11 - Gene expression in eukaryotes Flashcards
What are the two main differences between prokaryotic and eukaryotic genome organisation?
Prokaryotes have small genomes packaged in circular DNA, where as eukaryotes have large genomes consisting of linear DNA organised into chromatin
Why are eukaryotic genomes usually larger than prokaryotic genomes?
Eukaryotes have a larger number of genes (as they are more complex) and lots of non coding DNA
What is in non-coding DNA?
- gene regulatory sequences (such as promotors)
- introns
- sequences without a known function
2 types of repetitive sequences
- Interspersed repetitive DNA
2. Tandemly repetitive (satellite) DNA
- Interspersed repetitive DNA
- Repeated units scattered throughout the genome
- Single unit 100-10,000 bp
- Copies not necessarily identical, but closely related
- Makes up 25-40% of most mammalian genomes
- e.g. Alu elements (300 bp repeats) make up 5% of the mammalian genome
- Tandemly repetitive (satellite) DNA
- Can be broadly classified according to the length of a single repetitive region
- Much satellite DNA is located at telomeres and centromeres - suggesting a structural role
Chromatin structure
- Chromosomes are composed of chromatin (protein + DNA)
- Chromatin is an intricate form of packaging for DNA (10,000-fold compaction)
Chromatin
- DNA in a cell must be packed in an organized manner – to be accessible for transcription and replication
- This involves association with specific proteins (histones) and the formation of chromatin
- Eukaryotic chromatin is much more complex than prokaryotic chromatin
- Each human chromosome contains a single linear DNA double helix ~6 cm long
Heterochromatin
highly condensed during interphase, not actively transcribed
Euchromatin
less condensed during interphase, able to be transcribed
Histones
proteins with positively charged amino acids that bind to the negatively charged DNA
What is the first part of DNA packing?
Around 200 base pairs of DNA is wrapped around a core of 8 histone molecules to form a nucleosome bead.
Histone 1 sits on the outside and acts as a linker
What happens in the 2nd part of DNA packing (I.e. forming chromatin fibres)
the nucleosome beads are coiled together to create a 30nm chromatin fibre which is looped onto a protein scaffold to form euchromatin
Why does euchromatin have an open/looped structure?
To allow for transcription and replication
What is DNA methylation and what does it do?
The addition of methyl groups to certain DNA bases, which results in the chromatin compacting which reduces transcription and therefore gene expression.
It also accounts for genomic imprinting in mammals
What is histone acetylation and what does it do?
The addition of acetyl groups (-COCH3) to the histone tail which causes the histones to grip DNA less tightly which can in turn activate genes
What process is a form of long-term control of gene expression
cellular differentiation
Closed chromatin
DNA methylated;
histones not acetylated
Open chromatin
DNA unmethylated;
histones acetylated
3 different RNA polymerases
- RNA polymerase I (pol I): ribosomal RNA
- RNA polymerase II (pol II): messenger RNA (mRNA)
- RNA polymerase III (pol III): small RNAs e.g. tRNA
Promoter
DNA sequences adjacent to the gene (‘upstream’) that:
• Determine where the transcription of the gene is initiated
• Determine the rate of transcription
The TATA box
- A key part of the promoter
- Provides the site of initial binding of the transcription initiation machinery
- Located 10-35 bp upstream of the transcription start site
Describe the formation of the preinitiation complex
- TF2D binds to the TATA box
- TF2A and TF2B then bind as well
- TF2F and RNA polymerase 2 binds
- TF2E and TF2H to form the preinitiation complex
How do enhancers work?
When the activator binds to the enhancer sequence, the DNA folds to bring the enhancer and promoter together
