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ACS Synthetic Biology May 2022Rational design approaches for the regulation of gene expression are expanding the synthetic biology toolbox. However, only a few tools for regulating gene expression at...
Rational design approaches for the regulation of gene expression are expanding the synthetic biology toolbox. However, only a few tools for regulating gene expression at the translational level have been developed. Here, we devise an approach for translational regulation using the MS2 and PP7 aptamer and coat-protein pairs in . The aptamers are used as operators in transcription units that encode proteins fused to their cognate coat proteins, which leads to self-repression. RNA origami scaffolds that contain up to four aptamers serve as an alternate binder to activate translation. With this system, we demonstrate that the increase in expression of a reporter protein is dependent on both the concentration and number of aptamers on RNA origami scaffolds. We also demonstrate regulation of multiple proteins using a single MS2 coat protein fusion and apply this method to regulate the relative expression of enzymes of the branched pathway for deoxyviolacein biosynthesis.
Topics: Escherichia coli; Protein Binding; Protein Biosynthesis; RNA; Synthetic Biology
PubMed: 35438978
DOI: 10.1021/acssynbio.1c00608 -
FEBS Letters Jun 2011Since their introduction in therapy, antibiotics have played an essential role in human society, saving millions of lives, allowing safe surgery, organ transplants,... (Review)
Review
Since their introduction in therapy, antibiotics have played an essential role in human society, saving millions of lives, allowing safe surgery, organ transplants, cancer therapy. Antibiotics have also helped to elucidate several biological mechanisms and boosted the birth and growth of pharmaceutical companies, generating profits and royalties. The golden era of antibiotics and the scientific and economical drive of big pharma towards these molecules is long gone, but the need for effective antibiotics is increased as their pipelines dwindle and multi-resistant pathogenic strains spread. Here we outline some strategies that could help meet this emergency and list promising new targets.
Topics: Anti-Bacterial Agents; Bacteria; Drug Discovery; Humans; Protein Biosynthesis
PubMed: 21513713
DOI: 10.1016/j.febslet.2011.04.029 -
Molecular Cell Oct 2020Understanding the genetic design principles that determine protein production remains a major challenge. Although the key principles of gene expression were discovered... (Review)
Review
Understanding the genetic design principles that determine protein production remains a major challenge. Although the key principles of gene expression were discovered 50 years ago, additional factors are still being uncovered. Both protein-coding and non-coding sequences harbor elements that collectively influence the efficiency of protein production by modulating transcription, mRNA decay, and translation. The influences of many contributing elements are intertwined, which complicates a full understanding of the individual factors. In natural genes, a functional balance between these factors has been obtained in the course of evolution, whereas for genetic-engineering projects, our incomplete understanding still limits optimal design of synthetic genes. However, notable advances have recently been made, supported by high-throughput analysis of synthetic gene libraries as well as by state-of-the-art biomolecular techniques. We discuss here how these advances further strengthen understanding of the gene expression process and how they can be harnessed to optimize protein production.
Topics: Algorithms; Animals; Biotechnology; Genetic Code; Humans; Protein Biosynthesis; RNA Stability; Transcription, Genetic
PubMed: 33010203
DOI: 10.1016/j.molcel.2020.09.014 -
Journal of Applied Physiology... Aug 2019Skeletal muscle mass responds in a remarkable manner to alterations in loading and use. It has long been clear that skeletal muscle hypertrophy can be prevented by... (Review)
Review
Skeletal muscle mass responds in a remarkable manner to alterations in loading and use. It has long been clear that skeletal muscle hypertrophy can be prevented by inhibiting RNA synthesis. Since 80% of the cell's total RNA has been estimated to be rRNA, this finding indicates that de novo production of rRNA via transcription of the corresponding genes is important for such hypertrophy to occur. Transcription of rDNA by RNA Pol I is the rate-limiting step in ribosome biogenesis, indicating in turn that this biogenesis strongly influences the hypertrophic response. The present minireview focuses on ) a brief description of the key steps in ribosome biogenesis and the relationship of this process to skeletal muscle mass and ) the coordination of ribosome biogenesis and protein synthesis for growth or atrophy, as exemplified by the intracellular AMPK and mTOR pathways.
Topics: Animals; Humans; Hypertrophy; Muscle, Skeletal; Muscular Diseases; Protein Biosynthesis; RNA, Ribosomal; Ribosomes; Transcription, Genetic
PubMed: 31219775
DOI: 10.1152/japplphysiol.00963.2018 -
Bio Systems Jan 2013In this paper we discuss the entropy and information aspects of a living cell. Particular attention is paid to the information gain on assembling and maintaining a...
In this paper we discuss the entropy and information aspects of a living cell. Particular attention is paid to the information gain on assembling and maintaining a living state. Numerical estimates of the information and entropy reduction are given and discussed in the context of the cell's metabolic activity. We discuss a solution to an apparent paradox that there is less information content in DNA than in the proteins that are assembled based on the genetic code encrypted in DNA. When energy input required for protein synthesis is accounted for, the paradox is clearly resolved. Finally, differences between biological information and instruction are discussed.
