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Genetic recombination:

1.Homologous Recombination

2. Site-Specific Recombination

3. DNA Transposition

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Homologous Recombination at the Molecular

Level

DNA BREAKS ARE COMMON AND INITIATERECOMBINATION

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MODELS FOR HOMOLOGOUS

RECOMBINATION

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Strand invasion is a key step in homologous recombination

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Resolving Holliday junctions is a

key step to finishing genetic

exchange

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The double-strand break-repair

model describes many

recombination events

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HOMOLOGOUS RECOMBINATION PROTEIN

MACHINES

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RecBCD

RecA

RuvAB

RuvC

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The RecBCD helicase/nuclease processes broken DNA

molecules for recombination

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Structure of RecBCD

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Chi sites control RecBCD

(GCTGGTGG)

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RecA protein assembles on single-stranded DNA and

promotes strand invasion

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Three views of the RecA filament

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Polarity of RecA assembly

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Newly based-paired partners are established within RecA

filament

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RecA homologs are present in all organism

Human Rad51 RecA

Rad A of Archaea

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The RuvAB complex specifically recognizes Holliday

junctions and promote branch migration

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Branch migration can either enlarge heteroduplex regions or

release newly synthesized DNA as a single strand

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Structure of RuvA and model of RuvAB bound to Holliday junction

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Enzyme-catalyzed double branch migration at a Holliday junction.

InE. coli, a tetramer of the RuvA protein (green) and two hexamers of the RuvB

protein (pale gray)bind to the open form of the junction. The RuvB protein uses the

energy of ATP hydrolysis to move the crossover point rapidly along the paired DNA

helices, extending the heteroduplex region as shown. There is evidence that similar

proteins perform this function in vertebrate cells. (Image courtesy of P. Artymiuk;modified from S.C. West,Cell94:699701, 1998.)

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RuvC cleaves specific strands at the Holliday junction to

finish recombination

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Structure of RuvC and model of RuvC dimer bound to Holliday

junction

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RecBCD

RecA

RuvAB

RuvC

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HOMOLOGOUS

RECOMBINATION IN

EUKARYOTESHomologous recombination hasadditional functions in eukaryotes

It is required to pair homologouschromosomes in preparation for the

first nuclear division and for

segregation during meiosis

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Meiotic recombination between

homologous chromatids

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Meiotic recombination also frequently gives rise to crossing over

between genes on the two homologous parental chromosomes

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Programmed generation of double-stranded DNA breaks

occurs during meiosis

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MRX protein processes the

cleaved DNA ends for

assembly of the RecA-likestrand exchange proteins

(DMC1, Rad51)

DMC1 specifically functions

in meiotic recombination

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Many proteins function together to promote meiotic

recombination (Rad51, DMC1, Rad51 paralogs, Rad52,

Rad54)

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MATING-TYPE SWITCHING

Mating-type switching is initiated by a site-specific double-

strand break (synthesis-dependent strand annealing(SDSA))

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Mating-type switching is a gene conversion event and NOT

associate with crossing over (synthesis-dependent strand

annealing )

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GENETIC CONSEQUENCES OF THE

MECHANISM OF HOMOLOGOUS

RECOMBINATION

Hot and cold spots of recombination

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One cause of gene conversion is DNA repair during

recombination

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Site-Specific recombination & Transposition

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Three types of CSSR

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Structures involved in CSSR

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Recombination by a serine

recombinase

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Recombination by a tyrosine

recombinase

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The insertion of a circular bacteriophage

lambda DNA chromosome into the

bacterial chromosome.

Conservative site-specific recombination

was discovered in bacteriophage lambda

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The life cycle of bacteriophage

lambda.

The double-stranded DNA lambda

genome contains 50,000 nucleotide

pairs and encodes 5060 differentproteins. When the lambda DNA

enters the cell, the ends join to

form a circular DNA molecule.

This bacteriophage can multiply in

E. coliby a lytic pathway, which

destroys the cell, or it can enter a

latent prophage state. Damage to acell carrying a lambda prophage

induces the prophage to exit from

the host chromosome and shift to

lytic growth (green arrows). Both

the entrance of the lambda DNA to,

and its exit from, the bacterial

chromosome are accomplished bya conservative site-specific

recombination event, catalyzed by

the lambda integrase enzyme (see

Figure 580).

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DNA inversion by the Hin recombinase of Salmonella

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Transposition

Some genetic elements move to new chromosomal

locations by transposition

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The cut-and-paste

mechanism of transposition

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Replicative transposition

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S i t iti h i t th l

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Some virus use a transposition mechanism to move themselves

into host cell chromosomes

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Reverse transcriptase

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Poly-A retrotransposon

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