Inducible Suppression of Global Translation by Overuse of Rare Codons

ABSTRACTRecently, artificial gene networks have been developed in synthetic biology to control gene expression and make organisms as controllable as robots. Here, I present an artificial posttranslational gene-silencing system based on the codon usage bias and low tRNA content corresponding to minor codons. I engineered the green fluorescent protein (GFP) gene to inhibit translation indirectly with the lowest-usage codons to monopolize various minor tRNAs (lgfp). The expression oflgfpinterfered nonspecifically with the growth ofEscherichia coli,Saccharomyces cerevisiae, human HeLa cervical cancer cells, MCF7 breast cancer cells, and HEK293 kidney cells, as well as phage and adenovirus expansion. Furthermore, insertion oflgfpdownstream of a phage response promoter conferred phage resistance onE. coli. Such engineered gene silencers could act as components of biological networks capable of functioning with suitable promoters inE. coli,S. cerevisiae, and human cells to control gene expression. The results presented here show general suppressor artificial genes for live cells and viruses. This robust system provides a gene expression or cell growth control device for artificially synthesized gene networks.

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PVv3, a New Shuttle Vector for Gene Expression in Vibrio vulnificus

Applied and Environmental Microbiology ◽

10.1128/aem.03720-13 ◽

2013 ◽

Vol 80 (4) ◽

pp. 1477-1481 ◽

Cited By ~ 8

Author(s):

Karina Klevanskaa ◽

Nadja Bier ◽

Kerstin Stingl ◽

Eckhard Strauch ◽

Stefan Hertwig

Keyword(s):

Gene Expression ◽

Escherichia Coli ◽

Green Fluorescent Protein ◽

Vibrio Parahaemolyticus ◽

Fluorescent Protein ◽

Vibrio Vulnificus ◽

Shuttle Vector ◽

Content Type ◽

E Coli ◽

Green Fluorescent

ABSTRACTAn efficient electroporation procedure forVibrio vulnificuswas designed using the new cloning vector pVv3 (3,107 bp). Transformation efficiencies up to 2 × 106transformants per μg DNA were achieved. The vector stably replicated in bothV. vulnificusandEscherichia coliand was also successfully introduced intoVibrio parahaemolyticusandVibrio cholerae. To demonstrate the suitability of the vector for molecular cloning, the green fluorescent protein (GFP) gene and thevvhBAhemolysin operon were inserted into the vector and functionally expressed inVibrioandE. coli.

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Estrogen Regulates JNK1 Genomic Localization to Control Gene Expression and Cell Growth in Breast Cancer Cells

Molecular Endocrinology ◽

10.1210/me.2011-1158 ◽

2012 ◽

Vol 26 (5) ◽

pp. 736-747 ◽

Cited By ~ 11

Author(s):

Miao Sun ◽

Gary D. Isaacs ◽

Nasun Hah ◽

Nina Heldring ◽

Elizabeth A. Fogarty ◽

...

Keyword(s):

Breast Cancer ◽

Gene Expression ◽

Cell Growth ◽

Cancer Cells ◽

Breast Cancer Cells ◽

Control Gene ◽

Control Gene Expression

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Vibrio cholerae Triggers SOS and Mutagenesis in Response to a Wide Range of Antibiotics: a Route towards Multiresistance

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01549-10 ◽

2011 ◽

Vol 55 (5) ◽

pp. 2438-2441 ◽

Cited By ~ 107

Author(s):

Zeynep Baharoglu ◽

Didier Mazel

Keyword(s):

Escherichia Coli ◽

Antibiotic Resistance ◽

Vibrio Cholerae ◽

Fluorescent Protein ◽

Resistance Development ◽

Content Type ◽

E Coli ◽

Wide Range ◽

Green Fluorescent ◽

Regulated Promoters

ABSTRACTAntibiotic resistance development has been linked to the bacterial SOS stress response. InEscherichia coli, fluoroquinolones are known to induce SOS, whereas other antibiotics, such as aminoglycosides, tetracycline, and chloramphenicol, do not. Here we address whether various antibiotics induce SOS inVibrio cholerae. Reporter green fluorescent protein (GFP) fusions were used to measure the response of SOS-regulated promoters to subinhibitory concentrations of antibiotics. We show that unlike the situation withE. coli, all these antibiotics induce SOS inV. cholerae.

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LEC1 (NF-YB9) directly interacts with LEC2 to control gene expression in seed

Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms ◽

10.1016/j.bbagrm.2018.03.005 ◽

2018 ◽

Vol 1861 (5) ◽

pp. 443-450 ◽

Cited By ~ 10

Author(s):

C. Boulard ◽

J. Thévenin ◽

O. Tranquet ◽

V. Laporte ◽

L. Lepiniec ◽

...

