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Gene expression
Have you ever wondered how a frog grows?
http://www.teara.govt.nz/en/frogs
Native male frogs sit on the eggs and the young froglets hatch almost fully
formed.
http://www.erodent.co.uk/GardenPond/PondPhotos/Tadpole3w.jpg
Why are their parts where they are?
The answers lie in their cells.
In particular – the answer can be found in their genetic material.
A right handed helix
Let’s revisit how we know that DNA is important.
Streptococcus pneumoniae
S: smooth
(virulent)
R: rough
(avirulent)
DNA is the genetic material - Griffith 1928
Avery, MacLeod & McCarty 1944 Chemically characterised the ‘transforming extract’ from virulent cells
observed by Griffith Removed the protein from ‘transforming extract’
and it still transformed Removed RNA
extract still transformed Removed DNA
prevented transformation Therefore transforming substance was DNA
Biochemically characterised the transforming extract, all its properties were consistent with DNA: High Mw (centrifugation) High charge (electrophoresis) Characteristic UV absorbance Chemical analysis, ratio of nitrogen to phosphorus
Hershey-Chase 1952 Left, T2 virus protein
radioactively labelled 35S Radioactivity outside
Right: T2 virus DNA radioactively labelled 32P Radioactivity inside
THEREFORE genetic material infecting E.coli was DNA and not protein
Genes are made from DNA (except some RNA viruses)
Genes are encoded within DNA
What is a gene?
Classical molecular gene: “A stretch of DNA sequence that codes for a particular protein that has a particular function”.(10,11) This can be an interrupted sequence within a chromosome.
14
Evolutionary gene: “any portion of chromosomal material that potentially lasts for enough generations to serve as a unit of natural selection”.(39) As such a gene is an inherited unit which is somewhere between a nucleotide and a chromosome.
Systemic Concept: The gene is a combination of (one or more) nucleic acid (DNA or RNA) sequences, defined by the system (the whole cell, interacting with the environment, or the environment alone, in sub-cellular or pre-cellular systems), that gives origin to a product (RNA or polypeptide).(57)
Other definitions of a gene
The central dogma
RNA Precursors(ribonucleotides) RNA
Transcription
Protein Precursors(amino-acids) Protein
Translation
DNA Precursors(deoxyribonucleotides) Replication
DNA (Genotype)
(Phenotype)
http://vcell.ndsu.nodak.edu/animations/transcription/advanced.htm
Transcription is the first step in the process.
How many genes does it take to make a person?
20,000-25,000
26,000 19,000 13,000 6,000 4,000
18
DSCAM: one gene 38,016 mRNAsAn immunoglobulin (Ig) superfamily member
an axon guidance receptor
Drosophila melanogaster Dscam gene contains 115 exons spanning ~60,000 bp20 exons are constitutively spliced (open boxes) and 95 exons are alternatively spliced
(shaded boxes).Alternatively spliced exons are orgnized into 4 clusters (exons 4,6, 9, 17) that contain
12, 48, 33 and 2 alternative exons each.
Exons in each cluster are spliced in a mutually exclusive manner.
Celotto & Graveley 2001 Genetics 159:599-608
19
Immunoglobulin genes recombine within the DNA.
So what if nuclei are the wrong shape?
85 mm
16 mm
48129895 bp
249250621 bp
More or less matters!
DownEdwards
PatauKlinefelters
(XXY)
Turners(X)
22
Albert Einstein (1955)
23
30-35% of cells are aneuploid
~4% aneusomy for chr 21
Affects survival, proliferation potential, and protein imbalances
24Falchi et al. (2014) Nat Genetics 46:492-498
Copy number affects your weight
How big is a vertebrate cell?
http://medicalpicturesinfo.com/human-cell/
~6 µm
26
Progeria (Hutchinson-Gilford progeria)
Children with this disease typically have a stroke or heart attack caused by severe atherosclerosis at an average age of 13 years.
Nuclei in Progeria are the wrong shape.
Treating progeria cells with remodelin results in the nucleireverting to the ‘normal’ shape.
Shape directly affects the genes that are turned on.
31 of the altered genes affect vasculature and atherosclerosis
22 of the altered genes affect skeletal, limb and cartilage
Defects in transcription can cause problems. Who amongst us is lactose intolerant?
LCT gene 17 exons Chr 2q21
Lactase is encoded by a single gene
-24 Kb G-A
-14 Kb C-T
A single mutation (C changes to T) causes lactase persistence
Swallow; Annu Rev Genet 2003:37197-219
The frequency of lactase persistence varies dramatically in different
populations.
