The genome editing revolution: review Khalil, Ahmad M.
Journal of Genetic Engineering and Biotechnology,
10/2020, Letnik:
18, Številka:
1
Journal Article
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Background
Development of efficient strategies has always been one of the great perspectives for biotechnologists. During the last decade, genome editing of different organisms has been a fast ...advancing field and therefore has received a lot of attention from various researchers comprehensively reviewing latest achievements and offering opinions on future directions. This review presents a brief history, basic principles, advantages and disadvantages, as well as various aspects of each genome editing technology including the modes, applications, and challenges that face delivery of gene editing components.
Main body
Genetic modification techniques cover a wide range of studies, including the generation of transgenic animals, functional analysis of genes, model development for diseases, or drug development. The delivery of certain proteins such as monoclonal antibodies, enzymes, and growth hormones has been suffering from several obstacles because of their large size. These difficulties encouraged scientists to explore alternative approaches, leading to the progress in gene editing. The distinguished efforts and enormous experimentation have now been able to introduce methodologies that can change the genetic constitution of the living cell. The genome editing strategies have evolved during the last three decades, and nowadays, four types of “programmable” nucleases are available in this field: meganucleases, zinc finger nucleases, transcription activator-like effector nucleases, and the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR associated protein 9 (Cas9) (CRISPR/Cas-9) system. Each group has its own characteristics necessary for researchers to select the most suitable method for gene editing tool for a range of applications. Genome engineering/editing technology will revolutionize the creation of precisely manipulated genomes of cells or organisms in order to modify a specific characteristic. Of the potential applications are those in human health and agriculture. Introducing constructs into target cells or organisms is the key step in genome engineering.
Conclusions
Despite the success already achieved, the genome editing techniques are still suffering certain difficulties. Challenges must be overcome before the full potential of genome editing can be realized.
Synthetic biology is bringing together engineers and biologists to design and build novel biomolecular components, networks and pathways, and to use these constructs to rewire and reprogram ...organisms. These re-engineered organisms will change our lives over the coming years, leading to cheaper drugs, 'green' means to fuel our cars and targeted therapies for attacking 'superbugs' and diseases, such as cancer. The de novo engineering of genetic circuits, biological modules and synthetic pathways is beginning to address these crucial problems and is being used in related practical applications.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Curcumin is a phytochemical isolated from the dried rhizome of Curcuma longa L. family Zingiberaceae which possesses versatile biological activities and has hydrophobic properties. The current study ...was conducted to fabricate, and optimize curcumin loaded chitosan and Sodium tripolyphosphate (STPP) nanoparticles (NPs) to improve the bioavailability of curcumin. NPs were fabricated employing the Ionic gelation method. Four formulations were developed based on the selected variables like STPP and chitosan concentration, rotations per minute (rpm), temperature, and pH of chitosan solution. NPs were characterized for morphology, drug-polymer compatibility, yield, particle size, encapsulation efficiency, release behavior, anti-inflammatory and anti-arthritic activity compared to curcumin and standard drug. Fourier transform infrared spectroscopy (FTIR) shows nanoparticle compatibility. The maximum yield was 60%. Entrapment efficiency ranged from 45 to 65%. Curcumin NPs and standard drug 600 µg/ml shows 59% and 70% anti-inflammatory activity by HRB membrane stabilization method respectively which are greater than curcumin alone whereas anti-arthritic activity by protein denaturation method which is also comparable to standard drug and greater than curcumin was 66 and 70% respectively. Statistical analysis shows the mean significant difference at p ≤ 0.05. The study concluded that curcumin-loaded chitosan and STPP NPs formulated successfully by the Ionic gelation method, which increased curcumin absorption leading to a reduced dosing rate and improved patient compliance.
