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Banana Transposable Elements: The hAT DNA element story PAGXXIII

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W071 Transposable Elements and Their Evolution in Musa
Date: Tuesday, January 13, 2015
Pat Heslop-Harrison, University of Leicester, United Kingdom
Gerhard Menzel, Universität Dresden, Germany
Tony Heitkam, Faisal Nouroz,Trude Schwarzacher, Thomas Schmidt
Like other plant species, DNA transposable elements and retrotransposons represent a large fraction of the Musa genome. Mobile elements can be identified 1) by homology to known elements; 2) by characteristic sequence properties such are repeats and short duplications; and 3) by studying their mobility and insertions/deletions in comparisons of homologous or homoeologous chromosome sequences. We have used all three approaches in Musa and I will show results showing the nature of unselected mobile sequences in Musa accessions. Many of the active elements proved to belong to the hAT family of DNA transposons, where there has been limited information on their diversity, abundance and chromosomal localization in plants. Chromosomal in situ hybridization, in silico analysis of genomic sequences, Southern hybridization and biodiversity panels were used to show three major families of the elements in Musa, with some 70 complete autonomous elements, and abundant hAT-related MITEs (Miniature inverted tandem elements).MuhAT transposons and MuhMITEs were localized in subtelomeric, most likely gene-rich regions, of chromosomes. Variation between homologous chromosomes and transduplications of genomic sequences indicate activity of the transposons and suggest a role for the MITEs in modulation of genomic behaviour.
Further details from www.molcyt.com;
hAt element analysis: Chromosome Research 2015 DOI 10.1007/s10577-014-9445-5.

Published in: Education

Banana Transposable Elements: The hAT DNA element story PAGXXIII

  1. 1. Transposable element evolution and mobility in Musa genomes Gerhard Menzel, Tony Heitkam, Thomas Schmidt - Dresden Germany Faisal Nouroz, Trude Schwarzacher - Leicester UK Pat Heslop-Harrison phh4@le.ac.uk Pathh1: www.molcyt.com UserID/PW ‘visitor’ .Slideshare.com pathh1
  2. 2. Heslop-Harrison & Schwarzacher T. 2007. Domestication, genomics & future for banana. Annals of Botany 100(5):1073-1084.
  3. 3. Sequence components of a genome Tandem repeats Telomeres Genes
  4. 4. Sequence components of a genome Tandem repeats Telomeres Transposable elements Genes
  5. 5. Retrotransposons Class I transposable elements RNA intermediate DNA transposons Class II transposable elements Cut-and-paste + Transposable elements: Retrotransposons via RNA and DNA TEs
  6. 6. Transposable elements can be identified - by studying their mobility and insertions/deletions in comparisons of homologous or homoeologous chromosome sequences - by characteristic sequence properties such as repeats and short duplications - by homology to known elements
  7. 7. Figure M1-1: Dot plot of homoeologous BAC clones Musa balbisiana ‘MBP_81C12’ (horizontal) against Musa acuminata ‘MA4_82I11’ (vertical). The comparison of the BACs showed large homologous region with several gap-insertion pairs. The gaps showed transposon insertions present in one BAC and absent in others. Different TEs are encircled and named. Several small insertions are not highlighted here. Musa balbisiana (MBP_81C12) Musaacuminata(MA4_82I11) Transposed MaN-hAT2 MaN-hAT1 MaN-hAT2 MbN-hAT4 MbMITE4 MbN-hAT3 MBT MaMITE1 MAWA Microsatellite MaMUL1 Faisal Nouroz
  8. 8. Figure M1-1: Dot plot of homoeologous BAC clones Musa balbisiana ‘MBP_81C12’ (horizontal) against Musa acuminata ‘MA4_82I11’ (vertical). The comparison of the BACs showed large homologous region with several gap-insertion pairs. The gaps showed transposon insertions present in one BAC and absent in others. Different TEs are encircled and named. Several small insertions are not highlighted here. Transposed MaN-hAT2 MaN-hAT1 MaN-hAT2 MbN-hAT3 MBT MaMITE1 MAWA Microsatellite hAT superfamily 8-bp TSD Short variable TIRs of 5 to 27 bp Single gene coding transposase with TE class II-specific DDE amino acid motif
