11 Exchange of Genes: Transformation,

Conjugation and Transduction

1. Transformation
2. Conjugation
3. Transduction
4. Horizontal gene transfer
5. Evolution of bacteria
 Why do bacteria take up DNA ?

Three non-mutually exclusive models:

1. DNA for genetic diversity: uptake of metabolic

functions, virulence and antibiotic resistance
genes

2. DNA for repair

3. DNA as food: source of carbon, nitrogen and

phosphorous
1. Transformation
2. Conjugation
3. Transduction
4. Horizontal gene transfer
5. Evolution of bacteria
1. Transformation
1.1 Transformation of Gram-positive bacteria
Bacillus subtilis
Streptococcus pneumoniae
1.2 Transformation of Gram-negative bacteria
Neisseria gonorrhoeae
definition:

uptake of naked DNA by living cells

discovered:

by Griffith in 1927
1. Transformation
1.1 Transformation of Gram-positive bacteria
Bacillus subtilis
Streptococcus pneumoniae
1.2 Transformation of Gram-negative bacteria
Neisseria gonorrhoeae
1. General remarks
2. Binding of donor DNA
3. Fragmentation of donor DNA
4. Uptake (transportation) of donor DNA
5. Fate of the donor DNA within the cell
6. Genetic regulation of competence
1. General remarks
2. Binding of donor DNA
3. Fragmentation of donor DNA
4. Uptake (transportation) of donor DNA
5. Fate of the donor DNA within the cell
6. Genetic regulation of competence
What occurs at the end of the exponential growth
phase (transition phase) ?

1. synthesis of a complex motility and chemotaxis

system
2. secretion of degradative enzymes such as
proteases and amylases
3. production of antibiotics
4. development of genetic competence
5. formation of endospores
Schematic Representation of the Transfor-
mation Pathway

D. Dubnau (1997) Gene 192: 191-198.

1. General remarks
2. Binding of donor DNA
3. Fragmentation of donor DNA
4. Uptake (transportation) of donor DNA
5. Fate of the donor DNA within the cell
6. Genetic regulation of competence
Binding of donor DNA involves:

ComEA = receptor protein with two activities:

1. helix-hairpin-helix motif at its C-terminus: binds
donor DNA
2. flexible stretch (QQGGGG): delivers DNA to the
entrance of the pore

about 50 binding sites per competent cell

binding occurs without DNA specificity

1. General remarks
2. Binding of donor DNA
3. Fragmentation of donor DNA
4. Uptake (transportation) of donor DNA
5. Fate of the donor DNA within the cell
6. Genetic regulation of competence
Fragmentation of donor DNA involves:

NucA = endonuclease; part of the competence machine

• cleaves donor DNA to fragments between 15 –20 kb

• these fragments are taken up in single-stranded form
• the complementary strand is degraded and the nuc-
leotides released into the medium
1. General remarks
2. Binding of donor DNA
3. Fragmentation of donor DNA
4. Uptake (transportation) of donor DNA
5. Fate of the donor DNA within the cell
6. Genetic regulation of competence
 The Competence Machinery

pseudopilus

prepilin peptidase

I. Chen ad D. Dubnau (2004) Nature Rev. Microbiol. 2: 241-249.

Model for DNA Uptake
1. General remarks
2. Binding of donor DNA
3. Fragmentation of donor DNA
4. Uptake (transportation) of donor DNA
5. Fate of the donor DNA within the cell
6. Genetic regulation of competence
 What happens to the single-straned DNA
within the cytoplasm ?

• presumably associates with RecA

• recombination with the chromosomal DNA
• if plasmid DNA, circularization
1. General remarks
2. Binding of donor DNA
3. Fragmentation of donor DNA
4. Uptake (transportation) of donor DNA
5. Fate of the donor DNA within the cell
6. Genetic regulation of competence
 The Backbone Pathway of Competence
Regulation

Module 1 ComS Module 2 ComK

Transformation genes
 Regulation of Competence

Module 1

Module 2
1. Transformation
1.1 Transformation of Gram-positive bacteria
Bacillus subtilis
Streptococcus pneumoniae
1.2 Transformation of Gram-negative bacteria
Neisseria gonorrhoeae
 Protein Involved in DNA Uptake

• 33 – 75 uptake sites per cell

• DNA is nicked, nicks are about 6 kb apart
• uptake is initiated by a second break opposite the nick
• single-strand DNA enters with a 3' – 5' polarity
• uptake rate: 90- 100 nucleotides / sec at 31°C
• proteins forming the competence pseudopilus and
DNA translocation machinery present
• regulation involves the quorum sensing peptide CSP
and a two-component signal transduction system
Processing and Secretion of the Competence
Stimulating Peptide (CSP)
1. Transformation
1.1 Transformation of Gram-positive bacteria
Bacillus subtilis
Streptococcus pneumoniae
1.2 Transformation of Gram-negative bacteria
Neisseria gonorrhoeae
DNA uptake sequences:

Neisseria sp. 5'-GCCGTCTGAA-3'

Actinobacillus actinomycetemcomitans
5'-AAGTGCGGT-3'
 The Competence Machine of
Neisseria gonorrhoeae

DR = receptor

secretin channel pilot protein

pseudopilus

prepilin peptidase
Model for DNA Uptake
1. Transformation
2. Conjugation
3. Transduction
4. Horizontal gene transfer
5. Evolution of bacteria
 Visualization of Bacterial Sex by
Immuno-Fluorescence Microscopy

M. Kohiyama et al. (2003) Science 301: 802

1. Transformation
2. Conjugation
3. Transduction
4. Horizontal gene transfer
5. Evolution of bacteria
3. Transduction
3.1 Generalized transduction
3.2 Spezialized transduction
1. Transformation
2. Conjugation
3. Transduction
4. Horizontal gene transfer
5. Evolution of bacteria
1. Transformation
2. Conjugation
3. Transduction
4. Horizontal gene transfer
5. Evolution of bacteria
• the 3% rule: organisms that are more than 3%
different in 16S rRNA sequence are classified as
different species
• bacteria are a superorganism with a common gene
pool
• most bacteria are composites of genetic information
from many sources
• every cell is a patchwork quilt
• selection acts to create populations of cells that
share the same niche