repetitive elements significance evolutionary signposts passive markers for mutation assays actively...
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Repetitive elements
Significance
Evolutionary ‘signposts’ Passive markers for mutation assays Actively reorganise gene organisation by
creating, shuffling or modifying existing genes
Chromosome structure and dynamicsProvide tools for medical, forensic,
genetic analysis
Repetitive sequencesAAA, ATATATAT, CGTCGTCGT etc..
5 main classes
1) Tandem repeats2) Transposon-derived
repeats3) Segmental duplications4) Processed pseudogenes
1) Tandem repeats
Blocks of tandem repeats at subtelomeres pericentromeres Short arms of acrocentric
chromosomes Ribosomal gene clusters
Tandem / clustered
repeats
class Size of repeat
Repeat block
Major chromosomal
location
Satellite 5-171 bp > 100kb centromeric
heterochromatin
minisatellite 9-64 bp 0.1 – 20kb Telomeres
microsatellites 1-13 bp < 150 bp Dispersed
HMG3 by Strachan and Read pp 265-268
Broadly divided into 4 types based on size
SatellitesLarge arrays of repeats
Some examplesSatellite 1,2 & 3
- found in all chromosomes
Alphoid DNA) satellite
HMG3 by Strachan and Read pp 265-268
MinisatellitesModerate sized arrays of repeats
Some examplesHypervariable minisatellite DNA
- core of GGGCAGGAXG- found in telomeric regions- used in original DNA fingerprinting technique by Alec Jeffreys
HMG3 by Strachan and Read pp 265-268
MicrosatellitesVNTRs - variable number of tandem repeats, SSR - simple sequence repeats
1-13 bp repeats e.g. (A)n ; (AC)n
HMG3 by Strachan and Read pp 265-268
2% of genome (dinucleotides - 0.5%)Used as genetic markers (especially for disease mapping)
Individual genotype
Microsatellite genotyping
Fig 7.7 HMG3 by Strachan and Read pp 190
The most common way to detect microsatellites is to design PCR primers that are unique to one locus in the genome and that base pair on either side of the repeated portion
Therefore, a single pair of PCR primers will work for every individual in the species and produce different sized products for each of the different length microsatellites
Microsatellite genotyping.
CA repeat genotyping.
Fig 7.8 HMG3
A B C D E Allele typesA (3,6)B (1,5)C (3,5)D (2,5)E (3,6)
Marker D17S800
N.B. ‘stutters’ or shadow bands
Caused by strand slippage
strand slippage during replication
Fig 11.5 HMG3 by Strachan and Read pp 330
Fig 11.5 HMG3 by Strachan and Read pp 330
strand slippage during replication
2) Transposon-derived repeats
A.k.a. interspersed repeats45% of genomeArise mainly as a result of transposition either through a DNA or a RNA intermediate
4 main typesLINES, SINES, LTRs and DNA transposons
Repetitive elements…
Most ancient of eukaryotic genomes Autonomous transposition (reverse
trancriptase) ~6-8kb long Internal polymerase II promoter and 2 ORFs 3 related LINE families in humans
– LINE-1, LINE-2, LINE-3.
Believed to be responsible for retrotransposition of SINEs and creation of processed pseudogenes
LINEs (long interspersed elements)
Transposon-derived repeats…
Nature (2001) pp879-880 HMG3 by Strachan & Read pp268-272
Non-autonomous (successful freeloaders! ‘borrow’ RT from other sources such as LINEs)
~100-300bp long Internal polymerase III promoter No proteins Share 3’ ends with LINEs 3 related SINE families in humans
– active Alu, inactive MIR and Ther2/MIR3.
HMG3 by Strachan & Read pp268-272
SINEs (short interspersed elements)
Transposon-derived repeats…
Nature (2001) pp879-880
LINES and SINEs have preferred insertion sites
• In this example, yellow represents the distribution of mys (a type of LINE) over a mouse genome where chromosomes are orange. There are more mys inserted in the sex (X) chromosomes.
