Restriction point

The restriction point (R) is a point in G1 of the animal cell cycle at which the cell becomes 'committed' to the cell cycle and after which extracellular proliferation stimulants are no longer required. The restriction point (R) is a point in G1 of the animal cell cycle at which the cell becomes 'committed' to the cell cycle and after which extracellular proliferation stimulants are no longer required. Originally, Howard Martin Temin showed that chicken cells reach a point at which they are committed to replicate their DNA and are not dependent on extracellular signals. About 20 years later, in 1973, Arthur Pardee demonstrated that a single restriction point exists in G1. Previously, G1 had been defined simply as the time between mitosis and S phase. No molecular or morphological place-markers for a cell's position in G1 were known. Pardee used a double-block method in which he shifted cells from one cell cycle block (such as critical amino acid withdrawal or serum withdrawal) to another and compared each block's efficiency at preventing progression to S phase. He found that both blocks in all cases examined were equally efficient at blocking S phase progression, indicating that they must all act at the same point in G1, which he termed the 'restriction point', or R-point. In 1985, Zetterberg and Larsson discovered that, in all stages of the cell cycle, serum deprivation results in inhibition of protein synthesis. Only in postmitotic cells (i.e. cells in early G1) did serum withdrawal force cells into quiescence (G0). In fact, Zetterberg found that virtually all of the variability in cell cycle length can be accounted for in the time it takes the cell to move from the restriction point to S phase. Except for early embryonic development, most cells in multicellular organisms persist in a quiescent state known as G0, where proliferation does not occur, and cells are typically terminally differentiated; other specialized cells continue to divide into adulthood. For both of these groups of cells, a decision has been made to either exit the cell cycle and become quiescent (G0), or to reenter G1. A cell's decision to enter, or reenter, the cell cycle is made before S-phase in G1 at what is known as the restriction point, and is determined by the combination of promotional and inhibitory extracellular signals that are received and processed. Before the R-point, a cell requires these extracellular stimulants to begin progressing through the first three sub-phases of G1 (competence, entry G1a, progression G1b). After the R-point has been passed in G1b, however, extracellular signals are no longer required, and the cell is irreversibly committed to preparing for DNA duplication. Further progression is regulated by intracellular mechanisms. Removal of stimulants before the cell reaches the R-point may result in the cell's reversion to quiescence. Under these conditions, cells are actually set back in the cell cycle, and will require additional time (about 8 hours more than the withdrawal time in culture) after passing the restriction point to enter S phase. Mitogen Signaling Growth factors (e.g., PDGF, FGF, and EGF) regulate entry of cells into the cell cycle and progression to the restriction point.  After passing this switch-like “point of no return,” cell cycle completion is no longer dependent on the presence of mitogens.   Sustained mitogen signaling promotes cell cycle entry largely through regulation of the G1 cyclins (cyclin D1-3) and their assembly with Cdk4/6, which may be mediated in parallel through both MAPK and PI3K pathways. MAPK Signaling Cascade The binding of extracellular growth factors to their receptor tyrosine kinases (RTK) triggers a conformational change and promotes dimerization and autophosphorylation of tyrosine residues on the cytoplasmic tail of the RTKs. These phosphorylated tyrosine residues facilitate the docking of proteins containing an SH2-domain (e.g., Grb2), which can subsequently recruit other signaling proteins to the plasma membrane and trigger signaling kinase cascades. RTK-associated Grb2 binds Sos, which is a guanine nucleotide exchange factor that converts membrane-bound Ras to its active form (Ras-GDP ⟶ {displaystyle longrightarrow } Ras-GTP). Active Ras activates the MAP kinase cascade, binding and activating Raf, which phosphorylates and activates MEK, which phosphorylates and activates ERK (also known as MAPK, see also MAPK/ERK pathway).