Lecture 16 & 17 Flashcards
How big is the human genome in diploid cells
6 x 10^9 bp
2 m/cell
How is DNA protected
DNA is organized into histone octamers
Four histone proteins (H2A, H2B, H3, and H4) were found in equimolar amounts and H1 in a half molar amounts
describe the nucleosome structure
Beads on a string
histone octamer is composed of a tetramer of H3-H4 and two dimers of H2A-H2B
DNA (200bp) wrapped around the histone octamer composed of the core histones (H2A, H2B, H3, and H4)
Nucleosomes are linked together by DNA with one bound histone H1
Structure of the core histones
The core histones have long N-terminal histone tails that protrude from the histone octamer
Why are histone tails important
Important for forming higher-order, more compact, forms of chromatin.
Histone tails are subject to many post-translational modifications that affect the compaction level and chromatin which affects its accessibility to enzymes involved in replication and transcription.
Histone tails facilitate inter-nucleosome interactions that result in higher-order packaging/compaction of the DNA
What does the histone H1 do
Binds the nucleosome and organizes chromatin into a zig-zag structure. When H1binds it protect and additional 20-22 bp of DNA and now the DNA enters and exits the nucleosome on the same side
Describe what happens without H1.
Without H1 the nucleosome wraps
147 bp of DNA and the DNA enters and exits on different faces
Histone H1 allows assembly of chromatin into ____?
Histone H1 allows assembly of chromatin into a 30 nm fibre that increases the compaction afforded by the nucleosome by the factor of 35-40 times
what are the two things required for 30 nm fibre formation
H1 histone and N-terminal tails of the core histones
What are the two alternate arrangements of the 30nm fibre
Solenoid model (one-start helix)
Zigzag model (two-start model)
What are SMC complexes
30 nm fibres are arranged in 40-100 kbp loops within chromosome by SMC complexes that affords a much greater degree of compaction than the 30 nm fibre on its own
increases compaction of the fibre
What is gene expression
Series of events where the information in a DNA sequence is converted into an RNA product then to a protein product which performs biological function
Includes 5 things
What is a gene
It is the entire DNA sequence that is necessary for the synthesis of a protein or RNA molecule
This includes: the promoter, coding region, untranslated regions, introns, transcription termination signal
What is the amount of protein in a cell determined by
Abundance of mRNA
Efficiency of translation
Processing and stability of the protein
What is the abundance of mRNA determined by
Rate of its synthesis (transcription of the gene)
The rate of mRNA degradation (mRNA stability)
The availability of the RNA molecule (mRNA sequestration)
What is the purpose of prokaryotic gene control
Primarily to allow cells to optimize growth and division in response to changing environment
What is the purpose of eukaryotic gene control
Primarily to regulate a genetic program that underlies embryonic development and tissue differentiation, but also to respond to changes in the environment
What is a chromosomal scaffold
a proteinaceous scaffold that retains the overall structure of the chromosome after histones have been extracted
Pol, mRNA processing, Compatmentation, regulation, chromatin, intron
Prokaryotic vs eukaryotic gene transcription (6 things)
3 types of RNA polymerases (1 in prokaryotes)
mRNA undergoes significant processing (mRNA not processed in prokaryotes)
Compartmentation of transcription and translation; gene expression is slower! (no compartmentation in prokaryotes)
Regulation is more complex (regulation is much simpler in prokaryotes – polycistronic mRNAs – more than one protein can be translated from a single mRNA)
Presence of introns (no introns in prokaryotes)
DNA is packaged into chromatin (no histones in prokaryotes)
Which RNA polymerases drive the expressions of what classes of genes?
Pol I = pre-rRNA
Pol III = tRNAs and 5S rRNA
Pol II= everything else
What is the most abundant product of transcription
pre-rRNA (80% of all transcription in eukaryotic cells)
What is the pre-rRNA produced by RNAP I processed into
mature 5.8S, 18S and 28S rRNAs
Where the RNAP I transcription occur
In the subcompartment of the nucleus called the nucleolus
What are the two elements of the Pol I promoter
Core sequence and an upstream control element (UCE) located 100-150 bp upstream from the transcription start site
What kind of transcription levels can be maintained using RNAP I
Low basal levels of transcription can occur from the core sequence using RNAP I and selectivity factor 1 (SL1 = TBP + 3 TAFs). More robust transcription requires binding of the upstream binding factor (UBF) to the upstream control element (UCE).
Common features of Pol II core promoters
Pol II promoters share common features of
1) a TATA-box near -30
2) a TFIIB recognition element (BRE) upstream of TATA
3) an initiator sequence near the +1position and
4) downstream promoter element around the +30 position of the gene
Most pol II promoters also require other regulatory sequences called enhancers
What is the most complex RNAP
Pol III
two main types of Pol III promoters
1) for tRNA genes
2) 5S RNA genes
Pol III promoter elements include _____
box A-C elements that reside downstream of the transcription start site (+1) that recruit transcription factors which then recruit TFIIIB and TBP to the promoter elements upstream of +1.
These then recruit RNAP III to the promoter
Bacterial vs Eukaryotic RNAP II
Eukaryotic RNAP II must make mRNA
and miRNAs from a much larger genome than found in bacteria.
Eukaryotic genome is highly packaged in the form of chromatin, so is harder to access.
RNAP II is more complex than the bacterial RNAP.
Nonetheless, RNAP II, overall, is very similar in overall structure and function to the bacterial counterpart.
RBP1 & 2 are structurally similar to b’ and b, while RBP11 & RBP3 are similar to a1 and a2
5 steps of transcription at Pol II promoters
1) The pre-initiation complexis formed and recruits RNAP II to the promoter.
2) A transcription bubble is formed
3) the C-terminal domain (CTD) of RNAP II is phosphorylated and some pre-initiation complex factors are released
4) elongation proceeds as in bacteria
5) transcription is terminated, the RNAP II CTD is dephosphorylated and the cycle is ready to restart
Pol II transcription resembles bacterial transcription
True
What does Pol II transcription require
Requires a large set of transcription factors
Mediator complex
How does the mediator complex function
The mediator complex functions as an intermediary between the general transcription factors bound at the core promoter and more specific transcription factors
Mediator has ~20 subunits in yeast arranged into head, middle and tail domains -/+ a kinase complex that represses expression from a subset of genes. Mediator can facilitate the rate or efficiency of pre-initiation complex formation at the core promoter
How is TBP recruited to genes
Binds TATA-box
only 25% of genes of TATA-box sequences
Other genes must be recruited to TATA-less promoters by TBP-associated factors (TAFs)
