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.

Types of Changes in DNA

Coding Changes

• Changes can occur in coding sequences leading to:

  • Silent mutations (no effect on amino acid)

  • Missense mutations (change in one amino acid)

  • Nonsense mutations (premature termination of protein)

  • Frameshift mutations (shift in reading frame)

• Important to note that changes in DNA can also affect non-coding regulatory sites (promoters, enhancers).

Non-Coding and Regulatory Changes

• DNA modifications can impact protein-DNA interactions, influencing gene expression and function beyond mere open reading frames.

Misconceptions about Mutations

• Mutations are completely random with no inherent mechanism to choose beneficial or detrimental changes.

• They can be:

  • Neutral (no effect)

  • Negative (deleterious)

  • Positive (provide some advantage in specific conditions).

  • Example: Cancer cells can gain advantages through mutations allowing uncontrolled growth.

Mutation Rates and Causes

• Errors in DNA replication can occur with a chance of 1 in 10^7 bases.

• Factors leading to increased mutation rates include environmental damage and replication errors; deamination, and depurination are specific types of damage.

Specific Mutagenic Events

Deamination

• Removal of amine groups from nucleotides leads to:

  • Cytosine converting to Uracil, resulting in a substitution error as Uracil pairs with Adenine rather than Guanine.

  • Other bases can be similarly modified, creating different pairing partners and resulting in substitutions or coding changes based on position in codons.

  • Methylated Cytosine can change pairing due to deamination, affecting the mutation type experienced.

Depurination

• Removal of purine bases (Adenine, Guanine) can cause replication machinery to skip bases leading to deletions in the new DNA strand, likely causing frameshift mutations in coding sequences.

Oxidative Damage

• Caused by reactive oxygen species (e.g., hydrogen peroxide) leading to direct changes in nucleotide pairing, causing substitutions or blockage of the replication process by modifying Thymine to Thymine glycol.

Alkylation and UV Damage

• Alkaline solutions can damage DNA bases and the sugar-phosphate backbone leading to a range of possible outcomes.

• UV light exposure can cause thymine dimers, leading to replication errors and potential carcinogenic mutations due to alterations in 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.

Cancer Cells and DNA Damage

• The cells of cancer are constantly replicating, making the mutagens used to induce mutations a targeted method of chemotherapy, impacting rapidly dividing cancer cells more than resting normal cells.

Mismatch Repair Mechanism

• Mismatch repair occurs after DNA replication, significantly decreasing the error rate from 1 in 10^7 to 1 in 10^9 bases:

  • MutS detects mismatches based on recognition of unpaired bases.

  • MutL coordinates the response, and MutH makes cuts in the unmethylated newly synthesized strand.

  • Exonucleases remove the incorrectly incorporated sequence, and DNA polymerase fills in the gap before ligation.

• This process helps maintain integrity and accuracy of the genetic code despite replication errors.