⚡ Core Concept: High-Fidelity Semiconservative Replication & Genome Integrity
DNA replication is semiconservative, bidirectional, and semidiscontinuous. Polymerases synthesize solely in the $5′
ightarrow 3’$ direction, demanding coordinated continuous leading-strand synthesis alongside discontinuous lagging-strand synthesis of Okazaki fragments, coupled to multi-tiered DNA repair pathways maintaining error rates below $10^{-9}$.
1. The Eukaryotic Replication Fork & Polymerase Division of Labor
Initiation begins at Origins of Replication with the binding of the Origin Recognition Complex (ORC) and loading of the hexameric CMG (Cdc45-Mcm2-7-GINS) helicase during $G_1$, activated by S-CDK and DDK kinases during S-phase:
Figure 2: Molecular Architecture of the Asymmetric Bidirectional Replication Fork
Molecular Genetics
Figure 2: Leading and lagging strand replication fork machinery: CMG/DnaB helicase unwinding, topoisomerase supercoil relaxation, Pol delta/epsilon extension, and SSB/RPA stabilization.
2. The Three Primary Eukaryotic Replicative Polymerases
| Polymerase | Primary Strand / Function | Exonuclease Activity ($3′ ightarrow 5’$) | Processivity Factor |
|---|---|---|---|
| DNA Pol α / Primase | Initiates synthesis: synthesizes ~10 nt RNA primer + ~20 nt initiator DNA | None (No proofreading) | Low (dissociates after priming) |
| DNA Pol ε | Continuous synthesis of the Leading Strand | Intrinsic $3′ ightarrow 5’$ proofreading | High (associated with CMG helicase) |
| DNA Pol δ | Discontinuous synthesis of the Lagging Strand (Okazaki) | Intrinsic $3′ ightarrow 5’$ proofreading | PCNA homotrimeric sliding clamp |
3. Major DNA Repair Pathways & Human Disease Syndromes
- Mismatch Repair (MMR): MSH2/MSH6 detects replication slippage base mismatches; MLH1/PMS2 nicks the nascent strand. Defect: Hereditary Nonpolyposis Colorectal Cancer (Lynch Syndrome).
- Nucleotide Excision Repair (NER): XPA–XPG excises bulky UV-induced cyclobutane pyrimidine dimers (CPDs) and 6-4 photoproducts via a 24–32 nt oligonucleotide excision. Defect: Xeroderma Pigmentosum.
- Base Excision Repair (BER): DNA glycosylases (e.g., Uracil DNA Glycosylase) remove deaminated bases, creating an AP site cleaved by AP endonuclease 1 (APE1).
- Double-Strand Break Repair: Homologous Recombination (HR, error-free using sister chromatid via BRCA1/2, Rad51) vs. Non-Homologous End Joining (NHEJ, error-prone direct ligation via Ku70/80 and DNA-PKcs).
Telomere Biology & The End-Replication Problem
Because removal of the terminal lagging-strand RNA primer leaves an unreplicated 3′ overhang, linear chromosomes shorten with each cycle. Telomerase (a ribonucleoprotein reverse transcriptase with an internal TERC RNA template and TERT catalytic subunit) elongates telomeric TTAGGG repeats, conferring replicative immortality to germ cells and >85% of human cancers.
