class 13 microRNAs

norhtern blots

detect presence and abundance of RNAs

separate RNAs via gel electro.

use labeled complementary DNA to detect RNA you’re interested in base pairing and detecting the probe.

microRNA

turns off mRNAs by binding to them

microRNA biosynthesis

transcribed from genes in the genome by rna pol 2

initial transcript is pri microRNA which forms hairpin loop structure

then while still in the nucleus it is cleaaved by Drosha

now it is called pre microRNA

it can now move to cytoplasm (exportin 5)

in the cytoplasm DICER cleaves off the hairpin loop part, leaving a short double straneded microRNA

argonaute then binds, unwinds the double stranded and gets rid of one leaving only the guide strand

then argonaute and microRNA form RISC

how does RISC inactivate things

leads to cleavage

also blocks translation

microRNA - mRNA base pairing

imperfect usually

GU base pairing allowed

Short seed sequence is the initial anchor for the pairing and is usually an exact match. nucletodies 2-7/8 from the 5 prime end of miRNA.

seed sequence crucial for binding

everything else is chill

GU allowed!

consequences of miRISC binding

degradation - the main outcome

• removal of poly a tail and 5prime cap.

• recruits other proteins to do this

block translation

these are not exclusive, but sequential (first block translation) then over time it is degraded

mir mir* duplex

the product of the second cleavage reaction by dicer. not fully perfectly base paired together. some overhang on either side.

where on the mRNA do miRISCS usually bind

3’ UTR