G1 phase (G stands for gap) spans from the end of mitosis to the start of S phase. During G1 phase, cells check the external environment and the intracellullar state, and decide if they continue with the cell cycle or not. In multicellular organisms, the cell cycle progress is mainly influenced by extracellular signals, like cell adhesion, or molecules released by other cells, such as trophic factors. There is also internal information about the state of the cell that affects the progress of the cell cycle, like the health condition of the cell, if there is a correct number of cellular components after cell division, or if chromosomes were properly segregated. When external and internal signals are right, proliferating cells grow and get ready to enter S phase.
However, cells of multicellular organisms do not proliferate, but quit the cell cycle from G1 phase, either transiently or permanently. Stopping the cell cycle means that cells are going to differentiate, stay quiescent, go through senescence processes, or die by apoptosis (Figure 1). When cells remain in a quiescent state, it is said that they are in G0 phase. From G0 and differentiated states, some cell types are able to restart the cell cycle. Entering in quiescent stage means the expression of a particular set of genes and the repression of those genes that promote differentiation, senescence or apoptosis. Quiescent cells repressed genes that promote the cell cycle. From apoptosis or senescence, cells cannot return to the cell cycle. Thus, these are four possible decisions that cells may take during G1 phase, which depends on molecular complexes known as checkpoints. Cells must pass these checkpoints before entering S phase. If cells do not pass one of them, it is said that a decision has been taken. However, if cells do not stop in any of them, S phase will eventually start. This is the default process. The molecular mechanisms at the checkpoints have to be fast, complete, and irreversible.
Checkpoints are molecular complexes that control the progress of the cell cycle. At the core of checkpoints there are cyclin-dependent kinases (CdKs). Nine different CdKs have being found in eukaryotes. To be active, CdKs need to bind a regulatory protein known as cyclin, and be phosphorylated as well. Once activated, CdKs phosphorylate several substrates, such as inhibitors of the cell cycle, thus allowing the progress of the cell cycle. Cyclins are proteins synthesized periodically during the cell cycle. There are 16 different cyclins in eukaryote cells. Cyclins A, B, D, and E have more impact in the progress of the cell cycle. Cyclins D (there are 3) and E (there are 2) are important for G1 phase progress. CdK4/cyclin D (D/CDK4) and CdK2/cyclin E (E/CDK2) phosphorylate the transcription factor retinoblastoma, which is part of the last checkpoint of the G1 phase.
The checkpoint where retinoblastoma is phosphorylated is known as restriction point because, if this checkpoint is passed, cells immediately start the S phase. The concept of restriction point was introduced by A. Pardee in 1974. It is very important because there is no turning-back, once S phase is started, cells will divide and no external signals will stop the cell cycle. To reach the restriction point during G1 phase, cells need extracellular signals like mitogens, but they are not needed once this restriction point is passed. The sequence is as follows. Mitogens activate plasma membrane receptors that stimulate Ras-GTPases, which promote the expression of genes of transcription factors like c-Myc and others. Transcription factors boost the expression of cyclin D that in turn activate CdK4 and 6, which phosphorylate retinoblastome proteins that release the E2F factor (Figure 2).
At the core of the restriction point are CdK-cyclin, retinoblastoma and E2F (Figure 2). Retinoblastome is not phosphorylated during early G1 phase. In this form, retinoblastoma is bound to E2F and inhibits the expression of genes that promote the cell cycle progress. Retinoblastome is first phosphorylated by CdK4/cyclin D and then by CdK2/cyclin E, both in G1 phase. Retinoblastome have 16 phosphorylation sites, which indicates how complex is the regulation. Phosphorylation of all sites is successive, and each CdK phosphorylates specific sites. It seems that when 14 sites are phosphorylated, the affinity of retinoblastome for E2F decreases and, therefore, E2F factor may promote the expression of genes that favor the beginning of S phase.
In this molecular framework, the signals carrying information about the environment (nutrients, trophic signals, and others), DNA damages, or about the proper size for division are integrated. If everything is correct, the cell passes the restriction point and S phase begins. If something is wrong, there are several types of inhibitors that stop the cell cycle progress. One of them is p53, which is activated in many tumors. When DNA is damaged, or the cell is under stress, or there are changes in the pH, or any other potentially dangerous situations for the cell are present, p53 is over-expressed and activates the p21 gen, which in turn inhibits the phosphorylation of retinoblastoma, so that the cell cycle is arrested and does not start the S phase.
As we mentioned above, most cells of a multicellular organism are not in permanent proliferation. This is because there are inhibitors of Cdk/cyclins in G1 phase that make cells quit the cell cycle.
Blomen VA, Boonstra J. 2007. Cell fate determination during G1 phase progression. Cellular and molecular life sciences. 64:3084-3104.
Matson JP, Cook JG. 2017. Cell cycle proliferation decisions: the impact of single cell analyses. The FEBS journal. 284: 362-375.