RNA

RNA

1. tRNA

   • Also called Adaptor molecules: act as adapters for the translation of the sequence of nucleotides of the mRNA in to specific amino acids    Nucleotide length: 74-95  Contain a variety of rare bases in addition to C, G, A and U some of these are modified by methylation   There are at least 20 species of t RNA one corresponding to each of the 20 amino acids required for protein synthesis   
   • reads message of nucleic acids and translates these into proteins or amino acids. 
   • tRNA recognsies and binds to its corresponding codon in ribosme and transfers it to approrpriate amino acid to end of aa chain. these then continue to decode mRNA molecule until entire sequence translated into a protein.
   • AKA adaptor: Act as adapters for translation of sequence nucelotides of mRNA in to specific AA
   • 74-94 nucleotides long
     • contain C G A U
       • some modified by methylation
         • METHYLATION
   • at least 20 species
   • secondary structure
     • acceptor arm
       • 3' end (7bp)
       • 3' end, unpaired CCA
       • 3'OH group terminal of Adenine binds with Carboxyl group of AA
       • tRNA+AA = amino acyl tRNA
     • anticodon arm
       • recognises triplet codon in mRNA
       • complementary to base sequence of mRNA codon
       • binds specificially to mRNA by hydrogen bonds
     • DHU arm
       • recog site for aa tRNA synthase
         • adds aa to acceptor arm
           • ends in loop
     • TΨC arm
       • binding of tRNA to ribosomes
         • 4-5 bp stem containing sequence pseudo uridine
     • Extra Arm
       • 75% of tRNA posses sjprt extra arm
   • Tertiary Structure
     • Xray diffraction analysis: Ls shaped formed by further folding of clover leaf due to H bonds between T and D arms
     • 2 double helical columns continuous + perp to one another

2. rRNA

   • molecule in cells that forms part of the protein synthesising organelle called a ribosome. this is exported into cytoplasm and helps trasnlate infor in mRNA into protein. 
   • synthesised in nucleolus the proteins are synthesied in the cytoplasm and are transported to the nucelus for subassembly. 
   • secondary structure play active role in recognising conserved portions of mRNa and tRNA. 
   • primary constituent
   • protein manufacturing
   • mbine to form nuceloprotein called ribosome
   • majority of RNA found in eukaryotic cell
   • A site
   • P site
   • E site
   • performs peptodyl transferase activity and referred to as enzyme
     • incoming aminoacyl tRNA to A site, anticodon pairs with mRNa codon
       • peptide bonds formed
         • translocation: ribosome moves down mRNA and tRNA moves to P site

3. ncRNA

   • any that functions without being translated into a protein
     • produces RNA gene or non coding RNA

4. snRNA

   • involved in mRNA and gene regulation
     • involved in intron removal and processing of hnRNA into mRNA
     • U7 produced correct 3' ends of histone mRNA

5. RNA interference

   • conserved biological response to double stranded RNA that mediates resistance to both endogenous parasitic and exogneous. 
   • regulates expression of protein coding genes. Natural mechanism for gene silencing and has potential to revolutionise experimental bio. 
   • potential applications in genomics, therapeutic intervention and agriculture
   • inhibits gene expression at stage of translation
   • inhibits transcription of specific genes
   • aka co-suppression
   • targets RNA from viruses and transposons
   • WHY DO WE NEED INTERFERENCE?
     • Defense Mechanism
       • defends against infection by viruses
       • as defense mechanism to protect against transposons and insertional elements
     • genome wide regulation
       • RNAi plays role in regulating development and genome maintenance
   • Players
     • RNA
       • SiRNA: dsRNA
         • Small interfering RNAs that have an integral role in the phenomenon of RNA interference (RNAi), a form of post-transcriptional gene silencing 
         • most commononly used RNA interference tool for short-term silencing of a protein 
         • targets specific mRNA for degradation and is usually limited to cells that are emenable to tranfection of syntheic oligonucleotides
         • derived from longer double stranded RNAs which are either dleivered into the cell itself or delivered experimentally, intro is widely used to manip gene expression. 
         • often perfect complementarity to target sites, Argonaute ckeaves. these also guide RISC to mRNA
         • post transcriptional gene silencing
     • RISC
     • Enzymes

6. Micro RNAs

   • over 1000 miRNA produced from the human genome, these are transcribed by RNA polymerase, from indepedant genes or introns of protein coding genes 
   • microRNAs are repsonsible for gene regulation and they are a complementary to a part of one or more mRNAS to a site in 3' UTR 
   • non-coding  RNAs that act at post-transcriptional level they are transcribed by Drosha
   • single miRNa can regulate whole set of different mRNAs 
   • come from RNa transcribed in nucleus which fold and are processed before being transported to cytoplasm as 2x stranded precur miRNA, these bind to DICER which cuts its short degments (21 nucl long) these bind argo protein, one strand atsyas bound and gides, this combo is called RISC, these are directed to restirct specific RNA. 
   • 21-25bp long
   • 1. RNA Pol I/III in nucleus helps gene or intron transcribe to become pri-miRNA
     • 2. Drosha and DGCR8 cleave pri-miRNA and this becomes pre-microRNA
       • 3. now in cytoplasm and cleaved by dicer and TRBP which sees one strand degraded
         • 4. Ago2 is present and RISC formed, mature miRNA
           • mRNA target cleaved
           • Translational repression
           • mRNA deadenylation
   • DROSHA
     • core nuclease that executes the initiation step of microRNA (miRNA) processing in the nucleus.  
     • Pri-MiRNA to pre-miRNA
       • Drosha cleaves primary miRNAs to release hairpin shaped pre-miRNAs 
   • DICER role
     • after being cleaved by DROSHA, dicer cuts pre-miRNA to make mature miRNA
   • Mirtron Pathway
     • hairpins: defined by splicing machinery and they yield pre-miRNA 
     • alternate source of miRNA regulatory RNAs
   • Functions of miRNA
     • tumour supressor
       • annotated human miRNas map within fragile sites of chromosomes and these are areas of genome that are associated with cancers
   • uses include:
     • biomarkers
       • biomarker use as extremely stable and and currently being used in the form of panels to  help physicians determine the origins of cancer in disseminated tumors
       • challenges in delivery however

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