  9. 9. Alignment of two homologous Musa BACs shows in-dels in B genome M. balbisiana and A genome M. acuminata MA4_82I11 MBP_81C12 MuhAT 1 MuhAT2 a XX TE (SINGLE)XX TE MITE XX TE (AGNABI) MuhAT3 MuhAT4 MITE(MBIR ) XX TE XX TE (MBT) 272 bp 102,190 bp 26, 410 bp 128,068 bp DNA transposons hAT are particularly frequent 8 bp TSD, and short TIRs of 5–27 bp transposase (sometimes degenerate) including a DDE site. Non-autonomous (MITE) derivatives of hAT with deletion coding sequence
  10. 10. 8 bp TSDs - TCCCTGAG 30 bp TIRs - CAAGGTCTGcCATACCGtAcCGTACCGgCG 273 bp MaN-hAT1 TSD TIR TIR TSD M. balbisiana MBP_81C12 M.acuminataMA4_82I11 874 bp MaN-hAT2 8 bp TSD - GTGcTAaC 15 bp TIR - CAGTGATTTaAAaAG TSD TIR TIR TSD M. balbisiana MBP_81C12 M.acuminataMA4_82I11
  11. 11. (n=38) (n=20) (n=6) (n=6) TIR Terminal Inverted Repeat conservation In Musa hATs
  12. 12. TSD TIR TIR TSD 8 bp TSDs - GTTGCAAC 15 bp TIRs - CAAGGTctGCaTACC 1292 bp MbN-hAT3 M. balbisiana MBP_81C12 M.acuminataMA4_82I11 8 bp TSDs - TTCAAATG 9 bp TIRs - CAAGGTtTG 524 bp MbN-hAT4 TSD TIR TIR TSD M. balbisiana MBP_81C12 M.acuminataMA4_82I11
  13. 13. 100 100 100 97.4 9853.3 58.2 Hatvine-7 Hatvine-10 Hatvine-2 Tag1 Bg Tip100 Tam3 Hatvine-9 Ac Tag2 Hatvine-1 MuhAT III MuhAT II MuhAT I Menzel et al. Dec 2014. The diversification and activity of hAT transposons in Musagenomes. Chromosome Research 22: 559–571. see www.molcyt.org HMM identifies three abundant Musa hAT families
  14. 14. 100 100 100 97.4 9853.3 58.2 Hatvine-7 Hatvine-10 Hatvine-2 Tag1 Bg Tip100 Tam3 Hatvine-9 Ac Tag2 Hatvine-1 MuhAT III MuhAT II MuhAT I Hidden Markov Model: 274 transposase sequences, 114 (shown) span the five conserved hAT-specific amino acid blocks in the assembly (D’Hont et al. 2012) 61 + 47 +6 Complete and autonomous: 70 MuhAT I/II; no autonomous MuhAT III HMM identifies three abundant Musa hAT families
  15. 15. 15 Insertional polymorphisms of three Musa hATs in 48 Musa/banana accessions by PCR of flanking primers HP-1 A1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 A48 HP-1 B1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 B48 -1 A1 2 3 4 5 6 7 8 HP-1 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 HP1 25 26 27 28 29 30 31 32 3 3 34 35 36 37 38 39 40 HP1 41 42 43 44 45 46 47 A48
  16. 16. Figure M1-11: Sequence specific insertion polymorphisms of Musa hATs. a-b) Agarose gel electrophoresis illustrating MbN-hAT3 insertion sites in various Musa accessions: Long bands (1441bp) showed the amplified element and short bands amplifying the pre- insertion sites only. c-d) MbN-hAT4 amplification with degenerative primer pair MbNhAT4F and MbNhAT4R. Long bands (860-bp) showed the amplified MbN-hAT4 element and short bands amplifying the flanking sequences only. Numbers across the base are identifiers for individual specimens listed in table --- a and b. 0 HP1 A1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 A48 0 HP1 B1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 HP1 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 B48 0 HP1 A1 2 3 4 5 6 7 8 HP1 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 HP1 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 HP1 41 42 43 44 45 46 47 A48 ) HP1 B1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 HP1 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 HP1 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 B48 0
  17. 17. kb 0.5 1.0 2.0 3.0 10.0 6.0 kb 0.5 1.0 2.0 3.0 10.0 6.0 kb 0.5 1.0 2.0 3.0 10.0 6.0 1 2 3 4 5 6 7 8 9 10 11 12 Musa species (left 7 lanes) M. acuminata, M. balbisiana , Cavendish , M. obino l‘Ewai , M. textilis,# M. velutina and M. ornata ; Zea mays (lane 8), Glycine max , Beta vulgaris, Arabidopsis thaliana and the gymosperm Pinus elliottiii. DraI.digests M. acuminata-specific dimerization domain of the MuhAT I transposase 1 2 3 4 5 6 7 8 9 10 11 12 Full-length MuhMITE I Probe hAT abundance
  18. 18. v n=159 n=140 autonomous (2206-6814 bp) MITEs (646-1072 bp) v n=51 n=117 430-786 bp 116-286 bp 5‘-TSD5‘-TIR MuhAT I: autonomous and derived MITES transposase 5′- and 3′- untranslated regions
  19. 19. MuhAT transposases (top) and MuhMITEs I (lower): tending to be terminal on chromosomes
  20. 20. Retroelements Sequences which amplify through an RNA intermediate • 50% of all the DNA!
