Exchange of Genetic Information Dr. Jeffrey Patton Associate Professor Pathology, Microbiology, and Immunology USC-School of Medicine

TEACHING OBJECTIVES: 1. To explain the mechanisms of gene transfer in bacteria.

2. To describe the nature of transposable genetic elements and plasmids.

3. To discuss the significance of gene transfer, transposable genetic elements and plasmids.

Mutations in Bacteria • Mutations arise in bacterial populations – Induced – Spontaneous • Rare mutations are expressed – Bacteria are haploid – Rapid growth rate • Selective advantage enriches for mutants • Gene transfer occurs in bacteria

General Features of Gene Transfer in Bacteria • Unidirectional – Donor to recipient • Donor does not give an entire chromosome – Merozygotes • Gene transfer can occur between species

Transformation • Definition: Gene transfer resulting from the uptake of DNA from a donor.

• Factors affecting transformation – DNA size and state • Sensitive to nucleases – Competence of the recipient (

Bacillus, Haemophilus, Neisseria, Streptococcus

) • Competence factors • Induced competence

Transformation

• Steps – Uptake of DNA • Gram + • Gram – Recombination • Legitimate, homologous or general • recA, recB and recC genes • Significance – Phase variation in

Neiseseria

– Recombinant DNA technology

Transduction • Definition: Gene transfer from a donor to a recipient by way of a bacteriophage • Bacteriophage (phage): A virus that infects bacteria

Phage Composition and Structure • Composition – Nucleic acid • Genome size • Modified bases – Protein • Protection • Infection

Contractile Sheath

• Structure (T 4 ) – Size (80 X 100 nm)

Tail Fibers

– Head or capsid – Tail

Head / Capsid Tail Base Plate

Infection of Host Cells by Phages • Adsorption –Tail fibers – Receptor is LPS for T4 • Irreversible attachment – Base plate • Sheath Contraction • Nucleic acid injection • DNA uptake

Microbe Library, American Society for Microbiology www.microbelibrary.org

Types of Bacteriophage • Lytic or virulent – Phage that multiply within the host cell, lyse the cell, and release progeny phage (

e.g.

T4) • Lysogenic or temperate phage: Phage that can either multiply via the lytic cycle or enter a quiescent state in the bacterial cell. (

e.g.,

l ) – Expression of most phage genes repressed – Prophage – Phage DNA in the quiescent state – Lysogen – Bacteria harboring a prophage

Events Leading to Lysogeny • Circularization of the phage chromosome – Cohesive ends

Cohesive Ends Ligase Linear Double Stranded Opened Circle Closed Circle

Events Leading to Lysogeny • Site-specific recombination requires – Phage coded enzyme (Int, integrase) – Bacterial encoded IHF (Integration Host Factor)

gal

• Repression of the phage genome (maintains lysogeny) – Repressor protein (cl) – Specific – Immunity to superinfection by other l because of promoter repression

gal bio gal bio bio

Termination of Lysogeny • Induction – Adverse conditions (DNA damage, ie UV) • Role of proteases – recA protein is activated – Destruction of repressor cI

gal gal bio bio gal bio

• Gene expression (repression lifted) • Excision • Lytic growth

gal bio

Transduction • Definition: Gene transfer from a donor to a recipient by way of a bacteriophage • Resistant to environmental nucleases

Transduction • Types of transduction – Generalized - Transduction in which potentially any donor bacterial gene can be transferred

Generalized Transduction • Infection of Donor • Phage replication and degradation of host DNA • Assembly of phages particles • Release of phage • Infection of recipient • Homologous recombination Potentially any donor gene can be transferred

Transduction • Types of transduction – Generalized - Transduction in which potentially any dornor bacterial gene can be transferred.

– Specialized - Transduction in which only certain donor genes can be transferred

Specialized Transduction Lysogenic Phage • Excision of the prophage • Replication and release of phage • Infection of the recipient • Lysogenization of the recipient – Homologous recombination also possible

gal gal gal bio bio gal bio bio bio

Transduction • Definition • Types of transduction • Significance – Common in Gram+ bacteria – Lysogenic (phage) conversion • e.g.

Corynebacterium diptheriae

–

Toxin derived from lysogenic phage

toxin

Conjugation • Definition: Gene transfer from a donor to a recipient by direct physical contact between cells • Mating types in bacteria – Donor • F factor (Fertility factor) –

F (sex) pilus

– Recipient • Lacks an F factor

Dono r Recipient

Physiological States of F Factor • Autonomous (F + ) – Characteristics of F + crosses x F • F becomes F + while F + remains F + • Low transfer of donor chromosomal genes

F+

Physiological States

of F Factor

Integrated (Hfr) (High Frequency of Recombination) – Characteristics of Hfr x F crosses • F rarely becomes Hfr while Hfr remains Hfr • High transfer of certain donor chromosomal genes

F + Hfr

Physiological States of F Factor • Autonomous with donor genes (F’) – Characteristics of F’ x F crosses • F while F’ remains F’ becomes F’ • High transfer of donor genes on F’ and low transfer of other donor chromosomal genes

Hfr F’

Mechanism of F + x F Crosses • Pair formation – Conjugation bridge • DNA transfer – Origin of transfer – Rolling circle replication

F + F + F F + F + F + F F +

Mechanism of Hfr x F Crosses • Pair formation – Conjugation bridge • DNA transfer – Origin of transfer – Rolling circle replication • Homologous recombination

Hfr Hfr F F Hfr Hfr F F -

Microbe Library, American Society for Microbiology www.microbelibrary.org

Mechanism of F’ x F Crosses • Pair formation – Conjugation bridge • DNA transfer – Origin of transfer – Rolling circle replication

F’ F F’ F’ F’ F’ F F’

Conjugation

• Significance – Gram - bacteria • Antibiotic resistance • Exponential increase under selective pressure – Gram + bacteria • Production of adhesive material by donor cells

Transposable Genetic Elements • Definition: Segments of DNA that are able to move from one location to another • Properties – “Random” movement – Not capable of self replication (not a replicon) – Transposition mediated by site-specific recombination • Transposase – Transposition may be accompanied by duplication

Types of Transposable Genetic Elements • Insertion sequences (IS) – Definition: Elements that carry no other genes except those involved in transposition – Nomenclature - IS1 – Structure (flanking inverted repeats) – Importance

ABCDEFG

• Insertional Mutation •Plasmid insertion •Phase variation

Transposase GFEDCBA

Phase Variation in

Salmonella

H Antigens H1 gene IS H2 gene H1 flagella H2 flagella

Types of Transposable Genetic Elements • Transposons (Tn) – Definition: Elements that carry other genes in addition to those involved in transposition – Nomenclature - Tn10 – Structure • Composite Tns

IS IS Resistance Gene(s) Resistance Gene(s) IS IS

– Importance • Antibiotic resistance

Plasmids • Definition: Extrachromosomal genetic elements that are capable of autonomous replication (replicon) • Episome - a plasmid that can integrate into the chromosome

Classification

of

Plasmids

• Transfer properties – Conjugative – Nonconjugative • Phenotypic effects – Fertility – Bacteriocinogenic plasmid – Resistance plasmid (R factors)