Inducible CRISPR-targeted "knockdown" of human gut in gnotobiotic mice discloses glycan utilization strategies.

Understanding how members of the human gut microbiota prioritize nutrient resources is one component of a larger effort to decipher the mechanisms defining microbial community robustness and resiliency in health and disease. This knowledge is foundational for development of microbiota-directed therapeutics. To model how bacteria prioritize glycans in the gut, germfree mice were colonized with 13 human gut bacterial strains, including seven saccharolytic species.

Improving CRISPR-Cas9 Genome Editing Efficiency by Fusion with Chromatin-Modulating Peptides.

Bacterial-derived CRISPR-Cas9 nucleases have become a common tool in genome engineering. However, the editing efficiency by even the best-crafted Cas9 nucleases varies considerably with different genomic sites, and efforts to explore the vast natural Cas9 diversity have often met with mixed or little success. Here, we show that modification of the widely used Cas9 by fusion with chromatin-modulating peptides (CMPs), derived from high mobility group proteins HMGN1 and HMGB1, histone H1, and chromatin remodeling complexes, improves its activity by up to several fold, particularly on refractory target sites.

Improving CRISPR Gene Editing Efficiency by Proximal dCas9 Targeting.

Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and CRISPR-associated (Cas) systems function as an adaptive immune system in bacteria and archaea for defense against invading viruses and plasmids (Barrangou and Marraffini, 2014). The effector nucleases from some class 2 CRISPR-Cas systems have been repurposed for heterologous targeting in eukaryotic cells ( Jinek , 2012 ; Cong , 2013 ; Mali , 2013 ; Zetsche , 2015 ). However, the genomic environments of eukaryotes are distinctively different from that of prokaryotes in which CRISPR-Cas systems have evolved.

Targeted activation of diverse CRISPR-Cas systems for mammalian genome editing via proximal CRISPR targeting.

Bacterial CRISPR-Cas systems comprise diverse effector endonucleases with different targeting ranges, specificities and enzymatic properties, but many of them are inactive in mammalian cells and are thus precluded from genome-editing applications. Here we show that the type II-B FnCas9 from Francisella novicida possesses novel properties, but its nuclease function is frequently inhibited at many genomic loci in living human cells. Moreover, we develop a proximal CRISPR (termed proxy-CRISPR) targeting method that restores FnCas9 nuclease activity in a target-specific manner.

CYP709B3, a cytochrome P450 monooxygenase gene involved in salt tolerance in Arabidopsis thaliana.

Background: Within the Arabidopsis genome, there are 272 cytochrome P450 monooxygenase (P450) genes. However, the biological functions of the majority of these P450s remain unknown. The CYP709B family of P450s includes three gene members, CYP709B1, CYP709B2 and CYP709B3, which have high amino acid sequence similarity and lack reports elucidating biological functions.