DNA Replication (cell cycle overview)

Cell Cycle

• The cell cycle describes a cell's life, growth, DNA replication, and division.
• Essential for growth, repair, and reproduction in multicellular organisms.
• Two main phases: Interphase and M phase (mitotic phase).

Interphase

• A phase of growth and preparation for division, subdivided into three stages:
  • G1 Phase (Gap 1): Cell growth, protein synthesis, energy production; checks readiness for DNA replication.
  • S Phase (Synthesis): DNA replication – duplicates each chromosome.
  • G2 Phase (Gap 2): Further growth, protein synthesis; checks DNA replication accuracy.
• G0 Phase: A resting phase where some cells remain until signals trigger re-entry into the cell cycle.

M Phase (Mitotic Phase)

• Involves cell division, splitting one cell into two identical daughter cells.
  • Mitosis: Separates duplicated DNA into two identical sets. Divided into four substages:
    • Prophase: Chromosomes condense, spindle fibers form.
    • Prometaphase: Nuclear envelope breaks down, spindle fibers attach to chromosomes.
    • Metaphase: Chromosomes align in the center of the cell.
    • Anaphase: Sister chromatids separate to opposite poles.
    • Telophase: Chromatids arrive at poles, new nuclear membranes form.
  • Cytokinesis: Cytoplasm divides, forming two identical daughter cells.

DNA Replication

• The process of duplicating DNA before cell division, ensuring each new cell gets a complete set.
• Semi-conservative: Each new DNA molecule has one original and one new strand.

DNA Structure

• DNA is a double helix, like a twisted ladder.
• Nucleotides are the building blocks.
• Nucleotide Structure: Sugar (deoxyribose), phosphate group, nitrogenous base (A, T, C, G).
• Base Pairing: A with T, C with G.
• 5’ and 3’ ends: DNA strands have directionality; replication occurs 5' to 3'.

DNA Polymerase

• Enzymes that synthesize new DNA strands by adding nucleotides to an existing strand.
• 5' to 3' direction.
• Proofreading ability reduces errors.
• DNA Polymerase III (prokaryotes): Main enzyme for DNA synthesis; highly processive, meaning it can catalyze thousands of base pairings at a time.

Primase

• An enzyme that synthesizes RNA primers needed for DNA polymerase to initiate DNA synthesis.

DNA Replication Steps

• Initiation: DNA unwinding at origins of replication (one in prokaryotes, multiple in eukaryotes).
• Unwinding: Helicase unwinds the DNA. SSBPs stabilize the strands. Topoisomerase prevents supercoiling.
• Primer Binding and Synthesis: Primase adds short RNA primers.
• Elongation: DNA polymerase extends new strands from primers, synthesizing continuously (leading strand) and discontinuously (lagging strand) using Okazaki fragments.
• DNA Ligase joins Okazaki fragments on the lagging strand.

DNA Proofreading and Repair

• Proofreading by DNA polymerase corrects errors during replication.
• Repair mechanisms: Mismatch repair, Base excision repair (BER), Nucleotide excision repair (NER), and Double-strand break repair.

Telomeres and Cellular Aging

• Telomeres: Repetitive DNA sequences at chromosome ends, protecting from degradation during cell division.
• Telomerase: Enzyme lengthening telomeres in some cells, leading to continued cell division but not in most somatic cells (leading to senescence).

DNA Replication Models

• Semi-conservative model: Correct model; each DNA molecule has one new and one old strand.
• Meselson-Stahl experiment: Confirmed semi-conservative model using isotopes.
• Conservative and dispersive models were disproven.

Replisome

• Complex of proteins (helicase, primase, DNA polymerases, sliding clamp) coordinating DNA replication.

Key DNA Terminology

• Terms like chromosome, chromatid, centromere, sister chromatids, mitotic spindle, base pairing, hydrogen bonds, replication origin, template strand, antiparallel structure, leading strand, lagging strand, primers, proofreading, mutations, exonuclease activity, Okazaki fragments, DNA ligase, semi-conservative replication, replication bubble, telomeres, senescence, eukaryotes, prokaryotes, replication fork, telomerase.

Histones

• Positively charged proteins that bind to negatively charged DNA, allowing tight coiling.
• Five main types (H1, H2A, H2B, H3, H4) with specific roles in DNA organization.