Keyword(s):

Gene Expression ◽

Control Gene ◽

Control Gene Expression

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Nonhistone Proteins Control Gene Expression in Reconstituted Chromatin

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.71.12.5057 ◽

1974 ◽

Vol 71 (12) ◽

pp. 5057-5061 ◽

Cited By ~ 95

Author(s):

T. Barrett ◽

D. Maryanka ◽

P. H. Hamlyn ◽

H. J. Gould

Keyword(s):

Gene Expression ◽

Control Gene ◽

Nonhistone Proteins ◽

Control Gene Expression

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A Quantitative Optogenetic Tool to Control Gene Expression during Embryonic Development

Biophysical Journal ◽

10.1016/j.bpj.2020.11.2195 ◽

2021 ◽

Vol 120 (3) ◽

pp. 354a

Author(s):

Anand P. Singh ◽

Ping Wu ◽

Eric F. Wieschaus ◽

Jared E. Toettcher ◽

Thomas Gregor

Keyword(s):

Gene Expression ◽

Embryonic Development ◽

Control Gene ◽

Control Gene Expression

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Droplet- Rather than Aerosol-Mediated Dispersion Is the Primary Mechanism of Bacterial Transmission from Contaminated Hand-Washing Sink Traps

Applied and Environmental Microbiology ◽

10.1128/aem.01997-18 ◽

2018 ◽

Vol 85 (2) ◽

Cited By ~ 9

Author(s):

Shireen M. Kotay ◽

Rodney M. Donlan ◽

Christine Ganim ◽

Katie Barry ◽

Bryan E. Christensen ◽

...

Keyword(s):

Patient Care ◽

Sampling Methods ◽

Fluorescent Protein ◽

Model Organism ◽

Air Sampling ◽

Multidrug Resistant ◽

Hand Washing ◽

Content Type ◽

E Coli ◽

Green Fluorescent

ABSTRACT An alarming rise in hospital outbreaks implicating hand-washing sinks has led to widespread acknowledgment that sinks are a major reservoir of antibiotic-resistant pathogens in patient care areas. An earlier study using green fluorescent protein (GFP)-expressing Escherichia coli (GFP-E. coli) as a model organism demonstrated dispersal from drain biofilms in contaminated sinks. The present study further characterizes the dispersal of microorganisms from contaminated sinks. Replicate hand-washing sinks were inoculated with GFP-E. coli, and dispersion was measured using qualitative (settle plates) and quantitative (air sampling) methods. Dispersal caused by faucet water was captured with settle plates and air sampling methods when bacteria were present on the drain. In contrast, no dispersal was captured without or in between faucet events, amending an earlier theory that bacteria aerosolize from the P-trap and disperse. Numbers of dispersed GFP-E. coli cells diminished substantially within 30 minutes after faucet usage, suggesting that the organisms were associated with larger droplet-sized particles that are not suspended in the air for long periods. IMPORTANCE Among the possible environmental reservoirs in a patient care environment, sink drains are increasingly recognized as a potential reservoir to hospitalized patients of multidrug-resistant health care-associated pathogens. With increasing antimicrobial resistance limiting therapeutic options for patients, a better understanding of how pathogens disseminate from sink drains is urgently needed. Once this knowledge gap has decreased, interventions can be engineered to decrease or eliminate transmission from hospital sink drains to patients. The current study further defines the mechanisms of transmission for bacteria that colonize sink drains.

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Overexpressed Pituitary Tumor-Transforming Gene Causes Aneuploidy in Live Human Cells

Endocrinology ◽

10.1210/en.2003-0305 ◽

2003 ◽

Vol 144 (11) ◽

pp. 4991-4998 ◽

Cited By ~ 76

Author(s):

Run Yu ◽

Wenge Lu ◽

Jiandong Chen ◽

Chris J. McCabe ◽

Shlomo Melmed

Keyword(s):

Cancer Cells ◽

Chromosome Segregation ◽

Pituitary Tumor ◽

Fluorescent Protein ◽

Human Cancer ◽

Live Cells ◽

Pituitary Tumor Transforming Gene ◽

Transforming Gene

Abstract The mammalian securin, pituitary tumor-transforming gene (PTTG), is overexpressed in several tumors and transforms cells in vitro and in vivo. To test the hypothesis that PTTG overexpression causes aneuploidy, enhanced green fluorescent protein (EGFP)-tagged PTTG (PTTG-EGFP) was expressed in human H1299 cancer cells (with undetectable endogenous PTTG expression) and mitosis of individual live cells observed. Untransfected cells and cells expressing EGFP alone exhibited appropriate mitosis. PTTG-EGFP markedly prolonged prophase and metaphase, indicating that PTTG blocks progression of mitosis to anaphase. In cells that underwent apparently normal mitosis (35 of 65 cells), PTTG-EGFP was degraded about 1 min before anaphase onset. Cells that failed to degrade PTTG-EGFP exhibited asymmetrical cytokinesis without chromosome segregation (18 of 65 cells) or chromosome decondensation without cytokinesis (9 of 65 cells), resulting in appearance of a macronucleus. Fifty-one of 55 cells expressing a nondegradable mutant PTTG exhibited asymmetrical cytokinesis without chromosome segregation, and some (4 of 55) decondensed chromosomes, both resulting in macronuclear formation. During this abnormal cytokinesis, all chromosomes and spindles and both centrosomes moved to one daughter cell, suggesting potential chaos in the subsequent mitosis. In conclusion, failure of PTTG degradation or enhanced PTTG accumulation, as a consequence of overexpression, inhibits mitosis progression and chromosome segregation but does not directly affect cytokinesis, resulting in aneuploidy. These results demonstrate that PTTG induces aneuploidy in single, live, human cancer cells.

