Mutation

Mutation

• Means change• Definition: An event that gives rise to an

alteration in the genotype• It also can be the change itself• Mutation does not mean “bad”• Mutations are IMPORTANT for a variety of

reasons

Mutation ‘Levels’

• DNA Level

• Chromosomal Level

• Genome Level

DNA Level• At or below the gene level• 1) Addition or Deletion of one or two

nucleotides (+ and – frameshifts)• 2) Substitution of one nucleotide for

another• 3) Inversion within a gene• 4) Crossingover within a gene

– Equal–Unequal

• 1 and 2 are often referred to as POINT MUTATIONS

Chromosomal Level• 1) Duplications• 2) Deletions• 3) Translocations

–Reciprocal–Non-reciprocal

• 4) Inversions• ? ‘Standard’ crossing over ?• 3 and 4 may result in POSITION EFFECTS

Genome Level

• These result in ploidy changes• Aneuploidy• Plus or minus one or a few chromosomes

• Euploidy (true polyploidy)• Involves whole SETS of chromosomes

–Autopolyploidy–Allopolyploidy

Additional Points

• ALL genes can mutate• Observed levels are usually low• Some genes have very high rates of

mutation (Mutable Genes) • Some genes seem to increase the rate of

mutation in adjacent genes (Mutator Genes)

What Causes Mutations ?

• 1) Spontaneous (?????)• 2) Chemically induced• 3) Radiation induced• Most (if not all) really come down to a

chemical level• No such thing as a “spontaneous”

mutation• “Spontaneous” means…

Chemicals

• Many chemicals are mutagens; many others are promutagens

• Chemicals (just like radiation) can cause point mutations.

• Chemicals (just like radiation) can cause chromosome breakage (e.g. - LSD, Mercury, Caffeine --- at least in cell culture)

DNA Level Mutations

• + and – Frameshifts and their effects• + or – one nucleotide• + or – a whole codon• Multiple frameshifts• The effects depend on WHERE it occurs• The effects depend on what the change is• ?????? One change equals one change, or

does it ??????

DNA Level Mutations• Substitutions• Transitions (more common)• Purine (A, G) for a purine OR pyrimidine

(T, C) for a pyrimidine• Transversions (less common)• Purine for a pyrimidine (or pyrimidine for

a purine• GENERALLY have less chance of an effect

than frameshifts. WHY?

Other DNA Level Mutations

• Inversions within a gene• Crossingover within a gene

Substitution Mutations• Results from a temporary type of

isomerization (tautomerization) during replication

• Base enters a “rare” state (unstable)• Can be in a base already in the DNA strand• Can be in one of the incoming bases• We will look at both for a TRANSITION• Requires two divisions to get the “stable”

mutation

Substitution Mutations• One nucleotide is replaced by another• Transition --- insertion of a one purine base for

the other or one pyrimidine base for the other (results in an “odd” purine-pyrimidine pair)

• Transversion --- replacement of a purine base with a pyrimidine or vice versa (results in a temporary purine-purine pair or pyrimidine-pyrimidine pair)

Substitution Mutations

• Base analogs increase the rate of formation of substitution mutations

• 5-bromouracil (5BU)• Use of 5-fluorouracil

Radiation Induced Mutations• By non-ionizing radiation (UV)• Dimer formation (especially TT dimers)• Light Repair System (enzyme breaks dimers)

Radiation Induced Mutations• By non-ionizing radiation (UV)• Dimer formation (especially TT dimers)• Dark Repair System (excision repair)– Endonuclease, exonuclease,

DNA polymerase, ligase

Radiation Induced Mutations• By ionizing radiation• Mechanisms

- “Target Hypothesis”- “Physical-Chemical” Hypothesis

• Temperature, hydration, and oxygen contentall have an effect

Mechanism

Mechanism

Ionizing Radiation and Mutation

• Late 1920s• Stadler – working on barley and corn• Muller – working on Drosophila• Muller later received a Nobel Prize (1948)• Why H. Muller and not Stadler?

SOME GENERALIZATIONS FOR IONIZING RADIATION

• Mutation frequency is DIRECTLY proportional to the dose

• Doesn’t matter if given in one large dose or several small ones

• No lower limit (no threshold)• Safe amount?????

Additions and Deletions• Chemical basis• Caused by intercalation (insertion) of some

nucleotide like chemical into the chain• Before replication --- increases length• Results in a Plus (+) frameshift mutation• During replication --- decreases length• Results in a Minus (-) frameshift mutation• Acridines are capable of doing this

Additions and Deletions• Requires 2 divisions to get the “stable” mutation• Proflavin

• Acridine Orange (very potent)

Additions and Deletions

- frameshift

Intercalation of Proflavin

DURING DNA

replication

+ frameshift

Intercalation of Proflavin PRIOR TO

DNA replication

Chemicals Besides Acridines

• Too many to mention…but…• Mustard gas (adds a methyl group)• Formaldehyde• Phenol• Nitrous Oxide (replaces NH2 with O)

Mutation Concluded• Temperature has an effect• Carcinogens and Mutagens• Somatic vs. germinal mutations• Somatic are “dead-end” in animals• In plants somatic can be “passed on”

asexually by vegetative propagation(runners, bulbs, corms, cuttings, grafting)

• In plants somatic mutations can get into the germ line!

Testing for Mutagens & Promutagens

• Ames test• Uses a histidine deficient (dependent) strain

of Salmonella• Looks for reverse mutation to histidine

independence above the background rate• For suspected promutagens: first treat

substance with a liver (human or rat) extract and then do the test

Teratogenesis

• Teratogen – an agent that induces a non-inheritable defect during embryonic development (Proteratogen)

• In humans --- about 4% of births• How do they work?– Interfere with mitosis or cell migration– Interfere with differentiation (disturb gene

regulation, translation or activity of ultimate gene products)

Teratogens• Radiation• Viruses (Rubella and pregnant women)• Thalidomide (really a proteratogen)• Diethylstilbestrol (DES) (taken by pregnant

women - results in genital tract abnormalities in their children)

• Dioxins (found as a contaminant in herbicides such as Agent Orange)

• Hexachlorophene (a bacteriocide)

Teratogens• Smoking (CO reduces O2 available to the fetal

circulation; nicotine constricts blood vessels and reduces blood flow)

• Progestin (artificial progesterone-like compound; causes masculinization of female fetuses)

• Alcohol – microcephaly, low birth weight, some mental retardation)

• Caffeine, Aspirin and numerous legal and not so legal drugs

Some Other Birth Defects

• Cleft lip and/or cleft palate• Spina bifida• Club foot• Heart disease and malformations• Cataracts• Deafness• Intestinal tract disorders• Low birth weight