Swallow; Annu Rev Genet 2003:37197-219
Pastoralists and milk drinkers tend to have higher frequencies of lactase persistence
than nonpastoralists
Translation is important too.
tRNA: the adapter
What is the genetic code?
Redundant codons are all synonyms for the same protein building block.
Degenerate codons differ in their third positions; e.g. both GAA and GAG code for the amino acid glutamic acid.
SerCandida
Nons.Micrococcus Nons.
Micrococcus
Nons.Mycoplasma
TrpMycoplasmaSpiroplasma
CysEuplotes
Gln: Ciliates & Acetabularia
First position (5’ end)
Second Position
Third position (3’ end)
U C A G U UUU Phe UCU Ser UAU Tyr UGU Cys U
UUC Phe UCC Ser UAC Tyr UGC Cys C UUA Leu UCA Ser UAA Stop UGA Stop A UUG Leu UCG Ser UAG Stop UGG Trp G
C CUU Leu CCU Pro CAU His CGU Arg U CUC Leu CCC Pro CAC His CGC Arg C CUA Leu CCA Pro CAA Gln CGA Arg A CUG Leu CCG Pro CAG Gln CGG Arg G
A AUU Ile ACU Thr AAU Asn AGU Ser U AUC Ile ACC Thr AAC Asn AGC Ser C AUA Ile ACA Thr AAA Lys AGA Arg A AUG Met ACG Thr AAG Lys AGG Arg G
G GUU Val GCU Ala GAU Asp GGU Gly U GUC Val GCC Ala GAC Asp GGC Gly C GUA Val GCA Ala GAA Glu GGA Gly A GUG Val GCG Ala GAG Glu GGG Gly G
The genetic code is not always the same
Some people can’t use Glucose.Typically they have mutations in
GLUT1
37
These mutations occur because the codons are changed.
GLUT1 mutations are often in helix 4
38
R126H
R126L
R153L
Mitochondria in a mouse myoblast cell
39Image taken by James Markworth (Liggins Institute)
Mitochondria are red
Actin filaments are green
Nucleus is blue
40
Differences in the human nuclear and mitochondrial genetic codesFirst
position (5’ end)
Second Position
Third position (3’ end)
U C A G U UUU Phe UCU Ser UAU Tyr UGU Cys U
UUC Phe UCC Ser UAC Tyr UGC Cys C UUA Leu UCA Ser UAA Stop UGA Stop A UUG Leu UCG Ser UAG Stop UGG Trp G
C CUU Leu CCU Pro CAU His CGU Arg U CUC Leu CCC Pro CAC His CGC Arg C CUA Leu CCA Pro CAA Gln CGA Arg A CUG Leu CCG Pro CAG Gln CGG Arg G
A AUU Ile ACU Thr AAU Asn AGU Ser U AUC Ile ACC Thr AAC Asn AGC Ser C AUA Ile ACA Thr AAA Lys AGA Arg A AUG Met ACG Thr AAG Lys AGG Arg G
G GUU Val GCU Ala GAU Asp GGU Gly U GUC Val GCC Ala GAC Asp GGC Gly C GUA Val GCA Ala GAA Glu GGA Gly A GUG Val GCG Ala GAG Glu GGG Gly G
Met
Trp
Stop
42
Prions and Alzheimer’s disease are examples of
pathological protein misfolding
43
What’s different between these bears?
Polar bears and Brown BearsDivereged ~479-343 kya
Polar bears are adapted to high fat diets
Modified from Liu et al. Population genomics reveal recent speciation and rapid evolutionary adaptation in Polar bears. Cell (2014): 157, 785-794 and Ombostad, I. (2012) Relationships between persistent organic pollutants (POPs) and plasma clinical-chemical parameters in polar bears (Ursus maritimus) from Svalbard, Norway. Student thesis (Trondheim, Norway: Norwegian University of Science and Technology).
The environment is affecting the genes that Polar bears have.
Basic unit of muscle
Polar bears have seven missense substitutions in the LYST gene
These mutations occur in the same region as ones in
human disease
Chediak-Higashi syndrome Autosomal recessive
Mutation of lysosomal trafficking regulator (LYST)
What do Chimo and a Polar bear have in common?
Lyst affects melanosome maturation
The organelle that is involved in synthesis, storage and transport of
melanin
White fur is common in the arctic:
beluga whales, polar bears, arctic hare, arctic fox – but not all due to Lyst
mutations.
Confers a selective advantage?