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Elucidation of Regulatory Modes for Five Two-Component Systems in Escherichia coli Reveals Novel Relationships

mSystems ◽

10.1128/msystems.00980-20 ◽

2020 ◽

Vol 5 (6) ◽

Author(s):

Kumari Sonal Choudhary ◽

Julia A. Kleinmanns ◽

Katherine Decker ◽

Anand V. Sastry ◽

Ye Gao ◽

...

Keyword(s):

Gene Expression ◽

Escherichia Coli ◽

Target Gene ◽

Target Genes ◽

Rna Seq ◽

Content Type ◽

E Coli ◽

Component Systems ◽

Two Component ◽

Two Component Systems

ABSTRACT Escherichia coli uses two-component systems (TCSs) to respond to environmental signals. TCSs affect gene expression and are parts of E. coli’s global transcriptional regulatory network (TRN). Here, we identified the regulons of five TCSs in E. coli MG1655: BaeSR and CpxAR, which were stimulated by ethanol stress; KdpDE and PhoRB, induced by limiting potassium and phosphate, respectively; and ZraSR, stimulated by zinc. We analyzed RNA-seq data using independent component analysis (ICA). ChIP-exo data were used to validate condition-specific target gene binding sites. Based on these data, we do the following: (i) identify the target genes for each TCS; (ii) show how the target genes are transcribed in response to stimulus; and (iii) reveal novel relationships between TCSs, which indicate noncognate inducers for various response regulators, such as BaeR to iron starvation, CpxR to phosphate limitation, and PhoB and ZraR to cell envelope stress. Our understanding of the TRN in E. coli is thus notably expanded. IMPORTANCE E. coli is a common commensal microbe found in the human gut microenvironment; however, some strains cause diseases like diarrhea, urinary tract infections, and meningitis. E. coli’s two-component systems (TCSs) modulate target gene expression, especially related to virulence, pathogenesis, and antimicrobial peptides, in response to environmental stimuli. Thus, it is of utmost importance to understand the transcriptional regulation of TCSs to infer bacterial environmental adaptation and disease pathogenicity. Utilizing a combinatorial approach integrating RNA sequencing (RNA-seq), independent component analysis, chromatin immunoprecipitation coupled with exonuclease treatment (ChIP-exo), and data mining, we suggest five different modes of TCS transcriptional regulation. Our data further highlight noncognate inducers of TCSs, which emphasizes the cross-regulatory nature of TCSs in E. coli and suggests that TCSs may have a role beyond their cognate functionalities. In summary, these results can lead to an understanding of the metabolic capabilities of bacteria and correctly predict complex phenotype under diverse conditions, especially when further incorporated with genome-scale metabolic models.

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Universal attenuators and their interactions with feedback loops in gene regulatory networks

10.1101/074716 ◽

2016 ◽

Author(s):

Dianbo Liu ◽

Luca Albergante ◽

Timothy J Newman

Keyword(s):

Gene Expression ◽

Gene Regulatory Networks ◽

Gene Networks ◽

Regulatory Networks ◽

Human Cancer ◽

Cancer Cell Line ◽

Feedback Loops ◽

Human Cancer Cell ◽

E Coli ◽

Gene Regulatory

AbstractUsing a combination of mathematical modelling, statistical simulation and large-scale data analysis we study the properties of linear regulatory chains (LRCs) within gene regulatory networks (GRNs). Our modelling indicates that downstream genes embedded within LRCs are highly insulated from the variation in expression of upstream genes, and thus LRCs act as attenuators. This observation implies a progressively weaker functionality of LRCs as their length increases. When analysing the preponderance of LRCs in the GRNs of E. coli K12 and several other organisms, we find that very long LRCs are essentially absent. In both E. coli and M. tuberculosis we find that four-gene LRCs are intimately linked to identical feedback loops that are involved in potentially chaotic stress response, indicating that the dynamics of these potentially destabilising motifs are strongly restrained under homeostatic conditions. The same relationship is observed in a human cancer cell line (K562), and we postulate that four-gene LRCs act as “universal attenuators”. These findings suggest a role for long LRCs in dampening variation in gene expression, thereby protecting cell identity, and in controlling dramatic shifts in cell-wide gene expression through inhibiting chaos-generating motifs.In briefWe present a general principle that linear regulatory chains exponentially attenuate the range of expression in gene regulatory networks. The discovery of a universal interplay between linear regulatory chains and genetic feedback loops in microorganisms and a human cancer cell line is analysed and discussed.HighlightsWithin gene networks, linear regulatory chains act as exponentially strong attenuators of upstream variationBecause of their exponential behaviour, linear regulatory chains beyond a few genes provide no additional functionality and are rarely observed in gene networks across a range of different organismsNovel interactions between four-gene linear regulatory chains and feedback loops were discovered in E. coli, M. tuberculosis and human cancer cells, suggesting a universal mechanism of control.

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