategy to clear space from phage genomes in order to enable the introduction of other genes of interest that can potentiate the future treatment of P. aeruginosa infections.

THE PRODUCTION BY BACILLUS THURINGIENSIS BERLINER OF A HEAT-STABLE SUBSTANCE TOXIC FOR INSECTS

1959 ◽  
Vol 5 (2) ◽  
pp. 161-168 ◽  
Author(s):  
Ellicott McConnell ◽  
A. Glenn Richards
Keyword(s):  
Bacillus Thuringiensis ◽  
Inclusion Bodies ◽  
Galleria Mellonella ◽  
Culture Media ◽  
Water Soluble ◽  
Broth Culture ◽  
Toxic Principle ◽  
Heat Stable

Bacillus thuringiensis Berliner produces in vitro a heat-stable, dialyzable substance which is toxic for insects when injected. The same or a similar substance is produced in vivo. The toxic principle is of unknown composition. It is heat-stable, water-soluble, dialyzable, and resistant to low temperatures. It is probably neither a protein nor a lipid. Clearly it is distinct from the heat-labile inclusion bodies and from lecithinase. Growth-curve studies showed that the heat-stable toxin appeared in liver broth cultures during the active growth phase, prior to the formation of spores or inclusion bodies. An attempt to produce the toxic principle from culture media in the absence of bacteria was unsuccessful from sterile inocula both from in vivo and in vitro sources. The LD50 for larvae of Galleria mellonella injected with autoclaved supernatant from a 10-day-old liver broth culture of B. thuringiensis was determined to be 0.00036 ml per larva or 0.002 ml per gram of larvae. Approximately the same level of toxicity was found for another caterpillar, a fly larva, and cockroaches. After larvae of Galleria or Pyrausla have been dead for more than 2 days from infection with B. thuringiensis the bacillus could no longer be recovered. A sublethal amount of the heat-stable toxin injected into old larvae of Galleria delayed emergence of the adults by 30 to 40%. The non-pathogenic Bacillus cereus was found to produce a similar-acting, heat-stable toxin under the same conditions that one is produced by B. thuringiensis.

scholarly journals Nrg1 Is a Transcriptional Repressor for Glucose Repression of STA1 Gene Expression inSaccharomyces cerevisiae

1999 ◽  
Vol 19 (3) ◽  
pp. 2044-2050 ◽  
Author(s):  
Seok Hee Park ◽  
Sang Seok Koh ◽  
Jae Hwan Chun ◽  
Hye Jin Hwang ◽  
Hyen Sam Kang
Keyword(s):  
Gene Expression ◽  
Zinc Finger Protein ◽  
Glucose Repression ◽  
Transcriptional Repressor ◽  
Dependent Manner ◽  
Expression Of Genes ◽  
Dnase I Footprinting ◽  
Genes Encoding

ABSTRACT Expression of genes encoding starch-degrading enzymes is regulated by glucose repression in the yeast Saccharomyces cerevisiae. We have identified a transcriptional repressor, Nrg1, in a genetic screen designed to reveal negative factors involved in the expression of STA1, which encodes a glucoamylase. TheNRG1 gene encodes a 25-kDa C2H2zinc finger protein which specifically binds to two regions in the upstream activation sequence of the STA1 gene, as judged by gel retardation and DNase I footprinting analyses. Disruption of theNRG1 gene causes a fivefold increase in the level of theSTA1 transcript in the presence of glucose. The expression of NRG1 itself is inhibited in the absence of glucose. DNA-bound LexA-Nrg1 represses transcription of a target gene 10.7-fold in a glucose-dependent manner, and this repression is abolished in bothssn6 and tup1 mutants. Two-hybrid and glutathione S-transferase pull-down experiments show an interaction of Nrg1 with Ssn6 both in vivo and in vitro. These findings indicate that Nrg1 acts as a DNA-binding repressor and mediates glucose repression of the STA1 gene expression by recruiting the Ssn6-Tup1 complex.

scholarly journals YtxR, a Conserved LysR-Like Regulator That Induces Expression of Genes Encoding a Putative ADP-Ribosyltransferase Toxin Homologue in Yersinia enterocolitica

2006 ◽  
Vol 188 (23) ◽  
pp. 8033-8043 ◽  
Author(s):  
Grace L. Axler-DiPerte ◽  
Virginia L. Miller ◽  
Andrew J. Darwin
Keyword(s):  
Yersinia Enterocolitica ◽  
Transcription Initiation ◽  
Photorhabdus Luminescens ◽  
Expression Of Genes ◽  
Dnase I Footprinting ◽  
Genes Encoding ◽  
Adp Ribosyltransferase ◽  
Insight Into