Try the link below to do an online experiment which shows how an Alu insertion polymorphism has been used as a tool to reconstruct the human lineage
http://www.geneticorigins.org/geneticorigins/pv92/intro.html
Repeats on the same orientation on both sides of element
e.g. ATATATNNNNNNNATATAT
Autonomous or non-autonomous Autonomous retroposons encode gag, pol
genes which encode the protease, reverse transcriptase, RNAseH and integrase
Long Terminal Repeats (LTR)Transposon-derived repeats…
Nature (2001) pp879-880 HMG3 by Strachan & Read pp268-272
DNA transposons Inverted repeats on both sides of elemente.g. ATGCNNNNNNNNNNNCGTA
DNA transposons (lateral transfer?)
Transposon-derived repeats…
Nature (2001) pp879-880
FromGenesVII by Levin
Transposon derived repeats
class family size Copies*
% genome*
LINE LINE-1 (Kpn family)
~6.4kb 0.8x106 15.4
SINE Alu ~0.3kb 1.3x106 10.7
LTR e.g.HERV ~1.3kb 0.7x106 7.9
DNA
transposon
mariner ~0.25kb 0.4x106 2.7
major types
* Updated from HGP publications HMG3 by Strachan & Read pp268-272
3) Segmental duplications
Closely related sequence blocks at different genomic loci
Transfer of 1-200kb blocks of genomic sequence
Segmental duplications can occur on homologous chromosomes (intrachromosomal) or non homologous chromosomes (interchromosomal)
Not always tandemly arranged Relatively recent
Segmental duplicationsInterchromosomal segments duplicated
among non-homologous
chromosomes
Intrachromosomal duplications occur within a
chromosome / arm
Nature Reviews Genetics 2, 791-800 (2001);
Segmental duplicationsSegmental duplications in
chromosome 22
Segmental duplications - chromosome 7.
Nature Reviews Genetics 2, 791-800 (2001)
4) Pseudogenes - processed
Repetitive sequencesAAA, ATATATAT, CGTCGTCGT etc..
5 main classes1) Tandem repeats2) Transposon-derived repeats3) Segmental duplications4) Processed pseudogenes
7) Repeat content
a) Age distribution
b) Comparison with other genomes
c) Variation in distribution of repeats
d) Distribution by GC content
e) Y chromosome
Nature (2001) 409: pp 879-891
Insights from the HGP………
Repeat content…….
Most interspersed repeats predate eutherian
radiation (confirms the slow rate of clearance of
nonfunctional sequence from vertebrate genomes)
LINEs and SINEs have extremely long lives
2 major peaks of transposon activity
No DNA transposition in the past 50MYr
LTR retroposons teetering on the brink of extinction
a) Age distribution
overall decline in interspersed repeat activity in hominid lineage in the past 35-40MYr
compared to mouse genome, which shows a younger and more dynamic genome
a) Age distribution
b) Comparison with other genomes
Higher density of transposable elements in euchromatic portion of genome
Higher abundance of ancient transposons
60% of IR made up of LINE1 and Alu repeats
whereas DNA transposons represent only 6%
(a few human genes appear likely to have resulted from horizontal transfer from bacteria!!)
c) Variation in distribution of repeatsSome regions show eitherHigh repeat density
e.g. chromosome Xp11 – a 525kb region shows 89% repeat density
Low repeat density e.g. HOX homeobox gene cluster (<2% repeats)
(indicative of regulatory elements which have low tolerance for insertions)
High GC – gene rich ; High AT – gene poor
LINEs abundant in AT-rich regionsSINEs lower in AT-rich regions
Alu repeats in particular retained in actively transcribed GC rich regions E.g. chromosme 19 has 5% Alus compared to Y chromosome
d) Distribution by GC content
Unusually young genome (high tolerance to gaining insertions)
Mutation rate is 2.1X higher in male germline
Possibly due to cell division rates or different repair mechanisms
e) The Y chromosome !
Repeat content…….
• Working draft published – Feb 2001
• Finished sequence – April 2003
• Annotation of genes going on
ReferencesText: 1) Human Molecular Genetics 3 by Strachan
and Read – Chapter 9 pp 265-268
Optional Reading
1) Batzer MA, Deininger PL Alu repeats and human genomic diversity Nature Rev Genet 3 (5): 370-379 May 2002
2) BS Emanuel & TH Shaikh Segmental duplications: an 'expanding' role in genomic instability and disease Nature Reviews Genetics 2, 791-800 (2001)
3) Nature (2001) 409: pp 879-891