Chapter 14 – RNA molecules and RNA processing
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
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
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
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
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
Post-transcriptional modifications cont polyA tail At 3′ end there is at least one possible cleavage site where nucleotides are removed After removal, 50-250 adenine nucleotides are added Polyadenylation With associated proteins, stabilizes molecule
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
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
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
Transfer RNA (tRNA) 74-95 nucleotides long Can have chemically modified bases in addition to the normal 4 normally present in RNA
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
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
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