Eukaryotic Transcription Flashcards
Eukaryotic RNA polymerases
RNA Poly I
- synthesize pre 45S rRNA in single transcript containing 28S, 18S, and 5.8S rRNA
- functions in nucleolus
- insensitive to alpha amanitin
- uses class I promotor
RNA Poly II
- synthesizes pre-mRNA and snRNA
- functions in nucleoplasm
- strongly inhibited by alpha amanitin
- contains C-terminal called CDT
- can be regulated by phosphorylation of serine residues
- uses class II promotor
RNA Poly III
- synthesizes tRNA and 5.0S rRNA
- functions in nucleoplasm
- sensitive to high concentrations of alpha amanitin
- uses class III Promotors
Eukaryotic Class II Promotor
-Hogness Box or TATA box
-CAAT box
-GC box
contains class II genes that are transcribed by RNA Poly II and synthesizes premRNA and snRNA
Eukaryotic Class I promotor
contains class I genes that are transcribed by RNA poly I and synthesizes pre 45S rRNA in a single transcript that contains 28S, 18S, 5.8S -45S rRNA undergoes process/Modifications
Processing of eukaryotic rRNA
- pre 45S RNA gene are highly transcribed
- RNA poly I transcribes pre 45S rRNA in a single transcript which contains 28S, 18S, 5.8S
- undergoes modifications:
- methylate pre rRNA subunits (28s, 18s, 5.8s)
- unmethylated regions cleaved to form mature rRNA
-Humans contain 200 copies of pre 45S rRNA
Posttranscriptional modification of tRNA
class III genes transcribed by RNA polymerase III synthesize tRNA and 5.0S rRNA and tRNA undergoes modifications
- 5’ end cleaved by RNase P Ribozyme
- 3’ end cleaved by RNase and CCA added to 3’ end by nucleotidyltransferase and this is where amino acids attach
- introns removed containing 14 nucleotides to form anticodon which is a 3 nucleotide sequence that specifies which amino acid attaches to 3’ end
Processing of eukaryotic pre-mRNA
- pre-mRNA=primary Transcripts
- undergoes post transcriptional modifications
Capping-attachment of 7-methylguanisine using 5’ to 5’ triphosphate linkage to attach to 5’ end of mRNA
- involves guanylyltransferase (nuclear enzyme) which attaches guanine
- involves guanine-7-methyltransferase (cytoplasmic enzyme) which methylates guanine
Polyadenylation-attachment of 40 to several hundred adenine nucleotides to 3’ end of mRNA by Poly A polymerase
-pre-mRNA (or primary transcripts) contain a polyadenylation signal sequence (AAUAAA), which is cleaved by specific endonuclease and Poly A polymerase adds the PolyA tail
Splicing-removal of introns
- intron-intervening sequence-noncoding region of mRNA
- exons-coding region of mRNA
Splicing Types
1) TWO types of splicing:
- Spliceosome catalyzed
- Self splicing (Types I and II)
Splice Site: mRNA
mRNA splice site
- consensus sequence of splice site of intron
- GU on 5’ end
- AG on 3’ end
- Branch site “A” (In the middle) located 20-50 nucleotides upstream of the 3’ splice site
Splicing reaction/mechanism
Splicing Reaction: Two transesterification reactions
-Does NOT require Hydrolsis of ATP for E, thus number of phosphate bonds are conserved
FORMED PRODUCT OF LARIAT-3 bonds to adenine
Mech:
- 2’ OH of A in branch site attacks 5’ terminal phosphate of intron forming a 2’ to 5’ phosphodiester bond
- 3’ OH terminus of exon I attacks the phosphodiester bond between the introns and exon II-joing the two exons and FORMING the LARIAT
Spiceosome
- large structure (60s) that splices mRNA
- composed of snRNAs, protein splicing factors, and mRNAs being spliced
snRNA
snRNA (U1, U2, U4, U5, U6)
- involved in splicing
- small ribonucleotides
- highly conserved sequences in ONLY eukaryotes
- form complexes with proteins to form snRNPS (snurps)
class III promotor
class III genes transcribed by RNA polymerase III, synthesizes tRNA and 5.0S rRNA -tRNA undergoes postranscriptional modifications
tRNA
tRNA-transfer RNA
- small single stranded ribonucleotides
- adaptor between RNA and protein
- at least one tRNA per amino acid
- folds into clover leaf or L shape due to intramolecular Hydrogen Bonding between complementary base pairs
- modified bases found in tRNA
Post transcriptional modifications produces 3’ end CCA which amino acids attach to
Also contains a 5’ end
Anticodon:
- 3 nucleotide sequence specifies amino acid attachment to 3’end
- will H-bond to the complementary base seq on mRNA called CODON
mRNA
mRNA-messenger RNA
1)single stranded ribonucleotide
-in a heterogenous population the mRNA size varies by length of gene (protein encoded)
2) NO intramoleculer H-bonds
Translated into protein sequence
- read by ribosome and broken up into codons which anticodon of tRNA bind to during translation
Eukaryotic Initiation step of Transcription
Eukaryotic Transcription factors identify and bind to promotor and guides RNA polymerase into place
- additional transcription factors and enhances may increase rate of initiation
- unwind of dNA
