Engineering of CRISPR-Cas12b for human genome editing.

Nat Commun
Authors
Keywords
Abstract

The type-V CRISPR effector Cas12b (formerly known as C2c1) has been challenging to develop for genome editing in human cells, at least in part due to the high temperature requirement of the characterized family members. Here we explore the diversity of the Cas12b family and identify a promising candidate for human gene editing from Bacillus hisashii, BhCas12b. However, at 37 °C, wild-type BhCas12b preferentially nicks the non-target DNA strand instead of forming a double strand break, leading to lower editing efficiency. Using a combination of approaches, we identify gain-of-function mutations for BhCas12b that overcome this limitation. Mutant BhCas12b facilitates robust genome editing in human cell lines and ex vivo in primary human T cells, and exhibits greater specificity compared to S. pyogenes Cas9. This work establishes a third RNA-guided nuclease platform, in addition to Cas9 and Cpf1/Cas12a, for genome editing in human cells.

Year of Publication
2019
Journal
Nat Commun
Volume
10
Issue
1
Pages
212
Date Published
2019 01 22
ISSN
2041-1723
DOI
10.1038/s41467-018-08224-4
PubMed ID
30670702
PubMed Central ID
PMC6342934
Links
Grant list
DP1 HL141201 / HL / NHLBI NIH HHS / United States
R01 HG009761 / HG / NHGRI NIH HHS / United States
R01 MH110049 / MH / NIMH NIH HHS / United States