Bridge Recombinase Mechanism: The newly discovered gene-editing tool

Scientists have discovered a naturally existing next-generation genomic design method, called the bridge recombinase mechanism. Read here to learn more about it.

Gene editing technologies are powerful tools that allow scientists to make precise modifications to DNA, enabling advancements in fields such as medicine, agriculture, and biotechnology.

Bridge Recombinase Mechanism which exists naturally and has now been discovered, enhances the human ability to edit genomes beyond the capabilities and scope of CRISPR.

It utilises mobile genetic elements or “jumping genes”, which cut and paste themselves into genomes and are present in all forms of life, performing on-the-go DNA manipulation through all living beings.

Bridge Recombinase Mechanism

Bridge recombinase is a term used to describe a particular type of recombinase enzyme involved in genetic recombination, a fundamental process where DNA strands are broken and rejoined to introduce new combinations of genetic material.

• This mechanism plays a critical role in genetic diversity, DNA repair, and the evolution of genomes.

The recent discovery has been highlighted by two Nature studies–

• IS110 is a jumping gene that acts as a bridge between bits of unconnected DNA and exists naturally in humans. The RNA bridge can also be programmed.

What are jumping genes?

Jumping genes, also known as transposable elements or transposons, are sequences of DNA that can move or “jump” to different positions within the genome of a single cell.

This ability to move from one location to another within the genome can have significant implications for gene function and genome evolution.

Types of Transposable Elements:

1. Class I Transposons (Retrotransposons):
   • Mechanism: Copy and paste
   • Process: Transpose via an RNA intermediate. They are transcribed into RNA, which is then reverse-transcribed into DNA and inserted back into the genome.
   • Example: Long Interspersed Nuclear Elements (LINEs), Short Interspersed Nuclear Elements (SINEs)
2. Class II Transposons (DNA Transposons):
   • Mechanism: Cut and paste
   • Process: Transpose directly as DNA. They are excised from their original position and inserted into a new location within the genome.
   • Example: Activator/Dissociation (Ac/Ds) elements in maize

Example: Maize (Corn)

• The discovery of transposons is credited to Barbara McClintock, who observed their behaviour in maize (corn) in the 1940s.
• Her work demonstrated that certain genetic elements could move around within the genome, causing changes in the colouration of maize kernels.

Mechanism of Action of Bridge Recombinase Mechanism

The bridge recombinase mechanism can be described in several steps:

1. Recognition and Binding: The recombinase enzyme recognizes specific DNA sequences known as recombination sites. These sites are typically palindromic or have specific structural features that are identified by the recombinase.
2. Strand Cleavage: Once the recombinase binds to the recombination sites, it introduces single-strand or double-strand breaks in the DNA. This cleavage is usually staggered, creating overhangs or blunt ends.
3. Strand Exchange: The recombinase then facilitates the exchange of DNA strands between two different DNA molecules or within the same molecule. This exchange involves the pairing of homologous or similar sequences from the two DNA molecules.
4. Bridge Formation: In the context of bridge recombinase, a “bridge” is formed where the recombinase holds the cleaved DNA ends nearby, facilitating the exchange and rejoining of strands.
5. Strand Rejoining: The recombinase then catalyzes the rejoining of the DNA strands, completing the recombination process. This rejoining can result in crossover (where segments of DNA are swapped between molecules) or non-crossover products (where sequences are merely rearranged within a molecule).

Biological Importance of Bridge Recombinase Mechanism

• Genetic Diversity: By introducing new combinations of genetic material, bridge recombinase mechanisms contribute to genetic diversity within populations, which is essential for evolution and adaptation.
• DNA Repair: Recombination mechanisms, including those mediated by bridge recombinase, are crucial for repairing DNA damage, especially double-strand breaks. This repair mechanism maintains genomic stability.
• Immune System: In vertebrates, recombinases like RAG1 and RAG2 are involved in V(D)J recombination, a process that generates the diversity of antibodies and T-cell receptors, which are vital for the adaptive immune response.

