Euk translation regulation (mRNA) Flashcards
what are the main steps in translation in euk?
initiation
elongation
termination
what is translation?
synthesis of long chains of amino acids from a mRNA transcript using a ribosomes
describe a ribosome
they are large multisubunit complexes (ribonuceloproteins)
what are eIF?
eukaryotic initiation factors are proteins that can promotoe translation and transcription initiation
why is translation initation important?
Rate limiting step of translation so
the bulk of translation regulation occurs at the initiation phase
what are the steps of translation intiation in euk?
ribosome neeeds to be prepared
ribosomal entry point needs to be found
the start codon needs to be found
why is the 5’ m-G cap important in translation intiation
enables the capping binding complex to be formed which helps the ribosome align and dock onto the mRNA properly
how does ribosomal scanning work?
scanning model of initiation occurs so that the start codon can be found and elongation can begin
ribosome uses eiF4A/B helicase activity to unwind the mRNA secondary structure
what is the function of Kozak sequence?
the AUG start codon should best match the Kozak which helps with ribosomal recognition of the AUG
then allows 48S complex to form
before forming the 43 pre-initiation complex (40S+ternary complex), the 40S ribosome binds to other eIFs. Why is this?
binds to eIF1, 1A and 3
> this helps to seperate the 60S and 40S subunits and prevents reassociation of them until after the initiation codon has been found (scanning)
how is the 60S ribosomal subunit recruited
once 48S complex is formed, the eIF sneed to be released via GTP hydrolysis
then 60S can associate with 40S creating 80S initation complex
what 3 main things can influence translation initiation efficiency?
in Euk
5’ cap - if no there then complete absecne of tranlation
RNA hairpins - these riboswitches can pair strongly and block helicase activity
AUG - if strong or weak concensus can affect rate of synthesis
why are pico-viruses cool?
they are single.stranded RNA viruses and do not have a 5’ cap and contain several strong AUG codons
yet they can be translated!
what did picovirus research inform us about
it told us that ‘‘cap-Independent’’ translation initiation is another mechanism of tranlation :0
how does cap- independent translation intiation work?
they use IRES to recruit ribosome internally instead of having a 5’cap
(particualarly useful in pro with multiple ORF)
do Euk use cap-independent mechanism of translation?
yes!
often in times when cap-dependent is insufficient/ global translation is downregulated
e.g during mitosis
name some examples of when cap - independent translation would be used?
oncogene proteins
growth factors
apoptopsis process
(during mitosis)
hypoxia, heat stress, ox stress
(during cell stress)
how did we discover that cap - independent mechanism happened?
Experimental method
use a bicistronic construct using luciferase as a reporter.
> BI do 2 genes in tandem with stop codons
luciferase does not have 5’cap and instead under control of an IRES
what specific processes is cap-independent translation important for in animals?
very important for patterning and development of fly and mice
IRES acts as an extra layer of gene regulation
describe the closed loop model
model suggests that RNA is translated in a non-linear way
3’ interact with 5’ end
as ribosome in close proximity to 5’cap, reinitation is much quicker/efficient and ribosome is recycled
why is the transcriptome bigger than proteome?
there can be many PTMs increasing the diversity of mRNA transcripts produced
but not all become functional proteins
name 3 ways that translation can be globally downregulated in euk?
prevent ternary complex from forming which block the 48S formation
block capping binding complex forming
what is the main way/process that translation downregulation is controlled?
what molecules are used?
phosphorylation by kinases which influences binding and release of different eiF and e-BPs
when would translation downregulation be used?
during cell stress, stress granules appear in cytoplasm
they contain transcriptionally silent / stalled mRNAs allowing cell to respond to stress
what a p-bodies?
cytoplasmic foci of mRNA degradation, always present in cytoplasm
what is eiF2B? what is it role?
this is the guanine exchange factor for eIF2
it promotes GTP-GDP exchange and reactivation of ternary complex
how is eiF2B activity regulated
if eiF2 is phosphorlyated
then 2B can’t bind and activate ternary complex so translation downregulated
why would blocking eif4E downregulated translation in euk?
prevents formation of capping -binding complex which is NECCESSARY FOR translation initiaiton
how does eiF4E regulate translation downregulation?
eiF4E can bind to the extended n-terminal of 4G
but this interaction can be blocked by different eiF4E- BP which also have a concensus 4E binding site
what features do cancer cells have that can promote survival + proliferaiton?
there is hyperactivation of the cap-dependent pathway of translation intiaiton// there is no global downregulation
as they have hyperactivation of GF signalling pathways which results in 4E-BP phosphorylaiton enabling translation to take place
uncontrolled translation NOT downregulated
how can mRNA translation be LOCALLY regulated?
the UTRs are key regions to determine stability, localisation and translation of mRNA
as they are sites where RBP and microRNAs can bind
how do microRNAs influence local translation of mRNA?
they act to degrade mRNA completely or inhibit translation depending on the amount of complementarity they have
using the Risk complex
how can RBPs influcence local translation regulation?
they can recruit ei4E-BP or contain that concensus motif themself
so interact with eif4E directly or indirectly and can block translation happening
how can RBP translation regulation be overcome?
by phosphorlating the mRNA, polyApolymerase and BP can be recruited
and eiF4G can outcompete the RNA-BP allowing translation to happen by forming closed loop model
in times of low iron, how can translation of transferrin receptor be altered?
RNA-BP (IRP) will occupy the 3’UTR of the transferrin receptor mRNA
this stabilises it and allows it to be translated
in times of low iron how would ferritin mRNA be regulated?
ferritin stores iron so we dont want it translated
IRP will sterically block the 5’UTR of ferritin mRNA and prevent 43S complex to be recruited
and so prevents ferritin translation
in times of high iron how would transferrin receptor mRNA be regulated?
TransR will allow iron to enter cell - but don’t want that
IRP (the rna-bp) WONT occupy 3’ UTR of transferrin receptor
mRNA susceptible to degradation by endonuclease/unstable
