Figure 17–18 illustrates some of the molecular details underlying the control of the two steps in the initiation of DNA replication. A key player is a large multiprotein complex called the origin recognition complex (ORC), which binds to
Figure 17–17 Control of chromosome duplication. preparations for dna replication begin in late mitosis and g1, when the dna helicases are loaded by multiple proteins at the replication origin, forming the prereplicative complex (preRC). s-Cdk activation leads to activation of the dna helicases, which unwind the dna at origins to initiate dna replication. Two replication forks move out from each origin until the entire chromosome is duplicated. duplicated chromosomes are then segregated in m phase. s-Cdk activation in s phase also prevents assembly of new preRCs at any origin until the following g1—thereby ensuring that each origin is activated only once in each cell cycle.
replication origins throughout the cell cycle. In late mitosis and early G1, the proteins Cdc6 and Cdt1 collaborate with the ORC to load the inactive DNA helicases around the DNA next to the origin. The resulting large complex is the preRC, and the origin is now licensed for replication.
At the onset of S phase, S-Cdk triggers origin activation by phosphorylating specific initiator proteins, which then nucleate the assembly of a large protein complex that activates the DNA helicase and recruits the DNA synthesis machinery. Another protein kinase called DDK is also activated in S phase and helps drive origin activation by phosphorylating specific subunits of the DNA helicase.
At the same time as S-Cdk initiates DNA replication, several mechanisms prevent assembly of new preRCs. S-Cdk phosphorylates and thereby inhibits the ORC and Cdc6 proteins. Inactivation of the APC/C in late G1 also helps turn off preRC assembly. In late mitosis and early G1, the APC/C triggers the destruction of a Cdt1 inhibitor called geminin, thereby allowing Cdt1 to be active. When the APC/C is turned off in late G1, geminin accumulates and inhibits the Cdt1 that is not associated with DNA. Also, the association of Cdt1 with a protein at active replication forks stimulates Cdt1 destruction. In these various ways, preRC formation is prevented from S phase to mitosis, thereby ensuring that each origin is fired only once per cell cycle. How, then, is the cell-cycle control system reset to allow replication in the next cell cycle? At the end of mitosis, APC/C activation leads to the inactivation of Cdks and the destruction of geminin. ORC and Cdc6 are dephosphorylated and Cdt1 is activated, allowing preRC assembly to prepare the cell for the next S phase.
Chromosome duplication Requires duplication of Chromatin structure
The DNA of the chromosomes is extensively packaged in a variety of protein components, including histones and various regulatory proteins involved in the control of gene expression (discussed in Chapter 4). Thus, duplication of a chromosome is not simply a matter of replicating the DNA at its core but also requires the duplication of these chromatin proteins and their proper assembly on the DNA.
The production of chromatin proteins increases during S phase to provide the raw materials needed to package the newly synthesized DNA. Most importantly, S-Cdks stimulate a large increase in the synthesis of the four histone subunits that form the histone octamers at the core of each nucleosome. These subunits are assembled into nucleosomes on the DNA by nucleosome assembly factors, which typically associate with the replication fork and distribute nucleosomes on both strands of the DNA as they emerge from the DNA synthesis machinery.
Chromatin packaging helps to control gene expression. In some parts of the chromosome, the chromatin is highly condensed and is called heterochromatin, whereas in other regions it has a more open structure and is called euchromatin (discussed in Chapter 4). These differences in chromatin structure depend on a variety of mechanisms, including modification of histone tails and the presence of non-histone proteins. Because these differences are important in gene regulation,