15. + 16. Transposable elements in eukaryotes Flashcards
What is a transposable element?
Transposable element (TE) - jumping genes - mobile DNA fragments which can copy themselves around the genome
- can jump in/out -> affect expression of nearby genes
What are the types of TEs in eukaryotes?
TEs types:
Class I: retrotransposons / “Copy & Paste” / RNA->DNA
Class II: transposons / “Cut & paste” / DNA
Can TEs affect other gene expression?
Yes, TEs moving around the genome can change/block expression of nearby genes
Do TEs make up a large proportion of human genome?
Yes, TEs make up around 50% of human genome - mostly Alu (SINE) and L1 (LINE)
What are Alu and L1s TEs?
L1: LINE - encodes own reverse transcriptase
Alu: SINE - uses LINE reverse transcriptase for transposition
Are TEs proportion in genome constant between species?
No, proportion of TEs in genomes highly variable - corn up to 85% - C. elegans 15%
What are the reason why TEs proportions in genomes variable between species?
The reason underlying TEs genome proportion variability and ability to change % proporiton quickly within a species because of:
- different transposition rates (moving)
- different acquisition rates of new TEs (new TEs introduced)
- different efficiency in removing TEs
How do TEs spread in populations more quickly than normal genes?
TEs have different inheritance patterns compared to normal genes:
TEs spread more quickly in populations because are inherited at >50% frequency (cheat Mendelian genetics) - even if TE is harmful - if reproduction fitness is not reduced more than TE gain -> TE will spread in population
Explain how retrotransposons increase their copy numbers
Retrostransposons - “copy & paste”:
- TE transcribed as RNA
- reverse transcriptase RNA->DNA
- DNA incorporated into new location in the genome => new TE copy
Explain how transposons increase their copy numbers
Transposons - “cut & paste” - if only cut no extra copy created:
1) Copy by repair using sister chromatid
- after replication 2 chromatids - TE in one chromatid cut - jumps to new location
- ssDNA gap repaired using sister chromatid as template => 3 TE copies
2) Copy by moving ahead of replication
- during replication - when TE location doubled - 2 chromatids
- TE from one chromatid cut - jumps ahead of replication fork - replicated again - in both chromatids => 3 TE copies
What are the two mechanisms used by transposons to copy themselves?
Transposons use “cut & paste” method - need somehow copy:
1) Copy by repair using sister chromatid
2) Copy by moving ahead of replication fork
What are the possible consequences of transposition? Give examples
Transposition consequences for the host can be both:
- harmful: changed protein expression - disfunctional gene
- beneficial: acts instead of telomeres in Drosophila
Why are TEs transposing in the germline and not somatically?
Host-parasite relationship:
If TEs transposed somatically - bad for both host and TEs:
- somatic transposition (mutation) could harm the host
- TEs would not spread if hapenned somatically
=> bad for both
When TEs transpose in germline - bad only for the host:
- germline transposition doesn’t affect host - only passing of its genes
- TEs can spread if transpose in germline
=> good for TEs, bad for host
==> TEs transpose in germlines
How does transposition affect the host’s gene expression?
Transposition can:
- break coding genes by breaking the ORF / promoter
- TEs carry promoters / enhancers - affect neighbouring gene expressions
How is the effect of transposition been characterised in rodents?
Transposition effect event only strong for new insertions - fades over time => expression change of neighbouring genes is associated with TE insertion event
Give an example how TE can affect human behaviour?
HK2 - retrotransposon - recently active in human germline - present in 5% - HK2 transposition affects RASGRF2 expression level (associated with dopaminergic signalling) => people with this transposition respond to dopamine differently - carrying the allele doubles the chance of being chronic injection drug user
Can transposition cause disease?
Yes, certain TE insertion can cause severe developmental disorders
What is an ectopic recombination?
Ectopic recombination - atypical recombination event which happens between homologous sequences at non-allelic chromosome positions
–VS non-homologous allelic recombination (NHAR)
How TEs allow ectopic recombinations?
