L12 Eukaryotic Transcription Flashcards
Eukaryotic Cell
- has a nuclear membrane
- poses a problem during transcription and translation as mRNA needs to be transported from the nucleus
Genome size - paradox
More DNA doesn’t necessarily mean more genes
Size of genome doesn’t reflect number of genes you have
DNA during mitosis
See OneNote image
- highly compact DNA and protein, not easily transcribed
DNA during interphase
See OneNote image
- strands of chromatin
- DNA state where expression occurs
Chromatin
- composed of DNA and proteins: histones and non-histone proteins
Nuclear structure
See OneNote diagram
- nucleolus = where ribosomal RNA is made
DNA is not free flowing in nucleus, there are specific domains e.g. peripheral heterochromatin
Location in the nucleus and its correlation to expression/no expression
Nuclear localisation
See OneNote diagram
Expressed genes tends to be towards the middle
Non-expressed genes tends to be near the periphery
Nuclear localisation is not absolute
Localisation happens naturally for some genes or endogenous mechanism that control their location
E.g.
if in repressive domain: tether genes to periphery => not expressed
if in activating domain: regardless of where it is located, always expressed
Exome
Exome = all coding material in that chromosome, translated into proteins
Genome size
Factors influencing genome size:
- gene density (intergenic regions)
- introns (size and number)
- repeats
RNA polymerases
- DNA dependent RNA polymerase
- 3 types in eukaryotes
- core components are homologous with prokaryotic RNA pol except for the sigma factors
Genes transcribed by RNA pol 1,2,3
See OneNote table
Types of RNA
See OneNote table
Prokaryotic vs Eukaryotic genes
See OneNote diagram
Prokaryotic mRNA:
Polycistronic, codes for more than one protein
Operons that are co-regulated by 1 TF
Eukaryotic mRNA:
Encodes 1 protein
Protein has modified 5’ end and modified 3’ end
Intron splicing
Eukaryotic mRNA processing
5’ capping
RNA splicing
3’ polyadenylation
Export
Translation
Primary RNA transcript
See OneNote diagram
Encodes protein that does a generalised function?
After it mRNA has been translated, has slightly different domains and functions in different cells
Gene Organisation - Drosophila Adh region
See OneNote diagram
- osp (outspread wings) mutations mapped to both side of Adh
- Adh, Adhr, two other genes within introns of osp
Osp gene sits in the intron of the Adh genes
Can have entire genes sitting inside introns
Adh region has multiple start sites, polyA sites and differential splicing
RNA polymerase - conserved subunits of bacterial and eukaryotic RNA polymerases
See OneNote table
Transcription
- RNA pol 2 transcribes protein coding genes (mRNA)
Steps in transcription at Class 2 (RNA pol 2) promoters
- formation of pre-initiation complex
- promoter melting: separation of 2 DNA strands
- initiation: formation of the first phosphodiester bond in the nascent (freshly generated) mRNA
- promoter clearance
- elongation
- termination
Transcription initiation
See OneNote diagram
Inr = transcription initiation point
- which promoter present is dependent on the gene
- each promoter contains a subset of these elements e.g. TATA and Inr
Some genes lack apparent core promoter elements
- e.g. lack TATA, Inr, DPE
- these genes often transcribed at low rates
- are housekeeping genes
- have multiple transcription start points, not as specific (facilitate regulator gene expression, not as specific)
- contain CG-rich sequences (CpG islands)
Promoter Recognition
General transcription factor TF2D:
- TATA binding protein (TBP)
- TBP associated factors (TAFs)
- binds core promoter elements and initiates assembly of pre-initiation complex (TF2D, TF and RNA pol 2) on the promoter
Has motif that recognises TATA sequence
TAF2 recognises Inr sequences
Different components of TF2D recognises different sequences in the promoter => any combination of the interaction can lead to transcription initiation
Pre-initiation complex
TF2D, TF and RNA pol 2
TF2D
See OneNote diagram
- can be recruited by different promoter elements
- for each promoter, the”initiating” interaction between the promoter and TF2D may differ depending on the specific sequence of the elements
Transcription initiation - additional GTF
See OneNote diagram
GTF = general transcription factors
- in addition to TF2D, other GTFs required
