scholarly journals Transcriptional organization and regulation of the Pseudomonas putida flagellar system

2021 ◽  
Author(s):  
Antonio Leal-Morales ◽  
Marta Pulido-Sánchez ◽  
Aroa López Sánchez ◽  
Fernando Govantes
Keyword(s):  
Pseudomonas Putida ◽  
Flagellar Apparatus ◽  
Regulatory Elements ◽  
Gene Arrangement ◽  
Class Iii ◽  
Promoter Regions ◽  
Regulatory Circuit ◽  
Σ Factor ◽  
And Function

A single region of the Pseudomonas putida genome, designated the flagellar cluster, includes 59 genes potentially involved in the biogenesis and function of the flagellar system. Here we combine bioinformatics and in vivo gene expression analyses to clarify the transcriptional organization and regulation of the flagellar genes in the cluster. We have identified eleven flagellar operons and characterized twenty-one primary and internal promoter regions. Our results indicate that synthesis of the flagellar apparatus and core chemotaxis machinery is regulated by a three-tier cascade in which fleQ is the sole Class I gene, standing at the top of the transcriptional hierarchy. FleQ- and σ54-dependent Class II genes encode most components of the flagellar structure, part of the chemotaxis machinery and multiple regulatory elements, including the flagellar σ factor FliA. FliA activation of Class III genes enables synthesis of the filament, one stator complex and completion of the chemotaxis apparatus. Accessory regulatory proteins and an intricate operon architecture add complexity to the regulation by providing feedback and feed-forward loops to the main circuit. Because of the high conservation of the gene arrangement and promoter motifs, we believe that the regulatory circuit presented here may also apply to other environmental Pseudomonas.

scholarly journals CNBP controls transcription by unfolding DNA G-quadruplex structures

2019 ◽  
Vol 47 (15) ◽  
pp. 7901-7913 ◽  
Author(s):  
Aldana P David ◽  
Angélique Pipier ◽  
Federico Pascutti ◽  
Andrés Binolfi ◽  
Andrea M J Weiner ◽  
...  
Keyword(s):  
Transcriptional Control ◽  
Regulatory Elements ◽  
Transcription Start Sites ◽  
G4 Dna ◽  
Dna Strands ◽  
Living Organisms ◽  
And Function

Abstract Guanine-rich DNA strands can fold into non-canonical four-stranded secondary structures named G-quadruplexes (G4). Experimental evidences suggest that G4-DNA surrounding transcription start sites act as cis-regulatory elements by either stimulating or inhibiting gene transcription. Therefore, proteins able to target and regulate specific G4 formation/unfolding are crucial for G4-mediated transcriptional control. Here we present data revealing that CNBP acts in vitro as a G4-unfolding protein over a tetramolecular G4 formed by the TG4T oligonucleotide, as well as over the G4 folded in the promoters of several oncogenes. CNBP depletion in cellulo led to a reduction in the transcription of endogenous KRAS, suggesting a regulatory role of CNBP in relieving the transcriptional abrogation due to G4 formation. CNBP activity was also assayed over the evolutionary conserved G4 enhancing the transcription of NOGGIN (NOG) developmental gene. CNBP unfolded in vitro NOG G4 and experiments performed in cellulo and in vivo in developing zebrafish showed a repressive role of CNBP on the transcription of this gene by G4 unwinding. Our results shed light on the mechanisms underlying CNBP way of action, as well as reinforce the notion about the existence and function of G4s in whole living organisms.

scholarly journals Comparative Analysis of Super-enhancers Across Mammals Reveals Their High Function Conservation

2020 ◽  
Author(s):  
Yanling Peng ◽  
Yubo Zhang
Keyword(s):  
Cell Types ◽  
Regulatory Elements ◽  
Biological Processes ◽  
Orthologous Sequence ◽  
Orthologous Genes ◽  
Promoter Regions ◽  
Cell Tissue ◽  
Intergenic Regions ◽  
Regulation Function ◽  
And Function

