Transcription Biology Review
Bios 691 – Systems Biology
January 2008
Outline
• Gene structure
• Chromatin structure & modifications
• Transcription apparatus
• Transcription factors and cofactors
• Elongation and termination
• RNA capping, splicing, and adenylation
• RNA processing and miRNA’s
Chromosome Organization
• Mammalian chromosomes tend to fill discrete regions within the nucleus
• An elaborate network of fibrils maintains these arrangements
• RNA ‘factories’ at distinct locations do most of the transcription work
• Nucleoli are factories for rRNA
Chromatin Structure
• Protein scaffolds anchor the DNA
• Within the scaffold there are loops
• Most transcription happens on the loops
• Much chromatin is wrapped in 30nm ‘heterochromatin’
Fine Structure of Chromatin
• Heterochromatin – inaccessible– Bound with many proteins– Centromeres; telomeres; some other areas
• Euchromatin – accessible– Still needs to be opened
Telomeric Heterochromatin and Sirtuins Euchromatin: 30 nm & open
DNA Packaging & Nucleosomes
Gene Structure – Exons & Introns
Exon Size distribution
Gene Structure – Initiation Sites
• Most (~2/3) genes have multiple promoters
• Most promoters are either ‘sharp’:– Very narrow range– Usually TATA + Inr– Often tissue specific
• or ‘broad’:– Typically 70 bp range– Rarely TATA / Inr– Often widespread
Histones and Modifications
DNA contacts histones on their tails Histone tails can be modified
Histones can stay loose or assemble tightly
Proteins Modify Histones
DNA Methylation
Adding a Methyl to Cytosine
Cytosine methylation is passed on to daughter cells
Controlling Transcription
DNA-Binding Proteins• All proteins interact weakly
with DNA• Proteins with projecting
amino acids interact with the DNA major groove
• Hydrogen bonds stabilize position of proteins on DNA
• Proteins that line up several amino acid contacts bind strongly to specific DNA sequences
Transcription Factor Families
• Several structures line up amino acids– Helix-turn-Helix
(Homeodomain)– Helix-loop-helix– Zinc Finger
• Mostly dimers• These families have
proliferated because of their role in attracting transcription apparatus
Cofactors
• Frequently the effect of DNA-binding proteins depends on co-factors
• E.g. ER sits on the DNA but requires estrogen as a co-factor to function
• Myc requires Max as a co-factor to stimulate transcription
• If Max is coupled with Mad instead, the genes are repressed
Kick-starting Pol II & Elongation• Mediator protein
bridges TF proteins and RNA Pol II
• Contains kinase domains – may phosphorylate CTD of RNA Pol II
Initiating Transcription
TBP on a TATA Box
RNA Polymerase II
RNA (red) copied from DNA (blue) by RNA Polymerase II
RNA Polymerase II Structure The cycle of adding nucleotides
Terminating Transcription
RNA Processing
RNA Processing Steps
• Nucleus– capped, – spliced, – cleaved, – polyadenylated
• Exported• Cytoplasm
– stored– translated– degraded
Capping mRNA
The RNA factory
RNA Splicing
Poly-adenylating RNA
•Poly-A Polymerase adds ~100-150 Adenines to 3’ end•After export to cytoplasm, nucleases chop off ~10-20 A’s at a bite•Nucleases compete with ribosomes for mRNA’s•When ~30 A’s left degradation speeds up
RNA Export
• RNA has to be passed through nuclear pores to show up in the cytoplasm (where we measure it)
Micro RNA’s
P-Bodies• Loci where RNA accumulates and is degraded• Have their own structural proteins
Implications for Systems Biology
• Levels of TF’s on a promoter may not predict levels of transcripts
• Rate of transcription may not predict level of mRNA in the cytoplasm
• Levels of mRNA in cytoplasm may not predict levels of protein
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