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Molecular Biology: Transposition Robert F. Weaver

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77 views15 pages

Molecular Biology: Transposition Robert F. Weaver

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Lecture PowerPoint to accompany

Molecular Biology
Fourth Edition

Robert F. Weaver

Transposition

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Bacterial Transposons
• A transposable element (TE, transposon, or jumping
gene) is a DNA sequence that can change its position
within a genome
• Originally discovered in maize,
transposons have been found in all kinds
of organisms
– Bacteria
– Plants
– Humans
– These transposons are called insertion
sequences (ISs) 23-2
Insertion Sequences
• Insertion sequences are the simplest type of
bacterial transposon
• They contain only the elements necessary for
their own transposition
– Short inverted repeats at their ends
– At least 2 genes coding for an enzyme, transposase
that carries out transposition
• Transposition involves:
– Duplication of a short sequence in the target DNA
– One copy of this sequence flanks the insertion
sequence on each side after transposition
23-3
Generating Host DNA Direct
Repeats

23-4
Mobile genetic elements (transposons)

• DNA sequences that can replicate and insert copies of themselves at


other locations within genome
• Together with non-functional relics of old transposons, they account for
over half of total human DNA.
• Sometimes called "jumping genes",
• Some of these sequences represent endogenous retroviruses, DNA
copies of viral sequences that have become permanently integrated
into the genome.
• Mobile elements within the human genome can be classified into
• LTR retrotransposons (8.3% of total genome),
• SINEs (13.1% of genome) including Alu elements (50,000
copies)
• LINEs (20.4% of total genome, 100 copies)
• Class II DNA transposons (2.9% of total genome).
Complex Transposons
• The term “selfish DNA” implies that
insertion sequences and other
transposons replicate at the expense of
their hosts, providing no value in return
• Some transposons do carry genes that are
valuable to their hosts, antibiotic
resistance is among most familiar

23-6
Antibiotic Resistance and
Transposons
• Donor plasmid has
Kanr, harboring
transposon Tn3 with
Ampr
• Target plasmid has Tetr
• After transposition, Tn3
has replicated and
there is a copy in target
plasmid
• Target plasmid now
confers both Ampr, Tetr
23-7
Transposition Mechanisms
• Transposons are sometimes called “jumping genes”

• Non-replicative transposition
– “Cut and paste”
– Both strands of original DNA move together from 1
place to another without replicating
• Replicative transposition
– 1 copy remains at original site
– New copy inserts at the new site
– Replicative transposition
– “Copy and paste”

23-8
Replicative Transposition of Tn3
A 4957 base pair mobile genetic element, found in prokaryotes

• It encodes three proteins:


1. β-lactamase, an enzyme that confers resistance to β-lactam
antibiotics (and is encoded by the gene Bla).
2. Tn3 transposase (encoded by gene tnpA)
3. Tn3 resolvase (encoded by gene tnpR)

• In first step, 2 plasmids


fuse, forming a co-
integrate
• The co-integrate, breaks
down into 2 independent
plasmids, catalyzed by
resolvase gene product
23-9
Retrotransposons

• Retrotransposons replicate through an


RNA intermediate
• Retrotransposons resemble retroviruses
– Retroviruses can cause tumors in vertebrates
– Some retroviruses cause diseases such as
AIDS
• Before studying retrotransposons, look at
replication of the retroviruses

23-10
Retroviruses

• Class of virus is named for its ability to


make a DNA copy of its RNA genome
• This reaction is the reverse of the
transcription reaction – reverse
transcription
• Virus particles contain an enzyme that
catalyzes reverse transcription reaction

23-11
Retrovirus Replication

• Viral genome is RNA,


with long terminal repeats
at each end
• Reverse transcriptase
makes linear, ds-DNA
copy of RNA
• ds-DNA copy integrates
back into host DNA =
provirus
• Host RNA polymerase II
transcribes the provirus to
genomic RNA
• Viral RNA packaged into
a virus particle
23-12
Two Categories

 DNA transposons
 Retrotransposons

“cut-and-paste” “copy-and-paste”

 Most mobile elements in bacteria


is DNA transposons
 In contrast, most mobile
elements in eukaryotes are
retrotransposons, but eukaryotic
DNA transposons also occur.
Lodish et al., Molecular Cell Biology, 7th ed. Fig 10-8
Rearrangement of Immunoglobulin Genes
• Mammalian genes use a process that closely resembles
transposition for:
– B cell antibodies
– T cell receptors
• Enormous diversity of immune system is generated by 3
basic mechanisms:
– Assembling genes for antibody light chains and heavy
chains from 2 or 3 component parts
– Joining the gene parts by an imprecise mechanism that
can delete bases or add extra bases
– Causing a high rate of somatic mutations,

23-14
Rearrangement of Antibody Light Chain
Gene

23-15

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