Dr Hack's Lectures 2 Flashcards
What controls cAMP concentration?
glucose
what does cAMP bind to?
catabolite activator protein (CAP)
why is the lac promoter weak?
sequences at -10 and -35 are non-standard
cAMP-CAP complex binds to DNA next to which region? how does it affect DNA?
binds next to -35 region, bends DNA by 90 degrees
in what scenario is the lac operon transcribed?
CAP-cAMP binds to DNA
Lac repressor does not bind to DNA
What do catabolic operons contain genes for?
breakdown of metabolites
when are biosynthetic operons turned on?
when supply of end product is low
what does the trp operon contain genes for?
The biosynthesis of Tryptophan (an amino acid)
When does tryptophan repressor protein repress transcription of trp operon?
When tryptophan conc. in cell is HIGH
How does tryptophan stop transcription of trp operon?
tryptophan binds to repressor, which in turn binds to trp operator, blocking transcription
Is the trp operon the same as the lac operon?
No, tryptophan has an opposite effect on the operon than allolactose does on the lac repressor
What bacteria does Euprymna scolopes have a symbiotic relationship with?
Vibrio fischeri
When does bioluminescence occur?
when the bacteria are at high density
How is density measured and under which operon?
Quorum sensing, the lux operon
What signals cell density?
Acyl homoserine lactones (AHL)
what protein catalyses synthesis of AHL?
LuxI
How do bacteria use AHL?
They take up AHL from the cytosol
They produce AHL
What does AHL bind to and when?
When there is enough protein in the cell, AHL binds to LuxR protein
What does LuxR do?
With AHL bound to it, LuxR activates transcription of bioluminescence genes in the lux operon by binding to lux box in DNA
Where is the lux box in DNA?
next to lux promoter P
What kind of feedback does quorum sensing produce?
positive feedback - the system increases its own activity
What are other examples of quorum sensing?
‘Milky seas’
Biofilm formation
Pathogenic bacteria
Define genomics
the study of genome organisation and the identification of genes and their functions
Of the examples given in the lecture, which organism has the smallest genome size and which has the largest? in order (small –> large)
Yeast > Plant > Nematode > Fruit fly > Pufferfish > Chicken > Human
Where does the extra DNA come from within genes?
Introns
Where does the extra DNA come from between genes?
repetitive DNA
How are introns removed?
RNA Splicing
How is a lariat formed?
5’ cut, formation of loop with tail
3’ cut, joining of exons
What is intron excision catalysed by?
Spliceosomes
What do spliceosomes contain?
small, nuclear ribonucleoprotein complexes (snRNPs)
Where do snRNPs bind?
primary transcript
What is the lariat loop bond?
5’-2’ phosphodiester bond
what does alpha tropomyosin do?
helps regulate muscle contraction
stabilises actin filaments in cytoplasm
Introns can contribute to variation in gene size, give an example of two species with the same gene but in different sizes
humans and pufferfish - both have huntingtin gene but in Fugu it is much smaller
What is repetitive DNA?
DNA that occurs in many copies in the genome
How much of the human genome is made up of repetitive DNA
> 50%
What are simple sequence repeats?
tandem arrays of very short sequences, eg. CACACACACACA
How much of the human genome is made up of simple sequence repeats?
~5%
What is a transposon? Why is duplication so slow?
Mobile DNA elements, slow because it may cause mutations
What are the two classes of transposons?
LINEs and SINEs
What is the estimated current transposition rate in humans?
1 insertion for every 20-200 births
What are Alu elements? How big are they?
Members of the SINE class that make up 10% of human DNA, approx. 300bp long
What does Alu transposition depend on?
They are non-autonomous and depend on LINE elements
What is the current estimated transposition rate for Alu elements?
1 insertion for every 20 births
What kinds of genes are in the human beta-globin cluster?
5 genes and 1 non-functional pseudogene
How has the gene cluster evolved?
gene duplication
What is synteny?
comparison of genomes of different organisms shows order of genes in chromosomes is is partly conserved through evolution
What are the ends of chromosomes called?
Telomeres
What is the basic building block of chromatin?
nucleosomes
What are the copies of the histones called?
H2A, H2B, H3 and H4
What charge is a histone?
positive
The spread out parts of chromosomes in the nucleus are called…
euchromatin
Some parts of the chromosomes condensed in interphase are called….
heterochromatin
The structure of expressed genes is more ______ than others
open
How is the more open structure observed?
greater sensitivity to DNase
What are the two main processes involved with the opening of chromatin?
Histone modification (via acetylation) Chromatin remodelling complexes
What does RNA polymerase 1 transcribe?
ribosomal RNA genes
What does RNA polymerase 2 transcribe?
genes coding for proteins
What does RNA polymerase 3 transcribe?
genes coding for small RNAs
Name two basal factors needed for transcription by RNA polymerase 2
TFIIA, TFIIB etc
Where does transcription in eukaryotes start?
TATA box
What needs to bind to the TATA box for transcription to start? What does it do to DNA?
TATA box binding protein, binds DNA by 70 degrees
What is the complex called that is formed by TBP? What else has to bind to TBP to form this complex?
TFIID - TAFs must also be bound
Which TATA box binding protein does Pol 2 bind to, stabilising the complex?
TFIIF
What does TFIIH do?
Phosphorylates Pol 2
What does a cis-regulatory element do?
Increase/decrease basal level of transcription
Specify how gene expression responds to a particular signal
What does the metallothionein gene do?
Bind toxic metal ions
Store Zn2+
What induces transcription in humans?
Metal (Zn2+ ions)
Glucocorticoid hormone
Which cis-regulatory elements increase expression?
GC box
BLE
TRE
E-box
What are the response elements?
GRE
MRE
what is the principal glucocorticoid hormone? how does it work?
cortisol - enters the cell and exerts its effects directly
What does cortisol bind to?
glucocorticoid receptor (GR)
What protein keeps glucocorticoid receptor in cytosol? What kind of protein is it?
Hsp90 - chaperone protein
What are steroid hormone receptors?
Zinc finger proteins
What does AP1 do?
Increases general chromatin accessibility
When not bound, where is the activator protein?
Cytosol
What do fibroblast cells differentiate into?
Muscle cells
What are the transcription factors that are expressed in fibroblasts for them to differentiate into muscle cells?
MyoD, myogenin, Myf5, Mrf4
What are Somites?
Segmented blocks of cells that form along sides of the notochord in vertebrate embryos
What TF can reprogram fibroblasts into iPS celsl
Oct4, Sox2, Klf4, Myc
What TF can reprogram exocrine cells from the pancreas to islet beta cells?
Pdx1, Ngn3, MafA
If a cis-regulatory element is deleted from a gene, what happens to its effect on transcription?
it is eliminated
