Gene Expression Flashcards
What are isoforms
Why can they form
Why are splice donor and acceptor sequences very frequent.
Different proteins can be made from a single gene.
Alternative splice sites, start sites and polyadenylation sites.
They are only two bases
Optional exons and introns
Optional exons sit between two other exons.
They can either be spliced out with the introns or they can be included in the exon and not cut out.
This will make two possible mRNAs.
Optional introns are sometimes spliced out and sometimes kept in.
Mutually exclusive exons
If you have two optional sites only one will be included in the exon and the other will be spliced out.
The two possible mRNAs made will code for similar proteins.
Internal splice site
Half of one of the exons is optional.
Sometimes it will be included and sometimes it will be spliced out.
What affects the choice of splice site.
Other sequences in the RNA
Secondary structure
RNA binding proteins.
Cryptic splice site
The cell will favour one splice site but it will still use the others.
Dscam gene
Mutually exclusive exon and had 38000 possible variations.
What are the genes that are key regulators of male and female differentiation.
How does the male phenotype form
Sexlethal, transformer, doublesex
The sxl gene is spliced to give a non functional protein and so is tra. Males produce inactive isoforms of tra and sxl.
The dsx gene is spliced to give an active proteins that represses transcription of genes required for female development.
Causing male characteristics to develop.
How do female drosophila characteristics form
Because there are two X chromosomes it allows some functional sxl to be made.
Sxl will bind to its own transcript and will block splicing.
Blocking the specific splicing site will allow more of it’s functional form to be made.
sxl binds to the tra transcript and blocks splicing. This allows the production of the functional form of tra.
Functional tra binds to dsx RNA and stop it’s translation of the male isoform and allow translation of the female isoform.
This will repress male development and cause female characteristics
How does the site of polyadenylation affect the isoform made.
B lymphocytes
B lymphocytes produce two antibody isoforms by altering where the RNA is cleaved and polyadenylated.
One type is membrane bound and the other is diffusible.
This gene has two possible stop codons.
If the first stop codon is used half the intron won’t be transcribed and the acceptor splice site is missing so there is no splicing. This makes a secretory antibody.
If the second stop codon is used the intron will be spliced out and it will make a membrane bound antibody.
What is the optimal sequence to start translation
What is leaky scanning and how does it make many isoforms
What is common about isoforms made in this way.
What will eIF4F do to affect this
Kozak sequence ACCAUGG
The ribosome scans the mRNA with a Met attached seating for AUG.
It will skip some AUG codons if the sequence is not perfect.
Many isoforms can be made depending on which AUG the ribosome starts at.
The isoforms have equal sequences except for their N terminals.
High levels will cause the ribosome to favour the first AUG.
Basically how does HIV replicate
What type of mRNA can leave the nucleus and why is this a problem for HIV.
How do the overcome it
It enters the cell as RNA, which is reverse transcribed into DNA. which is integrated into the host genome.
This will then be transcribed into RNA and spliced. Only spliced RNA can leave the nucleus to be translated.
HIV needs the full length RNA to make new virons so it can’t do this if it’s been spliced. But it also can’t leave the nucleus without splicing.
The make Rev to bind to the introns and protect them from splicing and help them out of the nucleus.
Rev levels can be used to estimate how bad an infection is.
Untranslated regions of mRNA
Between the cap and the start codon there is an untranslated region of mRNA. there is another between the stop codon and polyA tail.
Stem loop structure often form here which can be detected by proteins.
A binding protein mat only be found in one half of the cell. So when the mRNA enters the cytoplasm it will bind and only be translated if it went to that half.
This gives rise to localised translation. Where all proteins made from the mRNA will be found on one half of the cell
where are Translational control elements on mRNA
Fe regulation example
In their untranslated regions.
Ferritin is a protein that stores Fe and reduces the amount in the cell.
Transferrin imports Fe into the cell and increases the amount in the cell.
If there is low Fe in the cell:
Aconitase binds to stem loops in ferritin 5’ RNA and blocks its translation to stop Fe storage.
Aconitase binds to stem loops on transferrin 3’ RNA and blocks it’s degradation so more Fe can enter the cell.
If there is high Fe in the cell:
Aconitase binds to the Fe and has a conformational change so it can’t bind to mRNA. so it can’t block ferritin translation.
Global/ non specific translation regulation and why it is useful
What a cell does with EIF2 to stop translation.
This controls all translation and is useful for when a cell is virally infected and doesn’t want to translate the virion RNA.
Or useful for when cells are low on energy.
EIF2 binds to Met to allow scanning for the first start codon.
EIF2 has to be bound to GTP to be active.
eiF2B binds and knocks off GDP to allow GTP to bind.
If a cell wants to stop translation it will phosphorylate EIF2 and cause it to bind to eiF2B very tightly and this will block its function and slow down translation.
