Jumping genes
Jumping genes
Dr. [Link]
Professor
Professor of Zoology
ManasagangotriDept. of Zoology
Manasagangotri,
Mysore – 570 006
Mysore – 570 006
Cell: 9448365799
Mendel's (1822-1884) Discovery – Factors –
With well designed experiments in pea plant
he proposed the laws of heredity
Rediscovery Hugo De Vries, Carl Correns,
and Tshermak - 1900
The word gene was coined by Johanssen
Gene was thought to be particulate matter
which is stable
Two years after the rediscovery - mutations
were discovered – gene
Linkage studies in Maize – Bateson and
Punnet – 1903
Chromosome theory of Heredity – Sutton –
1904
Genes occupy fixed position on
chromosome
Linkage and sex linkage in Drosophila –
[Link] (1910)
Linkage maps of Drosophila, Maize and
many other organisms
Although the experimental designs of
them were precise, Barbara
McClintok during 1940s suggested
that the genes are not always
stable.
• Barbara McClintock
(1902-1992)
• Cold Spring Harbor
Laboratory, NY
• Nobel Prize in
Physiology and
Medicine 1983
“for her discovery of
mobile genetic
elements”
• She discovered certain strains
with high frequency of
mutations
• They were thought to carry Mutator
and antimutator genes
• She also found number of unstable
elements in Maize
• Now they are called jumping
genes, moving genetic elements,
mobile genetic elements,
transposable elements, unstable
genetic elements, transposons,
lollipops, etc
• She found number of mutations,
deletions, transpositions in Maize
• She identified two specific genetic
regions dissociators (Ds) and
activators (Ac) which had variety of
effects on chromosomes
Definition
Transposons are segments of DNA that
can dislodge and move from one place
of the genome to another in a given cell
• The movement requires a recombination
• Transpositions may show sequence
specificity – insersion sites
• Nonhomologous recombination:
transposable elements insert into DNA
that has no sequence homology with
the transposon
• Those without sequence specificity
completely disrupt gene function
• Transposable elements cause genetic
changes and make important
contributions to the evolution of
genomes:
- Insert into genes.
- Position effect.
- Insert into regulatory sequences;
modify gene expression.
- Produce chromosomal mutations.
- they may act as delivery vectors.
• The movement can occur with or
without duplication of DNA
• Two types of transposons based on
duplication
– Conservative transposons – donot
duplicate
– Replicative transposons – duplicate
• Transposable elements are found in
the genomes of all organisms
• Normal and ubiquitous components
of prokaryote and eukaryote
genomes.
• Prokaryotes-transpose to/from cell’s
chromosome, plasmid, or a phage
chromosome
• Eukaryotes-transpose to/from same
or a different chromosome
• There are different types -
classified on the basis of structure
and recombination
[Link] transposons
[Link] transposons/
[Link]
DNA Transposons
They are DNA sequences serving as both
recombination sites as well the protein coding
sequences needed for recombination
Recombination sites have terminal repeats – 25
to few hundred bp long at both ends
They identify transposase needed for
transposition
Carry genes coding for transposase
Additional sites coding proteins that regulate
transcription or a function useful to the TE or
host cell
Flanking host DNA
Terminal
inverted repeat
Additional sites
25- few hundred
proteins needed for
bp identifies
transcription
transposase
Transposase
The element
Target site for
duplication
Structure of DNA transposon
Viral like and/or retrotransposons
• Retroviruses or viral like elements also are
transposable.
• They also carry inverted terminal repeats (P)
which serve as recombinase binding sites.
• The terminal inverted repeats are embedded
within longer direct repeat sequence (LTR),
that are organized at two ends.
• The viral like and/or retrotransposons
encode for two proteins needed for their
mobility.
Flanking host DNA
Additional sites
proteins needed for
P transcription
Intigrase &
LTR LTR
RTase
The element
Target site for
duplication
Structure of virus like transposon
Poly A retrotransposons
• Instead of terminal repeats they have two
ends with specific sequences.
• One end is called 5’ UTR (Untranslated
region) and another 3’ UTR followed by a
stretch of AT pairs.
• Therefore they are called poly A
retrotransposons.
