Mutants and Disease MUPGRET Workshop. Mutation Heritable change in the DNA sequence. Naturally...
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Transcript of Mutants and Disease MUPGRET Workshop. Mutation Heritable change in the DNA sequence. Naturally...
Mutations as a tool Associating a phenotype with a
gene. Understanding gene function. Studying protein interactions. Understanding cell lineage and
organ development.
Associating a phenotype with a gene Changes in the DNA sequence that
non-functional or reduced function proteins often cause a visible change in the appearance of the organism.
Some changes do not give visible phenotypes.
Often identified as an “off-type” in plant species.
Misspelled Genes: 3 Possible Outcomes
A misspelled gene
DNA
Cell may not be able to
follow damaged
instruction
Cell does not make the
protein
X
X
OR Spelling error may be harmless
Functional protein made
by the cell
OR Damaged protein is made
Damaged protein may or
may not be able to function in
the cell.
Dwarfing Gibberellic acid
(GA) is a plant hormone.
GA levels influence growth.
Mutants in genes for GA synthesis, reduce plant height.
Associating a phenotype #2 This is often the first step towards
understanding the function of a gene or to dissecting a biochemical pathway.
The mutation can be either a naturally occurring one or an induced one.
Can be targeted or random.
Understanding gene function “You don’t know how something
really works until you have to fix it.”
Disruptions of the gene can be either non-functional or “leaky”.
Often the “leaky” phenotypes will really help you understand how to gene works.
Understanding gene function In the case of targeted
mutagenesis where you know what the other genes in that would/could be co-regulated with the mutant are you can understand the pathway better by looking at expression of the co-regulated genes.
Understanding gene function In the case of site directed
mutagenesis where you can target particular sequences, you can dissect the part of the protein that is important for function.
Can help to ID the catalytic site or a site involved in protein-protein interactions or a site involved in transport, etc.
Protein Explorer Protein Explorer
http://molvis.sdsc.edu/protexpl/frntdoor.htm
Also available at Biology Workbenchhttp://workbench.sdsc.edu/
Tutorials at http://www.ornl.gov/sci/techresources/Human_Genome/posters/chromosome/pdb.shtml
Troubleshooting http://molvis.sdsc.edu/protexpl/troubles.htm
Studying protein-protein interaction Often use a series of alleles that have
defects in different parts of the gene to identify the site that is required for protein-protein interaction.
The series can be insertions, deletions, or point mutations and may come from nature or be induced or a combination of the two.
Understanding cell lineage Usually used with transposon
mutagenesis. Transposons are mobile pieces of DNA
that can insert into a gene and disrupt its function.
Insertion can happen throughout development and can be used to track where cells came from with visible marker.
Methods for detection mutations Alteration in electrophoretic
mobility Sequencing Protein trunctation test
Blazing a Genetic Trail It tells the story of how mutations
are involved in several different diseases.
http://www.hhmi.org/genetictrail/
Association Genetics Usually used for medical genetics. Recently applied to plant genetics.
Which genes were involved in domestication?
Is this gene responsible for part of the difference we see in a particular trait such as plant height?
Dwarf 8 Mutagenesis and
trait analysis suggested that d8 might influence flowering time and plant height.
D8 study Sequenced D8 in many ~100
maize lines. Measured flowering time and plant
height in the same material. Compare DNA sequence to
flowering time and plant height.
D8 summary Found several polymorphisms that
are associated with changes in flowering time.
Data also indicate that D8 has undergone selection. Compare synonymous vs.
nonsynonymous substitutions.
Plants as a Model for Disease Sometimes mutations in the same
gene in different organisms have similar phenotype.
This allows researchers to choose the organism with the best genetic resources to study the normal function of that gene.
This also allows researchers to identify prospective genes for a phenotype in one species, based on another.
Xeroderma pigmentosa Autosomal recessive. UV exposure damages DNA. Defect in DNA damage repair. Risks include cancer,
telangiectasia, disfigurement. Can be diagnosed before birth. Take total protection measures
from sun/fluorescent light.
Mechanisms of UV damage
Low penetration into tissues. Molecular fragmentation—proteins,
enzymes, and nucleic acids contain double bonds that can be ruptured by UV.
Free radical generation—molecules of susceptible tissues absorb UV and eject an electron, which is taken up by oxygen, then termed superoxide, a free radical.
Free radicals Are scavenged by superoxide
dismutase, vitamin C, vitamin E, glutathione peroxidase, carotene.
Prions and Disease Proteins that can change shape. And make other proteins change
their shape! As number of changed proteins
increases a phenotype is observed. Causal agent of mad cow disease,
scrapie in sheep and Creutzfeldt-Jakob disease in humans.
Prions II Previously thought only nucleic
acid encoded changes caused disease.
Stanley Prusiner discovered prion’s ability to change other protein’s structure and won the Nobel Prize.
Sup35 is a prion-like protein in yeast.
Sup35 Translation termination factor Carboxyl end binds to the ribosomal
complex to terminate translation. If Sup35 is converted to an alternate
conformation (infectious prion conformation) the shape change spreads throughout the cell and is passed to daughter cells.
Sup35 In prion conformation causes
ribosomes to read through stop codons altering shape and function of proteins.
Not adaptively advantageous so why is it maintained?