Abstract BackgroundSuper-enhancers (SEs) are key positive regulatory elements in defining cells/tissues identity in mammals, yet their similarities and differences of sequence and function across mammals have been poor studied. To allow sequence and function comparison across mammalian SEs, we employ H3K27ac ChIP-seq data to six cell types/tissues across human, pig, and mouse, which represent different lineages of mammals in the evolutionary tree.ResultsOverall, a median of 848 human SEs, 888 pig SEs and 503 mouse SEs are identified across cells/tissues. These SEs are largely distributed in promoter regions for human (91.9% in median) and mouse (63.4% in median), while mostly in distal intergenic regions for pig (66.1% in median). Extremely higher unique orthologous SEs frequency (91.6%~92.1%) has been detected for the same cell/tissue across species. Consistently, their overlapping rates are very low for the same cell/tissue across species (0.1%~0.5%). For the SE-associated orthologous genes, they also show high unique frequency for species (63.3%~83.9%) and low overlapping rates (0.8%~1.3%) at inter-species comparison. However, orthologous SEs function comparisons across species have shown similar biological processes related to cells/tissues identity in the top 15 significant enriched terms for the same cell/tissue. Meanwhile, common core transcription factors that determine cells/tissues identity are determined for the same cell/tissue across mammals.ConclusionsThis study highlights the differences of SEs genomic distribution across mammals. It reveals low orthologous sequence overlapping but high function conservation of SEs across mammals. It would improve our understanding of regulation function cis-regulatory elements in mammals.

scholarly journals Folding and Persistence Time of Intramolecular G-Quadruplexes Transiently Embedded in a DNA duplex

2021 ◽  
Author(s):  
Phong Lan Thao Tran ◽  
Martin Rieu ◽  
Samar Hodeib ◽  
Alexandra Joubert ◽  
Jimmy Ouellet ◽  
...  
Keyword(s):  
Single Molecule ◽  
Critical Role ◽  
Genetic Regulation ◽  
Regulatory Elements ◽  
Promoter Regions ◽  
Folding Rate ◽  
Persistence Time ◽  
G4 Dna

ABSTRACTG-quadruplex (G4) DNA structures have emerged as important regulatory elements during DNA replication, transcription or repair. While many in-vitro studies have focused on the kinetics of G4 formation within DNA single-strands, G4 are found in-vivo in double-stranded DNA regions, where their formation is challenged by pairing between the two complementary strands. Since the energy of hybridization of Watson-Crick structures dominates the energy of G4 folding, this competition should play a critical role on the persistence of G4 in vivo. To address this issue, we designed a single molecule assay allowing measuring G4 folding and persistence while the structure is periodically challenged by the complementary strand. We quantified both the folding rate and the persistence time of biologically relevant G4 structures and showed that the dynamics of G4 formation depends strongly on the genomic location. G4 are found much more stable in promoter regions and replication origins than in telomeric regions. In addition, we characterized how G4 dynamics was affected by G4 ligands and showed that both folding rate and persistence increased. Our assay opens new perspectives for the measurement of G4 dynamics, which is critical to understand their role in genetic regulation.

scholarly journals Human Somatostatin SST4 Receptor Transgenic Mice: Construction and Brain Expression Pattern Characterization

2021 ◽  
Vol 22 (7) ◽  
pp. 3758
Author(s):  
Balázs Nemes ◽  
Kata Bölcskei ◽  
Angéla Kecskés ◽  
Viktória Kormos ◽  
Balázs Gaszner ◽  
...  
Keyword(s):  
In Vivo Imaging ◽  
Regulatory Elements ◽  
Receptor Expression ◽  
Luciferase Reporter ◽  
Mouse Line ◽  
Humanized Mouse ◽  
Peripheral Tissues ◽  
And Function ◽  
The Brain

Somatostatin receptor subtype 4 (SST4) has been shown to mediate analgesic, antidepressant and anti-inflammatory functions without endocrine actions; therefore, it is proposed to be a novel target for drug development. To overcome the species differences of SST4 receptor expression and function between humans and mice, we generated an SST4 humanized mouse line to serve as a translational animal model for preclinical research. A transposon vector containing the hSSTR4 and reporter gene construct driven by the hSSTR4 regulatory elements were created. The vector was randomly inserted in Sstr4-deficient mice. hSSTR4 expression was detected by bioluminescent in vivo imaging of the luciferase reporter predominantly in the brain. RT-qPCR confirmed the expression of the human gene in the brain and various peripheral tissues consistent with the in vivo imaging. RNAscope in situ hybridization revealed the presence of hSSTR4 transcripts in glutamatergic excitatory neurons in the CA1 and CA2 regions of the hippocampus; in the GABAergic interneurons in the granular layer of the olfactory bulb and in both types of neurons in the primary somatosensory cortex, piriform cortex, prelimbic cortex and amygdala. This novel SST4 humanized mouse line might enable us to investigate the differences of human and mouse SST4 receptor expression and function and assess the effects of SST4 receptor agonist drug candidates.

