Lecture 2 – Repeat elements BB30055: Genes and genomes Genomes - Dr. MV Hejmadi (bssmvh@bath.ac.uk) Lecture 2 – Repeat elements

Lecture 2 – Repeat elements What are repeat elements? How did they originate? Why are they important?

Repetitive elements Main classes based on origin Tandem repeats Interspersed repeats Segmental duplications

1) Tandem repeats Blocks of tandem repeats at subtelomeres pericentromeres Short arms of acrocentric chromosomes Ribosomal gene clusters

Tandem / clustered repeats Broadly divided into 4 types based on size 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

Satellites Large arrays of repeats Some examples Satellite 1,2 & 3 a (Alphoid DNA) - found in all chromosomes b satellite Alphoid contains centromere binding protein CENPB HMG3 by Strachan and Read pp 265-268

Minisatellites Moderate sized arrays of repeats Some examples Hypervariable minisatellite DNA - core of GGGCAGGAXG - found in telomeric regions - used in original DNA fingerprinting technique by Alec Jeffreys Similar to ecoli chi sequence – hotspot for recombination HMG3 by Strachan and Read pp 265-268

Microsatellites VNTRs - Variable Number of Tandem Repeats, SSR - Simple Sequence Repeats /STR – short tandem repeats 1-13 bp repeats e.g. (A)n ; (AC)n 2% of genome (dinucleotides - 0.5%) Used as genetic markers (especially for disease mapping) A father might have a genotype of 12 repeats and 19 repeats, a mother might have 18 repeats and 15 repeats while their first born might have repeats of 12 and 15. Individual genotype HMG3 by Strachan and Read pp 265-268

Microsatellite genotyping design PCR primers unique to one locus in the genome a single pair of PCR primers will produce different sized products for each of the different length microsatellites .

2) Interspersed repeats A.k.a. Transposon-derived repeats ~ 45% of genome Arise mainly as a result of transposition either through DNA or RNA retrotransposons (retroposons) ‘copy and paste’ DNA transposons (‘cut & paste’) See lecture 3 for transposition

Interspersed repeats (transposon-derived) major types class family size Copy number % genome* LINE L1 (Kpn family) L2 ~6.4kb 0.5x106 0.3 x 106 16.9 3.2 SINE Alu ~0.3kb 1.1x106 10.6 LTR e.g.HERV ~1.3kb 0.3x106 8.3 DNA transposon mariner ~0.25kb 1-2x104 2.8 * Updated from HGP publications HMG3 by Strachan & Read pp268-272

Classes of transposable elements Classes of mobile elements. DNA transposons, e.g., Tc-1/mariner, have inverted terminal inverted repeats (ITRs) and a single open reading frame (ORF) that encodes a transposase. They are flanked by short direct repeats (DRs). Retrotransposons are divided into autonomous and nonautonomous classes depending on whether they have ORFs that encode proteins required for retrotransposition. Common autonomous retrotransposons are (i) LTRs or (ii) non-LTRs (see text for a discussion of other retrotransposons that do not fall into either class). Examples of LTR retrotransposons are human endogenous retroviruses (HERV) (shown) and various Ty elements of S. cerevisiae (not shown). These elements have terminal LTRs and slightly overlapping ORFs for their group-specific antigen (gag), protease (prt), polymerase (pol), and envelope (env) genes. They produce target site duplications (TSDs) upon insertion. Also shown are the reverse transcriptase (RT) and endonuclease (EN) domains. Other LTR retrotransposons that are responsible for most mobile-element insertions in mice are the intracisternal A-particles (IAPs), early transposons (Etns), and mammalian LTR-retrotransposons (MaLRs). These elements are not present in humans, and essentially all are defective, so the source of their RT in trans remains unknown. L1 is an example of a non-LTR retrotransposon. L1s consist of a 5'-untranslated region (5'UTR) containing an internal promoter, two ORFs, a 3'UTR, and a poly(A) signal followed by a poly(A) tail (An). L1s are usually flanked by 7- to 20-bp target site duplications (TSDs). The RT, EN, and a conserved cysteine-rich domain (C) are shown. An Alu element is an example of a nonautonomous retrotransposon. Alus contain two similar monomers, the left (L) and the right (R), and end in a poly(A) tail. Approximate full-length element sizes are given in parentheses Science 12 March 2004: Vol. 303. no. 5664, pp. 1626 - 1632

3) Segmental duplications Closely related sequence blocks (1-200kb) at different genomic loci Segmental duplications can occur on homologous chromosomes (intrachromosomal) or non homologous chromosomes (interchromosomal) Not always tandemly arranged Relatively recent

Segmental duplications Interchromosomal segments duplicated among non homologous chromosomes Prone to deletions/ duplications Intrachromosomal duplications occur within a chromosome / arm Prone to translocations Nature Reviews Genetics 2, 791-800 (2001);

Chromosome rearrangements originate from double strand break repair or homologous recombination between repeat sequences

Segmental duplications Segmental duplications in chromosome 22

Repeat elements How did they originate? Tandem repeats – replication slippage etc Interspersed repeats – transposition events Segmental duplications – strand exchange, recombination events

strand slippage during replication How are tandem repeats generated in the genome? strand slippage during replication Fig 11.5 HMG3 by Strachan and Read pp 330

strand slippage during replication Fig 11.5 HMG3 by Strachan and Read pp 330

Alu repeats (type of SINE) evolved from processed copies of the 7SL RNA gene

Repeat elements Why are they important? Evolutionary ‘signposts’ Passive markers for mutation assays Actively reorganise gene organisation by creating, shuffling or modifying existing genes Chromosome structure and dynamics Provide tools for medical, forensic, genetic analysis

Pathogenic potential of Short Tandem Repeats (STR) Reduction or expansion of STR can be pathogenic 1) Unstable expansion of short tandem repeats Characterised by anticipation Large expansions outside coding sequences Modest expansions within coding sequences FRAXA, FRAX E Huntington disease (HD) Myotonic dystrophy (DM1) SCA 1,2,3,6,7, 17 Friedrich ataxia (FA) Kennedy disease Spinocerebellar ataxia 8,11 Anticipation – lower age of onset and/or severity is worse over subsequent generations FA – Freidrich’s ataxia HD 10-30 CAG repeats – non pathogenic 40-200 repeats – unstable repeats cause huntingtin protein to aggregate and kill cells Pg 477 HMG3

Unstable deletions of STRs? STRs tend to be deletion hotspots Pg 358-HMG3

Interspersed repeats are susceptible to deletions/duplications E.g. Kearns-Sayre syndrome- encephalomyopathy External opthalmoplegia Ptosis Ataxia Cataract Common 4977bp deletion in mt DNA External opthalmoplegia Ptosis Ataxia Cataract Mt is recombination deficient therefore such deletions could arise due to replication slippage just like the STR

Pathogenic potential of segmental duplications Nature Reviews Genetics 2, 791-800 (2001)

References Chapters 9 and 11 Chapter 10: pp 339-348 HMG 3 by Strachan and Read Chapter 10: pp 339-348 Genetics from genes to genomes by Hartwell et al (2/e) Nature (2001) 409: pp 879-891