Jumping

s Gene
Mobile & Transposable Elements
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Ethnobotany

mosaic kernels

1938: Marcus Rhoades reported odd phenotypic ratios in corn.

Self pollination of a pigmented corn kernel yielded:

12 : 3 : 1
pigmented : dotted : colorless
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A hypothesis: Two mutations at unlinked loci: 1. pigment gene A1 mutated to colorless mutant a1, and 2. a dominant allele for dotting (Dt) appeared. The presence of the Dt allele caused spots of pigment to appear.

Barbara McClintock 1902-1992

Barbara McClintock 1902-1992

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Transposition = the movement of genetic information from one chromosomal location, the donor site, to another, the target site.
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DNA sequences that can change their genomic location intragenomically either autonomously or non-autonomously are called transposable elements.

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copy-and-paste cut-and-paste

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Retrotransposons can be divided into five orders on the basis of their mechanistic features, sequence organization, and reverse transcriptase phylogeny: LTR retrotransposons, DIRS-like elements, Penelope-like elements, LINEs, and SINEs.

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When a transposable element is inserted into a host genome, a small segment of the host DNA (usually 4-12 bp) is duplicated at the insertion site.
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Transposition = the movement of genetic information from one chromosomal location, the donor site, to another, the target site.
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DNA sequences that can change their genomic location intragenomically either autonomously or non-autonomously are called transposable elements.

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Classifications of Transposable Elements

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Transposition may be replicative or conservative. Replicative transposition will result in two copies of the element, one at the donor site and one at the target site. Following conservative transposition the transposable element will only be found at the target site, with no change in copy number.
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Conservative transposition = cut-and-paste transposition

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Duplicative transposition = copy-and-paste transposition

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DNA-mediated and RNA-mediated transposable elements: 1.Class I transposable elements (retrotransposons). 2.Class II transposable elements (DNA transposons).

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Autonomous and nonautonomous transposable elements Autonomous transposable elements encode all the components of the transposition machinery. Nonautonomous transposable elements appropriate the transposition machinery of autonomous transposable elements.

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Active and fossil transposable elements A transposable element is defined as active if it contains all the necessary sequence elements for either autonomous or nonautonomous transposition. Active elements may be rendered defective by different types of mutation, in which case they are referred to as fossil transposable elements.

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Active and fossil transposable elements A transposable-element family may contain different combinations of active autonomous, active nonautonomous, fossil autonomous, and fossil nonautonomous transposable elements. For example, the human genome contains approximately 50,000 fossil autonomous and 200,000 fossil nonautonomous DNA transposons.

Intriguingly, the human genome seems to contain NO active DNA transposons.

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According to the numbers and kinds of genes they contain, DNA-mediated transposable elements are divided into insertion sequences and transposons.

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Found in Escherichia coli and Shigella dysinteria. Length = 770 nucleotides, including two inverted terminal repeats, 23 bp each.

Contains two out-of-phase reading frames, insA and insB, from which a single protein is produced by translational frameshifting at a run of adenines. The N-terminal is an inhibitor of transposition; the Cterminal is a transposase, an enzyme that catalyzes the insertion of transposable elements into insertion sites.
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Carriers and non-carriers of the insertion sequence can be separated by centrifugation because the carriers are heavier. 33

INSERTION SEQUENCES (IS)

Insertion sequences were first discovered in the gal operon of E. coli.

Galactose (gal) operon

galE galT galK galM

galE = UDP-galactose 4-epimerase galT = galactose-l-phosphate uridylyltransferase galK = galactokinase galM = mutarotase
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Galactose (gal) operon

galE
IS

galT

galK

galM

Insertion of an IS affects only the transcription of the genes downstream from the insertion. For example, if the IS occurs in the galT gene, the galT, galK and galM genes will be disrupted, but galE will not be.

This phenomenon is known as a POLAR mutation.

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Stupidity
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Composite

Hypercomposite transposons contain two or more transposons.

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(symmetrical-inverted) Tn3 from E. coli confers streptomycin resistance. tnpR and bla are transcribed on one strand; tnpA on the other. Tn3 is flanked by 38-bplong inverted repeats.
(asymmetrical) Tn554 from Staphylococcus aureus lacks terminal repeats and contains 8 protein-coding genes. Three of the genes are transcribed as a unit and encode transposases (tnpA, tnpB, and tnpO). The spc and ermA genes confer spectinomycin and erythromycin resistance, 42 respectively.

Composite transposon Tn9 from Escherichia coli contains two copies of IS1 flanking the cat gene, which encodes a chloramphenicol-resistance protein.
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Some mobile elements can transpose themselves in all cells; others are cellspecific. Tc1 elements in the nematode Caenorhabditis elegans and P elements in Drosophila melanogaster are usually mobile only in germ cells.
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Transposition of many elements is regulated by developmental stage. From an evolutionary point of view, the developmental timing of transposition is particularly important, because it affects the propagation of the transposable element to future generations.
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LINE-1 transposable elements in mammals are particularly active during leptotene and zygotene, when DNA-strand breakages occur. This offers an opportunity for transposable elements to insert themselves into new sites.

