1. Transcription
AHMP 5406

2. Objectives: Describe the general process of DNA transcription
Discuss the function of RNA polymerases
Differentiate between messenger RNA, small nuclear RNA, ribosomal RNA, and transfer RNA
Explain the purpose of the promoter and terminator
Compare and contrast the three types of RNA polymerase in eukaryotic nuclei: RNA polymerase I, RNA polymerase II, and RNA polymerase III
Discuss RNA capping & RNA splicing
Explain the selective exportation of mRNAs from the nucleus
Discuss ribosomal RNA and its function

3. What is Transcription ? DNA does not direct protein synthesis directly
RNA is used as intermediate
Transcription = Copying DNA into RNA

4. RNA Ribonucleic Acids Polymers formed by A, G, C, and U
Contain ribose instead of deoxyribose (DNA)
Polymers formed by A, G, C, and U
U = uracil
Complementary to Thymine
But sometimes binds to G
Single stranded
Can bind to itself and form 3D structures

5. DNA to RNA Transcribed RNA is complementary to 1 DNA strand
Transcribe RNA = transcript
Transcripts can only reach a few thousand bases in length
Unidirectional
5’-3’

6. RNA Types Coding Non-coding Messenger RNA (mRNA) Ribosomal RNA (rRNA)
Transfer RNA (tRNA)
Small nuclear RNA (snRNA)
Small nucleolar RNA (snoRNA)

7. mRNA Codes for proteins
Can be modified by splicing, 5’ and 3’ additions

8. Non-Coding RNAs rRNA tRNA snRNA snoRNA
Form basic structure of ribosome
tRNA
Adaptors b/w mRNA and AA
snRNA
Several nuclear processes
Assist in splicing
snoRNA
Process and chemically modify rRNA

9. DNA sequences used as signals
Sequences signal transcription to start and stop
Promoters – start signals
s factor binds to promoter in bacteria
Terminators – stop signals
In bacteria A-T region causes fold

10. RNA polymerases RNA pols perform transcription
Immediate release of transcript allows many RNA to be synth. in a short time
More error prone than DNA pols
They have limited error correction abilities

11. Types of RNA polymerases
RNA pol I
Transcribe 5.8S, 18S and 28S genes
S = Svedberg units, sedimentation coefficient related to centrifugation
The bigger the number the bigger the molecule
RNA pol II
Transcribe protein-coding genes, snoRNA genes
Some snRNA genes
RNA pol III
Transcribe tRNA genes and some snRNA genes

12. Transcription in Euks.
Requires general transcription factors to initiate transcription
GTFs help position RNA pol II correctly
TFIIA, TFIIB, TFIID, TFIIE, TFIIF, TFIIH
Recognize specific sites on DNA

13. RNA pol start points TATA Box promoter Recognized by subunit of TFIID
TBP – TATA Box Binding protein
approx 25 bp upstream of initiation point
Causes conformational change in DNA

14. Other necessary proteins for initiation of transcription
Transcriptional activators
Attract RNA pol II to start site
Mediators
Allow communication between activator and GTFs
Chromatin remodeling proteins
Modify DNA conformation

15. Elongation Factors Increase affinity of RNA Pol II to DNA
Assist in moving through chromatin structure

16. mRNA modifications 5’ capping Splicing = removing introns
3’ Polyadenylation

17. 5’ Capping When RNA Pol II at 25 bases 5’ cap is added to transcript
Modified guanine
Three enzymes
Phosphatase = removes phosphate from 5’ end
Guanyl transferase = adds GMP
Reversed linkage 5’-5’
Methyltransferase = adds methyl group to guanosine

18. Splicing Removes introns (or sometimes shuffling)
Sequences signal where to cut
Spliceosome performs cutting
Complex of snRNAs (Us) and protein

19. Self Splicing Mechanisms
Group I
Bind free G nucleotide to specific site
Group II
Uses reactive A nucleotide
Unusual mechanisms

20. Poly A tail Cleavage stimulating factor F (CstF)
Cleavage and polyadenylation specificity factor (CPSF)

21. Export of mRNA Goes through nuclear pore complexes
Mature mRNA are modified and protein bound
Signal (passport) for transport of mRNA outside of nucleus
5’ cap proceeds first

22. rRNAs Transcribed by RNA Pol I rRNAs are made from a larger precursor
Sections are modified, cleaved and assembled into ribosomes in nucleolus

23. Subnuclear structures
Sites where snRNA processing machinery is assembled, stored and recycled
Cajal bodies
GEMS (Gemini of coiled-bodies)
Interchromatin granule structures