Transposone

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Transposone

  1. 2. <ul><li>Transposone </li></ul>Presented by: Salar Bakhtiyari
  2. 3. They are discrete sequence in the genome that are mobile they are able to transport themselves to other location. Other names: <ul><li>Jumping genes </li></ul><ul><li>Selfish DNAs </li></ul><ul><li>Molecular parasites </li></ul><ul><li>Controlling elements </li></ul>TEs are present in the genome all species of three domains Transposable Elements
  3. 4. What do we want to know about mobile genetics elements? <ul><li>1 – The history of mobile genetic elements </li></ul><ul><li>2 – The classification of TEs </li></ul><ul><li>3 – The structure of TEs </li></ul><ul><li>4 – The mechanism of transposition </li></ul><ul><li>5 – The effects of TEs on gene and genome </li></ul><ul><li>6 – The use of TEs as molecular tools </li></ul>
  4. 5. Why study mobile genetic elements? <ul><li>They are the major forces driving evolution </li></ul><ul><li>They can cause genome rearrangement (mutation , deletion and insertion ) </li></ul><ul><li>They have wide range of application potentials </li></ul>
  5. 6. The discovery of mobile genetic elements <ul><li>Transposable elements </li></ul><ul><li>Phage </li></ul><ul><li>Plasmid DNA </li></ul>
  6. 7. The discovery of transposable elements <ul><li>Barbara Mc Clintock discovered TEs in maize (1983) </li></ul><ul><li>Her work on chromosome breakage began by investigating genetic instability (1983) </li></ul><ul><li>Observing variegated patterns of pigmentation in maize plant and kernels </li></ul><ul><li>New kinds of genetic instability </li></ul><ul><li>She spent the next tree decades for this genetic elements </li></ul><ul><li>Controlling elements (1956) </li></ul>
  7. 8. Barbara Mc Clintock 1902  1980 ( noble in 1984)
  8. 9. Plasmid , phage <ul><li>Cell to cell conjugation </li></ul><ul><li>Bactriophage mediated transduction </li></ul><ul><li>Bill Hayes ( 1952 ) </li></ul><ul><li>Ellin Wollman and Francois Jancob , 1961 </li></ul><ul><li>Alan Campbell </li></ul>
  9. 10. Classification of transposable elements <ul><li>DNA transposons </li></ul><ul><li>Retrotransposons </li></ul>
  10. 11. Autonomous and non autonomous elements <ul><li>Both class are subdivided into distinct superfamilies and families </li></ul><ul><li>Structure feature , internal organization , the size of target site duplication , sequence similarities at the DNA and protein levels </li></ul><ul><li>Autonomous : they have the ability to excise and transpose </li></ul><ul><li>non autonomous elements </li></ul><ul><li>They don’t transpose </li></ul><ul><li>They become unstable only when an autonomous member of same family is present elsewhere in the genome </li></ul><ul><li>They are derived from autonomous elements </li></ul><ul><li>A family consists of single type of autonomous element accompanied by many varieties of non autonomous elements </li></ul>
  11. 13. DNA based elements <ul><li>Insertion sequence (IS) </li></ul><ul><li>The simplest (smallest) transposons are called IS </li></ul><ul><li>The IS elements are normal constituents of bacterial chromosome and plasmids </li></ul><ul><li>Spontaneous mutation of the lac and gal operons </li></ul><ul><li>They are autonomous units ,each of which codes only transposase </li></ul>
  12. 14. Structure of IS
  13. 15. Composite transposone <ul><li>One class of large transposons are called Composite transposons </li></ul><ul><li>They carring the druge marker is flanked on either side by arms that consist of IS elements </li></ul>IS modules - identical (both functional: Tn9; Tn903) or closely related (differ in functional ability: Tn10; Tn5) 1. A functional IS module can transpose either itself or the entire transposon
  14. 16. Mechanism of transposition <ul><li>The stugger between the cuts determines the length of the direct repeats. </li></ul><ul><li>The target repeat is characteristic of each transposon; reflects the geometry of the cutting enzyme </li></ul>Direct repeats are generated by introduction of staggered cuts whose protruding ends are linked to the transposon .