scholarly journals Regulatory Mechanisms at the MouseIgf2/H19 Locus

2001 ◽  
Vol 21 (23) ◽  
pp. 8189-8196 ◽  
Author(s):  
Christopher R. Kaffer ◽  
Alex Grinberg ◽  
Karl Pfeifer
Keyword(s):  
Gene Expression ◽  
Developmental Disorders ◽  
Expression Patterns ◽  
Regulatory Elements ◽  
Normal Function ◽  
Model Systems ◽  
Monoallelic Expression ◽  
Enhancer Element ◽  
And Function

ABSTRACT The closely linked H19 and Igf2 genes show highly similar patterns of gene expression but are reciprocally imprinted. H19 is expressed almost exclusively from the maternally inherited chromosome, while Igf2 expression is mostly from the paternal chromosome. In humans, loss of imprinting at this locus is associated with tumors and with developmental disorders. Monoallelic expression at the imprinted Igf2/H19 locus occurs by at least two distinct mechanisms: a developmentally regulated silencing of the paternal H19 promoter, and transcriptional insulation of the maternal Igf2 promoters. Both mechanisms of allele-specific silencing are ultimately dependent on a commoncis-acting element located just upstream of theH19 promoter. The coordinated expression patterns and some experimental data support the idea that positive regulatory elements are also shared by the two genes. To clarify the organization and function of positive and negative regulatory elements at theH19/Igf2 locus, we analyzed two mouse mutations. First, we generated a deletion allele to localize enhancers used in vivo for expression of both H19 and Igf2 in mesodermal tissues to sequences downstream of the H19 gene. Coincidentally, we demonstrated that some expression ofIgf2 is independent of the shared enhancer element. Second, we used this new information to further characterize an ectopicH19 differentially regulated region and the associated insulator. We demonstrated that its activity is parent-of-origin dependent. In contrast to recent results from Drosophilamodel systems; we showed that this duplication of a mammalian insulator does not interfere with its normal function. Implications of these findings for current models for monoallelic gene expression at this locus are discussed.

scholarly journals Role and Function of Class III LitR, a Photosensor Homolog from Burkholderia multivorans

2018 ◽  
Vol 200 (24) ◽  
Author(s):  
Satoru Sumi ◽  
Hatsumi Shiratori-Takano ◽  
Kenji Ueda ◽  
Hideaki Takano
Keyword(s):  
Constitutive Expression ◽  
Light Response ◽  
Binding Activity ◽  
Binding Domain ◽  
Class Iii ◽  
Promoter Regions ◽  
Content Type ◽  
Burkholderia Multivorans ◽  
Dna Binding Activity ◽  
And Function

ABSTRACT The LitR/CarH protein family is an adenosyl B12 (AdoB12)-dependent photoreceptor family with DNA-binding activity, and its homologs are widely distributed in the genomes of diverse bacterial genera. In this investigation, we studied the role and functions of a LitR homolog from a Gram-negative soil bacterium, Burkholderia multivorans, which does not possess an AdoB12-binding domain. Transcriptome analysis indicated the existence of 19 light-induced genes, including folE2, cfaB, litS, photolyase gene phrB2, and cryB, located in the region flanking litR. Disruption of litR caused constitutive expression of all the light-inducible genes, while mutation in the light-induced sigma factor gene, litS, abolished the transcription of the phrB2 operon and the cfa operon, indicating that LitR and LitS play a central role in light-inducible transcription. A gel shift assay showed that recombinant protein LitR specifically binds to the promoter regions of litR and the folE2 operon, and its binding was weakened by UV-A illumination. LitR absorbs light at maximally near 340 nm and exhibited a photocyclic response and light-dependent dissociation of multimer into tetramer. The litR mutant produced a 20-fold-higher intracellular level of folate than that of the wild-type strain. Thus, the evidence suggests that LitR light-dependently regulates the transcription of litR itself and the folE2 operon, resulting in the production of folate, and then the expressed RNA polymerase complex containing σLitS directs the transcription of the phrB2 operon and the cfa operon. These light-dependent characteristics suggest that class III LitR, in complex with a UV-A-absorbing molecule, follows a novel light-sensing mechanism. IMPORTANCE Members of the LitR/CarH family are adenosyl B12-based photosensory transcriptional regulator involved in light-inducible carotenoid production in nonphototrophic bacteria. Our study provides the first evidence of the involvement of a class III LitR, which lacks an adenosyl B12-binding domain in the light response of Burkholderia multivorans belonging to betaproteobacteria. Our biochemical analysis suggests that class III LitR protein exhibits features as a photosensor including absorption of light at the UV-A region (λmax = ca. 340 nm), photocyclic response, and light-dependent dissociation. This suggests that class III LitR associates with a UV-A-absorbing molecule, and it has a photosensing mechanism distinguishable from that of the B12-based type.