TEs sequences don’t diverge fast (at normal mutation rate) - when TEs move - homologous sequences in random chromosome places - recombination possible - within a chromosome / between 2 chromsomes
Explain ectopic recombination within one chromosome
Ectopic recombination within a single chromosome:
- two identical TEs on one chromosome - pair - form a loop
- recombine genetic info - chunk of the sequence is lost -> sequence DELETION (harmful)
Explain ectopic recombination between 2 chromsomes
Ectopic recombination between 2 chromosomes:
- two identical TEs on two chromosomes - pair - loops formed because alignment incorrect - based on TEs not chromosome lengths
- recombination ->
1) sequence DUPLICATION (not as harmful)
2) sequence DELETION (harmful)
Although sequence duplication because of TE ectopic recombination is not as harmful, what risk increases upon duplication?
When sequence duplicated in ectopic recombination because of TEs - # of TEs also increases => if TE # increases - chance of next ectopic recombination increases at (# TEs)^2
Can transposition have beneficial effects?
Yes, more rare than harmful - mutation introduced by TEs can be beneficial and go to fixation
Example: Doc non-LTR retrotransposon inserted upstream of TSS -> increased cytochrome P450 expression - higher detoxification of DDT - insecticide => flies with insertion became resistant - mutation selected for - spread in population
What is TE domestication?
TE domestication - adaptation of inserted TE within the genome to serve novel functions in a host cell
Give an exaple for TE domestication
In Drosophila telomeres have been lost in evolution -> TEs transpose on chromosome ends - repeats used as telomeres
How can TEs insertions be identified in the genome?
TE insertions can be identified using DNA sequencing - from reads and read-pairs:
- analyse reference sequence: cut - read - construct back reference genome
- analyse sample in terms of reference sequence: cut - read - see if match to reference - new reads = new TE inserts
How can TE insertion (transposition) rates be identified?
Estimate transposition rate by sequencing families (parents-offspring) over time - comparing genomes - figure out TE insertions from mutation accumulation lines
Are transposition rates identical within species?
Transposition rates vary between species / families / individuals - specific factors affect transposition rates in each individual + in each tissue
How are locations of new TE inserts identified?
Difficult to identify - remove host supression mechanism against TEs (selected against) - activate TEs - compare TE insertions with previous generations => identify new locations of TEs
What is the evidence that TE insertions is highly selected against?
Strong selection against TE insertion is proved - only low number is passed to offspring from de novo TEs in parent
What are the genomic locations where transposition occurs?
New TE inserts occur in:
- promoters
- exons
- introns - more rare
More common in promoters and exons - especially in TSS => transposition common in highly expressed genes
Why is transposition more common in promoters and exons than in introns?
TE insertion depends on chromatin accessibility - euchromatin - open chromatin more accessible for transposition => highly expressed genes are more common in new TE inserts
Differentiate between de novo TE insertions vs surviving TE insertions
De novo TEs - new TEs in host that have occurred in the host somatic/germline - common in highly expressed genes
Surviving TEs - TEs that are passed on to offspring (only from germline) - introns - more commonly allowed to be passed on because less risk of damage
Are there specific locations where TEs insert?
Some TEs have specific target sites for insertions - ex short sequence motifs
Usually TEs target low impact regions - reduce their cost of inserting into host - increase TE fitness to be passed on to offspring
Give two examples how TEs have specific target sites and why
- Ty LTR retroelements transpose in yeast upstream of tRNA genes - essential gene but there are many copies - if TE break one copy - little reduction in host fitness - will be passed on to offspring
- R2 non-LTR retrotransposons insert in animal 28s rRNA genes - ribososmal genes essential but there many copies - if TE insertion breaks one gene copy - little reduction in fitness of the host - TE can be passed on
What is the strcuture of Mariner-like TEs?
Mariner-like TEs - “cut & paste” DNA transposons consist of:
- single gene - encodes transposase
- inverted terminal repeats on both ends of the gene
Short direct repeats flanking TE
What is the mechanism of transposition in DNA “cut & paste” TE?
