Abstract
To study the molecular mechanism of complex biological systems, it is important to be able to artificially manipulate gene expression in desired target sites with high precision. Based on the light dependent binding of cryptochrome 2 and a cryptochrome interacting bHLH protein, we developed a split lexA transcriptional activation system for use in Drosophila that allows regulation of gene expression in vivo using blue light or two-photon excitation. We show that this system offers high spatiotemporal resolution by inducing gene expression in tissues at various developmental stages. In combination with two-photon excitation, gene expression can be manipulated at precise sites in embryos, potentially offering an important tool with which to examine developmental processes.
Highlights
Received: May 14, 2015Accepted: August 27, 2015Published: September 18, 2015
Significant GFP expression was observed in both cases after one-hour of continuous exposure to blue light (474nm, ~2.5mWcm-2) (Fig 1B)
A greater fold change in cell number and a lower basal activity in the dark was observed with the p65 activation domain (Fig 1C). This activation domain was used in our cryptochrome 2 (CRY2) constructs in all subsequent experiments
Summary
Data Availability Statement: All relevant data are within the paper and its Supporting Information files. Several promising optogenetic approaches have been developed that focus on light sensitive proteins These offer unmatched advantages in manipulating gene expression in vivo since light is non invasive and can be directed with exceptionally high spatiotemporal resolution. Light-Inducible Transcription System in Drosophila systems of this type already have been developed [4,5] These provide the advantage of rapid stimulation and reversibility. We demonstrated the adaptability and functionality of this system using GAL4 line to target different neural populations with GCaMP3.0 reporter and functional effectors like TrpA1 This system provides a novel approach to regulate gene expression and is especially suitable for neurodevelopmental studies that require precise spatial and temporal regulation
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
