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

3. Repetitive elements Significance 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

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

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

6. Tandem / clustered repeats classSize of repeat Repeat block Major chromosomal location Satellite5-171 bp> 100kbcentromeric heterochromatin minisatellite9-64 bp0.1–20kbTelomeres microsatellites1-13 bp< 150 bpDispersed HMG3 by Strachan and Read pp 265-268 Broadly divided into 4 types based on size

7. Satellites Large arrays of repeats Some examples Satellite 1,2 & 3  Alphoid DNA) - found in all chromosomes  satellite HMG3 by Strachan and Read pp 265-268

8. 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 HMG3 by Strachan and Read pp 265-268

9. Microsatellites VNTRs - 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

10. 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

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

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

13. 2) Interspersed repeats A.k.a. Transposon-derived repeats ~ 45% of genome Arise mainly as a result of transposition either through a DNA or a RNA intermediate

14. Interspersed repeats (transposon-derived) classfamilysizeCopy number % genome* LINEL1 (Kpn family) L2 ~6.4kb0.5x10 6 0.3 x 10 6 16.9 3.2 SINEAlu~0.3kb1.1x10 6 10.6 LTRe.g.HERV~1.3kb0.3x10 6 8.3 DNA transposon mariner~0.25kb1-2x10 4 2.8 major types * Updated from HGP publications HMG3 by Strachan & Read pp268-272

15. Most ancient of eukaryotic genomes  Autonomous transposition (reverse trancriptase)  ~6-8kb long, located mainly in euchromatin  Internal polymerase II promoter and 2 ORFs  3 related LINE families in humans – LINE-1, LINE-2, LINE-3. LINE-1 still active (~17% of human genme)  Believed to be responsible for retrotransposition of SINEs and creation of processed pseudogenes LINEs (long interspersed elements)

16. Nature (2001) pp879-880 HMG3 by Strachan & Read pp268-272

17.  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. SINEs (short interspersed elements) 100-300bp1,500,00013%

18. Alu repeats evolved from processed copies of the 7SL RNA gene

19. 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.

20. 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

21. Repeats on the same orientation on both sides of element e.g. ATATATnnnnnnnnnnnnnnATATAT contain sequences that serve as transcription promoters as well as terminators. These sequences allow the element to code for an mRNA molecule that is processed and polyadenylated. At least two genes coded within the element to supply essential activities for the retrotransposition mechanism. The RNA contains a specific primer binding site (PBS) for initiating reverse transcription. A hallmark of almost all mobile elements is that they form small direct repeats formed at the site of integration. Long Terminal Repeats (LTR)

22.  Autonomous or non-autonomous  Autonomous LTR encode retroviral genes gag, pol genes e.g HERV  Non-autonomous elements lack the pol and sometimes the gag genes e.g. MaLR Long Terminal Repeats (LTR) Nature (2001) pp879-880HMG3 by Strachan & Read pp268-272

23. DNA transposons Inverted repeats on both sides of element e.g. ATGCNNNNNNNNNNNCGTA DNA transposons (lateral transfer?) Nature (2001) pp879-880 From GenesVII by Levin

24. 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

25. 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);

26. Segmental duplications Segmental duplications in chromosome 22

27. Segmental duplications - chromosome 7.

28. Pathogenic potential of Short Tandem Repeats (STR) Reduction or expansion of STR can be pathogenic Large expansions outside coding sequences Modest expansions within coding sequences FRAXA, FRAX EHuntington disease (HD) Myotonic dystrophy (DM1)SCA 1,2,3,6,7, 17 Friedrich ataxia (FA)Kennedy disease Spinocerebellar ataxia 8,11 1) Unstable expansion of short tandem repeats Characterised by anticipation

29. Unstable deletions of STRs? STRs tend to be deletion hotspots

30. Interspersed repeats are susceptible to deletions/duplications External opthalmoplegia Ptosis Ataxia Cataract Common 4977bp deletion in mt DNA E.g. Kearns-Sayre syndrome- encephalomyopathy

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

32. References 1)Chapter 9 pp 265-268 HMG 3 by Strachan and Read 2)Chapter 10: pp 339-348 Genetics from genes to genomes by Hartwell et al (2/e) 3)Nature (2001) 409: pp 879-891