Source 1primary paper2025PPR
Toolkit/light-activatable gp41-1 intein
light-activatable gp41-1 intein
Taxonomy: Mechanism Branch / Architecture. Workflows sit above the mechanism and technique branches rather than replacing them.
Summary
The light-activatable gp41-1 intein is an engineered split intein system that enables blue light–dependent protein splicing to control Cre recombinase activity in mammalian cells. It is a multi-component switch derived from gp41-1 split variants engineered at unnatural split sites with reduced fragment affinity, allowing conditional reconstitution of Cre.
Usefulness & Problems
Why this is useful
This tool provides optical control over site-specific recombination in mammalian cells, enabling external regulation of Cre-dependent genetic programs. Reported applications include blue light–dependent activation of Cre and spatial control of apoptosis through localized expression of truncated BID and caspase-8.
Source:
Light-activatable gp41-1 and NrdJ-1 inteins enabled blue light–dependent control of Cre recombinase activity in mammalian cells.
Source:
The blue-light-controlled intein-Cre system was exploited to spatially control apoptosis via localized expression of truncated BID and caspase-8.
Problem solved
It addresses the problem of making split intein-mediated protein splicing conditional rather than constitutive, so that Cre recombinase activity can be triggered by blue light in mammalian cells. The cited work specifically frames this as regulated protein splicing using engineered split inteins with reduced spontaneous fragment association.
Source:
Light-activatable gp41-1 and NrdJ-1 inteins enabled blue light–dependent control of Cre recombinase activity in mammalian cells.
Source:
The blue-light-controlled intein-Cre system was exploited to spatially control apoptosis via localized expression of truncated BID and caspase-8.
Published Workflows
Objective: Engineer conditional split inteins for precise spatiotemporal protein regulation in mammalian cells using machine learning–guided split-site design and light-controlled fragment proximity.
Why it works: The workflow is based on first generating split intein variants with reduced mutual affinity, then restoring productive association conditionally by using a light-inducible heterodimerization system to control fragment proximity.
protein trans-splicing by split inteinslight-induced physical proximity of split intein fragmentsmachine learning–guided split-site predictionlight-inducible heterodimerization
Stages
- 1.ML-guided split-site prediction(in_silico_filter)
This stage identifies redesigned split configurations expected to weaken spontaneous fragment association while retaining function, creating a basis for conditional control.
Selection: Predicted unnatural split sites in gp41-1 and NrdJ-1 that would generate functional variants with reduced mutual fragment affinity.
- 2.Conditionalization by light-controlled proximity(functional_characterization)
This stage tests whether reduced-affinity split inteins can be switched on by externally controlling fragment proximity with light.
Selection: Engineered split intein variants were coupled to a light-inducible heterodimerization system to create conditional inteins.
- 3.Mammalian-cell functional validation with Cre(confirmatory_validation)
This stage confirms that the engineered light-activatable inteins can control a functional protein output in mammalian cells.
Selection: Blue light–dependent control of Cre recombinase activity in mammalian cells.
- 4.Downstream application to spatial apoptosis control(confirmatory_validation)
This stage demonstrates that the conditional intein system can drive a spatially resolved cellular phenotype downstream of Cre control.
Selection: Use of the blue-light-controlled system to spatially control apoptosis via localized expression of tBID and caspase-8.
Steps
- 1.Predict unnatural split sites in gp41-1 and NrdJ-1 using ML Int&indesign algorithm
Generate candidate split intein variants with reduced mutual fragment affinity.
Prediction is performed first to choose redesigned split configurations before building conditional light-responsive systems.
- 2.Create conditional inteins by coupling reduced-affinity fragments to a light-inducible heterodimerization systemengineered conditional split inteins
Make intein activity dependent on externally controlled fragment proximity.
This follows split-site redesign because reduced mutual affinity creates the opportunity to gate association through an inducible proximity module.
- 3.Test blue light–dependent control of Cre recombinase activity in mammalian cellsengineered switches under test
Validate that the engineered inteins can control a functional protein output in mammalian cells.
A functional cellular output is tested after conditional inteins are built to confirm that light-controlled splicing translates into regulated protein activity.
- 4.Exploit the system to spatially control apoptosis via localized expression of tBID and caspase-8upstream control modules enabling downstream phenotype
Demonstrate a spatially resolved downstream cellular application of the light-controlled intein system.
This application-level test follows Cre control validation to show that the system can drive a meaningful spatial cellular phenotype.
Taxonomy & Function
Primary hierarchy
Mechanism Branch
Architecture: A composed arrangement of multiple parts that instantiates one or more mechanisms.
Techniques
No technique tags yet.
Target processes
recombinationInput: Light
Implementation Constraints
The available evidence indicates use in mammalian cells and blue light as the input modality. The current summary states that the design uses gp41-1 split variants at unnatural split sites with reduced mutual fragment affinity, but the provided evidence does not specify construct architecture, illumination parameters, cofactors, or delivery format.
The supplied evidence is limited to a single 2025 preprint and brief application-level claims, so quantitative performance metrics, dynamic range, kinetics, leakiness, and generality across cargos are not available here. Independent replication and validation outside mammalian-cell Cre-based applications are not documented in the provided evidence.
Validation
Cell-freeBacteriaMammalianMouseHumanTherapeuticIndep. Replication
Supporting Sources
Ranked Claims
Light-activatable gp41-1 and NrdJ-1 inteins enabled blue light–dependent control of Cre recombinase activity in mammalian cells.
