TALE-LSD1 Fusion Proteins for Enhancer Chromatin Modulation

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Solution Overview

Problem

Current methods for mapping and understanding the functions of mammalian gene regulation, particularly involving tissue-specific enhancers, fail to determine which genes a given enhancer regulates and do not address the in vivo functions or chromatin states of putative enhancers.

Innovation Solution

Development of fusion proteins combining a DNA-binding domain, such as TALE or zinc finger, with a catalytic domain like LSD1 for histone demethylation, allowing for the specific targeting and modulation of enhancer-associated chromatin modifications, thereby regulating gene expression.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Quantity of substance

If mapping experiments are used to identify putative enhancers, then the number of identified enhancers increases, but the ability to determine in vivo functions and chromatin states is lost

Engineering Contradiction:

Improvenumber of identified enhancersVSAvoidin vivo functions and chromatin states information

Core Design Contradiction:

Quantity of substanceVSLoss of information

Solution Approach 1:

The patent introduces TALE-LSD1 fusion proteins as intermediary tools that bridge the gap between enhancer identification and functional characterization. These fusion proteins consist of a TALE DNA-binding domain that targets specific enhancer sequences and an LSD1 catalytic domain that modifies chromatin, thereby serving as a mediator to both identify and functionally test putative enhancers in vivo

Inventive Principle:

Principle #24Intermediary (Mediator)

Solution Approach 2:

The TALE-LSD1 fusion proteins enable self-service by allowing the identified enhancers to be directly tested for their in vivo functions through chromatin modification. The system uses the enhancers themselves as targets for the fusion proteins, allowing them to demonstrate their regulatory functions on proximal genes without requiring external validation methods

Inventive Principle:

Principle #25Self-service

2Measurement precision

If TALE-LSD1 fusion proteins are used to target enhancers, then chromatin modification specificity increases, but the complexity of the system increases

Engineering Contradiction:

Improvechromatin modification specificityVSAvoidsystem complexity

Core Design Contradiction:

Measurement precisionVSDevice complexity

Solution Approach 1:

The TALE-LSD1 fusion protein is segmented into distinct functional modules: a TALE repeat array for DNA recognition, a linker region for flexibility, and an LSD1 catalytic domain for chromatin modification. This segmentation allows each component to be independently optimized and characterized, managing complexity through modularity while achieving high specificity

Inventive Principle:

Principle #1Segmentation

Solution Approach 2:

The TALE-LSD1 fusion protein system serves multiple functions: it identifies target enhancers through specific DNA binding, modifies chromatin through histone demethylation, and regulates gene expression. This multi-functionality reduces the need for separate experimental systems, thereby managing overall system complexity while achieving precise chromatin modification

Inventive Principle:

Principle #6Universality (Multi-functionality)

3Loss of information

If enhancer chromatin is inactivated to characterize function, then gene regulation understanding improves, but control regions may be affected

Engineering Contradiction:

Improvegene regulation understandingVSAvoidcontrol regions affected

Core Design Contradiction:

Loss of informationVSObject-affected harmful factors

Solution Approach 1:

The TALE-LSD1 fusion proteins apply chromatin modification locally at the target enhancer through sequence-specific DNA binding. The TALE domain ensures that LSD1-mediated histone demethylation occurs only at the intended enhancer locus, creating a localized effect that spares other control regions and maintains their normal function

Inventive Principle:

Principle #3Local quality

Solution Approach 2:

The patent replaces broad, non-specific chromatin modification approaches with a targeted molecular recognition system. The TALE domain provides sequence-specific recognition that substitutes for non-specific chromatin targeting, allowing precise inactivation of only the intended enhancer while leaving other control regions unaffected

Inventive Principle:

Principle #28Mechanics substitution (Replace mechanical system)

Applied Scientific Principles

This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.

Function Achieved in This Case

These fusion proteins efficiently remove enhancer-associated chromatin modifications from target loci, down-regulating proximal genes and providing a tool for characterizing and modulating the activity of functional genomic elements with high specificity.

Implementation Method 1

a catalytic domain comprising a sequence that catalyzes histone demethylation

Methodology Applied

Scientific EffectHistone demethylation: Enzyme

Implementation Method 2

an engineered DNA-binding domain that binds specifically to a preselected target sequence

Methodology Applied

Scientific EffectDNA binding: Enzyme

Data Source

PatentUS20240254455A1Transcription activator-like effector (TALE) - lysine-specific demethylase 1 (LSD1) fusion proteins

Publication Date: 2024.08.01 THE GENERAL HOSPITAL CORP

AI summary

Fusion proteins comprising a DNA binding domain, e.g., a TAL effector repeat array (TALE) or zinc finger array, and a catalytic domain comprising a sequence that catalyzes histone demethylation, and methods of use thereof.