In this article, we first present a generalized methodology for deriving sensor brightness temperature sensor data records (SDR) from antenna temperature data records (TDR) applicable for Advanced ...Microwave Sounding Unit-A (AMSU-A) instruments. It includes corrections for antenna sidelobe contributions, antenna emission, and radiation perturbation due to the difference of Earth radiance in the main beam and that in the sidelobes that lie outside the main beam but within the Earth disk. For practical purposes, we simplify the methodology by neglecting the components other than the antenna sidelobe contributions to establish a consistent formulation with the legacy AMSU-A antenna pattern correction (APC) formula. The simplified formulation is then applied to the final AMSU-A instrument onboard the Metop-C satellite that was launched in November 2018, in order to compute APC coefficients for deriving SDR from TDR data. Furthermore, the performance of the calculated correction coefficients is validated by calculating the differences between the daily averaged AMSU-A (TDR and SDR) observations against radiative transfer model (O-B) simulations under clear sky conditions, and over open oceans. The validation results show that the derived temperature corrections are channel and scan position dependent, and can add 0.2-1.6 K to the antenna temperatures. In addition, the derived SDR O-B results exhibit a reduced and more symmetric scan angle-dependent bias when compared with corresponding TDR antenna temperatures.
Eukaryotic transcription factors (TFs) perform complex and combinatorial functions within transcriptional networks. Here, we present a synthetic framework for systematically constructing eukaryotic ...transcription functions using artificial zinc fingers, modular DNA-binding domains found within many eukaryotic TFs. Utilizing this platform, we construct a library of orthogonal synthetic transcription factors (sTFs) and use these to wire synthetic transcriptional circuits in yeast. We engineer complex functions, such as tunable output strength and transcriptional cooperativity, by rationally adjusting a decomposed set of key component properties, e.g., DNA specificity, affinity, promoter design, protein-protein interactions. We show that subtle perturbations to these properties can transform an individual sTF between distinct roles (activator, cooperative factor, inhibitory factor) within a transcriptional complex, thus drastically altering the signal processing behavior of multi-input systems. This platform provides new genetic components for synthetic biology and enables bottom-up approaches to understanding the design principles of eukaryotic transcriptional complexes and networks.
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► Zinc fingers can be used to wire orthogonal connections in yeast synthetic circuits ► Zinc finger TF design permits adjustable component properties for modulating outputs ► Protein-protein interactions can be used to engineer cooperativity in zinc finger TFs ► TF component properties can be combinatorially adjusted to reshape signal integration
A library of synthetic eukaryotic transcription factors constructed from modifiable component properties of artificial zinc fingers is capable of modulating outputs and reshaping signal integration.
Chemical modifications to DNA and histone proteins are involved in epigenetic programs underlying cellular differentiation and development. Regulatory networks involving molecular writers and readers ...of chromatin marks are thought to control these programs. Guided by this common principle, we established an orthogonal epigenetic regulatory system in mammalian cells using N6-methyladenine (m6A), a DNA modification not commonly found in metazoan epigenomes. Our system utilizes synthetic factors that write and read m6A and consequently recruit transcriptional regulators to control reporter loci. Inspired by models of chromatin spreading and epigenetic inheritance, we used our system and mathematical models to construct regulatory circuits that induce m6A-dependent transcriptional states, promote their spatial propagation, and maintain epigenetic memory of the states. These minimal circuits were able to program epigenetic functions de novo, conceptually validating “read-write” architectures. This work provides a toolkit for investigating models of epigenetic regulation and encoding additional layers of epigenetic information in cells.
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•A synthetic epigenetic regulatory system in human cells using m6A DNA modification•Engineered writers and readers of m6A enable construction of regulatory circuits•Read-write circuits drive spatial propagation and hallmarks of chromatin spreading•Read-write circuits enable epigenetic memory of transcriptional states
A synthetic, modular, and programmable read-write system allows isolated and orthogonal epigenetic control in mammalian cells.