Light-activatable gp41-1 and NrdJ-1 inteins enabled blue light–dependent control of Cre recombinase activity in mammalian cells.
Light-activatable gp41-1 and NrdJ-1 inteins enabled blue light–dependent control of Cre recombinase activity in mammalian cells.
Light-activatable gp41-1 and NrdJ-1 inteins enabled blue light–dependent control of Cre recombinase activity in mammalian cells.
Light-activatable gp41-1 and NrdJ-1 inteins enabled blue light–dependent control of Cre recombinase activity in mammalian cells.
Light-activatable gp41-1 and NrdJ-1 inteins enabled blue light–dependent control of Cre recombinase activity in mammalian cells.
Light-activatable gp41-1 and NrdJ-1 inteins enabled blue light–dependent control of Cre recombinase activity in mammalian cells.
Light-activatable gp41-1 and NrdJ-1 inteins enabled blue light–dependent control of Cre recombinase activity in mammalian cells.
The blue-light-controlled intein-Cre system was exploited to spatially control apoptosis via localized expression of truncated BID and caspase-8.
The blue-light-controlled intein-Cre system was exploited to spatially control apoptosis via localized expression of truncated BID and caspase-8.
The blue-light-controlled intein-Cre system was exploited to spatially control apoptosis via localized expression of truncated BID and caspase-8.
The blue-light-controlled intein-Cre system was exploited to spatially control apoptosis via localized expression of truncated BID and caspase-8.
The blue-light-controlled intein-Cre system was exploited to spatially control apoptosis via localized expression of truncated BID and caspase-8.
The blue-light-controlled intein-Cre system was exploited to spatially control apoptosis via localized expression of truncated BID and caspase-8.
The blue-light-controlled intein-Cre system was exploited to spatially control apoptosis via localized expression of truncated BID and caspase-8.
The blue-light-controlled intein-Cre system was exploited to spatially control apoptosis via localized expression of truncated BID and caspase-8.
ML Int&in predicted unnatural split sites in gp41-1 and NrdJ-1 that generated functional split intein variants with reduced mutual fragment affinity.
ML Int&in predicted unnatural split sites in gp41-1 and NrdJ-1 that generated functional split intein variants with reduced mutual fragment affinity.
ML Int&in predicted unnatural split sites in gp41-1 and NrdJ-1 that generated functional split intein variants with reduced mutual fragment affinity.
ML Int&in predicted unnatural split sites in gp41-1 and NrdJ-1 that generated functional split intein variants with reduced mutual fragment affinity.
ML Int&in predicted unnatural split sites in gp41-1 and NrdJ-1 that generated functional split intein variants with reduced mutual fragment affinity.
ML Int&in predicted unnatural split sites in gp41-1 and NrdJ-1 that generated functional split intein variants with reduced mutual fragment affinity.
ML Int&in predicted unnatural split sites in gp41-1 and NrdJ-1 that generated functional split intein variants with reduced mutual fragment affinity.
ML Int&in predicted unnatural split sites in gp41-1 and NrdJ-1 that generated functional split intein variants with reduced mutual fragment affinity.
Reduced mutual affinity in engineered split intein fragments was harnessed to create conditional inteins by controlling fragment proximity with a light-inducible heterodimerization system.
Reduced mutual affinity in engineered split intein fragments was harnessed to create conditional inteins by controlling fragment proximity with a light-inducible heterodimerization system.
Reduced mutual affinity in engineered split intein fragments was harnessed to create conditional inteins by controlling fragment proximity with a light-inducible heterodimerization system.
Reduced mutual affinity in engineered split intein fragments was harnessed to create conditional inteins by controlling fragment proximity with a light-inducible heterodimerization system.
Reduced mutual affinity in engineered split intein fragments was harnessed to create conditional inteins by controlling fragment proximity with a light-inducible heterodimerization system.
Reduced mutual affinity in engineered split intein fragments was harnessed to create conditional inteins by controlling fragment proximity with a light-inducible heterodimerization system.
Reduced mutual affinity in engineered split intein fragments was harnessed to create conditional inteins by controlling fragment proximity with a light-inducible heterodimerization system.
Reduced mutual affinity in engineered split intein fragments was harnessed to create conditional inteins by controlling fragment proximity with a light-inducible heterodimerization system.
Approval Evidence
1 source3 linked approval claimsfirst-pass slug light-activatable-gp41-1-intein
The resulting light-activatable gp41-1 and NrdJ-1 inteins enabled blue light–dependent control of Cre recombinase activity in mammalian cells.
Source:
application resultsupports
Light-activatable gp41-1 and NrdJ-1 inteins enabled blue light–dependent control of Cre recombinase activity in mammalian cells.
Source:
application resultsupports
The blue-light-controlled intein-Cre system was exploited to spatially control apoptosis via localized expression of truncated BID and caspase-8.
Source:
mechanism of controlsupports
Reduced mutual affinity in engineered split intein fragments was harnessed to create conditional inteins by controlling fragment proximity with a light-inducible heterodimerization system.
Source:
Comparisons
Source-backed strengths
The system was reported to support blue light–dependent control of Cre recombinase activity in mammalian cells. It was also exploited for spatial control of apoptosis via localized expression of truncated BID and caspase-8, indicating functional coupling to downstream cellular phenotypes.
Source:
ML Int&in predicted unnatural split sites in gp41-1 and NrdJ-1 that generated functional split intein variants with reduced mutual fragment affinity.
Ranked Citations
- 1.