ABSTRACT Yersinia enterocolitica causes human gastroenteritis, and many isolates have been classified as either “American” or “non-American” strains based on their geographic prevalence and virulence properties. In this study we describe identification of a transcriptional regulator that controls expression of the Y. enterocolitica ytxAB genes. The ytxAB genes have the potential to encode an ADP-ribosylating toxin with similarity to pertussis toxin. However, a ytxAB null mutation did not affect virulence in mice. Nevertheless, the ytxAB genes are conserved in many Y. enterocolitica strains. Interestingly, American and non-American strains have different ytxAB alleles encoding proteins that are only 50 to 60% identical. To obtain further insight into the ytxAB locus, we investigated whether it is regulated as part of a known or novel regulon. Transposon mutagenesis identified a LysR-like regulator, which we designated YtxR. Expression of ytxR from a nonnative promoter increased Φ(ytxA-lacZ) operon fusion expression up to 35-fold. YtxR also activated expression of its own promoter. DNase I footprinting showed that a His6-YtxR fusion protein directly interacted with the ytxA and ytxR control regions at similar distances upstream of their probable transcription initiation sites, identified by primer extension. Deletion analysis demonstrated that removal of the regions protected by His6-YtxR in vitro eliminated YtxR-dependent induction in vivo. The ytxAB locus is not present in most Yersinia species. In contrast, ytxR is conserved in multiple Yersinia species, as well as in the closely related organisms Photorhabdus luminescens and Photorhabdus asymbiotica. These observations suggest that YtxR may play a conserved role involving regulation of other genes besides ytxAB.

scholarly journals RhoGDIα-dependent balance between RhoA and RhoC is a key regulator of cancer cell tumorigenesis

2011 ◽  
Vol 22 (17) ◽  
pp. 3263-3275 ◽  
Author(s):  
T. T. Giang Ho ◽  
Audrey Stultiens ◽  
Johanne Dubail ◽  
Charles M. Lapière ◽  
Betty V. Nusgens ◽  
...  
Keyword(s):  
Tumor Growth ◽  
Cancer Progression ◽  
Dynamic Balance ◽  
Cellular Functions ◽  
Expression Of Genes ◽  
Genes Encoding ◽  
Interfering Rna

RhoGTPases are key signaling molecules regulating main cellular functions such as migration, proliferation, survival, and gene expression through interactions with various effectors. Within the RhoA-related subclass, RhoA and RhoC contribute to several steps of tumor growth, and the regulation of their expression affects cancer progression. Our aim is to investigate their respective contributions to the acquisition of an invasive phenotype by using models of reduced or forced expression. The silencing of RhoC, but not of RhoA, increased the expression of genes encoding tumor suppressors, such as nonsteroidal anti-inflammatory drug–activated gene 1 (NAG-1), and decreased migration and the anchorage-independent growth in vitro. In vivo, RhoC small interfering RNA (siRhoC) impaired tumor growth. Of interest, the simultaneous knockdown of RhoC and NAG-1 repressed most of the siRhoC-related effects, demonstrating the central role of NAG-1. In addition of being induced by RhoC silencing, NAG-1 was also largely up-regulated in cells overexpressing RhoA. The silencing of RhoGDP dissociation inhibitor α (RhoGDIα) and the overexpression of a RhoA mutant unable to bind RhoGDIα suggested that the effect of RhoC silencing is indirect and results from the up-regulation of the RhoA level through competition for RhoGDIα. This study demonstrates the dynamic balance inside the RhoGTPase network and illustrates its biological relevance in cancer progression.

scholarly journals Stigmasterol blocks cartilage degradation in rabbit model of osteoarthritis.

2012 ◽  
Vol 59 (4) ◽  
Author(s):  
Wei-Ping Chen ◽  
Chong Yu ◽  
Peng-Fei Hu ◽  
Jia-Peng Bao ◽  
Jing-Li Tang ◽  
...  
Keyword(s):  
Cruciate Ligament ◽  
Rabbit Model ◽  
Cartilage Degradation ◽  
Tissue Inhibitors Of Metalloproteinase ◽  
Expression Of Genes ◽  
Genes Encoding ◽  
Group 2 ◽  
Group 1

