Roche, P. | Search bioRxiv

bioRxiv2018

Receptor Mediated Delivery of Cas9-Nanobody Induces Cisplatin Synthetic Dose Sensitivity

The CRISPR/Cas9 system has shown great potential for precisely editing genomic DNA sequences by introducing site-specific DNA cuts that are subsequently repaired by the cell. However, delivery of the CRISPR ribonucleoprotein remains an understudied area and hinders realizing the full potential of the system. We prepared Cas9 ribonucleoprotein complexes chemically conjugated to the 7D12 nanobody and demonstrate receptor-mediated transfection of Cas9 into A549 non-small-cell lung cancer cells via binding to the epithelial growth factor receptor for subsequent cell internalization. We further show that transfection with a Cas9 ribonucleoprotein targeting the BRCA2 gene results in an enhanced sensitivity to the chemotherapeutic drug Cisplatin, and thereby induces a synthetic dose lethality in A549 cells.

biochemistry↗

bioRxiv2017

A divergent CheW stabilizes nucleoid-associated chemosensory arrays

Chemosensory systems are highly organized signaling pathways that allow bacteria to adapt to environmental changes. The Frz chemosensory system from M. xanthus possesses two CheW-like proteins, FrzA (the core CheW) and FrzB. We found that FrzB does not interact with FrzE (the cognate CheA) as it lacks the amino acid region responsible for this interaction. FrzB, instead, acts upstream of FrzCD in the regulation of M. xanthus chemotaxis behaviors and activates the Frz pathway by allowing the formation and distribution of multiple chemosensory clusters on the nucleoid. These results, together, show that the lack of the CheA-interacting region in FrzB confers new functions to this small protein.\n\nAUTHOR SUMMARYChemosensory systems are signaling complexes that are widespread in bacteria and allow the modulation of different cellular functions, such as taxis and development, in response to the environment. We show that the Myxococcus xanthus FrzB is a divergent CheW lacking the region involved in the interaction with the histidine kinase FrzE. Instead, it acts upstream of FrzCD to allow the formation of multiple distributed Frz chemosensory arrays at the nucleoid. The loss of the CheA-interacting region in FrzB might have been selected to confer plasticity to nucleoid-associated chemosensory systems. By unraveling a new accessory protein and its function, this work opens new insights into the knowledge of the regulatory potentials of bacterial chemosensory systems.

microbiology↗