DNA Replication and Repair Mechanisms

Class Logistics

• ### Microphone Update
  • Old microphone not functional; new microphone is more amplified.
• ### Exam Information
  • Exams will be conducted in person and online.
  • Students must come to class to receive a passport for exam access.
  • Exams can be accessed through laptops, cell phones, or tablets (recommend to use a fully charged device with internet connection).
  • Participants need to sign an Excel sheet with their signature and grade upon completion.
  • Early finishers can leave after signing the sheet.
  • Collaboration encouraged among students for assignments and exam preparation.

Previous Class Recap

• Topics Reviewed: DNA replication and epigenetic changes.
  • Importance of understanding DNA polymerase, primase, helicase, clamp proteins, and clamp loader in DNA replication process.
  • Importance of clarity on these concepts for understanding current class topics.

Current Class Objectives

• ### Main Question Addressed:
  • "How stable is the DNA?"
  • Mechanisms of DNA damage repair in case of mutations or errors.

Types of DNA Mutations

• ### Types of Mutations
  1. Incorporation of incorrect base during DNA replication:
  • Transitions and transversions.
  • DNA polymerase efficiency: can incorporate nucleotides rapidly but is not error-free.
  1. Chemical changes induced by external factors:
  • Three categories of changes:
    • Spontaneous (intrinsic to the DNA): Examples include incorporation of non-standard bases.
    • Chemical exposure (e.g., carcinogenics): Can adulterate DNA structure.
    • Radiation exposure (e.g., UV light, gamma, X-ray): High-energy light can disrupt DNA structure and function.

The Efficiency of DNA Polymerase

• ### Error Rates
  • DNA polymerase introduces approximately one incorrect base per 100,000 bases added, forming mismatches.
  • The human genome contains approximately $6 imes 10^9$ bases.
  • Each cell division may result in about 120,000 mismatch pairs, but only about 6 mismatches remain in daughter cells due to proofreading activity.

Proofreading Activity of DNA Polymerase

• DNA polymerase can recognize errors it makes, backtrack, and replace mismatched nucleotides.
• Mechanism operates effectively to correct approximately 99% of introduced errors.

Overview of Mutational Causes

• ### Types of Mutations
  • Spontaneous Mutations: Occur without external stimuli, characterized by base elimination.
  • Example:
    • Cytosine loses an amine group, forming uracil (not present in DNA).
    • Adenine loses an amine group, producing hypoxanthine.
  • Apurine Sites (Apocytes): Absence of purine due to nitrogen base elimination.
  • Alkylation: Addition of methyl group to guanine, leading to mispairing with thymine.
  • Induced Mutations: Caused by external factors like UV radiation leading to thymine dimers.
  • UV light causes covalent bond formation between adjacent thymine residues.
  • Result: DNA replication can be halted due to structural distortion, posing cancer risk.

Mechanisms of DNA Repair

Repair Scenarios

1. Permanent mutation in genome if DNA is not repaired.
2. Excision and repair of the template strand leading to partial repair.
3. Only new strands are repaired via excision.

Types of DNA Repair Mechanisms

1. Direct Reversal of Damage: Enzymatic correction of spontaneous mutations and UV-induced damage.

• Examples include photoreactivation in prokaryotes (not present in eukaryotes) via enzyme photolyase; nicked thymine dimers corrected by light activation.

1. Excision Repair: Removes and replaces damaged DNA segments, categorized further into:

   • Base Excision Repair (BER): Removes single, damaged bases.
     • Utilizes glycosylases for base removal, followed by endonucleases, DNA polymerase replacement, and ligation.
   • Nucleotide Excision Repair (NER): Removes longer DNA segments containing damage.
     • Two forms:
     • Global Genomic NER: Constant surveillance by proteins like XPC.
     • Transcription-Coupled NER: Repair occurs if RNA polymerase is stalled at a damaged site.

2. Mismatch Repair: Corrects mismatched bases that escape proofreading. It involves:

   • MutS and MutL proteins in prokaryotes identify and excise mismatches, followed by replacement by DNA polymerase.
   • Genomic implications include association with hereditary cancers like Lynch syndrome.

3. Translesion Synthesis: A mechanism that permits replication across damaged DNA without correction. It involves specialized polymerases, allowing cells to continue division despite errors.

Conclusion

Take-Home Messages

1. Importance of recognizing specific DNA damage mechanisms and respective repair pathways for maintaining genomic integrity.
2. Spontaneous versus induced DNA mutations and their implications in diseases such as cancer.
3. Understanding key roles of DNA polymerases throughout replication and repair processes, along with the implications of repair mechanisms for genetic stability and longevity of cellular function.