Stigmasterol has been shown exhibit anti-osteoarthritic properties in vitro studies. However, the in vivo effects of stigmasterol on cartilage are still unclear. This study investigated the anti-osteoarthritic properties of stigmasterol on cartilage degradation in a rabbit model of osteoarthritis (OA). Twenty rabbits underwent bilateral anterior cruciate ligament transection (ACLT) to induce OA. Five rabbits were used as normal control. Two weeks after operation, the rabbits were randomly divided into two groups. Each group of 10 rabbits received intra-articular injection with 0.3 ml of stigmasterol in left knees and vehicle in right knees, once weekly. Group 1 was killed 6 weeks after ACLT and 2 were sacrificed 9 weeks after ACLT. The knee joints were assessed by gross morphology, histology and gene expression analysis. We found that expression of genes encoding matrix metalloproteinases (MMPs) was significantly higher while tissue inhibitors of metalloproteinase (TIMP)-1 was significantly lower in the both joints of the two OA groups compared to normal controls. Stigmasterol reduced the cartilage degradation as assessed by histological analysis and markedly suppressed MMPs expression both in group 1 and group 2. Our results suggest that stigmasterol may be considered as a possible therapeutical agent in the treatment of OA.

scholarly journals Transcriptional Modulation of Genes Encoding Structural Characteristics of Differentiating Enterocytes During Development of a Polarized Epithelium In Vitro

2007 ◽  
Vol 18 (11) ◽  
pp. 4261-4278 ◽  
Author(s):  
Jennifer M. Halbleib ◽  
Annika M. Sääf ◽  
Patrick O. Brown ◽  
W. James Nelson
Keyword(s):  
Gene Expression ◽  
Cell Polarity ◽  
Structural Characteristics ◽  
Expression Patterns ◽  
Expression Of Genes ◽  
Genes Encoding ◽  
Functional Features ◽  
Cell Cell

Although there is considerable evidence implicating posttranslational mechanisms in the development of epithelial cell polarity, little is known about the patterns of gene expression and transcriptional regulation during this process. We characterized the temporal program of gene expression during cell–cell adhesion–initiated polarization of human Caco-2 cells in tissue culture, which develop structural and functional polarity similar to that of enterocytes in vivo. A distinctive switch in gene expression patterns occurred upon formation of cell–cell contacts between neighboring cells. Expression of genes involved in cell proliferation was down-regulated concomitant with induction of genes necessary for functional specialization of polarized epithelial cells. Transcriptional up-regulation of these latter genes correlated with formation of important structural and functional features in enterocyte differentiation and establishment of structural and functional cell polarity; components of the apical microvilli were induced as the brush border formed during polarization; as barrier function was established, expression of tight junction transmembrane proteins peaked; transcripts encoding components of the apical, but not the basal-lateral trafficking machinery were increased during polarization. Coordinated expression of genes encoding components of functional cell structures were often observed indicating temporal control of expression and assembly of multiprotein complexes.

scholarly journals A Peptide Found in Human Serum, Derived from the C-Terminus of Albumin, Shows Antifungal Activity In Vitro and In Vivo

2020 ◽  
Vol 8 (10) ◽  
pp. 1627
Author(s):  
Tecla Ciociola ◽  
Pier Paolo Zanello ◽  
Tiziana D’Adda ◽  
Serena Galati ◽  
Stefania Conti ◽  
...  
Keyword(s):  
Antifungal Activity ◽  
Human Serum ◽  
Fungicidal Activity ◽  
Galleria Mellonella ◽  
Serum Proteins ◽  
Morphological Alterations ◽  
C Terminus ◽  
Different Sources

The growing problem of antimicrobial resistance highlights the need for alternative strategies to combat infections. From this perspective, there is a considerable interest in natural molecules obtained from different sources, which are shown to be active against microorganisms, either alone or in association with conventional drugs. In this paper, peptides with the same sequence of fragments, found in human serum, derived from physiological proteins, were evaluated for their antifungal activity. A 13-residue peptide, representing the 597–609 fragment within the albumin C-terminus, was proved to exert a fungicidal activity in vitro against pathogenic yeasts and a therapeutic effect in vivo in the experimental model of candidal infection in Galleria mellonella. Studies by confocal microscopy and transmission and scanning electron microscopy demonstrated that the peptide penetrates and accumulates in Candida albicans cells, causing gross morphological alterations in cellular structure. These findings add albumin to the group of proteins, which already includes hemoglobin and antibodies, that could give rise to cryptic antimicrobial fragments, and could suggest their role in anti-infective homeostasis. The study of bioactive fragments from serum proteins could open interesting perspectives for the development of new antimicrobial molecules derived by natural sources.

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