Efficient photoactivatable Dre recombinase for cell type-specific spatiotemporal control of genome engineering in the mouse

Abstract

Precise genetic engineering in specific cell types within an intact organism is intriguing yet challenging, especially in a spatiotemporal manner without the interference caused by chemical inducers. Here we engineered a photoactivatable Dre recombinase based on the identification of an optimal split site and demonstrated that it efficiently regulated transgene expression in mouse tissues spatiotemporally upon blue light illumination. Moreover, through a double-floxed inverted open reading frame strategy, we developed a Cre-activated light-inducible Dre (CALID) system. Taking advantage of well-defined cell-type–specific promoters or a well-established Cre transgenic mouse strain, we demonstrated that the CALID system was able to activate endogenous reporter expression for either bulk or sparse labeling of CaMKIIα-positive excitatory neurons and parvalbumin interneurons in the brain. This flexible and tunable system could be a powerful tool for the dissection and modulation of developmental and genetic complexity in a wide range of biological systems.

Stable Transgenic Mouse Strain with Enhanced Photoactivatable Cre Recombinase for Spatiotemporal Genome Manipulation

“…Therefore, 48 h of pulse illumination (1 min pulse every 5 min) was approximately 8 h of continuous illumination which was shorter than a previous study which used 16 h of continuous illumination. [ 12 ] In addition, users can shorten the illumination time via increasing the illumination strength, extending the illumination pulse, or designing new illumination strategies by using an optical fiber [ 15 , 23 , 24 ] or two‐photon. [ 13 ] In summary, the data above demonstrated that the recombinase activity was activated efficiently and dramatically in various tissues upon blue light induction in ePA‐Cre: Ai14 mice without obvious background activity.…”

“…To date, several elegant photoactivatable recombinases have been developed mainly based on the CRY2‐CIB1 [ 10 ] and Magnets system, [ 28 ] such as PA‐Cre, PA‐Flp, and PA‐Dre, offering promising approaches for high‐resolution spatiotemporal transgenesis. [ 10 , 11 , 12 , 23 , 24 ] In this study, we optimized the PA‐Cre2.0 system leading to ePA‐Cre, which exhibited enhanced efficiency and reduced leakage compared with other tested PA‐Cre systems. A transgenic mouse strain containing the ePA‐Cre system was generated, and the Cre activity was stringently responsive to blue light illumination in multiple mouse tissues.…”

Plug-and-Display Photo-Switchable Systems on Plant Virus Nanoparticles

“…Further controls showed that antibody binding does not occur in the absence of BphP1-mCherry-His 6 ( Figure S2B,C,E,F ) and that neither far-red nor red light affects the particle morphology ( Figure S2G,H ). The BphP1/QPAS1 system has previously been used to control gene expression and protein activity in vivo [ 5 , 16 , 72 ] but again this is the first study to demonstrate the functionality of BphP1/QPAS1 on the surface of nanoparticles.…”

Optogenetics for transcriptional programming and genetic engineering

“…The in vivo feasibility was elegantly demonstrated in a rodent model to drive Cav3.1 silencing in the medial septum neurons to increase object-exploration behaviors [108]. Recently, photoactivatable Dre recombinases, named as PA-Dre and iDreV, were independently developed by two groups by using Magnet or VVD-based optical dimerizers for light-inducible restoration of Dre activity [109,110]. Both tools showed light-dependent DNA recombination events in rodent models.…”