CHAPTER
10
A
gene mutation is when a hereditary change occurs in the gene message causing a
particular phenotype. It typically cannot be visualized microscopically. It is
usually referred to as a point mutation.
It typically involves a single base or a small cluster of nearby bases. Point
mutations may be:
Base
substitutions-
where one base pair is replaced by another. A base substitution may be a transition
(purine replaces purine and pyrimidine replaces pyrimidine), or a transversion
(a purine replaces a pyrimidine or vice versa).
Additions
or Deletions-
a base pair is added or eliminated from the previous message.Often these are
referred to as Indel mutations. These usually lead to frame shift mutations
which scramble the message downstream from the location of the addition or
deletion. Often this causes a translational termination (stop) signal to arise
prematurely in the mRNA message.
Mutations
are described as synonymous if
they represent redundant codons for the same amino acid, missense
if they represent codons for different amino acids, or nonsense
if they replace a codon for an amino acid with a stop signal.
Conservative
substitutions replace one amino acid by another with similar properties and
are likely to have less severe phenotypic effects than nonconservative
substitutions.
Mutations
may affect coding regions of DNA or noncoding regions (e.g., regulatory
regions).
If
mutations occur in response to a known environmental cause or due to treatment
with a mutagen they are classified as induced mutations. Mutations that can
not be attributed to known mutagens reflect background mutational levels and
are referred to as spontaneous mutations.
Spontaneous mutations occur at the rate of 10-5 to 10-8.
When researchers attempt to induce mutations they assume that all mutations
that occur are the result of induction and disregard the low levels of
mutations that are spontaneous.
Tautomerization
Each
base can occur in either of two states and relative frequency of these two
states is in equilibrium for each base. The most significant thing to remember
about these states is that the rare forms will have different pairing
properties than the common forms, BUT the rare form will be a purine if the
common form is a purine and a pyrimidine if the common form is a pyrimidine.
Therefore, if a base spontaneously takes on its rare tautomeric
form during replication a transition mutation results. Figure 10-6.
Base
Analogues
are compounds that are sufficiently similar to normal nitrogenous bases that
they may be incorporated in nucleotides in the place of the normal nitrogenous
bases. These base analogues often will undergo tautomeric shifts with greater
frequency than the normal bases leading to transitional mutations. These are
commonly used for inducing mutations. See 5-bromouracil
(5-Bu) in Figure 10-8 and 2-Aminopurine
(2-AP) in Figure 10-9. %-Bu in its common form displaces Thymine,
but in its tautomeric form pairs with Guanine. 2-AP will displace Adenine and
miss pair with Cytosine.
Base
Alteration.
Some mutagens are not incorporated into the DNA, but alter the existing base. Alkylating
agents (e.g., ethylmethanesulfonate
(
Intercalating
agents:
(e.g., proflavin, acridine orange)
fit into the DNA double helix between adjacent base pairs leading to frame
shifts.
Base
damage
can prevent replication under some circumstances. This must be avoided at all
costs by organisms or they will be doomed. Some bacteria have an SOS system
that responds to such situations and reduce the fidelity of fidelity of
positioning complementary bases opposite the sites with damage. UV
radiation induces dimer formation and may stimulate the SOS
response. Aflotoxin B1 (Figure
10-15) causes guanine to
break off from the deoxyribose sugar thereby creating an apurinic
site (Figure 10-16).
The SOS response is elicited in this case and typically places an adenine
opposite the apurinic site (a transversion).
Deamination
of cytosine yields uracil which will pair with adenine leading to a transition
mutation. However, uracil-DNA glycosylase usually recognizes uracil in the DNA
and removes them. Often cytosine is methylated in eukaryotes forming
5-methylcytosine. When this is deaminated it pairs with thymine and the
consequent transition. Figure 10-17. Methylated cytosine represents a hot spot
for mutations because thymine is not recognized as an inappropriate base in
DNA by any DNA glycosylase (Figure 10-18).
Some
oxygen species can cause oxidative damage to bases (Figure 10-19). These may
lead to transversions.
Trinucleotide
repeats
are common and they are frequently associated with human genetic diseases in
individuals having too many copies. Often the parents have more copies of the
trinucleotide repeats than is typical and this leads to slippage during
replication producing many more copies in affected individuals.
