Replication licensing--defining the proliferative state?

Abstract

The proliferation of eukaryotic cells is a highly regulated process that depends on the precise duplication of chromosomal DNA in each cell cycle. Regulation of the replication licensing system, which promotes the assembly of complexes of proteins termed Mcm2-7 onto replication origins, is responsible for preventing re-replication of DNA in a single cell cycle. Recent work has shown how the licensing system is directly controlled by cyclin-dependent kinases (CDKs). Repression of origin licensing is emerging as a ubiquitous route by which the proliferative capacity of cells is lowered, and Mcm2-Mcm7 proteins show promise as diagnostic markers of early cancer stages. These results have prompted us to propose a functional distinction between the proliferative state and the non-proliferative state (including G0) depending on whether origins are licensed.

Figure 1

Replication licensing and CDK activity through the cell cycle. A small segment of chromosomal DNA is depicted at different cell cycle stages, with bound (red hexagons, M) or unbound (blue hexagons, M) Mcm2-7. The activity of the replication licensing system (RLS, purple) and cyclin-dependent kinases (CDK, green) at the different stages are show in the central circle. Inset, the sequence of events occurring as each origin binds Mcm2-7 and becomes licensed. The loading of ORC (purple), Cdt1 (yellow) and Cdc6 (green), and then Mcm2-7 (red, M) is shown. At some time after licensing is complete, ORC, Cdc6 and Cdt1 may become inactivated, as indicated by grey shading.

Figure 2

Different routes by which cellular DNA content may increase. A small segment of chromosomal DNA is depicted at different cell cycle stages. Red hexagons show Mcm2-7 bound to origins in G1 and S. Outer ring (black arrows): the normal sequence of cell cycle events. Red arrows show routes by which cellular DNA content is increased (endocycles and origin re-firing). CDK activity in S, G2 and M is denoted by green colouring.

Figure 3

Pathways for CDK inhibition of origin licensing. The ways that CDKs inhibit the activity of ORC, Cdc6, Cdt1 and Mcm2-7 in S. cerevisiae, S. pombe, Xenopus and humans.

Figure 4

Mcm2 protein expression in differentiating colonic epithelial cells. Indirect immunoperoxidase staining of colon with an anti-Mcm2 antibody. Anatomical regions containing stem cells, rapidly proliferating cells and terminally differentiated cells are indicated. Reproduced from ref. with permission.

Figure 5

Entry and exit of cycling cells into quiescence and terminal differentiation. On the left is a cartoon of cells passing through four phases of the cell division cycle. The presence of licensed origins is shown in red, and the presence of active CDKs is shown in green. When cells pass from G1 into G0 or terminal differentiation, origins become unlicensed. When G0 cells are stimulated to re-enter G1 their origins become re-licensed. It is currently unclear whether CDK reactivation of the E2F transcription system is required for this re-licensing to occur.