Cell Cycle Control of DNA Replication

Replication of DNA in S phase is subject to strict control in the cell cycle (review Stillman, 1996), resulting in the following observations  

A main control of DNA replication occurs at the level of initiation of replication. The replication of a DNA sequence starts at specific sequence sections of the DNA, known as replication origins. In yeast, the initiation sites of DNA replication have been very well defined and characterized at the sequence level. In higher eucaryotes, in contrast, initiation has a broad initiation zone and it has not been possible to identify a defined initiation sequence. The size of the genome in eucaryotes necessitates the use of many replication origins, which can be activated in a defined time sequence and position-specific order. 

The most important components in cell cycle control of origin activity have been identified, although the questions of the spatial coordination and time sequence of origin activity are stiU imanswered. 

Control of origin activity occiu s via specific protein complexes that are boimd at certain times of the cell cycle to a replication origin. For replication initiation, two states of this protein complex are important, known as the pre-rephcation complex and the post-replication complex. 

The pre-replication complex (pre-RC) is formed diming anaphase and is inherited by the sister chromatids. Upon entry into S phase, the pre-RC must be disrupted for initiation to occur. If initiation begins, the pre-RC changes to the post-rephcation complex state, which does not permit further initiation. 

---

D Urso, G., Marraccino, R. L., Marshak, D. R., and Roberts, J. M. (1990). Cell cycle control of DNA replication by a homologue from human cells of the p34cprotein kinase. Science 250 786-791. 

Stillman, B. Cell cycle control of DNA replication (1996) Science 274,1659-1664... 

Stillman, B. Cell Cycle Control of DNA Replication. Science 274, 1659-1663 (1996). [A description of how eukaryotic replication is controlled and linked to cell division.]... 

Nishitani, H. lygerou, Z. (2002) Control of DNA replication licensing in a cell cycle. Genes Cells 7, 523-534. 

Hutchison, C. J., Cox, R., and Ford, C. C. (1988). The control of DNA replication in a cell-free extract that recapitulates a basic yield cycle in vitro. Development (Cambridge, UK) 103,553-566. 

Minichromosome maintenance protein 2 (MCM2) is involved in the control of DNA replication. The expression ofMCM2 starts in early Gi and is maintained throughout the cell cycle. MCM2 also has been shown to represent a suitable marker for cell proliferation [12, 16]. Aside from using markers that were expressed by the dividing cells, the visualization of experimentally administered bromodeoxyuridine (BrdU an S phase-specific marker, which incorporates into the DNA) allows the detection of newly formed cells. BrdU administration is mainly performed by intraperitoneal injection. 

Enoch, T and Nurse, P. (1990). Mutation of the fission yeast cell cycle control genes abolishes dependence of mitosis on DNA replication. Cell 60 665-673. 

However, cells of Chlamydomonas treated with ethidium bromide are able later to regenerate their chloroplast DNA. This result has been interpreted to mean that there may be one or a few "master copies" of chloroplast DNA in specially protected locations. The result should also be considered in relationship to the following observation. Although nuclear and organelle DNA molecules replicate at different times in the cell cycle, constant proportions of the organelle and nuclear DNA tend to be maintained. Thus, there must be some kind of control mechanism leading to a coupling of DNA replication in nuclei, mitochondria, and chloroplasts.465... 

Figure 24.24. Retinoblastoma and cell cycle. Rb regulates the movement of cells through Gi into the S phase of the cell cycle. Regulation of Gi — S phase by Rb represents a key control point in the cell cycle because once the cell has passed the restriction point in Gi (a point in late G, the cell moves into S phase and is now irreversibly committed to replicate its DNA and progress through the entire cell cycle. In early GI phase, Rb binds to E2F and blocks E2F s ability to transcribe S-phase genes. Phosphorylation of Rb by Cdk4/6 and Cdk2 releases E2F from Rb and allows E2F to transcribe S-phase genes. The cell then enters S phase. (Adapted S. Ortega et al. Biochimica et Biophysica Acta. 1602, 73-87, 2002.)...

Ribonucleotide reductase plays a critical role in the life cycle of all living organisms. By catalysing the conversion of ribonucleotides to deoxyribonucleotides, it holds a unique position at the biological crossroads between RNA synthesis and DNA synthesis. Its control of DNA synthesis and cell proliferation is mediated both by providing all precursors for replication and by keeping a balanced supply between them. Throughout the years, the metabolic key-role of RNR has been explored successfully in a variety of antiviral and antiproliferative therapies. 

Figure 1. Cellular response to DNA damage. Sensor proteins are able to detect the presence of DNA damage and alert the cellular machinery so that the cell can face the stress and protects itself. The sensor then activates a transducer and through a series of proteins activation by phosphorylations, DNA integrity checkpoints (red arrow), in charge ofthe cell cycle control are activated and play their role which is to stop the cell cycle progression In order to allow repair to fully operate. Indeed, DNA replication should not occur on damaged DNA because of possible mistakes when copying modified bases, so the G l/S interphase checkpoint prevents the cell to enter into S-phase and to start replication. The G2 checkpoint verifies DNA for the presence of double strand breaks and incomplete replication, so that an altered DNA copy is not transmitted to the daughter cell.

---