  21. 21. Retroelements • Homologous BAC sequences from Calcutta 4 Homologous over the full length • except for a 5kb insert • a Ty1-copia retroelement
  22. 22. Diploid 2n=2x=22 Musa / banana metaphase probed red with retrotransposable element Teo & Schwarzacher
  23. 23. A-genome specific MuhAT1 on ABB Musa M. balbisiana (BB) f) Musa (ABB)
  24. 24. MuhAT transposases (top) and MuhMITEs I (lower): tending to be terminal on chromosomes
  25. 25. MuhAT transposons and MuhMITEs Associated with genes – introns and exons Family specificity 0 10 20 30 40 50 60 70 80 90 100 Portionofcopies[%] Exon Intron ≤ 500 bp ≤ 1000 bp ≤ 1500 bp > 1500 bp 33 12 6 96 1512
  26. 26. hAT transposons and derived MITEs evolution in Musa – Three major families – Hundreds of copies of hATs – Thousands of copies of MITEs • Subtelomeric, gene-rich regions • Species (A and B genome) -specific mobility of MuhMITEs • MuhMITE II showing transduplications of genomic sequences – active MuhAT transposons – MuhMITEs as modulators of genome evolution Gerhard Menzel, Thomas Schmidt, Faisal Nouroz Pat Heslop-Harrison phh4@le.ac.uk www.molcyt.com Twitter pathh1
  27. 27. http://molcyt.org/2014/11/10/the- diversification-and-activity-of-hat- transposons-in-musa-genomes/ • Menzel G, Heitkam T, Seibt KM, Nouroz F, Müller-Stoerme M, Heslop-Harrison JS, Schmidt T. 2014. The diversification and activity of hAT transposons in Musagenomes. Chromosome Research 22: 559–571. DOI 10.1007/s10577-014-9445-5 and Pubmed link ID: 25377178And author print hATs in Musa _2014_CR_MenzelEtAlAuthorVersion2014. • Sequencing of plant genomes often identified the hAT superfamily as largest group of DNA transposons. Nevertheless, detailed information on the diversity, abundance and chromosomal localization of plant hAT families are rare. By in silico analyses of the reference genome assembly and BAC sequences, respectively, we performed the classification and molecular characterization of hAT transposon families in Musa acuminata. Musa hAT transposons are organized in three families MuhAT I, MuhAT II and MuhAT III. In total, 70 complete autonomous elements of the MuhAT I and MuhAT II families were detected, while no autonomous MuhAT III transposons were found. Based on the terminal inverted repeat (TIR)-specific sequence information of the autonomous transposons, 1722 MuhAT I- and MuhAT II-specific miniature inverted repeat transposable elements (MuhMITEs) were identified. Autonomous MuhAT I and MuhAT II elements are moderately abundant in the sections of the genus Musa, while the corresponding MITEs exhibit an amplification in Musa genomes. By fluorescent in situhybridization, autonomous MuhAT transposons as well as MuhMITEs were localized in subtelomeric, most likely gene-rich regions of M. acuminata chromosomes. A comparison of homoeologous regions of M. acuminata and Musa balbisiana BACs revealed the species-specific mobility of MuhMITEs. In particular, the activity of MuhMITEs II showing transduplications of genomic sequences might indicate the presence of active MuhAT transposons, thus suggesting a potential role of MuhMITEs as modulators of genome evolution of Musa. • Keywords Musa acuminata, Musa balbisiana, genome assembly, BAC, hATtransposons, FISH
  28. 28. https://pag.confex.com/pag/xxiii/web program/Paper14993.html • W071Transposable Elements and Their Evolution in Musa • Date: Tuesday, January 13, 2015 Time: 11:10 AM • Room: Pacific Salon 6-7 (2nd Floor) • Pat Heslop-Harrison , University of Leicester, Leicester, Leic, United Kingdom • Gerhard Menzel , Institute of Botany, Technische Universität Dresden, Dresden, Germany • Tony Heitkam , Institute of Botany, Technische Universität Dresden, Dresden, Germany • Faisal Nouroz , University of Leicester, Leicester, United Kingdom • Trude Schwarzacher , University of Leicester, Leicester, United Kingdom • Thomas Schmidt , Dresden University of Technology, Dresden, Germany • Like other plant species, DNA transposable elements and retrotransposons represent a large fraction of the Musa genome. Mobile elements can be identified 1) by homology to known elements; 2) by characteristic sequence properties such are repeats and short duplications; and 3) by studying their mobility and insertions/deletions in comparisons of homologous or homoeologous chromsome sequences. We have used all three approaches in Musa and I will show results showing the nature of unselected mobile sequences in Musa accessions. Many of the active elements proved to belong to the hAT family of DNA transposons, where there has been limited information on their diversity, abundance and chromosomal localization in plants. Chromosomal in situ hybridization, in silico analysis of genomic sequences, Southern hybridization and biodiversity panels were used to show three major families of the elements in Musa, with some 70 complete autonomous elements, and abundant hAT-related MITEs (Miniature inverted tandem elements).MuhAT transposons and MuhMITEs were localized in subtelomeric, most likely gene-rich regions, of chromosomes. Variation between homologous chromosomes and transduplications of genomic sequences indicate activity of the transposons and suggest a role for the MITEs in modulation of genomic behaviour.Further details fromwww.molcyt.com; hAt element analysis: Chromosome Research 2015 DOI10.1007/s10577-014-9445-5.