Abstract Long non-coding RNAs have emerged as highly versatile players in the regulation of gene expression in development and human disease, particularly cancer. Hundreds of lncRNAs become ...dysregulated across tumor types, and multiple lncRNAs have demonstrated functions as tumor-suppressors or oncogenes. Furthermore, studies have demonstrated that dysregulation of lncRNAs results in alterations of the epigenome in cancer cells, potentially providing a novel mechanism for the massive epigenomic alterations observed in many tumors. Here, we highlight and provide some illustrious examples of lncRNAs in various epigenetic regulatory processes, including coordination of chromatin dynamics, regulation of DNA methylation, modulation of other non-coding RNAs and mRNA stability, and control of epigenetic substrate availability through altered tumor metabolism. In light of all these known and emerging functions in epigenetic regulation of tumorigenesis and cancer progression, lncRNAs represent attractive targets for future therapeutic strategies in cancer.
The traditional view of protein aggregation as being strictly disease-related has been challenged by many examples of cellular aggregates that regulate beneficial biological functions. When coupled ...with the emerging view that many regulatory proteins undergo phase separation to form dynamic cellular compartments, it has become clear that supramolecular assembly plays wide-ranging and critical roles in cellular regulation. This presents opportunities to develop new tools to probe and illuminate this biology, and to harness the unique properties of these self-assembling systems for synthetic biology for the purposeful manipulation of biological function.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Decapping represents a critical control point in regulating expression of protein coding genes. Here, we demonstrate that decapping also modulates expression of long noncoding RNAs (lncRNAs). ...Specifically, levels of >100 lncRNAs in yeast are controlled by decapping and are degraded by a pathway that occurs independent of decapping regulators. We find many lncRNAs degraded by DCP2 are expressed proximal to inducible genes. Of these, we show several genes required for galactose utilization are associated with lncRNAs that have expression patterns inversely correlated with their mRNA counterpart. Moreover, decapping of these lncRNAs is critical for rapid and robust induction of GAL gene expression. Failure to destabilize a lncRNA known to exert repressive histone modifications results in perpetuation of a repressive chromatin state that contributes to reduced plasticity of gene activation. We propose that decapping and lncRNA degradation serve a vital role in transcriptional regulation specifically at inducible genes.
► lncRNAs are degraded by a DCP2-dependent decapping pathway ► Many lncRNAs targeted to decapping are associated with inducible gene loci ► Decapping of lncRNAs is required for robust activation of GAL gene expression ► lncRNA degradation contributes plasticity to gene regulatory mechanisms
Bacteriostatic and bactericidal antibiotic treatments result in two fundamentally different phenotypic outcomesâthe inhibition of bacterial growth or, alternatively, cell death. Most antibiotics ...inhibit processes that are major consumers of cellular energy output, suggesting that antibiotic treatment may have important downstream consequences on bacterial metabolism. We hypothesized that the specific metabolic effects of bacteriostatic and bactericidal antibiotics contribute to their overall efficacy. We leveraged the opposing phenotypes of bacteriostatic and bactericidal drugs in combination to investigate their activity. Growth inhibition from bacteriostatic antibiotics was associated with suppressed cellular respiration whereas cell death from most bactericidal antibiotics was associated with accelerated respiration. In combination, suppression of cellular respiration by the bacteriostatic antibiotic was the dominant effect, blocking bactericidal killing. Global metabolic profiling of bacteriostatic antibiotic treatment revealed that accumulation of metabolites involved in specific drug target activity was linked to the buildup of energy metabolites that feed the electron transport chain. Inhibition of cellular respiration by knockout of the cytochrome oxidases was sufficient to attenuate bactericidal lethality whereas acceleration of basal respiration by genetically uncoupling ATP synthesis from electron transport resulted in potentiation of the killing effect of bactericidal antibiotics. This work identifies a link between antibiotic-induced cellular respiration and bactericidal lethality and demonstrates that bactericidal activity can be arrested by attenuated respiration and potentiated by accelerated respiration. Our data collectively show that antibiotics perturb the metabolic state of bacteria and that the metabolic state of bacteria impacts antibiotic efficacy.
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