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Spacer acquisition from RNA mediated by a natural reverse transcriptase-Cas1 fusion protein associated with a type III-D CRISPR-Cas system in Vibrio vulnificus. | LitMetric

Spacer acquisition from RNA mediated by a natural reverse transcriptase-Cas1 fusion protein associated with a type III-D CRISPR-Cas system in Vibrio vulnificus.

Nucleic Acids Res

Structure, Dynamics and Function of Rhizobacterial Genomes, Grupo de Ecología Genética de la Rizosfera, Department of Soil Microbiology and Symbiotic Systems, Estación Experimental del Zaidín, Consejo Superior de Investigaciones Científicas, C/ Profesor Albareda 1, 18008 Granada, Spain.

Published: November 2019

AI Article Synopsis

  • Recent studies reveal that reverse transcriptases (RTs) can help the CRISPR-Cas system acquire RNA-derived spacers, but how this works is not fully understood.
  • Researchers examined a specific RT-Cas1 fusion protein in a type III-D system from Vibrio vulnificus, showing that it could function effectively in a different host and that mutations in the RT active site reduced spacer acquisition.
  • The study identified that two Cas2 proteins are essential for the spacer acquisition process, which has no specific bias toward the ends of coding sequences, indicating a complex mechanism for integrating RNA-derived spacers into the CRISPR array.

Article Abstract

The association of reverse transcriptases (RTs) with CRISPR-Cas system has recently attracted interest because the RT activity appears to facilitate the RT-dependent acquisition of spacers from RNA molecules. However, our understanding of this spacer acquisition process remains limited. We characterized the in vivo acquisition of spacers mediated by an RT-Cas1 fusion protein linked to a type III-D system from Vibrio vulnificus strain YJ016, and showed that the adaptation module, consisting of the RT-Cas1 fusion, two different Cas2 proteins (A and B) and one of the two CRISPR arrays, was completely functional in a heterologous host. We found that mutations of the active site of the RT domain significantly decreased the acquisition of new spacers and showed that this RT-Cas1-associated adaptation module was able to incorporate spacers from RNA molecules into the CRISPR array. We demonstrated that the two Cas2 proteins of the adaptation module were required for spacer acquisition. Furthermore, we found that several sequence-specific features were required for the acquisition and integration of spacers derived from any region of the genome, with no bias along the 5'and 3'ends of coding sequences. This study provides new insight into the RT-Cas1 fusion protein-mediated acquisition of spacers from RNA molecules.

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Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6821258PMC
http://dx.doi.org/10.1093/nar/gkz746DOI Listing

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