Topic 2 - Mutation and Mutants

Mutations

Heritable alteration to the DNA sequence of an organism

-Classified as being spontaneous or induced

Spontaneous Mutations - Occur due to errors in replication and repair defects, rare

Induced Mutations - Use/apply mutagenic agents (mutagens) to increase frequency of mutations

Mutagen

Agents used to induce mutations, organized into three broad classes

Classes of Mutagens

Chemical Agents - Ethyl Methanesulfonate (EMS)

Physical Agents - UV light (non-ionizing), X-rays/Gamma-rays/Fast neutrons (ionizing)

Biological (insertional) - Transposable elements/moveable DNA

Administration of Chemical Mutagens

Topically, injection, ingestion, inhalation, soaking (ex; Soaking Seeds)

Type of Mutations Caused by Chemical Mutagens

-Point Mutation - Focal events including Single-Base point Substitution (SBS) and indels (insertions/deletions)

-Single-Base Point substitution is either Transversion (Purine → Pyramidine) or Transition (Purine → Purine)

Chemical Mutagen Alteration Generation

1. Base Modification - Alters base pairing properties

-Ex; Ethylation of a Guanine results in it binding to Thymine with two bonds

2. Intercalates in DNA backbone, altering physical structure

-Alteration creates replication difficulties, tends to generate indels

3. Mimics normal bases, isomers have altered base pairing properties

-Typically referred to as Base Analogs, suited for point-mutations

Administration of Physical Mutagens

Irradiation (ionizing or non-ionizing)

Typical Mutations Caused by Physical Mutagens

-Non-ionizing radiation (UV) results in point mutations (SBS)

-Ionizing Radiation (x-rays, etc) results in inversions, translocations, large deletions

Reciprocal Translocation - moving of genetic material in both directions

Physical Mutagen Alteration Generation

-UV results in pyrimidine dimers

-Ionizing radiation results in double-stranded breaks in DNA

Administration of Biological Mutagens

-Transformation of an exogenous Transposable element

-Mobilize an endogenous transposable element

Type of Mutations Caused by Biological Mutagens

Insertions and Deletions

Biological Mutagen Alteration Generation

-Transformation or stimulation of mobilization of our Transposable Element

Ex; Wrinkled pea seeds have mutant alleles that have resulted from an insertion of a transposable element

Mobilization by Biological Mutagens

1. Replicative Transposition (Copy and Paste) via an RNA intermediate

• Original element transcribed into RNA, then reverse transcribed and pasted into new location

2. Non-replicative Transposition (Cut and Paste) as a DNA molecule

Mutations Can be Described at the DNA Level

-Transition vs Transversion (SBS)

-Indel (addition/deletion fo 3 or fewer base pairs)

-Inversion - Sequence of DNA inverted by 180 degree

-Reciprocal Translocation - Movement of sequence of DNA to different chromosome

Description of Mutations in Coding Region

1. Silent Mutation - No change in the amino acid

2. Non-Conservative Missense - Change of an amino acid into a different family

3. Conservative Missense - Change of amino acid into the same family

4. Non-Sense Mutation - Premature stop-codon

5. Frameshift Mutation - Shift of every codon downstream of insertion or deletion

Classification of Mutations

Translation Start Site (ATG) - A is referred to as nucleotide (1)

Nucleotides in Intron - Either highest number of nearest exon + 1, or lowest number of nearest exon - 1

Upstream of Start-Codon (5’ UTR) - Referred with negative numbers

Downstream of Stop-Codon (3’ UTR) - Referred with an asterisk

Nomenclature for Mutations

C -> Coding DNA

3 -> Position of nucleotide

C.3 -> Third nucleotide of the exon (coding-region)

C.X+Y -> Nucleotide in an intron 

C.X-Y -> Nucleotide in an intron

C.-X -> 5’ UTR

C.*X -> 3’ UTR

Additional Nomenclature

p.X uses protein sequence (amino acid) as the reference point

g.X uses genomic DNA as the reference point

“X” is used as a symbol for a stop codon

Ex; p.A400X (400th codon of a protein sequence, change from Glu to stop codon)

Non-Coding Sequence Mutations

Mutations outside of the coding sequence can also impact gene expression:

-Promoters/Enhancers

-Termination signals

-Splice donor/acceptor sites

-Ribosome binding site

Muller’s Scheme (Muller’s Morphs)

Based on the mutation’s effect on gene product activity/function

-Amorph (nullomorph), Hypomorph, Hypermorph, Antimorph (dominant-negative), Neomorph

Amorphs (Nullomorph)

Mutants with no remaining gene product function/activity (null alleles)

-Loss of Function (lof) mutations are recessive

-Wildtype allele is “Haplosufficient” single copy is sufficient to maintain wildtype phenotype

Amorphs lack phenotypic difference between homozygous Amorph mutation (m/m) and m/Df (deficiency/deletion of allele)

Hypomorph

Mutants with reduced gene product function/activity

-Mutations are loss of function mutations and are recessive (wildtype allele is haplosufficient)

Differentiating feature from amorph is that hypomorph phenotype (m/m) is not the same as m?df

-m/Df is more sever phenotype than m/m

Increasing the copy number of a hypomorphic allele (ex; m/m/m) should reduce severity of a hypomorphic mutation

Common Mutations

Most mutations are recessive, thus most mutations encountered are Amorphs/Hypomorphs

Hypermorphs

Mutations with increased gene product function/activity

-mutant gene product either more active, or an excess produced

Gain of Function (gof) mutations are dominant, wildtype is haploinsufficient

Defining characteristic is that the phenotype of M/Df is less sever ethan M/M

Antimorph

Mutant whose gene product function/activity poisons the wildtype allele product’s function/activity

-Antimorph mutations impact genes whose product have dimer, trimer or tetrahimer shapes (proteins binding to each other), not those who function as a polypeptide chain

-Are loss of function mutations (lof) that are dominant

Can lessen the impact by increasing dosage of the wildtype allele (swamp-out)

Neomorphs

Mutants whose gene product has a new/novel function/activity

-Gain of function mutations that are dominant

-Often due to ectopic (improper) expression that may be temporal (time-based) or spatial (place-based)

-Correct activity at an incorrect time/place

Neomorphs do not poison the function of the wildtype allele gene product

Allelic Series

Nullimorphic and hypomorphic alleles can be arranged into an allelic series

-Arranged from most severely affected (strongest allele) to least severely affected