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Transposone And Retrotransposone

Transposone And Retrotransposone






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    Transposone And Retrotransposone Transposone And Retrotransposone Presentation Transcript

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