The Varble Lab is pleased to be able to share our research and findings.
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2022
The CRISPR-Cas system of Streptococcus pyogenes: function and applications
Andrew Varble, Luciano Marraffini
Here we review how the Cas9 nuclease mediates anti-phage immunity and how it can be repurposed for the genetic engineering of human cells and other eukaryotic organisms.
Cleavage of viral DNA by restriction endonucleases stimulates the type II CRISPR-Cas immune response
Pascal Maguin, Andrew Varble, Joshua W Modell, Luciano A Marraffini
We found that restriction endonucleases provide a short-term defense, which is rapidly overcome through methylation of the phage genome. In a small fraction of the cells, however, restriction results in the acquisition of spacer sequences from the cleavage site, which mediates a robust type II-A CRISPR-Cas immune response against the methylated phage
2021
Andrew Varble, Edmondo Campisi, Chad W Euler, Pascal Maguin, Albina Kozlova, Jessica Fyodorova, Jakob T Rostøl, Vincent A Fischetti, Luciano A Marraffini
Here we demonstrate that the ΦAP1.1 temperate phage utilizes an alternative approach to antagonize the type II-A CRISPR response in Streptococcus pyogenes. Immediately after infection, this phage expresses a small anti-CRISPR protein, AcrIIA23, that prevents Cas9 function, allowing ΦAP1.1 to integrate into the direct repeats of the CRISPR locus, neutralizing immunity.
RecT Recombinase Expression Enables Efficient Gene Editing in Enterococcus spp
Victor Chen, Matthew E Griffin, Pascal Maguin, Andrew Varble, Howard C Hang
The dissection of E. faecium functions and mechanisms has been restricted by inefficient gene-editing methods. To address these limitations, here, we report that the expression of E. faecium RecT recombinase significantly improves the efficiency of recombineering technologies in both commensal and antibiotic-resistant strains of E. faecium and other Enterococcus species such as E. durans and E. hirae.
Type III-A CRISPR immunity promotes mutagenesis of staphylococci
Charlie Y Mo, Jacob Mathai, Jakob T Rostøl, Andrew Varble,, Dalton V Banh, Luciano A Marraffini
Here we show that the non-specific DNase activity of the staphylococcal type III-A CRISPR-Cas system increases mutations in the host and accelerates the generation of antibiotic resistance in Staphylococcus aureus and Staphylococcus epidermidis.
2019
Three New Cs for CRISPR: Collateral, Communicate, Cooperate
Andrew Varble, Luciano A Marraffini
Recent work in the field has revealed unexpected features of the CRISPR-Cas mechanism: (i) collateral, nonspecific, cleavage of host nucleic acids; (ii) secondary messengers that amplify the immune response; and (iii) immunosuppression of CRISPR targeting by phage-encoded inhibitors. Here, we review these new and exciting findings.
Andrew Varble, Sean Meaden, Rodolphe Barrangou, Edze R Westra, Luciano A Marraffini
Although CRISPR-cas loci are widely distributed throughout microbial genomes and often display hallmarks of horizontal gene transfer10-12, the drivers of CRISPR dissemination remain unclear. Here, we show that spacers can recombine with phage target sequences to mediate a form of specialized transduction of CRISPR elements
2018
Gregory W Goldberg, Elizabeth A McMillan, Andrew Varble, Joshua W Modell, Poulami Samai, Wenyan Jiang, Luciano A Marraffini
Here we show that maintenance of conditionally tolerant type III-A systems can produce fitness costs within populations of Staphylococcus aureus lysogens.
Broad Targeting Specificity during Bacterial Type III CRISPR-Cas Immunity Constrains Viral Escape
Nora C Pyenson, Kaitlyn Gayvert, Andrew Varble, Olivier Elemento, Luciano A Marraffini
Here we show that targeting by the Staphylococcus epidermidis type III-A CRISPR-Cas system does not require PAM or seed sequences, and thus prevents viral escape via single-nucleotide substitutions. Instead, viral escapers can only arise through complete target deletion.