• BMB Reports
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• v.57 no.1
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• pp.40-49
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• 2024

Prokaryotes encode clustered regularly interspaced short palindromic repeat (CRISPR) arrays and CRISPR-associated (Cas) genes as an adaptive immune machinery. CRISPR-Cas systems effectively protect hosts from the invasion of foreign enemies, such as bacteriophages and plasmids. During a process called 'adaptation', non-self-nucleic acid fragments are acquired as spacers between repeats in the host CRISPR array, to establish immunological memory. The highly conserved Cas1-Cas2 complexes function as molecular recorders to integrate spacers in a time course manner, which can subsequently be expressed as crRNAs complexed with Cas effector proteins for the RNA-guided interference pathways. In some of the RNA-targeting type III systems, Cas1 proteins are fused with reverse transcriptase (RT), indicating that RT-Cas1-Cas2 complexes can acquire RNA transcripts for spacer acquisition. In this review, we summarize current studies that focus on the molecular structure and function of the RT-fused Cas1-Cas2 integrase, and its potential applications as a directional RNA-recording tool in cells. Furthermore, we highlight outstanding questions for RT-Cas1-Cas2 studies and future directions for RNA-recording CRISPR technologies.

• BMB Reports
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• v.53 no.7
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• pp.341-348
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• 2020

The targeted nuclease clustered, regularly interspaced short palindromic repeats/CRISPR-associated proteins (CRISPR/Cas) system has recently emerged as a prominent gene manipulation method. Because of its ease in programming targeted DNA/protein binding through RNA in a vast range of organisms, this prokaryotic defense system is a versatile tool with many applications in the research field as well as high potential in agricultural and clinical improvements. This review will present a brief history that led to its discovery and adaptation. We also present some of its restrictions, and modifications that have been performed to overcome such restrictions, focusing specifically on the most common CRISPR/Cas9 mediated non-homologous end joint repair.

• The Plant Pathology Journal
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• v.38 no.2
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• pp.102-114
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• 2022

Pectobacterium carotovorum subsp. carotovorum (Pcc) is a gram-negative, broad host range bacterial pathogen which causes soft rot disease in potatoes as well as other vegetables worldwide. While Pectobacterium infection relies on the production of major cell wall degrading enzymes, other virulence factors and the mechanism of genetic adaptation of this pathogen is not yet clear. In the present study, we have performed an in-depth genome-wide characterization of Pcc strain ICMP5702 isolated from potato and compared it with other pathogenic bacteria from the Pectobacterium genus to identify key virulent determinants. The draft genome of Pcc ICMP5702 contains 4,774,457 bp with a G + C content of 51.90% and 4,520 open reading frames. Genome annotation revealed prominent genes encoding key virulence factors such as plant cell wall degrading enzymes, flagella-based motility, phage proteins, cell membrane structures, and secretion systems. Whereas, a majority of determinants were conserved among the Pectobacterium strains, few notable genes encoding AvrE-family type III secretion system effectors, pectate lyase and metalloprotease in addition to the CRISPR-Cas based adaptive immune system were uniquely represented. Overall, the information generated through this study will contribute to decipher the mechanism of infection and adaptive immunity in Pcc.