Exam 2: Lecture .5

Question 1

DNA Transposable Elements (Structure)

Answer 1

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

Question 2

Transposase

Answer 2

• 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

Question 3

DNA Transposable Elements (Increase Elements)

Answer 3

• 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

Question 4

DNA Transposable Elements (What?)

Answer 4

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

Question 5

Transposons in Disruption of Gene Function in Drosophilia

Answer 5

-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

Question 6

Thomas Morgan Turn of 20th Century

Answer 6

• 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)

Question 7

1930’s and 1940’s

Answer 7

• 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

Question 8

Discovery of Transposable Elements (and Modes of Movement)

Answer 8

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

Question 9

Gerald Rubin

Answer 9

• 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

Question 10

Transposable Elements Today

Answer 10

-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

Question 11

Retrotrasposons

Answer 11

• 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)

Question 12

Reverse Transcriptase (RT)

Answer 12

• can synthesize a DNA strand from a mRNA template.

- normal cellular DNA polymerases cannot do this task.

Question 13

Transcription of retrotransposons

Answer 13

• 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

Question 14

cDNA

Answer 14

• 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

Question 15

RT Does Not…

Answer 15

• bind and copy retrotransposon mRNA transcripts exclusively

- will also attach itself and copy other cellular mRNA transcripts

Question 16

Core Promoter

Answer 16

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

Question 17

Enhancer Elements

Answer 17

• 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.

Question 18

Introns

Answer 18

• 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

Question 19

Intergenic Regions

Answer 19

• DNA sequences found between genes

- contain neither regulatory nor coding sequences

Question 20

Exon Sequences

Answer 20

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