Design, Science and Technology

Researchers set out to develop a robust off-switch for the highly efficient Cas3 system they had previously discovered from Neisseria lactima, a bacterium that lives harmlessly in the human upper respiratory tract. Examining all known anti-CRISPRs that have been reported in the literature as inhibitors of other Cas3 variants from distinct bacterial organisms, they found two, AcrIC8 and AcrIC9, with a strong cross-reactive effect against Neisseria Cas3. Using genetic and biochemical studies at UM and cryogenic electron microscopy analyses at Cornell, they determined the mechanism of action and structure of AcrlC8 and AcrlC9 at the molecular level. Both proteins prevent the CRISPR-Cas3 machine from binding to its DNA target site, but by slightly different mechanisms. Finally, the team provided key proof-of-concept that each of these anti-CRISPR proteins can act as a switch for CRISPR-Cas3 in human cells. They can almost completely block two versions of CRISPR-Cas3 technologies, one for deletion of important genomes and the other for gene activation, making them the first switches developed for any CRISPR-Cas3 gene editor. 

CRISPR/Cas9 method can lead to unintended DNA mutations that can have negative effects. Recently, Japanese researchers have developed a new gene-editing technique that is as effective as CRISPR/Cas9, yet significantly reduces these unintended mutations. In a new study published in Nature Communications , researchers led by Osaka University have introduced a new technique called NICER, based on the creation of several small cuts in single DNA strands by an enzyme  Cas 9 nickase. For their first experiments, the research team used human lymphoblastic cells with a known heterozygous mutation in a gene called TK1. When these cells were treated with nickase to induce a single cut in the TK1 region, TK1 activity was recovered at a low rate. However, when nickase induced multiple cuts in this region on both homologous chromosomes, the efficiency of gene correction was increased approximately seventeen-fold via activation of a cellular repair mechanism. Because the NICER method does not involve DNA double-strand breaks or the use of exogenous DNA, this technique appears to be a safe alternative to conventional CRISPR/Cas9 methods.