Iterative, multiplexed CRISPR-mediated gene editing for functional analysis of complex protease gene clusters.

Elucidation of gene function by reverse genetics in animal models frequently is complicated by the functional redundancy of homologous genes. This obstacle often is compounded by the tight clustering of homologous genes, which precludes the generation of multigene-deficient animals through standard interbreeding of single-deficient animals. Here, we describe an iterative, multiplexed CRISPR-based approach for simultaneous gene editing in the complex seven-member human airway trypsin-like protease/differentially expressed in a squamous cell carcinoma (HAT/DESC) cluster of membrane-anchored serine proteases.

Matriptase zymogen supports epithelial development, homeostasis and regeneration.

Background: Matriptase is a membrane serine protease essential for epithelial development, homeostasis, and regeneration, as well as a central orchestrator of pathogenic pericellular signaling in the context of inflammatory and proliferative diseases. Matriptase is an unusual protease in that its zymogen displays measurable enzymatic activity.

Results: Here, we used gain and loss of function genetics to investigate the possible biological functions of zymogen matriptase.

An essential role for bacterial nitric oxide synthase in Staphylococcus aureus electron transfer and colonization.

Nitric oxide (NO) is a ubiquitous molecular mediator in biology. Many signalling actions of NO generated by mammalian NO synthase (NOS) result from targeting of the haem moiety of soluble guanylate cyclase. Some pathogenic and environmental bacteria also produce a NOS that is evolutionary related to the mammalian enzymes, but a bacterial haem-containing receptor for endogenous enzymatically generated NO has not been identified previously.