|Negatively charged DNA coiled around histones. Credit: Thomas Splettstoesser (from PDB 1EQZ)|
Mutations are permanent changes in the genetic material passed down from one generation to the next. In order for the continuation of the species, each generation of an organism must faithfully replicate their genomes. In order for the species to evolve and adapt, some mutations must be allowed to occur. As the substrate of evolution, mutations are especially crucial for the maintenance of genetic variation in microbial populations, which do not rely on sex or meiosis.
In the laboratory, mutagens allow geneticists to study loss-of-function mutations and elucidate gene function. For instance, the function of the gene products involved in DNA repair was elucidated by introducing mutations in their respective genes and comparing mutants’ sensitivity to chemical and ionizing mutagen to the sensitivity of the wild-type organism.
|Nitrous acid. Don't drink it.|
For example, mitomycin C and 1,2,7,8-diepoxyoctane cause deletions while nitrous acid causes oxidative deamination of adenine and thymine to hypoxanthine and xanthine, respectively. Diepoxybutane affects G:C base pairing. N-methyl-N-nitrosourea and N-methyl-N’-nitro-N-nitrosoguanidine methylate bases, causing base-pair substitutions and baseless deoxyriboses. Ethidium bromide has a ring structure that intercalates bases and stretches the duplex, leading to frameshift mutations. Incorporation of base analogs, with higher rates of tautomerization, into DNA during replication will also lead to mutations. Ultraviolet irradiation is also mutagenic, and comes in three forms (in order of increasing energy) : UVA, UVB, UVC. UVC is the most lethal to bacteria, but all of them will induce the formation of thymine dimers. UV formation of a cyclobutane ring between the two adjacent thymines, called a thymine dimer. Without chemical and ionizing mutagens, the rate of mutagenesis would be so low, mutant yield would be low in the laboratory, and isolating them would be impractical.
Spontaneous mutations can occur when bases react with water or other natural species in the cellular environment. For example, cytosine deaminates to uracil, methylcytosine deaminates to thymine, adenine . Bases can tautomerize, switching from keto to enol form. Each form has a different base pairing property. For example, a guanosine in its keto form base pairs with cytosine, but a guanosine in its enol form will base pair with thymine. Thus, a switch from the keto to the enol form results in a G:C to A:T transition in one of the daughter cells. Mutagenic base analogs like 5-bromouracil tautomerize and cause transitions this way. However, DNA damage can involve more than just incorrectly incorporated bases. When double strand breaks or gaps occur, they are repaired either by non-homologous end joining (before replication) or homologous recombination (after replication, when sister chromatids can provide a template). In NHEJ, an exonuclease process the single stranded ends of the broken DNA, and ligases then directly join them together. The digestion of ends may result in the loss of nucleotides and mutations. Homologous recombination is less mutagenic because sister chromatids or other homologous regions are used as a template for repairing the gap.
Given that these changes to the DNA do not result in mutations unless they are fixed after a round of replication, the cell must repair the DNA before replication ends. This Monday night, we'll go over how the cell detects aberrations and repairs them!