  29. 29. Some Musa / banana papers from Pat Heslop-Harrison and www.molcyt.com • 311. Menzel G, Heitkam T, Seibt KM, Nouroz F, Müller-Stoerme M, Heslop-Harrison JS, Schmidt T. 2014. The diversification and activity of hAT transposons in Musagenomes. Chromosome Research 22: 559– 571. DOI 10.1007/s10577-014-9445-5 and Pubmed link ID: 25377178And author print hATs in Musa _2014_CR_MenzelEtAlAuthorVersion2014. • 300. D’Hont A, … Heslop-Harrison P, … Wincker P. 2012. The banana (Musa acuminata) genome and the evolution of monocotyledonous plants. Nature 488 (7410), 213-217. DOI: http://dx.doi.org/10.1038/nature11241 • 303. Nair AS, Heslop-Harrison P, Schwarzacher T. 2013. Production of haploid tissues and SNP analysis of the genome in Musa acuminata cv.‘Matti’ (AA). Plant Mutation Reports 3(1): 18-24. PMR Vol3 No1 • 250. Heslop-Harrison JS, Schwarzacher T. 2007. Domestication, genomics and the future for banana. Annals of Botany 100(5):1073-1084.doi:10.1093/aob/mcm191 • 238. Nair AS, Teo CH, Schwarzacher T, Heslop-Harrison JS. 2005 Genome classification of banana cultivars from South India using IRAP markers.Euphytica 144: 285-290. DOI: 10.1007/s10681-005-7321-2 • 237. Teo CH, Tan SH, Ho CL, Faridah QZ, Othman YR, Heslop-Harrison JS , Kalendar R , Schulman AH. 2005. Genome constitution and classification using retrotransposon-based markers in the orphan crop banana. Journal of Plant Biology 48(1): 96-105 • 184. Harper G, Osuji JO, Heslop-Harrison JS, Hull R. 1999. Integration of banana streak badnavirus into the Musa genome: molecular and cytogenetic evidence. Virology 255: 207- 213. doi:10.1006/viro.1998.9581 • 168. Osuji JO, Harrison G, Crouch J, Heslop-Harrison JS. 1997. Identification of the genomic constitution of Musa L. lines (bananas, plantains and hybrids) using molecular cytogenetics. Annals of Botany 80: 787- 793.
  30. 30. From Chromosome to Nucleus Pat Heslop-Harrison phh4@le.ac.uk www.molcyt.com
  31. 31. MuMITE I-8 81,400 81,800 82,200 82,600 83,000 83,400 (B) AC226051 MuMITE II-24 MuMITE I-9 MuMITE II-8 MuMITE II-7 MuMITE II-9 MuMITE II-13 MuMITE II-12 MuMITE II-11 MuMITE II-10 2000 3000 4000 5000 6000 7000 8000 9000 10,000 11,000 12,000 13,000 14,000 15,000 16,000 17,000 18,000 19,000 (A) AC226036
  32. 32. Genome evolution How do genomes evolve? – Gene mutation very rarely (human: 10−8 /site/generation) – Chromosome evolution – Polyploidy and genome duplication (ancient and modern) – Repetitive sequences: mobility & copy number (10−4 /gen.in µsat) – Recombination – Epigenetic aspects – centromeres & expression How can we exploit knowledge of genome evolution? – Biodiversity – Chromosome and genome engineering – Breeding – Markers

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