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DNA Transposable Elements (Structure)

-2 key features
-inverted repeat sequences at terminal ends of element
-gene that encodes an enzyme called transposase



-can bind to terminal repeats
-can excise entire element fromgenome and insert it into a new location (called precise excision)
-does not involve adjacent DNA
-sometimes will take some flanking DNA with it (imprecise excision) and can cause a mutation or can lead to duplication


DNA Transposable Elements (Increase Elements)

-can do this by moving out of hemi-methylated DNA (newly replicated) into dually methylated DNA (yet to be replicated).
-Cell will repair gap left during excision using sister-chromatid as template. Number of transposable elements can be doubled this way


DNA Transposable Elements (What?)

-short pieces of DNA that are able to move around the genome


Transposons in Disruption of Gene Function in Drosophilia

-transposable elements insert themselves randomly in the genome
-if an element drops into and disrupts a coding sequence (exon) then the translated protein will be defective
, if the element lands within and disrupts and enhancer element then the binding of developmentally regulated transcription factors could be inhibited
-last two scenarios transcription of the affected gene is inhibited while insertion of transposons into coding exons results in blocks in translation


Thomas Morgan Turn of 20th Century

-starting to screen adult fruit flies for mutants
-mutagens did not exist and so the mutants that were found were the results of spontaneous mutations (ie replication errors and degradation of nucleotides)


1930's and 1940's

-chemical mutagenesis and radiation were being used to generate single base changes and deletions respectively
-chemicals and radiation generated large numbers of mutants it was difficult and time-consuming to map the mutation to a particular position on the chromosome


Discovery of Transposable Elements (and Modes of Movement)

-provided an opportunity to use this element as a biological mutagen


Gerald Rubin

-determined sequence of P-element
-molecular methods could be employed to determine the location of the P-element in the genome
-once an inserted P-element had been located the flanking genomic regions could be sequenced
-helped reveal identity of disrupted genes


Transposable Elements Today

-used used to generate mutations in a wide range of organisms including but not limited to bacteria, yeast, fruit flies, mice and several species of plants



-structurally similar to DNA transposons: the ends of the element contain repeated DNA sequences and the body of the element contains a gene that codes for a transposase/integrase enzyme
-contain a second gene that codes for a special polymerase called Reverse Transcriptase (RT)


Reverse Transcriptase (RT)

-can synthesize a DNA strand from a mRNA template.
-normal cellular DNA polymerases cannot do this task.


Transcription of retrotransposons

-done by normal cellular machinery thereby generating mRNA (100's if not 1000's produced) templates that can be translated by host to produce RT and transposase/integrase enzymes
-RT copies the mRNA transcripts into single DNA strands which in turn are replicated by cellular DNA polymerase
-complementary DNA strands are zipped shut to form hundreds/thousands of double-stranded DNA segments called cDNA (copy)
-transposase/integrase enzyme is then capable of binding to the inverted repeats of the cDNA retrotransposon and inserting it into the genome
-leads to a profound increase in the number of transposable elements within the genome



-looks similar to the genomic version of the retrotransposon with the exception that it lacks all intronic sequences
-creation and insertion of cellular cDNAs into the genome is one basis for pseudogene formation
-lack core promoters and enhancer elements
-cannot be transcribed and instead simply sit idle within the genome
-accumulate random mutations over time and contribute to the “junk” DNA that litters the genome


RT Does Not...

-bind and copy retrotransposon mRNA transcripts exclusively
-will also attach itself and copy other cellular mRNA transcripts


Core Promoter

-site where RNA polymerase and general transcription factors bind and initiate transcription


Enhancer Elements

-sites where transcriptional activators and repressors bind and enforce temporal and spatial control of transcription
-in general eukaryotic genes will contain multiple enhancer elements
-some can control expression in multiple tissues while in some cases multiple enhancers can direct expression to single tissue
-can be found upstream of transcriptional start site (TSS), within intronic sequences and even downstream of the 3' untranslated region.
-many located within 5kb of core promoter, some can function at significant distances away from TSS.



-non-coding sequences that physically separate exons
-numbers vary from gene to gene
-some genes contain dozens and others are completely lacking
-vary in size
-many examples of genes in which total combined size of all introns can be larger than that of coding exons


Intergenic Regions

-DNA sequences found between genes
-contain neither regulatory nor coding sequences


Exon Sequences

-only parts of gene that will be both transcribed int mRNA and then translated into protein