Transcript Messenger RNA

Messenger RNA
Todd Metzger
BMS 265
May 8, 2009
Nomenclature
• Messenger RNAs (mRNA) describes
cytoplasmic product
• Pre-mRNA (part of hnRNA) in nucleus is
processed and exported
• Describe by gene/protein, including isoform
– or NCBI sequence (NM, natural; XM, in silico)
– New methods for describing spliced mRNAs
(Sammeth 2008)
1955 - Goldstein and Plaut
6 min
62 hr
Discovery
• Volkin and Astrachan discovered a new type of
RNA synthesized following infection of
bacteria by T2 Bacteriophage (Volkin 1956)
– Newly synthesized RNA mirrored composition of
T2 DNA
• Brenner, Jacob, and Meselson found that only
RNA, and not ribosomes, were newly
synthesized following infection (1961)
– The term messenger RNA was coined
Translation Leads to Linear Protein
Synthesis
• Dintzis 1961
• Reticulocytes analyzed
– Make ~90% hemoglobin
• Radiolabeled amino acids incorporated to Cterminus only
1970 - Messenger RNA Visualized
Miller et al
Biogenesis
• Enzymatic Reaction by RNA Polymerase
– RNA Pol II in eukaryotes
• Involves Initiation Factors (TFIIB,D,E,F,H) and
Mediator Complex
• Core RNA polymerase II is 12 subunits, 500
kDA, highly conserved
– 10 subunit catalytic core
Woychik 2002, Sims III 2004
Yeast RNA pol II
Catalytic core
Positively charged cleft
Active site occurs between the wall and clamp (shown closed - elongation)
Transcription
• Template DNA, 5 GTFs, and RNA pol II
associated to form pre-initiation complex
– TFIID contains the TATA-box binding protein (TBP)
– TFIIE and -H melt DNA
• CTD phosphorylation is required for
dissociation of initiation factors and entrance
to the elongation phase
• TFIIS allows stalled transcription to continue
– Reverse procession followed by cleaved of RNA
TFIIS
Mediator
• Connects transcription factors to basal
machinery
Casamassimi 2007
mRNA Secondary Structure
• Many mRNAs can code for the same protein
• Made program to scramble codons while
preserving base content
• Looked at 50 bp structures
Katz 2003
mRNA Secondary Structure
- No significant differences for human sequences
mRNA Vaccinations - Direct
Hess et al 2006
mRNA Vaccinations - Indirect
Hess et al 2006