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CRISPR01:59

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Genome editing technologies allow scientists to modify an organism’s DNA via the addition, removal, or rearrangement of genetic material at specific genomic locations. These types of techniques could potentially be used to cure genetic disorders such as hemophilia and sickle cell anemia. One popular and widely used DNA-editing research tool that could lead to safe and effective cures for genetic disorders is the CRISPR-Cas9 system. CRISPR-Cas9 stands for Clustered Regularly Interspaced...
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The CRISPR-Cas system serves as a bacterial defense mechanism against invading genetic elements such as viruses and plasmids, forming the foundation for its adaptation as a powerful genome-editing tool. Originally discovered in prokaryotes, this system has been repurposed to revolutionize genetic engineering across a wide range of organisms, including plants, animals, and humans. The core component, Cas9, is an endonuclease derived from Streptococcus pyogenes, capable of introducing...
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Bacteria and archaea are susceptible to viral infections just like eukaryotes; therefore, they have developed a unique adaptive immune system to protect themselves. Clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins (CRISPR-Cas) are present in more than 45% of known bacteria and 90% of known archaea.
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The thermodynamic processes can be classified into reversible and irreversible processes. The processes that can be restored to their initial state are called reversible processes. It is only possible if the process is in quasi-static equilibrium, i.e., it takes place in infinitesimally small steps, and the system remains at equilibrium However, these are ideal processes and do not occur naturally. An ideal system undergoing a reversible process is always in thermodynamic equilibrium within...
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A diode is reverse-biased when the positive terminal of an external voltage source is connected to the n-type material and the negative terminal to the p-type material. This configuration opposes the natural direction of current flow through the diode, effectively increasing the width of the depletion region and the barrier potential. The reverse bias condition produces a minimal leakage current, primarily due to minority charge carriers. This leakage becomes significant when the reverse...
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CRISPR Epigenome Editing in Human Cells using Plasmid DNA Transfection and mRNA Nucleofection Delivery
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Reversing Mechanoinductive DSP Expression by CRISPR/dCas9-mediated Epigenome Editing.

Jing Qu1, Lanyan Zhu1,2, Zijing Zhou1,2

  • 11 Division of Pulmonary, Allergy, and Critical Care Medicine, Department of Medicine, University of Alabama at Birmingham, Birmingham, Alabama.

American Journal of Respiratory and Critical Care Medicine
|June 21, 2018
PubMed
Summary

Matrix stiffness increases desmoplakin (DSP) gene expression in lung cells. Epigenetic editing using CRISPR technology reversed this overexpression, offering a potential therapeutic strategy for idiopathic pulmonary fibrosis (IPF).

Keywords:
CRISPR/dCas9desmoplakinepigenome editinglung fibrosismatrix stiffness

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Area of Science:

  • Cellular mechanobiology
  • Epigenetics
  • Pulmonary fibrosis research

Background:

  • Desmoplakin (DSP) is a key component of desmosomes, crucial for epithelial integrity.
  • DSP is implicated in idiopathic pulmonary fibrosis (IPF) pathogenesis, with higher expression in IPF patients.
  • Understanding DSP regulation in fibrotic lung environments is critical.

Purpose of the Study:

  • To investigate the mechanisms regulating DSP expression in the context of lung fibrosis.
  • To explore the role of matrix stiffness in DSP gene regulation.
  • To evaluate the potential of epigenome editing for controlling DSP expression.

Main Methods:

  • Simulated lung stiffness using polyacrylamide gels of varying elasticity.
  • Assessed transcription factor binding to the DSP promoter via ChIP and EMSA.
  • Utilized CRISPR/dCas9-mediated Dnmt3A for targeted DNA methylation and epigenome editing.

Main Results:

  • Stiff matrices significantly increased DSP gene expression in lung epithelial cells.
  • DSP promoter demethylation correlated with increased DSP expression, mediated by EGR1.
  • CRISPR/dCas9-mediated epigenome editing successfully inhibited stiff matrix-induced DSP overexpression.

Conclusions:

  • DSP functions as a mechanosensitive gene regulated by matrix stiffness.
  • Epigenome editing can reverse DSP overexpression by restoring epigenetic control.
  • This approach offers a novel tool for studying DSP's role in lung fibrosis.