Genetic Recombination Overview

Genetic Recombination

• Genetic recombination is the exchange of genetic information between DNA molecules
• This process occurs through an intricate series of enzymatic reactions
• It involves pairing of DNA molecules and breaking/rejoining phosphodiester bonds

Types of Genetic Recombination

• Homologous recombination: Involves regions of nearly identical DNA sequences
• Site-specific recombination: Occurs at specific DNA sequences
• Illegitimate recombination: Doesn't require specific DNA sequences or homology

Homologous Recombination Substrates

• DNA sequences that will be recombined are called homology
• Proteins involve:
  • Identical or similar DNA sequences
  • Ability to form hydrogen bonds
  • Enzymes like endonucleases, exonucleases, RecA, and ligase
• Products involved are 50-100 base pairs or the entire genome

Chi Sites

• Crossover hotspot instigator (Chi)
• About 1000 Chi sites in E. coli
• RecBCD complex: 330 kDa, containing exonuclease V, helicase, and ATPase
• 10 to 50 RecBCD molecules per cell
• RecA binds per 5 base pairs of DNA
• It is a DNA-dependent ATPase
• Facilitates strand unwinding by breaking hydrogen bonds, thereby promoting recombination with other DNA molecules

RecA Loading and Reciprocal/Non-reciprocal Break-Join

• RecA loading facilitates strand exchange
• Reciprocal break-join is a type of homologous recombination. Non-reciprocal break-copy is another type of homologous recombination.

Earlier Models of Recombination

• Breakage and rejoining: Two DNA molecules break at homolgous positions and rejoin.
• Copy choice: DNA polymerase switches between homologous DNA molecules, acting as replication templates.

Holliday Model

• Start with two homologous dsDNA molecules
• Single-strand breaks occur at the same site in both strands of same polarity.
• Dissociation and assimilation
• Ligation forms Holliday junction
• Branch migration
• Isomerization

Single-Strand Invasion Model (Meselson and Radding)

• Single-strand breaks occur in one strand
• Dissociation and assimilation
• DNA polymerase I fills in gap
• D-loop formation (due to displaced strand)
• D-loop digestion
• Ligation
• Resolution by nicking to separate joined DNA molecules

Site-Specific Recombination

• Occurs between specific DNA sites
• Cutting catalyzed by integrase, then exchange and ligation.
• Site is a target sequence that can be recognized by integrase, and the excision and integration of DNA requires the participation of integrase and site-specific proteins.

Illegitimate Recombination

• Occurs in the absence of RecA and DNA homology or specific sites
• Leads to spontaneous DNA rearrangements, involving deletions and duplications
• Associated with Short Sequence Repeats (SSRs)

Significance of Genetic Recombination in Bacteria

• Genetic diversity: Primary source of diversity in bacterial populations, crucial for adaptation to varied environments (temperature, nutrient availability, toxins).
• Evolutionary innovation: Allows bacteria to acquire new genes for traits like antibiotic resistance and virulence factors.
• Horizontal gene transfer (HGT): Movement of genetic material between organisms, not via parent-offspring transmission.

Mechanisms of Gene Transfer Among Bacteria

• Vertical transmission: Plasmids are passed from parent to offspring during replication.
• Horizontal transmission: Three key mechanisms:
  • Transformation: Direct uptake of free DNA from the environment.
  • Transduction: Transfer of DNA via bacteriophages (viruses that infect bacteria).
  • Conjugation: Transfer of genetic material between bacteria through a pilus.

Transduction (specific components)

• Generalized transduction: Bacteriophage can package random pieces of bacterial DNA instead of just its own.
• Specialized transduction: Bacteriophage integrates into bacterial chromosome, then excision occurs causing phage to only pick specific host genes during the process.

Transformation

• Bacteria take up free DNA from environment
• DNA binding proteins, nucleases, free nucleotides, and RecA proteins are involved.

Natural Competence

• Induced when cell density increases, or optimal to poor growth conditions are shifted
• Cell pathways monitor nutrient depletion.
• Signals cells to induce competency genes

Quorum Sensing System

• ComX and ComQ proteins modify precursor ComX into smaller active peptides.
• Active ComX is excreted, affecting cell density, and is involved in cell-cell communication

Conjugation

• Requires donor and recipient cells
• Donors must contain a conjugative plasmid encoding conjugation genes.
• One conjugative E. coli plasmid is called an F plasmid (conjugative plasmid).

Map of Drug Resistance Plasmid

• Shows various genes on the drug-resistance plasmid (e.g., transfer region, origin, resistance genes)

F cell and Hfr cell

• F+ cells donate to F-cells via rolling circle replication
• Hfr cell is a high-frequency recombination cell
• F’ cells donate F-plasmid which also contains chromosomal DNA

Notes on specific sections as needed (e.g., from certain pages).