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Genomic locations of target sites for transposition:

Exclusive genomic preference: In the vast majority
of cases IS4 incorporates itself in the galactosidase operon of Escherichia coli, and thus each bacterium contains mostly one copy of IS4.

Complete randomness: Bacteriophage Mu transposes

itself at random within the genome.

Intermediate genomic preference: 40% of all Tn10

transposons in E. coli are found in the lacZ gene, which constitutes a minute fraction of the host genome.
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Genomic locations of target sites for transposition: Affinity for a particular nucleotide composition: IS1 favors AT-rich sites. Affinity for a particular sequence: IS630 has a
special affinity for 5'CTAG3' sequences.

Chromosomal preference: TRIM elements in

Drosophila miranda exhibit a preference for the Y chromosome.
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Hotspots for P element insertion in the X chromosome of Drosophila melanogaster

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Genomic locations of target sites for transposition:

The DIRS-1 transposable element in the slime mold Dictyostelium discoideum

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Genomic locations of target sites for transposition:

Self affinity: DIRS-1 preferentially inserts itself into
other DIRS-1 sequences. D. discoideum contains, on average, ~40 intact copies of DIRS-1 and ~300 fragments.

oldest

newest

oldest

Active DIRS

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Species specificity:
DIRS in Dictyostelium discoideum only.

mariner moves from species to species, even if the species belong to different taxonomic kingdoms.
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Retroelements are sequences that contain a gene for reverse transcriptase, which catalyzes the synthesis of cDNA from an RNA template.
Not all retroelements possess the intrinsic capability to transpose. Therefore, not all retroelements are transposable elements. Retroelements that transpose do so by retroposition.

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Central Dogma

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Modified Central Dogma

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Not the Central Dogma

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That would have been nice

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That would have been nice

Joanna Masel

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Retroelements Retroelements are DNA or RNA sequences that contain a gene encoding the enzyme reverse transcriptase, which catalyzes the synthesis of DNA from an RNA template. The resulting DNA molecule is called complementary DNA (cDNA). Not all retroelements are transposable or mobile. Retroelements can be divided into three categories: (1) transposable elements that move within a genome by replicative RNA-mediated transposition (but may also move intergenomically) (2) mobile nontransposable elements that only move intergenomically (3) non-mobile elements
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Masayori Inouye Rutgers University

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Retrons are widely distributed among bacterial species. Within each bacterial species retrons tend to be rare. Natural populations of retron-carrying genomes possess a single retron copy, either in the intergenic part of the genome or inside a prophage (a viral genome that had became integrated into the bacterial chromosome).
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multicopy single-stranded DNA (msDNA)

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TERT genes Human telomeres consist of the sequence TTAGGG tandemly repeated many thousand times. Because of asymmetrical DNA replication, a few of these repeats are lost from the tips of the chromosomes each replication cycle.

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TERT genes
Telomerases are nucleoproteins whose function is to add DNAsequence repeats to the 3' end of the DNA strands in the telomeres at the ends of linear eukaryotic chromosomes. The de novo addition of TTAGGG repeats by the enzyme telomerase partially or wholly compensates for telomere shortening. Telomerases in all eukaryotic species share at least two components essential for catalytic activity: a telomerase reverse transcriptase protein (TERT) and a telomerase RNA. The TERT encoding gene is a retroelement. In humans, this retroelement is located on chromosome 5.
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Retroplasmids
Extragenomic DNA and RNA molecules (plasmids) are frequently detected in fungal mitochondria. They can be divided into: - Genomically derived plasmids (similar in sequence to the mitochondrial sequence) - Autonomously-replicating true plasmids that exhibit no sequence similarity with the host mitochondrial genome.

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Group-II introns Group-II introns are a subclass of self-splicing introns. Some group II introns contain protein-coding genes for endonuclease and reverse transcriptase. The latter may act as retrotransposable genetic elements.

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Group-II introns
Group-II introns can integrate into the homologous position of an intronless allele of the same gene (homing), and at much lower frequencies into other sites (retroposition).

Retroposition

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Twintrons are introns-within-introns excised by sequential splicing reactions. Group II twintrons have presumably been formed by the insertion of a group II intron into an existing group II intron.

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Group-III introns are short ORF-less introns found in a small number of protist eukaryotes, such as Euglena gracilis. They appear to be group-II introns from which the central ORF-containing portion has been removed.
Thus, group-III introns are essentially nonautonomous group-II introns. Group III twintrons are known.