DNA “cut & paste” (P-element, Mariner-like transposons, piggyBac):
- TE expresses transposase
- transposase cuts TE out (with terminal inverted ends)
- OH on TE attack target site - TE inserts
- staggered ends - DNA repair needed
- DNA repair fills in gaps - new DNA = direct repeats
- ligation
Explain Mariner transposition mechainism in detail
Mariner and Mariner-like transposons - DNA “cut & paste” (the mechanism follows the overview of DNA “cut & paste” transposition mechanism):
- TE expresses transposase
- transposase binds to TE - cuts at terminal inverted repeats (leave them out)
- second transposase binds - dimerize - TE sequence forms a loop
- transposases cut again - loop released - bound by 2 transposases
- OH attacks target sites in the genome - TE integrates
- gaps - repaired - regenerate direct repeats
What is the main differences between transposases activity?
Main difference - where transposase cuts TE - for different TE - different transposase
Are all DNA transposons “cut & paste”?
No, not all DNA TE are of “cut & paste” transposition mechanism - cut only one ssDNA - leave other to repair - cut out migrates
What are therecently discovered groups of DNA TEs?
Recently discovered DNA TEs groups:
- helitrons
- polintons
- cryptons
Explain how helitrons transpose
Helitron transposition - very different from Mariner transposition:
- transposase expressed
- transposase cuts only one ssDNA from two
- the left ssDNA gap repaired - dsDNA TE remains
- the excised ssDNA TE circularizes
- rolling circle replication -> multiple TE copies:
1) integrate into new site
2) further replicate
Explain how polintons are thought to transpose
Polintons - larger TEs - encode polymerase, integrase - have particular end sequences
Only model for transposition - not sure yet:
- one ssDNA excised - forms a loop
- replicated into dsDNA
- integrate into genome
What are the subclasses of RNA retrotransposons?
RNA retrotransposons:
- non-LTRs
- LTRs
LTR - long terminal repeat
What are RNA retrotransposons?
RNA retrotransposons - “copy & paste” TEs that retrotranspose through RNA intermediate and using reverse transcriptase
Explain what are non-LTRs
Non-LTRS - RNA retrotransposons that don’t have long terminal repeats (LTRs)
Express mRNA-like product encoding reverse transcriptase
What is the structure of a non-LTR retrotransposon?
Structure of non-LTR retrottransposon:
-1-2 ORFs + TSS
- not long terminal repeats
- could have polyA tail
What is non-LTR mechanism of retrotransposition?
Non-LTR retrotransposition involves transcription, nuclear export, translation, ribonucleoparticle formation (RNP)
- expressed RNA binds at cleaved target site - base pairs with one ssDNA
- OH of cleaved DNA - target site of DNA synthesis using RNA as a template - DNA synthesis
- top strand cleavage:
1) downstream of primery cut: - synthesised DNA bends and base pairs to protruding cut end of ssDNA
- exposed OH - for second DNA strand synthesis using first ssDNA as template
- RNA released - gaps left - repair+ligation - target site duplication
=> new TE insertion
2) upstream of primary cut:
- ssDNA base pairs to protruding end - RNA released - overlapping flaps of original sequence left - degraded - target site deletion
=> new TE insertion
Explain RNA transposon LTR subclass
LTRs - RNA retrotransposons - TEs - have long terminal repeats (LTRs)
What is the strcuture of an LTR?
LTR - RNA retrotransposon - “copy & paste” strtcture:
- 2-3 ORFs
- TSS
- long terminal repeats at both ends (LTR)
- some may have additional codon for envelope proteins
LTRs encode pol gene (integrase, reverse trancriptase, RNAse)
Explain how LTRs retrotranspose
LTR retrotransposition mechanism
What is the currently new proposed update on LTR retrotransposition mechanism?
LTR replication occurrs via circular intermediate
Why do TE have to synthesise their own proteins?
Because host cell doesn’t have necessary proteins for transposition - ex reverse transcriptase (RNA->DNA)
Why is nuclear export and then import needed for TE genes?
