RNA Interference

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What does RNA interference do?
Silences gene expression - transcriptionally or post transcriptionally
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Ways small regulatory RNA's silence gene expression
Bindng to mRNA's by complimentary base pairing, causing destruction or bloackage of translation. Or enter the nucleus and bind to gene.
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Sources of long double stranded RNA
RNA viruses. Two promotors close in the genome in opposite orientations. Mobile genetic elements like transposons and reteroelements. self annealing of mRNA
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How is siRNA formed?
Dicer cuts the long double stranded RNA into siRNA duplexes between 21 and 25 nucleotides in length
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What happens after dicer has sliced the long dsRNA
Dicer joins with a Rna Inducing silencing complex (RISC), which includes argonaut and TRBP
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What happens after the RISC has bound?
Argonaut cleaves the passenger strand and this is ejected from the complex
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Next step after the argonaut has ejected the passenger
Argonaut uses the remaining guide RNA to bind to the target mRNA. Argonaut then catalyses the splicing of the target
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What happens to the sliced target mRNA?
Degraded by cellular RNases
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Why does dicer cut fragments to 21-24bp?
Dicer has two RNase domains, separated by 65A, which corresponds to 24bps of dsRNA
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Are all dicers the same in all species?
NO- they have different spacing between RNase domains - species-specific sizes of siRNA and miRNA
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What are primary miRNAS
Molecules produced in the nucleus from endogenous RNA transcripts.
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The structure of primary miRNAs
65-70 nucleotide stem-loop structures.
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What gets cleaved from pri-miRNA and by what
a hairpin loop known as pre-miRNA. By the microprocessor complex that consists of RNaselll - Drosha, and a double stranded RNA binding protein called DGCR8.
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What happens to pre-miRNA then
It's exported to the cytoplasm by the export receptor exoportin-5
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In comes Dicer. What does it do?
cuts the pre-mirna into the miRNA-miRNA* duplex.
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Then what happens to the duplex?
Dicer is joined by RISC which includes TRBP and Argonaute and the miRNA-miRNA* duplex is transferred to Argonaute.
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After argonaut ejects the passenger strand, what does the guide strand do?
bind to the 3' utr of it's target mrna.
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How does miRNA binding prevent translation?
promotion of the removal of polyA tails which leads to mRNA degredation
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The two parts of pri-miRNA that get cleaved. Draw it.
Lower stem, 11bp. Terminal loop. Upperstem remains as mRNA
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Where are most microRNA genes found?
in intergenic regions or anti-sense orientations to genes. But with their own promotor
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What does the presence of similar miRNAs in C.elegans tell us?
use of miRNAs is a common evolutionary strategy
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How many miRNAs in the human genome, what percentage of human genes are regulated by miRNAs
over 1000, 30%
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What is the effect of a mutation involving miRNA
Decreasing or increasing levels of native miRNA, leading to many complex diseases
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Genes frequenly amplified in melanoma
miR-182, miR-15/16.
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Genes frequently deleted in melanoma
Let 7a/b
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What is the first step in chromatin-dependent gene silencing?
Double stranded RNA produced from transcription of opposing primers is cut into siRNA by dicer.
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What happens to this siRNA
It is loaded onto the argonaute containing complex called the RITS - RNA-induced transcriptional silencing complex.
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What does the siRNA-RITS complex do then?
binds to RNAs as soon as they are produced by RNA polymerase II
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What two protiens are recruited and what do they do?
Clr4 and Swi6 - modify the nucleosomes by methylation of the lysine 9 residue of the histone h3.
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Card 2

Front

Ways small regulatory RNA's silence gene expression

Back

Bindng to mRNA's by complimentary base pairing, causing destruction or bloackage of translation. Or enter the nucleus and bind to gene.

Card 3

Front

Sources of long double stranded RNA

Back

Preview of the front of card 3

Card 4

Front

How is siRNA formed?

Back

Preview of the front of card 4

Card 5

Front

What happens after dicer has sliced the long dsRNA

Back

Preview of the front of card 5
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