CRISPR-Cas9 genomic target locus (CRISPR target)

Cas9–guide-RNA–specified double-stranded DNA loci are the fundamental genomic targets for CRISPR-Cas9-based therapeutic interventions (Jinek et al., 2012). These loci are defined by the base-pairing of a chimeric single-guide RNA (sgRNA) to a complementary protospacer sequence in the host genome, typically 20 nucleotides in length and immediately upstream of a protospacer adjacent motif (PAM) (Doudna & Charpentier, 2014). The Cas9 endonuclease undergoes a conformational change upon binding, positioning its RuvC and HNH nuclease domains to generate a site-specific double-strand break (DSB). This DSB triggers endogenous DNA repair pathways, such as non-homologous end joining (NHEJ) for gene disruption or homology-directed repair (HDR) for precise sequence correction. Clinically, these targets are leveraged to silence disease-causing genes or restore functional protein expression, as seen in the FDA-approved treatment for sickle cell disease (Frangoul et al., 2021). For example, Exagamglogene autotemcel targets the BCL11A enhancer to induce fetal hemoglobin. The primary challenge in targeting these loci remains the potential for off-target activity at homologous sites, necessitating advanced computational prediction and high-throughput sequencing for safety validation (Fu et al., 2013).

Other names

ProtospacergRNA-targeted DNACas9 target siteGenomic target sequenceCas9-gRNA-DNA complex

Targeted induction of double-strand DNA breaks to facilitate gene knockout, correction, or insertion via cellular repair pathways.