Abstract
Inducible gene expression systems are valuable tools for studying biological processes. We previously developed an optogenetic gene expression system called TAEL that is optimized for use in zebrafish. When illuminated with blue light, TAEL transcription factors dimerize and activate gene expression downstream of the TAEL-responsive C120 promoter. By using light as the inducing agent, the TAEL/C120 system overcomes limitations of traditional inducible expression systems by enabling fine spatial and temporal regulation of gene expression. Here, we describe ongoing efforts to improve the TAEL/C120 system. We made modifications to both the TAEL transcriptional activator and the C120 regulatory element, collectively referred to as TAEL 2.0. We demonstrate that TAEL 2.0 consistently induces higher levels of reporter gene expression and at a faster rate, but with comparable background and toxicity as the original TAEL system. With these improvements, we were able to create functional stable transgenic lines to express the TAEL 2.0 transcription factor either ubiquitously or with a tissue-specific promoter. We demonstrate that the ubiquitous line in particular can be used to induce expression at late embryonic and larval stages, addressing a major deficiency of the original TAEL system. This improved optogenetic expression system will be a broadly useful resource for the zebrafish community.
Competing Interest Statement
L.B.M-M. and K.H.G were co-founders of Optologix, Inc., which developed light-gated transcription factors for research applications. As of September 2020, Optologix, Inc. has ceased business.
Footnotes
As suggested by peer reviewers, we have made the following revisions: 1. Clarified our reporting of statistical error in our analysis of the basal activity of the C120 promoter (p. 12, p. 13, and Fig. 6). 2. Revised the text and accompanying figure for our time course data to more accurately describe our experimental design, data analysis, and interpretation (p. 9-10 and Fig. 3). 3. Added a discussion of other optogenetic expression systems and how TAEL 2.0 compares to them (p. 15-16). 4. Revised panels A and B in Figures 1 and 2 to improve readability. 5. Revised our discussion of basal promoter choice to better highlight previous work done in this area (p. 14-15). 6. Included information about plasmid distribution (p. 5). 7. Ensured that all reported genetic elements adhere to the Zebrafish Nomenclature Conventions established by ZFIN. 8. Expanded our discussion of the ectopic liver fluorescence observed in the Tg(C120F:GFP) line (p. 11).