(8) DNA Repair

DNA Replication and Repair Mechanisms

DNA Polymerases and Proofreading

  • DNA Polymerases: Enzymes responsible for DNA replication that possess proofreading ability, which minimizes base-pair misincorporation (mutations).

  • Despite proofreading, some misincorporations still occur due to the vast number of bases involved (approximately 3.2 billion in the human genome).

DNA Damage During Replication

  • Causes of DNA Damage:

    • Bases can become exposed to water when the DNA duplex is unwound during replication, leading to potential damage.

    • Types of Damage:

      • Depurination: The glycosidic bond in A and G bases is vulnerable, leading to loss of these bases.

      • Deamination: Alters bases, changing cytosine to uracil, etc.

      • Environmental Mutagens: Such as UV light and radiation can induce DNA damage.

    • T-T Dimers: Formed by UV light, they block replication and transcription.

Mutation and Genetic Variation

Significance of Mutations

  • Limited mutations can lead to genetic variation within a population, providing raw material for natural selection and evolution.

  • Most mutations within an individual are typically non-advantageous and can lead to deleterious effects.

DNA Repair Mechanisms

  • DNA Repair Enzymes include:

    • Excision Repair: Repairs damaged bases like those from deamination and depurination.

    • Mismatch Repair: Corrects mis-paired nucleotides post-replication.

    • Double Strand Break Repair: Repair mechanisms for double-stranded breaks.

Excision Repair Details

  • Damage Covered: Addresses deamination, depurination, and T-T dimers.

  • Process Steps:

    • Repair Endonuclease: Cuts the DNA on either side of the damage (typically within 29 bp).

    • DNA Helicase: Unwinds the DNA between nicks.

    • DNA Polymerase: Fills the gap using the complementary strand as a template.

    • DNA Ligase: Seals the nicks to restore DNA integrity.

Xeroderma Pigmentosum

  • Cause: Mutations in nucleotide excision repair enzymes.

  • Consequence: Individuals with this condition are highly susceptible to UV-induced skin cancer.

Mismatch Repair Process

  • Function of Mismatch Repair Enzymes: Fix incorrectly paired nucleotides that are common after DNA replication.

  • Key Proteins:

    • MutS: Scans the genome and binds to mismatched base pairs.

    • MutL/MutH: Identify nearby nicks and unmethylated cytosines to guide repair.

    • Exonucleases: Excise the flanking bases for correction.

    • DNA Polymerase & Ligase: Refill the gap and seal nicks, respectively.

Implications of Mismatch Repair Loss

  • Loss of mismatch repair proteins leads to genome-wide mutations, contributing to diseases such as Hereditary Colon Cancer (HNPCC), where individuals inherit a defective copy of the MutS or MutL/H gene.

Double Strand Breaks and Repair Mechanisms

  • Repair Pathways:

    • Non-Homologous End Joining (NHEJ): Directly joins broken DNA ends.

    • Homologous Recombination (HR): Uses a homologous sequence to repair breaks, often involving proteins like BRCA1.

Proper Chromosome Replication

  • Successful DNA replication results in two identical sister chromatids, preparing for mitosis:

    • DNA molecules consist of original and newly synthesized strands.

Cell Cycle and Mitosis

  • DNA Replication Phase: Precedes mitosis; the cell cycle is tightly controlled to check for DNA damage and ensure proper repair.

Phases of Mitosis

  1. Prophase: Chromosomes condense; spindle apparatus begins to form.

  2. Metaphase: Chromosomes align at the metaphase plate; spindle fibers attach.

  3. Anaphase: Sister chromatids are pulled apart to opposite poles.

  4. Telophase & Cytokinesis: Nuclear envelopes form around each set of chromatids; cytoplasm divides.

Role of Proteins in Chromosome Structure

  • Condensin: Compacts sister chromatids into chromosomes during prophase.

    • Functions: Requires ATP; involved in maintaining chromosome integrity.

  • Cohesin Complexes: Tether sister chromatids together, ensuring equitable distribution during anaphase.

Microtubule Dynamics During Mitosis

  • Spindle apparatus composed of three types of microtubules includes:

    • Kinetochore Microtubules: Attach to chromosomes and aid in separation.

    • Polar Microtubules: Push apart during anaphase.

    • Astral Microtubules: Anchor to the plasma membrane for further separation.

Chromosome Movement in Anaphase

  • Movement is mediated by motor proteins such as dynein which pulls chromosomes towards poles, complemented by the depolymerization of kinetochore microtubules.

Stabilization of Spindle Microtubules

  • Spindle microtubules stabilize when:

    • Bound to kinetochores.

    • Tension is equalized across microtubules.

Aneuploidy and Consequences

  • Aneuploidy describes abnormal chromosome separation leading to altered DNA content in daughter cells:

    • Examples include Turner Syndrome (monosomy X) and Down’s Syndrome (Trisomy 21).

Cytokinesis

  • Occurs after mitosis and is characterized by:

    • Formation of a contractile ring via actin microfilaments.

    • Cleavage furrow divides the cell along the spindle equator, distributing organelles equally between daughter cells.

Cell Cycle Variability

  • The length of the cell cycle varies based on cell type and environmental factors.