  15. 17. Mechanism of transposition 1- Replicative transpositon <ul><li>Replicative : </li></ul><ul><li>Transposon is duplicated ; a copy of the original element is made at a recipient site(TnA); donor keeps original copy </li></ul><ul><li>Transposition- an increase in the number of Tn copies </li></ul><ul><li>ENZs: transposase (acts on the ends of original Tn) and resolvase (acts on the duplicated copies) </li></ul>
  16. 18. Mechanism of transposition 2 - Nonreplicative <ul><li>Nonreplicative : </li></ul><ul><li>Transposon moves from one site to another and is conserved; breaks in donor repaired </li></ul><ul><li>b) IS and Tn10 and Tn5 use this mechanism; no Tn copy increase </li></ul><ul><li>c) ENZs: only transposase </li></ul>
  17. 19. <ul><li>The first stages of Both replicative and non-replicative transpositio are semilar </li></ul><ul><li>IS elements, prokaryotic eukaryotic transposons, and bacteriophage Mu . </li></ul>Donor cut 1. Synapsis stage - two ends of transposon are brought together 3. . Nicked ends joine crosswise;covalent connection between the transposon the target 2. Transposon nicked at both ends; target nicked at both strands
  18. 20. cuts in trans transfers in trans 22 bp Mu integrates by nonreplicative transposition; during lytic cycle- number of copies amplified by replicative transposition - MuA binds to ends as tetramer forming a synapsis . - MuA subunits act in trans to cut next to R1 and L1 (coordinately; two active sites to manipulate DNA). - MuA acts in trans to cut the target site DNA and mediate in trans strand transfer
  19. 21. The chemistry of Donor and target cut The 3’-ends ends groups released from flanking DNA by donor cut reaction They are nuclophile that attack phosphodiester bonds in target DNA <ul><li>Cutting of both ends </li></ul>3 ‘ OH 3 ‘ OH 3 ‘ OH 3 ‘ OH <ul><li>Cutting of 3 ‘ end only </li></ul>
  20. 22. <ul><li>The product of these reaction is strand transfer complex </li></ul><ul><li>In strand transfer complex transposon is connected to the target site through one strand at each end </li></ul><ul><li>Next step differs and determines the type of transposition: </li></ul><ul><li>Strand transfer complex can be target for replication (replicative transposition) or for repair (nonreplicative transposition; breakage & reunion ) </li></ul>transposon target Strand transfer complex
  21. 23. Molecular mechanism of transposition (I) Replicative transposition Replicative transposition proceeds through a cointegrate . Transposition may fuse a donor and recipient replicon into a cointegrate. Resolution releases two replicons-each has copy of the transposon
  22. 24. Replicative transposition Ligation to target ends 3. 3’-ends prime replication The crossover structure contains a single stranded region at each of the staggered ends= pseudoreplication forks that provide template for DNA synthesis Donor and target cut cointegrate .
  23. 25. Non-replicative Replicative additional nicking common structure Breakage & reunion
  24. 26. Retrotransposon ( retroposons ) <ul><li>Use of an RNA Intermediate </li></ul><ul><ul><li>element is transcribed </li></ul></ul><ul><ul><li>reverse transcriptase produces a double-stranded DNA copy for insertion at another site </li></ul></ul><ul><ul><li>they as other elements generating short direct repeat </li></ul></ul>
  25. 27. Types of Retrotransposons <ul><li>1 – viral superfamily (autonomousretrotransposon) </li></ul><ul><ul><ul><ul><li>retrovirus </li></ul></ul></ul></ul><ul><ul><ul><ul><li>LTR- retrotransposon </li></ul></ul></ul></ul><ul><ul><ul><ul><li>LINES </li></ul></ul></ul></ul><ul><li>2 – nonviral superfamily </li></ul><ul><li>(non autonomous retransposons) </li></ul><ul><li>SINES </li></ul>non LTR- retrotransposon
  26. 28. retrovirus RNA reverstranscriptase Liner DNA Integration provirus Transcription RNA
  27. 29. LTR - retrotrasposon pol Reverse transcriptase (RT) Integrase (IN) Ribonuclease H (RH) gag env ?