scholarly journals Confined migration induces heterochromatin formation and alters chromatin accessibility

2021 ◽  
Author(s):  
Chieh-Ren Hsia ◽  
Jawuanna McAllister ◽  
Ovais Hasan ◽  
Julius Judd ◽  
Seoyeon Lee ◽  
...  
Keyword(s):  
Chromatin Accessibility ◽  
Cellular Functions ◽  
Promoter Regions ◽  
Collagen Matrices ◽  
Heterochromatin Formation ◽  
Damage Response ◽  
Intergenic Regions ◽  
And Function ◽  
Extensive Deformation

During migration, cells often squeeze through small constrictions, requiring extensive deformation. We hypothesized that the nuclear deformation associated with such confined migration could alter chromatin organization and function. Studying cells migrating through collagen matrices and microfluidic devices that mimic interstitial spaces in vivo, we found that confined migration results in increased H3K9me3 and H3K27me3 heterochromatin marks that persist for several days. This "confined migration-induced heterochromatin" (CMiH) was distinct from heterochromatin formation during migration initiation. CMiH predominantly decreased chromatin accessibility at intergenic regions near centromeres and telomeres, suggesting heterochromatin spreading from existing heterochromatin sites. Consistent with the overall decrease in chromatin accessibility, global transcription was decreased during confined migration. Intriguingly, we also identified increased accessibility at promoter regions of genes linked to chromatin silencing, tumor invasion, and DNA damage response. Inhibiting CMiH reduced migration speed, suggesting that CMiH promotes confined migration. Together, our findings indicate that confined migration induces chromatin changes that regulate cell migration and other cellular functions.

scholarly journals Bordetella pertussisCan Be Motile and Express Flagellum-Like Structures

mBio ◽  
2019 ◽  
Vol 10 (3) ◽  
Author(s):  
Casandra L. Hoffman ◽  
Laura A. Gonyar ◽  
Federico Zacca ◽  
Federico Sisti ◽  
Julieta Fernandez ◽  
...  
Keyword(s):  
Whooping Cough ◽  
Stop Codon ◽  
Genetic Material ◽  
Flagellar Apparatus ◽  
Growth Conditions ◽  
Content Type ◽  
Flagellar Biosynthesis ◽  
Gene And Protein Expression ◽  
And Function

ABSTRACTBordetella bronchisepticaencodes and expresses a flagellar apparatus. In contrast,Bordetella pertussis, the causative agent of whooping cough, has historically been described as a nonmotile and nonflagellated organism. The previous statements thatB. pertussiswas a nonmotile organism were consistent with a stop codon located in the flagellar biosynthesis gene,flhA, discovered when theB. pertussisTohama I genome was sequenced and analyzed by Parkhill et al. in 2003 (J. Parkhill, M. Sebaihia, A. Preston, L. D. Murphy, et al., Nat Genet, 35:32–40, 2003, https://doi.org/10.1038/ng1227). The stop codon has subsequently been found in all annotated genomes. Parkhill et al. also showed, however, thatB. pertussiscontains all genetic material required for flagellar synthesis and function. We and others have determined by various transcriptomic analyses that these flagellar genes are differentially regulated under a variety ofB. pertussisgrowth conditions. In light of these data, we tested forB. pertussismotility and found that both laboratory-adapted strains and clinical isolates can be motile. Upon isolation of motileB. pertussis, we discovered flagellum-like structures on the surface of the bacteria.B. pertussismotility appears to occur primarily in the Bvg(−) phase, consistent with regulation present inB. bronchiseptica. Motility can also be induced by the presence of fetal bovine serum. These observations demonstrate thatB. pertussiscan express flagellum-like structures, and although it remains to be determined ifB. pertussisexpresses flagella during infection or if motility and/or flagella play roles during the cycle of infection and transmission, it is clear that these data warrant further investigation.IMPORTANCEThis report provides evidence for motility and expression of flagella byB. pertussis, a bacterium that has been reported as nonmotile since it was first isolated and studied. As withB. bronchiseptica,B. pertussiscells can express and assemble a flagellum-like structure on their surface, which in other organisms has been implicated in several important processes that occurin vivo. The discovery thatB. pertussisis motile raises many questions, including those regarding the mechanisms of regulation for flagellar gene and protein expression and, importantly, the role of flagella during infection. This novel observation provides a foundation for further study ofBordetellaflagella and motility in the contexts of infection and transmission.

