Transposons

A transposon "jumping gene" is a DNA sequence that can change its position within the genome. This may create or reverse mutations as well as latering genome size. 

Two families of transposons;

• RNA transposons - use a copy and paste mechanism which increases number of copies of that DNA sequence in genome. Involves insertion of cDNA created from RNA. 
• DNA transposons - use a cut and paste mechanism which mantains copy number (except if it occurs during replication). Where DNA sections are cut out and inserted somewhere else in genome.

Section of DNA inlcudes genes usually (but not always) a transposase gene. Section is flanked by inverted repeats (a sequence plus its reverse complement) which is where the transposase recognition site is. The transposon also has direct repeats at each end generated from host during transposition process. 

Example of DNA transposon: Ac/Ds in maize 

Ac - "activator" as it is autonomous and has own transposase gene. Ds - "Dissociation" as nonautonomous and need to use transposase enzyme from Ac. Both transposons have same inverted repeats (same transposon family). 

Enzyme is transcribed and translated using host machinery. It then binds to inverted repeats at the end of transposon sequence and cuts the DNA to remove the transposon from its location and creates a break at the target site to allow insertion of the transposon. Some transposons have a preferred target site others insert at random.

Insertion creates gaps due to staggered cutting at insertion site. These gaps are filled by DNA repair enzymes and create direct repeats. These repeats remian after transposon has moved again. 

Used in mutating bacterial genomes and have insertion sequences (transposase flanked by inverted repeats) which work in a similar way to DNA transposons in eukaryotes. Also create short repeats at target sites. 

Also known as retrotransposons or retroposons. Use copy and paste mechanism and "jump" via and RNA intermediate. 

Three main families: LTR elements, LINES and SINES. 

Long Terminal Repeats (LTRs) are important for the reverse transcription mechansims. These elements also contain a pol gene (of viral origin) which encodes proteins required for transposition: reverse transcriptase, RNAse H and intergrase. It may also conatin other viral genes such as gag and env but these arent required for transpostion and arent under selection pressures. The process of transpostion creates directs repeats as with DNA transposons. 

Mechanism: Generation of an RNA and protein products by host mahcinery (exported to cytoplasm normally). Complex mechanism involving reverse transcriptase (creates cDNA from mRNA) and RNAase H (degrades RNA template) create cDNA. Integrase binds to LTRs of cDNA and guides it to nucleus and the DNA inserts into genome. 

Examples: 

Mammals: ERV = endogenous retrovirus. 8% of the human genome but often only LTRs are left as rest is lost following homologous recombination between two LTRs 

Yeast: Ty elements ~35 copies of Ty1 in haploid yeast genome

These are non LTR transposons and are long interspersed elements (LINEs). They can be up to 6kbp but full length ones arent seen very often. There are 3 in the huamn genome (L1,L2, L3) which makes us 21% of the total genome, however only L1 is ever still functional. Have 2 open reading frames ORF1 contains an RNA- binding protein and ORF2 contains a reverse transcriptase and DNA endonuclease. 

Mechansim: 

• Transposon transcribed and translated by host machinery and is polyadenylated
• ORF1 binds LINE RNA and ORF2 binds LINE polyA (in cytoplasm) 
• RNA transported to the nucleus 
• ORF2/polyA complex binds to complementary polyT strand on DNA sequence somewhere in the genome
• Endonuclease activty of ORF2 nicks DNA 
• OFR2 reverse transcriptase activty (primed by poly T region on host DNA but using RNA template) - often this process doesn't reach the end and the transposon is truncated at about 900bp
• OFR2 continues synthesis, now using host DNA as a template  (LINE DNA has inserted into gap created from nicks and this synthesis uses host DNA template creating the direct repeats due to staggered nicks) 
• A second strand of DNA is made by host enzymes 

These are another form of Non LTR transposon known as short interspersed elements (SINEs). They are non-autonomous so require LINEs enzymes to function. They are AT-rich sequences that bind to ORF1 and OFR2. 

Example: Alu element 

common in primate genomes (over 1 mil in human genome) but many truncated. The consensus sequence is 282 bp and the structure itself has two similar halves. Its name comes from the AluI restriction site it contains but thi shas no relevance to its function. 

Different genomes contain different types of transposons e.g Humans have lots of LINE transposons and Maize have lots of LTRs. 

Transpositions only happen once in hundreds of cell divisons as cellular defence mechanisms come into play to reduce them as much as possible. They are only fixed in germline genome too but over evolutionary time they are important in shaping genomes. 

1) Due to crossing over between different transposons in different parts of genomes: 

Exons of different genes taht are both flankes by the same transposon are swapped over when there is recombination in the transposition sequence (2 unrelated genes double crossover) Most of the time its a disaster but sometimes it's functional. 

2) Due to 'mistakes' in transposition

An exon in between two DNA transposons might be excised from the genome and inserted into a new location as it may use one end of each transposon thinking there is only one. 

A LINE transposon might use a polyA signa of a neighbouring gene instead of its own, thus adding the exon onto its normal transcript. 

Three main mechanisms: replication slippage, unequal crossing over (both like in satellite sequences) and retrotransposition of an mRNA. Less than half the genes in multiceullar eukaryotic organisms are solitary genes, duplication is common. 

Unequal crossing over is where two transposons align incorrectly on parental chromosomes (may align with another of the same transposon either upstream or downstream from where it should). This causes one recombinant chromosome to have no copies of the gene and one to have two copies of the gene. 

Orthologues: evolved by speciation. Genes have been evolving seperately since the divergence of the two species. 

Paralogues: evolved by duplication. Genes have been evolving seperately since gene duplication event. 

Tandemly repeated genes: All copies are consevred due to high amounts of the gene needing to be transcribed e.g rRNA gene now has over 350 copies in the human genome. Non transcribed spacer sequences in between genes can be divergent but genes are well consevred. 

Two copies - less selective pressure:

Degradation can occur where an accumulation of mutations occur and the gene loses ability to code for a functional proetin and becomes a pseudogene. 

Neofunctionalization can occur where one copy of the gene gains a new function. 

Subfunctionalization occurs where each copies specialises (e.g protein may have two domains and the two copies are better at the different domains) 

Often duplication is simply just lost. 

Conventional pseudogene: has accumulated mutations which prevent the gene from functioning, e.g frameshifts and point mutations creating stop codons etc. If there is no selection pressure due to the presence of another copy of the gene more mutations occur and the gene loses function. Often in gene duplication one gene function whilst the other becomes a pseudogene. 

Processed pseudogene: generated by reverse trnascription of a functional mRNA and insertion of the cDNA into the genome by LINE proteins. Inserted sequences don't have processing signals such as ribosome binding sites and promoter regions so sequences aren't functional. 

Globin gene family 

Human haemoglobin: transports oxygen in blood and has two alpha family and two beta family chains. Members of the globin family expressed varies throughout prenatal development. 

This family is an example of sub-functionalization. The family contains a number of members generated by gene duplication events which have then evolved slightly different properties. For example fetal haemoglobin needs a higher affinity for oxygen than adult haemoglobin so the mother can pass on oxygen to the developing fetus. 

Reminder of process that occured: Unequal crossing over between two transposons. Chromosome with two beta-globin genes passed onto germline cells. Two copies evolved independently to generate paralogues.