• These elements are also flanked by short
target site duplications.
• Retrotransposons carry two genes ORF 1
and ORF 2, the first encoding an RNA
binding protein and second encodes a
protein with both reverse transcriptase
activity and an endonuclease activity.
• The poly A transposons can exist as both
autonomous and non autonomous form.
• Some of the retrotransposons exist as
truncatated forms, unable to transpose.
5’UTR ORF1 ORF2 3’UTR
Target site Target site
for replication for replication
Poly A retrotransposon
Prokaryotic transposons
Insertion sequences (IS elements):
These were the first transposable
elements discovered in bacteria.
They are the simplest transposons.
IS element
Defective IS
Defective Tetracycline resistance Transposase
Transposase genes gene
gene
Pout
IS10 left Pin
IS10 right
A composite element with IS
Elements of [Link]
IS elements have a central region of about 700-
1500 bp surrounded by an inverted repeat of
about 10 - 30 bp.
The inverted repeats signal the transposing
enzyme to act at the ends of the IS element.
The central region of the IS element contains a
gene needed for transposition.
The target site that the transposable element
moves to is not a specific sequence as with
the att site of λ.
During insertion the target site is cut in a
staggered fashion, leaving single
stranded ends.
The IS element is then inserted between
the single stranded ends.
Repair process converts the two single
stranded tails to double stranded
segments and hence to direct flanking
repeats.
When DNA is sequenced, the pattern of a
direct flanking repeat surrounding the
inverted repeat with a segment in
between signals, suggests the
existence of a transposable element.
More than 15 families including a total of
over 500 known members of IS
elements have been identified.
After the discovery of IS elements, more
complex transposons were discovered.
Such a composite transposon consists
of a central region surrounded by two
IS elements.
The central region usually contains
bacterial genes, frequently an antibiotic
resistance gene.
The best example for composite
transposon is Tn10 which contains
genes for transposase, resolvase and
the bacterial gene - lactamase which
7
confers resistance to ampicillin.
The arrangements of composite
transposons vary.
Transposition of Tn10 occurs at a rate 10
־per element per generation.
IS element
Defective IS
Defective Tetracycline resistance Transposase
Transposase genes gene
gene
Pout
IS10 left Pin
IS10 right
IS Elements of [Link]
Each bacteria contains large number of
sites for transposition which are spread
at least one per one thousand base
pairs.
These sites have the sequence GCTAGC
which may exist as duplicated sets.
• Transposition for Tn10 occurs as
follows;
• Among the two IS 10 units, one codes
for transposase, and the left one is
non-coding.
• There are two promoters in the IS 10 of
right, each separated by 33 base pairs.
• Although IS 10 on the left does not
encode for transposase, its RNA can
hybridize with that coded by the right
and works as transacting inhibitor of
transposase expression.
• The promoter pout has stronger action
than PIN.
• Transposase on the other hand is
cisacting and has a preference for
hemimethylated ends of the
transposon.
• This results in the increase in
transposition frequency.
• Phage Mu transposons:
• Phages like lamda are lysogenic.
• Their genomes work like a large DNA
transposon.
• This phage uses transposition to insert its
DNA into the genome of the host cell
during infection and in this way is similar to
the retroviruses.
• Mu also uses multiple rounds of replicative
transposition to amplify its DNA during lytic
growth.
• During the lytic cycle, Mu completes about
100 rounds of transposition per hour,
making it the most efficient transposon
known.
• Even when present as quiscent lysogen,
the Mu is short for mutator and stems from
this ability to transposase promiscuously.
• Cells carrying an inserted copy of Mu DNA
frequently accumulate new mutations due
to insertion of the phage DNA into cellular
genes.
• The Mu genome is about 40 kb and
carries more than 35 genes, but only two
encode proteins with dedicated roles in
transposition.
• These are the A and B genes, which
encode the proteins MuA and MuB.
• MuA is the transposase.
• MuB is an ATPase that stimulates MuA
activity and controls the choice of DNA
target site.
• Eukaryotic transposons :
• P- Elements in Drosophila : P element is a
transposon that is present in the fruit fly,
[Link].