Examples of Bridge Recombinases

• RAG1 and RAG2: These enzymes are involved in V(D)J recombination, crucial for generating the diverse repertoire of antibodies and T-cell receptors in the immune system.
• Cre Recombinase: Derived from bacteriophage P1, Cre recombinase is widely used in genetic engineering to induce site-specific recombination between loxP sites in DNA.
• FLP Recombinase: Derived from the yeast Saccharomyces cerevisiae, FLP recombinase mediates recombination between FRT sites and is also commonly used in genetic manipulation.

Different gene editing technologies

Here are some of the most prominent gene editing technologies:

1. CRISPR-Cas9

CRISPR-Cas9 is the most widely used gene-editing technology due to its efficiency, precision, and ease of use.

• Mechanism: Uses a guide RNA to target a specific DNA sequence, and the Cas9 enzyme to create double-strand breaks in the DNA, allowing for modifications.
• Applications: Used for gene knockout, gene insertion, and correction of genetic mutations in research, medicine, and agriculture.
• Advantages: High specificity, versatility, and cost-effectiveness.
• Limitations: Potential off-target effects and ethical concerns regarding germline editing.

2. TALENs (Transcription Activator-Like Effector Nucleases)

TALENs are proteins engineered to bind to specific DNA sequences and create double-strand breaks.

• Mechanism: Composed of a DNA-binding domain and a nuclease domain, TALENs recognize specific DNA sequences and induce targeted breaks.
• Applications: Used for gene disruption, correction, and regulation in various organisms.
• Advantages: High specificity and versatility in targeting different DNA sequences.
• Limitations: More complex and time-consuming to design and produce compared to CRISPR-Cas9.

3. ZFNs (Zinc Finger Nucleases)

ZFNs are synthetic proteins that combine a zinc finger DNA-binding domain with a nuclease domain to create targeted DNA breaks.

• Mechanism: Zinc fingers bind to specific DNA sequences, and the nuclease domain induces double-strand breaks at these sites.
• Applications: Used in gene therapy, functional genomics, and the creation of genetically modified organisms.
• Advantages: High specificity in targeting DNA sequences.
• Limitations: Designing and engineering ZFNs is labour-intensive and expensive.

4. Base Editing

Base editing allows for direct, irreversible conversion of one DNA base pair to another without creating double-strand breaks.

• Mechanism: Uses a modified CRISPR-Cas9 enzyme coupled with a deaminase enzyme to convert specific DNA bases (e.g., C to T or A to G).
• Applications: Correcting point mutations and studying genetic diseases at the base level.
• Advantages: High precision and reduced risk of unintended mutations.
• Limitations: Limited to certain types of base conversions and potential off-target effects.

5. Prime Editing

Prime editing is a versatile technology that allows for targeted insertions, deletions, and base conversions without double-strand breaks.

• Mechanism: Combines a modified Cas9 enzyme with a reverse transcriptase and a guide RNA to introduce specific edits directly into the DNA.
• Applications: Wide range of genetic modifications, including correcting mutations and introducing precise genetic changes.
• Advantages: High precision, versatility, and reduced off-target effects.
• Limitations: Requires optimization for different targets and may have limitations in efficiency.

6. RNA Editing

RNA editing involves the modification of RNA sequences after transcription, allowing for changes in gene expression without altering the underlying DNA.

• Mechanism: Uses enzymes like ADAR (adenosine deaminase acting on RNA) to edit specific RNA bases.
• Applications: Potential treatments for genetic diseases, regulation of gene expression, and studying RNA functions.
• Advantages: Reversible and does not involve permanent changes to the genome.
• Limitations: Currently less efficient and specific compared to DNA editing technologies.

Conclusion

Each gene editing technology has its strengths and limitations, and the choice of which to use depends on the specific application and the desired outcome.

Advances in these technologies continue to expand their potential uses in medicine, agriculture, and beyond, while ongoing research aims to improve their precision, efficiency, and safety.

Read: MicroRNA: The Nobel Prize in Physiology or Medicine 2024

-Article by Swathi Satish