Source 1primary paper2022International Journal of Molecular Sciences
Toolkit/VP16-EL222 light-responsive transcription factor
VP16-EL222 light-responsive transcription factor
Also known as: EL222-VP16-based transcription factor, light-responsive element, TF
Taxonomy: Mechanism Branch / Architecture. Workflows sit above the mechanism and technique branches rather than replacing them.
Summary
VP16-EL222 is a chimeric blue light-responsive transcription factor composed of the photosensitive protein EL222 fused to the transcriptional activator VP16. In Yarrowia lipolytica, it functions with a promoter containing (C120)5 and the minimal promoter CYC102 to drive blue light-induced GFPMut3 expression.
Usefulness & Problems
Why this is useful
This construct provides a single-component optogenetic transcription system for regulating gene expression with blue light in Yarrowia lipolytica. It is useful where inducible transcription is needed without relying on a multi-component light-sensing circuit, although the supplied evidence is limited to reporter activation.
Problem solved
It addresses the problem of achieving light-controlled transcriptional induction in Yarrowia lipolytica using a defined transcription factor-promoter pair. The reported system links blue light input to activation of a target promoter driving GFPMut3 expression.
Problem links
provides the light-responsive transcriptional control element for the system
LiteratureIt solves the need for a genetically encoded light-responsive transcription factor that can drive inducible expression. In the paper it underlies the blue-light sensor used to activate reporter and functional genes.
Source:
It solves the need for a genetically encoded light-responsive transcription factor that can drive inducible expression. In the paper it underlies the blue-light sensor used to activate reporter and functional genes.
Published Workflows
Objective: Construct and characterize a single-component blue-light-induced gene expression system in Yarrowia lipolytica and demonstrate its ability to control expression of reporter and functional proteins.
Why it works: The workflow couples a blue-light-responsive transcription factor to a responsive promoter and first verifies inducible reporter output before applying the system to a functional protein, allowing the authors to establish both controllability and practical expression capability.
blue-light-responsive transcriptional activation via an EL222- and VP16-based transcription factoroperator/promoter-mediated induction using (C120)5 and minimal promoter CYC102genetic system constructionreporter-based characterizationlight dose and periodicity testingfunctional protein expression verification
Stages
- 1.System construction(library_build)
This stage creates the genetic system needed for subsequent light-response testing in Yarrowia lipolytica.
Selection: Assembly of a blue-light-induced expression system in Yarrowia lipolytica using an EL222- and VP16-based transcription factor and responsive promoter elements.
- 2.Reporter-based light response characterization(functional_characterization)
This stage verifies that the constructed system functions as a blue-light-inducible transcriptional sensor before applying it to a functional protein.
Selection: Ability of the TF plus (C120)5 and minimal promoter CYC102 to respond to blue light and initiate GFPMut3 expression.
- 3.Light dose and periodicity testing(secondary_characterization)
This stage assesses how illumination parameters affect system control properties beyond basic reporter induction.
Selection: Effects of light dose and periodicity on system behavior.
- 4.Functional protein validation with BleoR(confirmatory_validation)
This stage confirms that the system can control expression of a functional non-reporter protein, not just a fluorescent reporter.
Selection: Ability of the light-controlled system to drive synthesis and functional verification of BleoR.
Steps
- 1.Construct the blue-light-induced expression system in Yarrowia lipolyticaengineered optogenetic system and core components
Build the EL222/VP16-based light-responsive transcription system and responsive promoter architecture in the host yeast.
The system must be constructed before its light responsiveness and controllability can be tested.
- 2.Test blue-light-induced GFPMut3 expression from the responsive sensorsystem under test
Verify that the constructed sensor responds to blue light and initiates reporter expression.
Reporter expression provides an initial functional readout before more application-oriented validation.
- 3.Investigate effects of light dose and periodicity on system behaviorsystem under characterization
Assess spatial and temporal controllability of the light-controlled system.
After establishing basic reporter induction, illumination parameters are varied to characterize control behavior.
- 4.Use the light-controlled system for BleoR synthesis and functional verificationexpression control system
Demonstrate that the system can drive expression of a functional protein beyond a reporter.
Functional protein validation follows reporter and controllability characterization to confirm application utility.
Taxonomy & Function
Primary hierarchy
Mechanism Branch
Architecture: A reusable architecture pattern for arranging parts into an engineered system.
Techniques
No technique tags yet.
Target processes
transcriptionInput: Light
Implementation Constraints
cofactor dependency: cofactor requirement unknownencoding mode: genetically encodedimplementation constraint: context specific validationimplementation constraint: spectral hardware requirementoperating role: sensor
The reported implementation uses a chimeric transcription factor built from EL222 and VP16 together with a promoter containing five C120 elements upstream of the minimal promoter CYC102. Validation was reported in Yarrowia lipolytica using GFPMut3 as the output under blue light.
The supplied evidence only documents reporter induction in one host organism and does not provide broader validation across targets, conditions, or organisms. Quantitative performance characteristics, reversibility, dynamic range, kinetics, and off-state leakiness are not described in the provided evidence.
