Eukaryotic DNA Replication

DNA replication follows a semiconservative mechanism: within the double helix, the two parental strands separate from each other, and each individual parental strand then serves as the template on which a brand-new, complementary strand is synthesised, so that every resulting daughter DNA molecule is a hybrid conserving exactly one of the two original parental strands. In eukaryotes, replication does not start from a single point; it begins simultaneously at many distinct chromosomal sites called origins of replication -- the yeast Saccharomyces cerevisiae, for example, has roughly 400 such origins. Each origin is activated by the assembly of a large, fourteen-protein prereplication complex, part of which is a six-protein subassembly called the origin recognition complex (ORC), which functions as the actual initiator of replication; in yeast, the origins themselves are specific DNA sequences called ARS elements (Autonomously Replicating Sequences), to which ORC binds directly. The physical site where the parental double helix is being actively unwound and the two new daughter strands are being synthesised is called the replication fork; because eukaryotic chromosomes have many origins, many replication forks are active simultaneously along one chromosome. At each fork, the enzyme helicase unwinds the double helix by breaking the hydrogen bonds between the two parental strands, while replication protein A (RPA) coats the newly separated single strands to stop them from reannealing to each other prematurely, and topoisomerase relieves the positive supercoiling that unwinding generates just ahead of the fork by transiently breaking and resealing DNA's covalent backbone. Because DNA polymerase can only extend a strand from a free 3' hydroxyl end, replication must first be primed: the enzyme DNA polymerase alpha, acting together with primase, lays down a short stretch of RNA primer on both strands. DNA synthesis then proceeds only in the 5' to 3' direction, which has an important consequence for the two strands: the leading strand, whose synthesis runs in the same direction as the fork itself is moving, can be synthesised continuously, while the lagging strand, whose synthesis must run in the direction opposite to fork movement, must instead be synthesised discontinuously, in short pieces called Okazaki fragments (discovered by Reiji Okazaki and colleagues in the 1960s), which DNA ligase subsequently joins together into one continuous strand by forming a phosphodiester bond between a 3' hydroxyl and a 5' phosphate group at each remaining nick. Three distinct DNA polymerase enzymes carry out nuclear DNA replication in eukaryotes: DNA polymerase alpha synthesises the short RNA primers; DNA polymerase delta is the main replicating enzyme of the cell nucleus; and DNA polymerase epsilon extends the DNA strands at the replication fork (a separate enzyme, DNA polymerase beta, plays no role in normal replication at all, and instead removes incorrect bases from damaged DNA during base excision repair). The raw material and energy for the whole process come from the four deoxyribonucleoside triphosphates, dATP, dGTP, dCTP and dTTP, which serve both as the literal building-block substrates incorporated into the new strand and as the energy source that drives the polymerisation reaction forward. In plants specifically, telomere maintenance differs from the pattern seen in vertebrate somatic cells: …