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Retrotransposons are transposable elements that use RNAmediated transposition, but do not construct virion particles, i.e., they lack the env (envelope) gene, and so, unlike retroviruses, cannot independently transport themselves across cells.
Initially, the retrotransposons were divided into LTR retrotransposons and non-LTR retrotransposons (or retroposons) according to whether or not their coding sequences were flanked by long terminal repeats (LTRs). Subsequent evolutionary studies indicated that while most nonLTR retrotransposons constitute a monophyletic group, the LTR-retrotransposons are paraphyletic. Some LTR-retrotransposons have secondarily acquired env-like reading frames that may enable them to move from cell to cell (i.e., 72 they are in practice viruses).

LTR retrotransposons

Non-LTR retrotransposons

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Integrase

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Pararetroviruses:
1. HepaDNAviruses, e.g., hepatitis B virus

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Pararetroviruses:
2. Caulimoviruses, e.g., cauliflower mosaic virus

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Pararetroviruses are not transposable elements

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RETROSEQUENCES
Restrosequences (or retrotranscripts) are genomic sequences that have been derived through the reverse transcription of RNA and subsequent integration of the resulting cDNA into the genome.
Retrosequences lack the ability to produce reverse transcriptase, and have been produced through the use of a reverse transcriptase from a retroelement.
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Diagnostic features of mRNA derived retrosequences

1. lack of introns 2. precise boundaries coinciding with the transcribed regions 3. stretches of poly(A) at the 3 end 4. short direct repeats at both ends 5. truncations 6. posttranscriptional modifications
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many
many few
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Retrosequences:
Retrogenes
Processed genes Semiprocessed genes

Retropseudogenes
Processed pseudogenes Semiprocessed pseudogenes
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Number of human retropseudogenes and number of parental functional genes __________________________________________________________________________ Number of Number of Gene genes retropseudogenes __________________________________________________________________________ argininosuccinate synthetase 1 14 -actin 1 ~20 -tubulin 2 15-20 Cu/Zn superoxide dismutase 1 >4 cytochrome c 2 20-30 dihydrofolate reductase 1 ~5 G3PD 1 ~25 lactate dehydrogenase A 1 10 lactate dehydrogenase B 1 3 lactate dehydrogenase C 1 6 laminin 1 >20 nonmuscle tropomyosin 1 >3 nucleophosmin B23 1 7-9 phosphoglycerate kinase 1 2 prohibitin 1 >4 prothymosin 1 >5 ribosomal protein L32 1 ~20 triosephosphate isomerase 1 5-6 93 __________________________________________________________________________

Genes tend to bombard the genome with dead copies of themselves. The Vesuvian paradigm

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How do you get reverse transcribed sequences to become incorporated into the germline genome if the gene itself is not transcribed in the germline?
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Pseudogenes are affected by two evolutionary processes:

Compositional assimilation: The accumulation of mutations which obliterate the similarity between the pseudogene and its functional paralogue. The nucleotide composition of the pseudogene will come to resemble its surroundings, eventually blending into it.

Abridgment: Due to the excess of deletions over insertions, pseudogenes become increasingly shorter compared to the functional gene.
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It takes on average 400 million years for a mammalian retropseudogene to lose half of its length.
Mammals are ~200 million years old and, therefore, the mammalian genome is expected to contain reptilian pseudogenes. These ancient pseudogenes have by now become unrecognizable.

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Mammalian processed pseudogenes are created at a much faster rate than the rate by which they are obliterated by deletion.

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Processed pseudogenes are abundant in mammals.

Processed pseudogenes are rare in amphibians, rarer in birds, and even rarer in Drosophila.

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Creation rates

rates Deletion
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Hypothesis: Retroposition occurs mainly in the female germline Spermatogenesis is similar among animals. Oogenesis in mammals differs from that in the other animals by a prolonged lambrush stage (= suspended animation) that lasts from birth to ovulation (up to 40 years in humans).

Lampbrush chromosome

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Creation rates are determined by the length of the suspended animation during oogenesis

20-40 years in humans.

2-4 months in amphibians. Less than 3 weeks in birds.

Less than 1 day in Drosophila.

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Prediction:
Retrosequences should be found in high numbers on the X chromosome, in intermediate numbers on autosomes, and be rare on the Y chromosome.

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Prediction: Retrosequences should be found in high numbers on the X chromosome, in intermediate numbers on autosomes, and be rare on the Y chromosome.
Density of Processed Pseudogenes in Human Chromosomes (from Bischof et al. 2006)

Mean Density in Autosomes

2.28 0.40

Density in X-chromosome
3.01

Density in Y-chromosome
0.74
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Differences among organisms in numbers of retropseudogenes can also be due to deletion rate.

DNA loss in Drosophila is ~75 times faster than that in mammals.

This high rate may explain the dearth of pseudogenes in Drosophila.

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