Topics: Cells; DNA; Entropy; Genetic Code; Information Theory; Models, Theoretical; Probability; Protein Biosynthesis; Proteins; Thermodynamics
PubMed: 23159919
DOI: 10.1016/j.biosystems.2012.10.005 -
Journal of Applied Physiology... Aug 2019The ribosome is typically viewed as a supramolecular complex with constitutive and invariant capacity in mediating translation of mRNA into protein. This view has been... (Review)
Review
The ribosome is typically viewed as a supramolecular complex with constitutive and invariant capacity in mediating translation of mRNA into protein. This view has been challenged by recent research revealing that ribosome composition could be heterogeneous, and this heterogeneity leads to functional ribosome specialization. This review presents the idea that ribosome heterogeneity results from changes in its various components, including variations in ribosomal protein (RP) composition, posttranslational modifications of RPs, changes in ribosomal-associated proteins, alternative forms of rRNA, and posttranscriptional modifications of rRNAs. Ribosome heterogeneity could be orchestrated at several levels and may depend on numerous factors, such as the subcellular location, cell type, tissue specificity, the development state, cell state, ribosome biogenesis, RP turnover, physiological stimuli, and circadian rhythm. Ribosome specialization represents a completely new concept for the regulation of gene expression. Specialized ribosomes could modulate several aspects of translational control, such as mRNA translation selectivity, translation initiation, translational fidelity, and translation elongation. Recent research indicates that the expression of is markedly increased, while that of is highly reduced during mouse skeletal muscle hypertrophy. Moreover, overexpression impairs the growth and myogenic fusion of myotubes. Although the function of and in the ribosome remains to be clarified, these findings suggest that ribosome specialization may be potentially involved in the control of protein translation and skeletal muscle size. Limited data concerning ribosome specialization are currently available in skeletal muscle. Future investigations have the potential to delineate the function of specialized ribosomes in skeletal muscle.
Topics: Animals; Humans; Hypertrophy; Muscle Development; Muscle Fibers, Skeletal; Muscle, Skeletal; Protein Biosynthesis; Protein Processing, Post-Translational; RNA, Messenger; RNA, Ribosomal; Ribosomal Protein L3; Ribosomal Proteins; Ribosomes
PubMed: 30605395
DOI: 10.1152/japplphysiol.00946.2018 -
Nucleic Acids Research May 2021Ribosomes are evolutionary conserved ribonucleoprotein complexes that function as two separate subunits in all kingdoms. During translation initiation, the two subunits...
Ribosomes are evolutionary conserved ribonucleoprotein complexes that function as two separate subunits in all kingdoms. During translation initiation, the two subunits assemble to form the mature ribosome, which is responsible for translating the messenger RNA. When the ribosome reaches a stop codon, release factors promote translation termination and peptide release, and recycling factors then dissociate the two subunits, ready for use in a new round of translation. A tethered ribosome, called Ribo-T, in which the two subunits are covalently linked to form a single entity, was recently described in Escherichia coli. A hybrid ribosomal RNA (rRNA) consisting of both the small and large subunit rRNA sequences was engineered. The ribosome with inseparable subunits generated in this way was shown to be functional and to sustain cell growth. Here, we investigated the translational properties of Ribo-T. We analyzed its behavior during amino acid misincorporation, -1 or +1 frameshifting, stop codon readthrough, and internal translation initiation. Our data indicate that covalent attachment of the two subunits modifies the properties of the ribosome, altering its ability to initiate and terminate translation correctly.
Topics: Codon, Terminator; Frameshifting, Ribosomal; Peptide Chain Initiation, Translational; Peptide Chain Termination, Translational; Protein Biosynthesis; RNA, Transfer; Ribosomes
PubMed: 33950196
DOI: 10.1093/nar/gkab259 -
Toxins Aug 2021Arthropod venoms offer a promising resource for the discovery of novel bioactive peptides and proteins, but the limited size of most species translates into minuscule...
Arthropod venoms offer a promising resource for the discovery of novel bioactive peptides and proteins, but the limited size of most species translates into minuscule venom yields. Bioactivity studies based on traditional fractionation are therefore challenging, so alternative strategies are needed. Cell-free synthesis based on synthetic gene fragments is one of the most promising emerging technologies, theoretically allowing the rapid, laboratory-scale production of specific venom components, but this approach has yet to be applied in venom biodiscovery. Here, we tested the ability of three commercially available cell-free protein expression systems to produce venom components from small arthropods, using U-sicaritoxin-Sdo1a from the six-eyed sand spider as a case study. We found that only one of the systems was able to produce an active product in low amounts, as demonstrated by SDS-PAGE, mass spectrometry, and bioactivity screening on murine neuroblasts. We discuss our findings in relation to the promises and limitations of cell-free synthesis for venom biodiscovery programs in smaller invertebrates.
Topics: Biotechnology; Cell-Free System; Protein Biosynthesis; Spider Venoms; Synthetic Biology
PubMed: 34437446
DOI: 10.3390/toxins13080575 -
Cancer Cell Aug 2013The master regulator of the classical cytoprotective "heat shock" response, heat shock factor 1 (HSF1), is increasingly implicated in cancer pathogenesis, but the...
The master regulator of the classical cytoprotective "heat shock" response, heat shock factor 1 (HSF1), is increasingly implicated in cancer pathogenesis, but the mechanisms remain poorly understood. A recent study connects increased protein translation to activation of HSF1 in malignant cells and demonstrates the therapeutic benefit of targeting this link.
Topics: Animals; DNA-Binding Proteins; Humans; Neoplasms; Protein Biosynthesis; Ribosomes; Transcription Factors
PubMed: 23948296
DOI: 10.1016/j.ccr.2013.07.017 -
Cell Cycle (Georgetown, Tex.) 2015
Topics: Adaptor Proteins, Signal Transducing; CDC2 Protein Kinase; Cell Cycle; Cell Cycle Proteins; Cyclin-Dependent Kinases; Humans; Mitosis; Phosphoproteins; Phosphorylation; Protein Biosynthesis
PubMed: 26496165
DOI: 10.1080/15384101.2015.1084192