Lecture 14

Importance of Understanding DNA Replication Mistakes

DNA replication mistakes are critical because changes at the DNA level are permanent, unlike RNA which is transient. Mistakes can have significant consequences depending on their location in the sequence, potentially leading to various diseases, including cancer.

Types of Changes in DNA Coding

Changes can occur in coding sequences leading to:

  • Silent mutations: No effect on the amino acid sequence, often occurring in the third position of codons due to the redundancy of the genetic code.

  • Missense mutations: Change in one amino acid, which can alter protein function or stability depending on the role of that amino acid in the protein structure.

  • Nonsense mutations: Premature termination of protein translation, dramatically shortening the protein and often resulting in loss of function.

  • Frameshift mutations: Occurs when insertions or deletions shift the reading frame, leading to widespread changes in the amino acid sequence downstream of the mutation.

It is important to note that changes in DNA can also affect non-coding regulatory sites (e.g., promoters, enhancers), which can profoundly influence gene regulation and expression levels, potentially leading to diseases.

Non-Coding and Regulatory Changes

DNA modifications can impact protein-DNA interactions, influencing gene expression and function beyond mere open reading frames. For instance, mutations in enhancers may drastically alter the expression of genes critical for development or cellular function.

Misconceptions about Mutations

Mutations are often perceived as completely random with no inherent mechanism to choose beneficial or detrimental changes. They can be classified as:

  • Neutral mutations: No effect on the organism's fitness.

  • Negative mutations: Deleterious effects that diminish survivability or reproductive success.

  • Positive mutations: Provide some advantage under specific environmental conditions, such as antibiotic resistance in bacteria or cancer cell growth advantages.

Example

Cancer cells can gain advantages through mutations allowing uncontrolled growth and survival in unfavorable conditions. The accumulation of mutations can lead to tumor heterogeneity, making treatment challenging.

Mutation Rates and Causes

Errors in DNA replication occur with a chance of approximately 1 in 10^7 bases. Factors leading to increased mutation rates include environmental damages such as radiation and chemicals, along with inherent errors in the DNA replication machinery. Specific types of damage, such as deamination and depurination, can lead to permanent mutations if not correctly repaired.

Specific Mutagenic Events

Deamination

Removal of amine groups from nucleotides, such as cytosine converting to uracil, results in a substitution error since uracil pairs with adenine rather than guanine. Other bases can also be modified, creating different pairing partners, leading to substitutions or coding changes based on position in codons. Methylated cytosine can change pairing due to deamination, leading to transitions that alter the normal pairing mechanism.

Depurination

The removal of purine bases (adenine, guanine) can cause the DNA replication machinery to skip bases, leading to deletions in the new DNA strand. This is highly likely to cause frameshift mutations in coding sequences, drastically impacting gene function and leading to disease in some cases.

Oxidative Damage

This type of damage is caused by reactive oxygen species (e.g., hydrogen peroxide), leading to direct changes in nucleotide pairing and causing substitutions or blockage of replication processes by modifying thymine to thymine glycol.

Alkylation and UV Damage

Alkylating agents can damage DNA bases as well as the sugar-phosphate backbone, resulting in a range of possible consequences. Furthermore, exposure to UV light can cause thymine dimers, leading to replication errors and potential carcinogenic mutations due to disruptions in normal base pairing patterns.

Breaks from X-rays

X-ray exposure can result in single or double-strand breaks in DNA, adversely affecting replication if the backbone is disrupted, further complicating cellular repair processes and increasing the likelihood of mutations.

Cancer Cells and DNA Damage

The cells of cancer are constantly replicating, which makes the mutagens used to induce mutations a targeted method of chemotherapy, impacting rapidly dividing cancer cells more than resting normal cells. This treatment approach exploits the higher mutation rates and the reduced ability of cancerous cells to correct DNA damage.

Mismatch Repair Mechanism

Mismatch repair occurs after DNA replication and significantly decreases the error rate from 1 in 10^7 to 1 in 10^9 bases. The process involves:

  1. MutS: Detects mismatches based on differences in base pairing.

  2. MutL: Coordinates the repair response to the detected mismatch.

  3. MutH: Makes cuts in the unmethylated newly synthesized strand.

  4. Exonucleases: Remove the incorrectly incorporated nucleotides and repair DNA polymerase fills in the gap.

  5. Ligation: Seals the repaired DNA strand, thus maintaining the integrity and accuracy of the genetic code despite potential replication errors.