  28. 30. mechanism of transposition Integrase acts on both the retrotransposon line DNA and target DNA <ul><li>The integrase bring the ends </li></ul><ul><li>of the linear DNA together </li></ul><ul><li>Generate 2 base recessed 3’ -ends </li></ul><ul><li>and staggered end in target DNA </li></ul>3’-ends 5’-ends
  29. 31. Non – LTR retrovirus <ul><li>LINES = long interspersed elements </li></ul><ul><li>SINES = short interspersed elements </li></ul><ul><li>don’t terminate in the LTRs </li></ul><ul><li>they are significant part of relatively short sequence of mammalian genomes . </li></ul>
  30. 32. Effect of transposabli elements on gene and genome <ul><li>TEs cause a varity of change in the genome of their hosts </li></ul><ul><li>this ability to induce mutation depend on their of capability of transposing </li></ul><ul><li>some arrangement can be beneficial they can advantageous for adaptation to new environment </li></ul><ul><li>play important role in evolution . </li></ul><ul><li>they have the ability to rearrange genomic information in several ways </li></ul><ul><li>1 – Modification of gene expression </li></ul><ul><li>2 – Alternation gene sequence </li></ul><ul><li>3 – Chromosomal structural changes </li></ul>
  31. 33. Modification of gene expression <ul><li>insertion of a TE within or adjacent to a gene </li></ul><ul><li>the element blocks or alters the pattern of transcription . </li></ul><ul><li>i nsertion of Fot1 in a intron of niad ( F . oxysporum ) </li></ul><ul><li>different mutant transcripts all were shorter </li></ul><ul><li>They result from: </li></ul><ul><li>- presence of termination signal </li></ul><ul><li>- presence of an alternative promotor </li></ul>
  32. 34. Alternation gene sequence <ul><li>cut and pate mechanism often produce variation when they excise . </li></ul><ul><li>the excision process may result in addition of a few base pair ( footprint ) at donor site . </li></ul><ul><li>these footprint cause diversification of nucleotide sequence and new functional alleles </li></ul><ul><li>Example : Fot1 and Impala generally leave 4 bp – ( 108 ) or 5 – ( 63 ) foot prints </li></ul><ul><li>excision of the Asco - 1 transposon in A .immersus </li></ul><ul><li>Deletions of a a few to up to 1713 nucleotide in b2 gene </li></ul><ul><li>larger deletion led to variety of phenotypes in spore coloration </li></ul>
  33. 35. Chromosomal structural changes <ul><li>TEs can produce a series of genome rearrangment through ectopic recombination </li></ul><ul><li>deletion , duplication , inversion and translucation mediate by TEs ( Drosophila , Yeast , human ) </li></ul><ul><li>karyoptypic variation in natural isolate in fungai </li></ul><ul><li>high level of chromosome – length polymorphism ( Magnoporthe grisea , F. oxysporum ) </li></ul><ul><li>translocation tox1 of Cochliobolus heterostrophus </li></ul><ul><li>appearance of new virulence alleles in M . grisea </li></ul>
  34. 36. Use as strain specific diagnostic tools <ul><li>TEs are often restricted to specific strains </li></ul><ul><li>identify specific pathogen in plant pathology </li></ul><ul><li>Fot1 ( F. oxysporum f sp. albedians ) provide PCR targets </li></ul><ul><li>a sensitive detection thechnique to prevent the introduction of pathogenic form </li></ul><ul><li>- race of F. oxysporum responsible of carnation wilt </li></ul><ul><li>- date palm pathogen </li></ul>Use of TEs as molecular tools
  35. 37. Use of TEs as molecular tools <ul><li>MGR 586 ( Magneporthe grisea ) </li></ul><ul><li>oryza : 30 – 50 wheat and other ( 1 – 2 ) </li></ul><ul><li>they have used to distinguish genetically divergent population </li></ul><ul><li>fingerprinting of isolates pathogenic to oil palm tree. ( F. oxysporum , palm ) </li></ul>Tools for the analysis of population structure
  36. 38. Gene tagging with transposable elements <ul><li>arise mutant phenotype </li></ul><ul><li>Disrupt target gene </li></ul>Use of TEs as molecular tools <ul><li>jumping into coding region </li></ul>Target gene can easily determined by PCR methods
  37. 39. Thanks for attention

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