@article{nethery_hidalgo-cantabrana_roberts_barrangou_2022, title={CRISPR-based engineering of phages for in situ bacterial base editing}, volume={119}, ISSN={["1091-6490"]}, url={https://doi.org/10.1073/pnas.2206744119}, DOI={10.1073/pnas.2206744119}, abstractNote={Significance Microbial consortia possess a wealth of genetic information; however, current approaches to deciphering genetics in a community context are largely constrained by low delivery efficiencies and a breadth of targeting specificities. Here, we describe a phage-based approach for precision editing of an individual gene within a target host organism in a community context. Engineering bacteriophage λ to contain a cytosine base editor, we inactivate host chromosomal and plasmid-based genetic targets through the strategic introduction of single-nucleotide mutations without Cas-based double-strand DNA cleavage. Next, we establish species- and site-specific editing within a synthetic soil community by using a fabricated ecosystem. Phage-assisted delivery, together with base editing, offers an important in situ editing approach for the genetic interrogation of microbial community members.}, number={46}, journal={PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA}, author={Nethery, Matthew A. and Hidalgo-Cantabrana, Claudio and Roberts, Avery and Barrangou, Rodolphe}, year={2022}, month={Nov} }
 @article{roberts_nethery_barrangou_2022, title={Functional characterization of diverse type I-F CRISPR-associated transposons}, volume={11}, ISSN={["1362-4962"]}, url={https://doi.org/10.1093/nar/gkac985}, DOI={10.1093/nar/gkac985}, abstractNote={Abstract CRISPR-Cas systems generally provide adaptive immunity in prokaryotes through RNA-guided degradation of foreign genetic elements like bacteriophages and plasmids. Recently, however, transposon-encoded and nuclease-deficient CRISPR-Cas systems were characterized and shown to be co-opted by Tn7-like transposons for CRISPR RNA-guided DNA transposition. As a genome engineering tool, these CRISPR-Cas systems and their associated transposon proteins can be deployed for programmable, site-specific integration of sizable cargo DNA, circumventing the need for DNA cleavage and homology-directed repair involving endogenous repair machinery. Here, we selected a diverse set of type I-F3 CRISPR-associated transposon systems derived from Gammaproteobacteria, predicted all components essential for transposition activity, and deployed them for functionality testing within Escherichia coli. Our results demonstrate that these systems possess a significant range of integration efficiencies with regards to temperature, transposon size, and flexible PAM requirements. Additionally, our findings support the categorization of these systems into functional compatibility groups for efficient and orthogonal RNA-guided DNA integration. This work expands the CRISPR-based toolbox with new CRISPR RNA-guided DNA integrases that can be applied to complex and extensive genome engineering efforts.}, journal={NUCLEIC ACIDS RESEARCH}, author={Roberts, Avery and Nethery, Matthew A. and Barrangou, Rodolphe}, year={2022}, month={Nov} }
 @article{pan_morovic_hidalgo-cantabrana_roberts_walden_goh_barrangou_2022, title={Genomic and epigenetic landscapes drive CRISPR-based genome editing in Bifidobacterium}, volume={119}, ISSN={["1091-6490"]}, url={https://doi.org/10.1073/pnas.2205068119}, DOI={10.1073/pnas.2205068119}, abstractNote={Significance Genome engineering in Bifidobacterium remains challenging, despite successful CRISPR deployment in other bacteria, hindering the comprehensive understanding of the molecular mechanisms of health-promoting effects. In this study, we adapted existing CRISPR effectors and editing strategies to generate a variety of editing outcomes in B. animalis subsp. lactis. We also showed that the editing efficiency and genome accessibility are shaped by the genomic and epigenetic landscapes, despite their monomorphic genomes. Together, these findings emphasize that strain-to-strain variation can impact the deployment of genome editing and lead to phenotypic and functional differences. This study expands the genome editing platform of Bifidobacterium to enhance their probiotic efficacy and opens opportunities for the engineering of bacterial biotherapeutics.}, number={30}, journal={PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA}, author={Pan, Meichen and Morovic, Wesley and Hidalgo-Cantabrana, Claudio and Roberts, Avery and Walden, Kimberly K. O. and Goh, Yong Jun and Barrangou, Rodolphe}, year={2022}, month={Jul} }