1. Chapter 14 – RNA molecules and RNA processing

2. Gene organization Francis Crick – 1958
Nucleotide sequence of a gene directly codes for amino acid sequence of polypeptide
Gene contains interruptions of nucleotides that do not code for amino acids
Eukaryotic genes

3. Gene organization
DNA and RNA transcripts within the nucleus are larger than transcripts found in the cytoplasm
Exons are coding regions
Introns are intervening sequences in eukaryotic genes
Occasionally seen in prokaryotes as well
Are spliced out of pre-mRNA/primary transcript before leaving the nucleus

4. Messenger RNA (mRNA) Structure 5′ untranslated region
Bacteria contain Shine-Dalgarno sequence
Serves as ribosome attachment site
Protein coding region
Codes for amino acids
Codon – 3 nucleotide sequence that codes for one amino acid
3′ untranslated region
Aids in stability of molecule

5. Prokaryotic mRNA
Since introns are rare, mRNA can begin to be translated before transcription is complete
Ribosome associates with Shine-Dalgarno sequence, and moves down mRNA molecule in 5′→3′ direction

6. Eukaryotic mRNA
mRNA requires post-transcriptional modifications before exiting nucleus to cytoplasm (site of translation)
5′ cap
Guanine nucleotide added 5′ to 5′
Several nucleotides at the 5′ end are methylated
Stabilizes molecule; aids in initiation of translation

7. Post-transcriptional modifications cont
polyA tail
At 3′ end there is at least one possible cleavage site where nucleotides are removed
After removal, adenine nucleotides are added
Polyadenylation
With associated proteins, stabilizes molecule

8. Post-transcriptional modifications cont
RNA splicing
Splicing out of introns requires 5′ splice site, 3′ splice site, and branch point
Spliceosome
5 snRNPs and several other proteins
Process of splicing
5′ end of intron is cut and folds back on itself to attach to branch point sequence
Forms a lariat
3′ end of intron is cut and intron is released
3′ of exon #1 is ligated to 5′ end of exon #2
Intron reverts to linear form and is degraded

9. Alternative processing
Approx. 30,000 genes in genome; 120,000 polypeptides
A single pre-mRNA molecule can give rise to different mature mRNA – each results in a different polypeptide
Alternative splicing
In addition to intron removal, exons may be removed as well

10. Alternative processing cont
Multiple 3′ cleavage sites
Cleavage may occur at different sites before polyA tail is added
Any exons not included will yield a different polypeptide

11. Transfer RNA (tRNA) 74-95 nucleotides long
Can have chemically modified bases in addition to the normal 4 normally present in RNA

12. tRNA cont
Complementary base pairs form a cloverleaf shape (folds into an “L” 3D)
3′ end is the acceptor arm – where a specific amino acid attaches
Anticodon arm contains 3 nucleotides (anticodon) that recognize codon of mRNA
Initial transcript contains introns that are removed

13. Ribosomal RNA (rRNA)
Original transcript for ribosomal RNA is cleaved by snoRNA
snoRNA + proteins form snoRNPs
Sizes of rRNA measured in Svedberg untis (S) – how fast substances sediment out in a centrifugal field)
Based on molecular weight and structure – not cumulative
Ribosomes
1 or more molecules of rRNA and approx 50 proteins
Complete ribosome consists of 2 subunits – large and small

15. RNA interference (RNAi)
miRNA (microRNA) and siRNA (small interfering RNA)
Both arise from double-stranded RNA, which is cute by enzyme Dicer – fragments are miRNA and siRNA
Small fragments bind to mRNA
miRNA inhibits translation
siRNA – degrades mRNA