RNA

History

• mRNA and tRNA worked out in the 1960s• Purification of RNA lead to 23S, 16S, and 4S

species and accounted for 85% of all RNA• tRNA accounted for about 15% of all RNA• Hybridization experiments did not match DNA• mRNA accounts for only 1-4% of RNA

Types of RNA

• mRNA• tRNA• rRNA• snRNA• snoRNA• siRNA• microRNA

Primary RNA

• mRNA (messenger RNA)– Transcribed from DNA– Is then translated into protein

• rRNA (ribosomal RNA)– Necessary for helping ribosomes make protein

(catalysis)• tRNA (transfer RNA)– Mediates amino acid recognition with codons

Other RNA

• snRNA – small nuclear• snoRNA – small nucleolar• microRNA – micro • siRNA – short interferingThese RNA species have a variety of functions and not all of them have been worked out yet.

RNA and Rfam

• Pfam is a database we discussed to determine groups of proteins

• Rfam is a similar database for families of RNA• www.rfam.sanger.ac.uk

tRNA and folding

• tRNA have a unique fold• There are multiple

websites to determine RNA folding

• http://rna.tbi.univie.ac.at/cgi-bin/RNAfold.cgi

Ribosomal RNA

• Has a structural and functional role in ribosomes

• Prokaryotes: 70S which is 30S & 50S, which is 16S, 23S, 5S

• Eukaryotes: 80S which is 45S which is 18S, 28S, and 5.8S

snRNA

• Found in nucleus• Involved with splicing and telomer

maintenance• 5 classes U1, U2, U4, U5, U6

snoRNA

• Modify both rRNA and snRNA• 2 Classes– C/D Box – methylate rRNA– H/ACA Box – convert uridine to peudouridine in

rRNA

miRNA

• Recent discovery• Small approximately 22 nucleotides• Function to down regulate protein expression.• http://www.mirbase.org/search.shtml

RNAi (siRNA)

• Double stranded RNA• Small, artificial• Degradation by RISC and Dicer• Protective mechanism for plants and animals

mRNA

• Coding RNA• Central Dogma “one gene – one protein”– No longer valid– Alternative splicing, regulation, introns, etc.

• Altered expression by:– Region, development, environment, disease, gene

activity

mRNA

• Key research areas– mRNA expression database• There is a DNA database, but is an mRNA database

equally good?• Potential problems with and mRNA database• How to measure mRNA expression

mRNA expression

• Steady-state mRNA (polyA isolation)– Transcriptional control– RNA processing control– RNA export control– RNA surveillance control

est database

• How to study mRNA– Highly unstable, need to make cDNA• Turn mRNA into DNA with reverse transcriptase• Clone into a vector• Sequnce (this is called an express sequence tag EST)

– ESTs can be isolated from specific cells at specific times under specific conditions.

– This can aid in finding when types of proteins MAY be expressed