• It is widely used for mutagenesis and the
creation of genetically modified flies used
for genetic research.
• They seem to have first appeared in the
species only in the middle of the twentieth
century.
• Within 50 years they have spread through
every wild population of the species, so
that only older laboratory stocks lack them.
• Some naturally collected flies have no p-
elements - Empty strains – Primitive
• Some natural population contain a few p-
elements, but others contain upto 50
copies.
• P-elements are small transposons
with 31 bp inverted repeats and the
element generates 8 bp direct repeats
of target DNA sequences upon
insertion.
• The complete element consists of
2907 bp and is autonomous because
it can code for tansposase.
• Incomplete member of p-elements
have lost the transposition ability
because the transposase has been
mutated.
• But if a complete element exists in the
same cell the incomplete element can
transpose.
• The laboratory stock maintained as
empty strain (collected in 1950) is still
available.
• It is suggested that the p-element
might have entered the species by
the viruses which naturally infect
them.
p-element movement within the fly is a
maternally inherited trait.
It leads to a phenomenon called hybrid
dysgenesis
The movement is suppressed in the fly with p-
cytotype, but permissible in the M cytotype.
The p-cytotype flies contain p-elements, where
as M-cytotype do not.
When movement occurs (- it occurs in the germ
line -) in those cells which produce gametes.
This movement of p-element creates many
different changes in the phenotype but few
are lethal.
The progeny are said to be dysgenic, which
refers to biological deficiencies that are a
result of a p-element movement.
Movement is suppressed in somatic cells
where the effects would be more damaging.
This condition of germline abnormalities which
generates mutations, chromosomal
aberrations and sterility is called p-M hybrid
dysgenesis.
• Quite apart from their interest as a genetic
phenomenon, the p-elements have
become major tools of the modern
Drosophila geneticist, being used to tag
genes for cloning and to insert genes
transgenically.
LINES : The autonomous poly – A
retrotransposon, also known as LINE’s are
abundant in the genomes of vertebrate
organisms.
In human genome, 20% is composed of
LINEs.
These elements were first recognized as a
family of repeat sequences.
Their name is derived from this initial
identification – Long interspersed nuclear
element.
L1 is one of the best understood LINEs in
human
In addition to promoting their own mobility
LINES also donate the protein needed to
reverse transcribe and integrate another
class of repeat sequences the non
autonomous poly A – retrotransposons,
known as short interspersed nuclear
elements (SINEs).
Genome sequences reveal the presence of
huge number of these elements, which are
typically only between 100 to 400 bp long.
The Alu sequence is an example of a wide
spread SINE in the human genome.
A comparison of structures of typical LINE
and SINE is given in the figure.
LINE
5’UTR 3’UTR
ORF1 ORF2
SINE
Target site
duplication
• Uses of transposons : -
1. Mutagenesis : Mutagenise a strain by
taking a strain carrying the transposon
and looking for cases where the Tn has
moved and generated an interesting
mutation.
2. Selection for a mutant phenotype : Most
mutations (large mutations) leading to the
loss of gene function do not have
selectable phenotype. A mutation caused
by a transposon therefore has a distinct
advantage, since it has two phenotypes,
the loss of function of the affected gene
and the drug resistance of the transposon.
The latter is selectable, but brings the
farmer along.
3. Organisms specificity : Because most
transposons borrow some host machinery
during transposition, they tend to be limited
to hosts with compatible machinery. Most
transposons found in Gram-ve bacteria
therefore tend to be specific for that group
and fail to function in Gram+ve.
4. Genetically altered transposons : An
increasing number of genetically altered
transposons are being produced with
useful amusing properties.
5. Physically detectable homology : When a
transposon is used to generate a mutant,
the mutated region can be physically
isolated using probes for the transposon
sequence. The mutated region can then
be used in turn as a probe for the wild type
sequence.
6. Transposon tagging : Here the gene with
the insertion is tagged (marked) with
antibiotic resistance phenotype of the
transposon.
Once the gene has been tagged, by
transposon insertion, it can be used for
genetic mapping because the phenotype
resulting from the presence of the tagged
gene is antibiotic resistant which is easily
identified and selected.
