CRISPR Converts Microbes to Tape Recorders

GEN News Highlights, 27 Nov 2017, https://www.genengnews.com/gen-news-highlights/crisprcas-used-to-create-microscopic-data-recorder/81255201

Scientists have created a CRISPR/Cas system in E. coli that allows for the recording of events and the time at which they occurred.  This was done by having the cell insert CRISPR spacers at regular occurring sets of time, however when a signal was detected the bacteria would insert a signal sequence instead.  This locus can then be read using pre-existing sequencing tools providing a readout of events.  The researchers hope this technology could be used to record biological events.

CRISPR May Not Revolutionize Mitochondrial Genome Editing

Gammage, P.A.  et. al. (2017) Trends in Genetics. https://www.ncbi.nlm.nih.gov/pubmed/29179920

Mitochondrial DNA has increased in prominence in both biology and medicine as a major player for various diseases.  Attempts to alter the genetic information of mitochondria have made significant advances using zinc finger and TALEN nucleases.  These protein only nucleases are easily delivered into the mitochondria through existing machinery.  While CRISPR has revolutionized genome editing, it is struggling in mitochondria due to the lack of an endogenous mechanism for nucleic acid transport into the mitochondria, preventing its adoption.

New Cas9 Cryo-EM Structure Solved

Huai, C. et. al. (2017) Nature Communications. 8:1375. https://www.ncbi.nlm.nih.gov/pubmed/29123204

Researchers have solved a 5.2 Å cryo-EM structure of Cas9 complexed with sgRNA and target DNA.  The structure found that the HNH domain moves closer to the DNA than previously reported and suggests that this new structure resembles a DNA cleavage-activating structure of Cas9.

Identification of a Thermostable Cas9

Harrington L.B. et. al. (2017) Nature Communications 8:1424. https://www.ncbi.nlm.nih.gov/pubmed/29127284

The CRISPR/Cas systems used for genome editing to date have come from mesophilic bacteria, preferring temperatures of 20-45°C, preventing their use at higher temperatures.  Harrington et. al. have identified a Cas9 protein from the thermophilic bacterium Geobacillus stearothermophilus (GeoCas9) that is active in temperature up to 70°C, providing a much wider range of possibilities.  Additionally, GeoCas9 showed greater stability as an RNP complex in human plasma, opening the door to possible therapeutic uses.

Could CRISPR/Cas Claims Inhibit Innovation?

Gray, B.N and Spruill, W.M. (2017) Nature Biotechnology 35:630-633. https://www.ncbi.nlm.nih.gov/pubmed/28700549

The ongoing patent battle between the Broad Institute and the University of California-Berkeley provides difficulties for researchers and companies wishing to develop CRISPR/Cas technology, though this is not the only barrier.  This article describes the broad claims that have been granted or that are being investigated and presents the argument that these claims are overly broad and could limit the genome editing field.

Using Gene Drives to Study Fungal Pathogenicity

GEN News Highlights, 30 October 2017, https://www.genengnews.com/gen-news-highlights/crispr-drives-out-fungal-resistance/81255106

Gene drives have been described as a way to eliminate pests, notably the mosquito, from the environment. However, they are a powerful research tool as well.  Candida albicans can be notoriously difficult to study due to its diploid nature.  By combining a newly discovered haploid C. albicans and CRISPR/Cas gene drive technology, researchers have been able to rapidly create diploid knockouts for study (https://www.ncbi.nlm.nih.gov/pubmed/29062088).  The creation of these mutants could serve to increase the pace of drug discovery to combat this and other fungal pathogens.

Model Predicts Relationship Between Editing Speed and Off-Target Effects

David Ruth, 17 October 2017, PHYS.org, https://phys.org/news/2017-10-genome-efficient.html

Researchers at Rice University have used computational models to predict the speed at which the CRISPR/Cas system identifies and cleaves the targeted location.  The research, published in the Biophysical Journal (https://www.ncbi.nlm.nih.gov/pubmed/28978436), determined that by allowing CRISPR to cut at off-target sites the system could quickly find and cleave the targeted site.  By limiting the system’s ability to cleave off-target sites, the dissociation of Cas9 from DNA greatly decreased the speed at which on-target sites were identified.

Gold Nanoparticles Deliver CRISPR in Muscular Dystrophy Treatment

Sophia Ktori, GEN News Highlights, 04 October 2017, http://www.genengnews.com/gen-news-highlights/crispr-nanoparticles-repair-duchenne-muscular-dystrophy-gene/81255009

Scientists working on CRISPR delivery have developed a gold nanoparticle that encapsulates the CRISPR/Cas machinery for delivery to cells.  This new technique, coined CRISPR-Gold, was published in Nature Biomedical Engineering (https://www.nature.com/articles/s41551-017-0137-2).  In the paper the authors demonstrated CRISPR-Gold’s ability to correct the mutated dystrophin gene in a mouse model, with mice receiving CRISPR-Gold treatment displaying two-fold improvement in hanging time in a four-limb hanging test, compared to control mice.

Chinese Research Team Fix Genetic Disorder in Human Embryos

David Cyranoski, 02 October 2017, Nature News, http://www.nature.com/news/chinese-scientists-fix-genetic-disorder-in-cloned-human-embryos-1.22694

A new report in Protein and Cell (https://www.ncbi.nlm.nih.gov/pubmed/28942539) is the latest in a string of human embryo CRISPR publications.  Using a modified CRISPR/Cas9 system tethered to a second enzyme that can swap individual DNA bases, the researchers targeted an A to G point mutation that results in β-thalassemia.  Eight of the 20 cloned embryos contained a corrected copy of the gene, possibly curing the recessive disorder.  The scientists were careful to point out that not all cells in the embryo were modified, which could have unintended consequences.

New Cas9 Mutations Increase Accuracy

Chen et. al. (2017) Nature. https://www.ncbi.nlm.nih.gov/pubmed/28931002

Researchers using FRET to study previously engineered high-fidelity Cas9 (SpCas9-HF1) and enhanced Cas9 (eSpCas9), identified that these versions are trapped in an inactive state when bound to off-target sites.  Using this observation and rational protein engineering, the researchers made additional modifications to the REC3 domain to prevent activation of the HNH nuclease domain unless the guide RNA and target DNA match is very close.  This new Cas9, coined Hyper Cas9 (HypaCas9) maintains the native Cas9 on target efficiency, but decreases the number of off-target events.