TE genes code for proteins not present in host cells - express mRNA - export out of nucleus into cytoplasm - get translated into proteins - have to re-enter the nucleus to act on TE sequences in the genome
Why are TEs and viruses alike?
Features making TEs (retroelements) and viruses alike - same gene at same order:
- capsid protein production
- ability to integrate into the host genome
- specific activation in certain tissues
- high mutability
- existence of virophages propagating only with another virus - similar to non-autonomous transposons - use expression products of autonomous
How TEs and viruses differ?
Features making TEs and viruses different:
- LTR elements - no horizontal transmission
- envelope genes
Did viruses evolve from TE or TEs from retroviruses?
Retroviruses likely evolved from LTR elements (RNA retrotransposones)
However - endogenous retroviruses, polintons - not known which way
What is the classification of TEs
What is Gag gene?
Gag gene encodes Gag proteins - LTR transcripts - act in virus-like particle formation
What is Env gene?
Env gene encodes Env proteins - for envelope formation - LTR retroelement (Gypsy) - cross-species transmission
Gag+Env proteins form virus-like particles
What role do Gag+Env proteins play?
Gag+Env expressed proteins form virus-like particles - formed by Gypsy retroelement - structure very similar to LTR => makes question if LTRs <-> retroviruses evolved one from another
What is the structure of retroviruses, ex Gypsy?
Structure very similar to LTR:
- long terminal repeats
- Gag proteins
+Env gene added
Whate are endogenous retroviruses? Give example
Endogenous retroviruses - TE in genomes that highly resemble retroviruses - encode capsid
Ex: KoRV-A endogenous retrovirus - resembles Koala KoRV virus - can be horizontally transmitted even without infection - KoRV-A can make viral particles
Why are polintons considered a potential virus cross-over?
Because polintons (DNA “cut&paste” TE) are:
- large sequences
- widely distributed in eukaryotes
- have virus-like DNA pol - synthesise own DNA
- many polintons encode capsid proteins
=> polintons - viral origin in genome? (not determined)
Are polintons derived from retroviruses?
Not determined - two possible mechanisms:
1) unknown virus donated capsid - polintons gained capsid proteins (can horizontally transmit)
2) virus integrated into genome - lost capsid function - became polinton
What are autonomous vs non-autonomous elements in genomes?
Autonomous elements: transpose themselves - perform excision + integration proteins
Non-autonomous elements: need help from autonomous element to transpose - needs enzymes => TEs that lost all structures (genes) due to mutation - except inverted terminal repeats - important in transposition to get help from autonomous element protein - ex MITEs
What are SINE elements?
Short interspearsed nuclear elements (SINEs) - non-autonomous - not because lost genes - because gained inverted terminal repeats to be transposed by autonomous elements - ex Alu elements in human genome
What are Alu elements?
Alu elements - SINEs in human genome 10.6%:
- derived from RNA of SRP
- has RNA pol II promoter - transcription initiation
- 3’ end A-rich region - used in transposition to mimic LINEs - use LINE mechanism to transpose
How does the host defend againts TEs? What are the mechanisms?
Immune system against TEs - protect against parasites - several mechanisms:
- KRAB domain zinc fingers
- piRNA pathway
Explain KRAB domain zinc fingers as defence mechanism against TEs
Kruppel-associated box zinc-finger proteins (KRAB-ZFPs) - largest TF family - bind DNA via tandem zinc-fingers - form complexes - recruits repressors - together induce inactive chromatin (accessibility of chromatin important for transposition)
Explain piRNA pathway as defence mechanism against TEs
piRNA - RNA interference supresses TEs in animals:
1) piRNA generating cluster
2) piRNA maturation in somatic cells / Ping-pong amplification in germline
3) TE transcription silencing by mature piRNA
Long transcripts produced from piRNA generating clusters: neighbouring + non-functional TEs in heterochromatin - sequence used in recognising TE -> methylating
What is the piRNA maturation process in somatic cells?
What is the piRNA Ping pong amplification process in germline?
In Drosophila germline:
What is the piRNA induced transcriptional silencing of TEs?
Common in both somatic and germline:
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