Move over Cas9; here comes yet another CRISPR-associated player. The CRISPR pioneers in the lab of Feng Zhang at MIT continue to discover the power of CRISPR , bringing us a fancy new part for genome editing. Recently, they changed things up with a slimmer Cas9 from Staphylococcus pyogenes (spCas9) but now it seems they’ve found a new star of a whole new type.
CRISPR from Prevotella and Francisella 1 (Cpf1) is a unique CRISPR effector. In the bacterial immune system, Cpf1 doesn’t require the tracRNA, which means only the cRNA is needed. When it comes to genome editing, this means a smaller sgRNA molecule is required (~42 nt compared ~100 nt), on top of the fact that Cpf1 is smaller than spCas9. However, the new features don’t end there.
Here’s what Cpf1 has to offer to genome editing:
- It recognizes a T-rich protospacer-adjacent motif (PAM) as opposed to the G-rich PAM of Cas9, which enables new targeting possibilities in the genome.
- When editing, Cpf1 creates sticky ends (4-5 nt), rather than the blunt ends of Cas9, which could give Cpf1 a leg up over Cas9 when it comes to ensuring proper orientation during the tricky insertions of non-homologous end joining (NHEJ).
- This could also be advantageous in non-dividing cell types that don’t like to do the homology-directed repair (HDR) route that is Cas9’s claim to fame.
- Interestingly, when Cpf1 cleaves, it does so further away from PAM than Cas9, which is also further away from the target site, leaving open the potential for a second round of cleavage if the correct repair pathway doesn’t happen the first time.
- After a bit of codon optimization, two Cpf1 orthologs (from Acidaminococcus and Lachnospiraceae) were used to efficiently target DNMT1 in human cells.
Zhang concludes that “We are committed to making the CRISPR-Cpf1 technology widely accessible. Our goal is to develop tools that can accelerate research and eventually lead to new therapeutic applications. We see much more to come, even beyond Cpf1 and Cas9, with other enzymes that may be repurposed for further genome editing advances.”
Go check out next-gen CRISPR-based genome editing in Cell, September 2015