Osseointegration interface of calcified bone and endosseous implants

1992 ◽  
Vol 50 (2) ◽  
pp. 1096-1097
Author(s):  
K.E. Krizan ◽  
J.E. Laffoon ◽  
M.J. Buckley
Keyword(s):  
Structure And Function ◽  
Titanium Implants ◽  
En Bloc ◽  
Endosseous Implants ◽  
Ultrastructural Studies ◽  
The Past ◽  
Cacodylate Buffer ◽  
One Year ◽  
And Function

With increase use of tissue-integrated prostheses in recent years it is a goal to understand what is happening at the interface between haversion bone and bulk metal. This study uses electron microscopy (EM) techniques to establish parameters for osseointegration (structure and function between bone and nonload-carrying implants) in an animal model. In the past the interface has been evaluated extensively with light microscopy methods. Today researchers are using the EM for ultrastructural studies of the bone tissue and implant responses to an in vivo environment. Under general anesthesia nine adult mongrel dogs received three Brånemark (Nobelpharma) 3.75 × 7 mm titanium implants surgical placed in their left zygomatic arch. After a one year healing period the animals were injected with a routine bone marker (oxytetracycline), euthanized and perfused via aortic cannulation with 3% glutaraldehyde in 0.1M cacodylate buffer pH 7.2. Implants were retrieved en bloc, harvest radiographs made (Fig. 1), and routinely embedded in plastic. Tissue and implants were cut into 300 micron thick wafers, longitudinally to the implant with an Isomet saw and diamond wafering blade [Beuhler] until the center of the implant was reached.

Functional characterization of human brown adipose tissue metabolism

2020 ◽  
Vol 477 (7) ◽  
pp. 1261-1286 ◽  
Author(s):  
Marie Anne Richard ◽  
Hannah Pallubinsky ◽  
Denis P. Blondin
Keyword(s):  
Adipose Tissue ◽  
Brown Adipose Tissue ◽  
Functional Characterization ◽  
Human Infants ◽  
Positron Emission ◽  
Brown Adipose ◽  
Treatment Of Obesity ◽  
And Function

Brown adipose tissue (BAT) has long been described according to its histological features as a multilocular, lipid-containing tissue, light brown in color, that is also responsive to the cold and found especially in hibernating mammals and human infants. Its presence in both hibernators and human infants, combined with its function as a heat-generating organ, raised many questions about its role in humans. Early characterizations of the tissue in humans focused on its progressive atrophy with age and its apparent importance for cold-exposed workers. However, the use of positron emission tomography (PET) with the glucose tracer [18F]fluorodeoxyglucose ([18F]FDG) made it possible to begin characterizing the possible function of BAT in adult humans, and whether it could play a role in the prevention or treatment of obesity and type 2 diabetes (T2D). This review focuses on the in vivo functional characterization of human BAT, the methodological approaches applied to examine these features and addresses critical gaps that remain in moving the field forward. Specifically, we describe the anatomical and biomolecular features of human BAT, the modalities and applications of non-invasive tools such as PET and magnetic resonance imaging coupled with spectroscopy (MRI/MRS) to study BAT morphology and function in vivo, and finally describe the functional characteristics of human BAT that have only been possible through the development and application of such tools.

Sign in / Sign up

Export Citation Format

Share Document