Transposable
elements can
disrupt the normal coding messages for polypeptides if they are
inserted within the coding region of a gene and alternatively they can modify
the rate at which a gene is transcribed when inserted in the regulatory region
for a gene.
Biological
Repair mechanisms
When
DNA is being replicated initially incorrect base pairing is not uncommon.
During DNA replication, mismatched nucleotides are incorporated at the rate of
10-5 per template nucleotide per round of replication. DNA
polymerase has an additional function beyond synthesizing the polymer; it has proof
reading responsibility. It has 3'
to 5' exonuclease activity
that permits it to eliminate most incorrectly positioned bases soon after they
are added to the growing sugar phosphate backbone. About 99% of the initially
mismatched nucleotides are corrected via proof reading. Mismatch repair (see
below) corrects 99.9% of the remaining mismatches.
Another
method for preventing errors is via the enzyme superoxide
dismutase which converts superoxide radicals to hydrogen peroxide
(H2O2) which is then converted to water (H2O) by the enzyme catalase.
Lesions
to DNA consisting of pyrimidine dimers
can be repaired by the enzyme photolyase
in bacteria and lower eukaryotes. Photolyase can only function if it is
exposed to visible light during repair.
Alkyltransferases
are enzymes that reverse lesions caused by alkylating agents.
The
general excision repair system
can recognize significant DNA lesions (e.g., thymine dimers) and use
endonuclease activity to break the sugar phosphate phosphodiester bonds up and
down stream from the lesion. This allows the problem area to be excised from
the DNA. The excised gap is 12 or 13 nucleotides in prokaryotes, 27-29
nucleotides in eukaryotes. This strand is resynthesized using DNA polymerase I
and the strand is sealed using a ligase.
The
specific excision repair
involves repair of less obvious lesions in the DNA. DNA glycosylases recognize
altered bases and cleave the base from its sugar thereby creating an AP site.
AP sites are recognized by AP endonucleases which then excise the AP site and
replace it with an appropriate nucleotide using DNA polymerase I and a ligase.
Mismatch
repair
occurs soon after the DNA is replicated. Incorrectly matched base pairs are
recognized and the incorrect nucleotide is excised and replaced by a correct
nucleotide. The difficulty in this system is identifying which strand contains
the incorrect nucleotide. This is done in prokaryotes by taking advantage of
the fact that when a DNA sequence 5'GATC3' occurs, the adenine will be
methylated in prokaryotes. However, this methylation occurs post-replication.
Therefore, there is a short period of time during which only one of the two
DNA strands is methylated and this represents the older strand and the one
with the correct sequence. The methylation site does not have to exist
extremely near the mismatch.
Recombinational
repair
is a process that occurs post replication. When a strand containing a major
DNA lesion is to be replicated the complement is synthesized in such a way
that a gap is left opposite the region containing the lesion. The details
concerning how this is done by DNA polymerase have yet to be worked out. After
replication, a region of the other DNA strand in the original DNA molecule is
used to fill in the missing gap. Now the other original strand is missing a
gap. This gap is synthesized by using its newly synthesized complement as a
template. See Figure 10-41.
Mutations
may be somatic or germinal
in nature. We are most concerned about germinal mutations because they are
passed from generation to generation. Somatic mutations may affect inheritance
in plants when they occur in branches that give rise to flowers that become
part of the reproductive system of the organism. They also can be important
for asexual species.
Most
mutations are recessive. They can only be recognized if they do not coexist
with a dominant form in a diploid individual. Therefore, to recognize that a
mutation has been induced in a subject often that subject is test crossed.
This type of mutation detection is called a specific-locus
test.
A
mutation from wild type to mutant form is referred to as a forward
mutation. A mutation from mutant form to wild type is a back (or
reverse) mutation.
A
conditional mutant allele causes
the mutant phenotype only in a certain environment, called the restrictive
condition. The wild type phenotype is produced in other
environments (permissive conditions).
Some mutant alleles can only influence phenotypes if the individual exists in
the restrictive condition during a particular period of its development (sensitive
period).