Identifying and Understanding the Function of CRISPR Leader-Repeat Sequences

Pushya Krishna, Andrew Santiago-Frangos and Dr. Blake Wiedenheft, Department of Microbiology and Immunology, Montana State University, 100 Culbertson Hall, Bozeman MT 59717

Clustered Regularly Interspersed Short Palindromic Repeats (CRISPR) and associated genes (cas) are essential components of diverse adaptive immune systems that defend bacteria and archaea from viral and plasmid infection. During CRISPR adaptation, newly encountered viral and plasmid DNA is integrated as a new spacer into the “leader” end of the CRISPR locus. In Type I-E CRISPR systems, the Integration Host Factor (IHF) binds to and kinks the leader, recruiting an upstream motif that helps dock the Cas1-2 integrase complex onto the first repeat of the CRISPR locus. To determine the prevalence of this IHF-dependent mechanism of CRISPR expansion, I used bioinformatic methods to analyze 15,274 CRISPR leader sequences for the presence of conserved IHF binding sites and upstream motifs. Results from these experiments demonstrate phased distribution of IHF and upstream motif sites in subsets of I-E, I-F, I-C and II-C leaders. Furthermore, in vitro biochemical assays reveal that motif phase, rather than motif proximity to the leader-repeat junction, is critical to CRISPR evolution.

Additional Abstract Information

Presenter: Pushya Krishna

Institution: Montana State University Bozeman

Type: Poster

Subject: Microbiology

Status: Approved

Time and Location

Session: Poster 8
Date/Time: Tue 5:00pm-6:00pm
Session Number: 5628