CreLite: An Optogenetically Controlled Cre/loxP System Using Red Light.
Abstract:
Precise manipulation of gene expression with temporal and spatial control is essential for functional analysis and determining cell lineage relationships in complex biological systems. The Cre-loxP system is commonly used for gene manipulation at desired times and places. However, specificity is dependent on the availability of tissue- or cell-specific regulatory elements used in combination with Cre. Here we present CreLite, an optogenetically-controlled Cre system using red light in developing zebrafish embryos. Cre activity is disabled by splitting Cre and fusing with the Arabidopsis thaliana red light-inducible binding partners, PhyB and PIF6. Upon red light illumination, the PhyB-CreC and PIF6-CreN fusion proteins come together in the presence of the cofactor phycocyanobilin (PCB) to restore Cre activity. Red light exposure of zebrafish embryos harboring a Cre-dependent multi-color fluorescent protein reporter injected with CreLite mRNAs and PCB resulted in Cre activity as measured by the generation of multi-spectral cell labeling in several different tissues. Our data show that CreLite can be used for gene manipulations in whole embryos or small groups of cells at different developmental stages, and suggests CreLite may also be useful for temporal and spatial control of gene expression in cell culture, ex vivo organ culture, and other animal models. This article is protected by copyright. All rights reserved.
