L10 RNA degradation in Ribosomes

why do we need RNA degradation

• damaged RNA needs to be removed

• incorrectly transcribed/processed RNA

• control gene expression- cell can switch pattern of genes being expressed

what’s the range of RNA half-life

minutes to days

where is casein mRNA expressed

mammary gland

how much does casein mRNA increase on stimulation by prolactin

~70fold

• half-life increases dramatically

what is prolactin

hormone promoting milk production

casein transcription only increases by how much on stimulation by prolactin

~2fold

why does casein mRNA increase on stimulation by prolactin

increase in half-life increases the length of the poly-(A) tail

• 3’UTR of RNA binds proteins which aid the stabilisation of the poly(A)tail

• more stabilisation = more casein made

circular mRNA during translation monitors mRNA integrity…

it will not be circular if it has lost cap or poly(A)

circular mRNA during translation brings ribosomes ending translation close to the…

AUG

• ribosomes my ‘recycle’ onto the 5’ end

the closed loop must be… before exonucleases can gain access

broken

• (either getting rid of the cap or the poly(A)tail)

what are exonucleases

main enzymes involved in RNA turnover

what’s deanylation dependent decay

CCR4-NOT complex or PARN enzyme

• trim gradually the poly(A)tail in any RNA -wont form a circular RNA

• leads to further decay in the 3’ end from the exosome (3’ to 5’ ribonuclease)

• 5’ cap is now more susceptible to decapping from the mRNA driven by DCP1 and 2- RNA susceptible to 5’ to 3’ exonucleases (XRN1)

what does the exosome do

decay the 3’ end (3’ to 5’ ribonuclease)

what decap’s the 5’ cap from mRNA

DCP1 and DCP2

what 5’ to 3’ exonuclease is decapped RNA susceptible to

XRN1

what does DCPS do

also removes the cap

what are the endonucleases

• argonaute

• Swt1

• Smg6

what are the deadenylases

Ccr/NOT complex

what are the decapping enzymes

• DCP1

• DCP2

describe the exosome

• 3’ to 5’ exonuclease in the cell

• multisubunit complex

what is the exosome involved in

RNA turnover and processing

what are the multiple nuclease activities of the exosome

• RRP6

• RRP44

where is RRP6

the nucleus

where is RRP44

cytoplasm

what is XNR1

5’ to 3’ exonuclease

what is XNR1 involved in

• RNA turnover and processing

• transcriptional termination

what does XRN1 function after

decapping of the mRNA

what process is deadenylation-dependent decay

constitutive

• happens all the time

what does deadenylation-dependent decay not do in all mRNAs

take place at the same rate

what do AU-rich elements or ARE’s do

bind specific proteins that stimulate the deadenylase complex to initiate decay

where are AU-rich elements (AREs) found

3’ untranslated regions of mRNAs

what is nonsense mediated decay (NMD)

mistakes in the RNA are detected, RNA is targeted for degradation

what do Premature Stop Codons (PTCs) result from errors in

• transcription

• splicing

• editing

• polyadenylation

• mutations

1/3 of all inherited disorders are caused by…

nonsense or frameshift mutations that introduce PTCs (premature stop codons)

nearly all properly spliced mRNAs have what in the final exon

stop codon

what happens in the first round of translation during NMD (nonsense mediated decay)

EJCs are removed from the mRNA by the ribosome

when ribosomes reach the PTC, an EJC remains downstream, what specific factors that are part of the EJC or are recruited to it, interact with the RNA degradation machinery

UPF proteins

when ribosomes reach the PTC, what remains downstream in nonsense mediated decay

exon junction complexes (EJCs)

what is the nonsense mediated decay process known as

surveillance

what is RNAi

RNA interference

what is siRNA

small inhibitory RNA

what is miRNA

micro RNA

what is RISC

RNA induced silencing complex

how many nucleotides long is siRNA

21-23 nucleotide RNAs

how complementary is siRNA to target RNA

perfect

what is the function of siRNA

viral defense mechanism

what does siRNA lead to the degradation of

the target RNA

how many nucleotides long is miRNA

21-23 nucleotide RNAs

how complementary is miRNA to target RNA

imperfect

what is the function of miRNA

key gene regulatory mechanism in the cell

what does miRNA lead to

a block in translation

what protein cleaves mRNA

Argonaute protein

describe translational repression

inhibition of initiation, also leads to RNA degradation

during embryonic development, how does 3’ UTR length change

gets longer

mRNAs in proliferating cells tend to have…3’ UTRs

shorter

what is 3’ UTR length change driven by

use of alternate polyadenylation sites

what do longer 3’ UTRs have more possibility of

binding sites for miRNAs (and proteins)

siRNA use in research

take experiments from looking at one gene at a time to looking at families of proteins

what are the clinical uses of siRNAs

6 approved

• many more in trial

• summarises the currently prescribable RNAs targeted at particular diseases

• Huntington’s Gene Therapy

what does the (viral) gene therapy vector encode in Huntington’s Gene Therapy

a microRNA targeting the Huntingtin mRNA

what does a microRNA targeting the Huntingtin mRNA lead to

a less mutant Huntingtin protein being synthesied

how does Huntington’s gene therapy work?

• Huntington’s mutation leads to toxic proteins in brain cells

• gene therapy infused into the brain

• brain cells become their own drug factory

• lowers levels of toxic proteins in the brain

how long are short RNAs

21-23 nts long

where does the RISC complex assemble

miRNAs and siRNAs

what is the RISC complex

nuclease involved in blocking translation