Validation
Cell-freeBacteriaMammalianMouseHumanTherapeuticIndep. Replication
Supporting Sources
Ranked Claims
The light-controlled system was used for synthesis and functional verification of Bleomycin resistance protein (BleoR).
The light-controlled system was used for synthesis and functional verification of Bleomycin resistance protein (BleoR).
The light-controlled system was used for synthesis and functional verification of Bleomycin resistance protein (BleoR).
The light-controlled system was used for synthesis and functional verification of Bleomycin resistance protein (BleoR).
The light-controlled system was used for synthesis and functional verification of Bleomycin resistance protein (BleoR).
The system showed good spatial and temporal controllability based on light dose and periodicity experiments.
The system showed good spatial and temporal controllability based on light dose and periodicity experiments.
The system showed good spatial and temporal controllability based on light dose and periodicity experiments.
The system showed good spatial and temporal controllability based on light dose and periodicity experiments.
The system showed good spatial and temporal controllability based on light dose and periodicity experiments.
The TF plus (C120)5 and minimal promoter CYC102 responds to blue light and initiates GFPMut3 expression.
The TF plus (C120)5 and minimal promoter CYC102 responds to blue light and initiates GFPMut3 expression.
The TF plus (C120)5 and minimal promoter CYC102 responds to blue light and initiates GFPMut3 expression.
The TF plus (C120)5 and minimal promoter CYC102 responds to blue light and initiates GFPMut3 expression.
The TF plus (C120)5 and minimal promoter CYC102 responds to blue light and initiates GFPMut3 expression.
The core light-responsive transcription factor of the system is constructed from EL222 and VP16.
The core light-responsive transcription factor of the system is constructed from EL222 and VP16.
The core light-responsive transcription factor of the system is constructed from EL222 and VP16.
The core light-responsive transcription factor of the system is constructed from EL222 and VP16.
The core light-responsive transcription factor of the system is constructed from EL222 and VP16.
With four copies of the responsive promoter and reporter gene assembled, the system produced a 128.5-fold higher fluorescent signal than dark conditions after 8 hours of induction.
fluorescent signal increase versus dark 128.5 fold
With four copies of the responsive promoter and reporter gene assembled, the system produced a 128.5-fold higher fluorescent signal than dark conditions after 8 hours of induction.
fluorescent signal increase versus dark 128.5 fold
With four copies of the responsive promoter and reporter gene assembled, the system produced a 128.5-fold higher fluorescent signal than dark conditions after 8 hours of induction.
fluorescent signal increase versus dark 128.5 fold
With four copies of the responsive promoter and reporter gene assembled, the system produced a 128.5-fold higher fluorescent signal than dark conditions after 8 hours of induction.
fluorescent signal increase versus dark 128.5 fold
With four copies of the responsive promoter and reporter gene assembled, the system produced a 128.5-fold higher fluorescent signal than dark conditions after 8 hours of induction.
fluorescent signal increase versus dark 128.5 fold
A blue-light-induced expression system based on EL222 was constructed in Yarrowia lipolytica.
A blue-light-induced expression system based on EL222 was constructed in Yarrowia lipolytica.
A blue-light-induced expression system based on EL222 was constructed in Yarrowia lipolytica.
A blue-light-induced expression system based on EL222 was constructed in Yarrowia lipolytica.
A blue-light-induced expression system based on EL222 was constructed in Yarrowia lipolytica.
Approval Evidence
1 source2 linked approval claimsfirst-pass slug vp16-el222-light-responsive-transcription-factor
The core of the blue light-induced system, the light-responsive element (TF), is constructed based on the blue photosensitive protein EL222 and the transcription activator VP16.
Source:
functional responsesupports
The TF plus (C120)5 and minimal promoter CYC102 responds to blue light and initiates GFPMut3 expression.
Source:
mechanism of actionsupports
The core light-responsive transcription factor of the system is constructed from EL222 and VP16.
Source:
Comparisons
Source-backed strengths
The reported construct is genetically compact in concept because the core light-responsive transcription factor is a fusion of EL222 and VP16. Its function was demonstrated in Yarrowia lipolytica by blue light-dependent activation of GFPMut3 from a promoter built from (C120)5 and minimal CYC102.
Compared with 4pLRE-cPAOX1
VP16-EL222 light-responsive transcription factor and 4pLRE-cPAOX1 address a similar problem space because they share transcription.
Shared frame: same top-level item type; shared target processes: transcription; same primary input modality: light
Compared with blue-light-activated DNA template ON switch
VP16-EL222 light-responsive transcription factor and blue-light-activated DNA template ON switch address a similar problem space because they share transcription.
Shared frame: same top-level item type; shared target processes: transcription; same primary input modality: light
Compared with triple brake design
VP16-EL222 light-responsive transcription factor and triple brake design address a similar problem space because they share transcription.
Shared frame: same top-level item type; shared target processes: transcription; same primary input modality: light
Ranked Citations
- 1.