12.8 Regulation of the Cell Cycle by Protein Kinases in Chapter 12 Biochemical Signaling

12.8 Regulation of the Cell Cycle by Protein Kinases

One of the most dramatic manifestations of signaling pathways is the regulation of the eukaryotic cell cycle. During embryonic growth and later development, cell division occurs in virtually every tissue. In the adult organism, cells of most tissues stop dividing, becoming quiescent. A cell’s “decision” to divide or not is of crucial importance to the organism. When the regulatory mechanisms that limit cell division are defective and cells undergo unregulated division, the result is catastrophic — cancer. Proper cell division requires a precisely ordered sequence of biochemical events that assures every daughter cell a full complement of the molecules required for life. Investigations into the control of cell division in diverse eukaryotic cells have revealed universal regulatory mechanisms. Signaling mechanisms much like those discussed above are central in determining whether and when a cell undergoes cell division, and they also ensure orderly passage through the stages of the cell cycle.

The Cell Cycle Has Four Stages

Cell division accompanying mitosis in eukaryotes occurs in four well-defined stages (Fig. 12-35). In the S (synthesis) phase, the DNA is replicated to produce copies for both daughter cells. In the G2 phase (G indicates the gap between divisions), new proteins are synthesized and the cell approximately doubles in size. In the m phase (mitosis), the maternal nuclear envelope breaks down, paired chromosomes are pulled to opposite poles of the cell, each set of daughter chromosomes is surrounded by a newly formed nuclear envelope, and cytokinesis pinches the cell in half, producing two daughter cells (see Fig. 24-22). In embryonic or rapidly proliferating tissue, each daughter cell divides again, but only after a waiting period (G1). In animal cells in the laboratory, the entire process takes about 24 hours.

A piece graph shows descriptions and ties of the four stages of the eukaryotic cell cycle: G 1, S, G 2, and M. — FIGURE 12-35 The eukaryotic cell cycle. The durations (in hours) of the four stages vary, but those shown are typical.

The G 1 phase is at the 1 o’clock position to almost the 6 o’clock positions and is 6 to 12 h. Accompanying text reads, G 1 phase: R N A and protein synthesis. No D N A synthesis. A short bar crosses the graph at just past the 5 o’clock position and is labeled, Restriction point: A cell that passes this point is committed to pass into S phase. The S phase is from almost the 6 o’clock position to just past the 10 o’clock position and is labeled 6 to 8 h. Accompanying text reads, S phase: D N A synthesis doubles the amount of D N A in the cell. R N A and protein are also synthesized. The G 2 phase is at just past the 10 o’clock position to the 12 o’clock position and is 3 to 4 h. Accompanying text reads, G 2 phase: No D N A synthesis. R N A and protein synthesis continue. The M phase is at the 12 o’clock to 1 o’clock positions and is 1 h. Accompanying text reads, M phase: Mitosis (nuclear division) and cytokinesis (cell division yield two daughter cells. Arrows pointing left and right from just past the 1 o’clock position indicate that the cell can transition to G 0 at this point. Accompanying text reads, G 0 phase: Terminally differentiated cells withdraw from cell cycle indefinitely. Additional text reads, Reentry point: A cell returning from G 0 enters at early G 1 phase.

After passing through mitosis and into G1, a cell either continues through another division or ceases to divide, entering a quiescent phase (G0) that may last hours, days, or the lifetime of the cell. When a cell in G0 begins to divide again, it reenters the division cycle through the G1 phase. Differentiated cells such as hepatocytes or adipocytes have acquired their specialized function and form; they remain in the G0 phase. Stem cells retain their potential to divide and to differentiate into any of a number of cell types.

Levels of Cyclin-Dependent Protein Kinases Oscillate

The timing of the cell cycle is controlled by a family of protein kinases with activities that change in response to cellular signals. By phosphorylating specific proteins at precisely timed intervals, these protein kinases orchestrate the metabolic activities of the cell to produce orderly cell division. The kinases are heterodimers with a regulatory subunit, a cyclin, and a catalytic subunit, a cyclin-dependent protein kinase (CDK). In the absence of the cyclin, the catalytic subunit is virtually inactive. When the cyclin binds, the catalytic site opens up, a residue essential to catalysis becomes accessible, and the protein kinase activity of the catalytic subunit increases 10,000-fold. Animal cells have at least 10 different cyclins (designated A, B, and so forth) and at least 8 CDKs (CDK1 through CDK8), which act in various combinations at specific points in the cell cycle.

In a population of animal cells undergoing synchronous division, some CDK activities show striking oscillations (Fig. 12-36). These oscillations are the result of four mechanisms for regulating CDK activity: phosphorylation or dephosphorylation of the CDK, controlled degradation of the cyclin subunit, periodic synthesis of CDKs and cyclins, and the action of specific CDK-inhibiting proteins. The precisely timed activation and inactivation of a series of CDKs produces signals serving as a master clock that orchestrates the events in normal cell division and ensures that one stage is completed before the next begins.

A graph plots kinase activity against time and shows four curves with respect to the stages of the eukaryotic cell cycle: cyclin D-C D K 4-6, cyclin E- C D K 2, cyclin A- C D K 2, and cyclin B- C D K 1. — FIGURE 12-36 Variations in the activities of specific CDKs during the cell cycle in animals. Early in G1, the activity of cyclin D–CDK4-6 rises slowly, then drops sharply as G1 ends. Near the end of G1, cyclin E–CDK2 activity rises and peaks near the G1 phase–S phase boundary, when the active enzyme triggers synthesis of enzymes required for DNA synthesis (see Fig. 12-40). Cyclin A–CDK2 activity rises during the S and G2 phases, then drops sharply in the M phase, as cyclin B–CDK1 peaks. [Data from P. Icard et al., *Trends Biochem. Sci.* 44:490, 2019, Fig. 3.]

The horizontal axis is labeled time and the vertical axis is labeled kinase activity. The horizontal axis is divided into five color-coded ranges labeled G 1, S, G 2, M, and G 1. G 1 extends across the first quarter of the graph, S extends from one-quarter to one-half of the distance across the horizontal axis, and G 2 extends from one-half to three-quarters of the distance across the horizontal axis. The remaining quarter of the horizontal axis is divided in half with M on the left and G 1 on the right. The orange curve for cyclin D-C D K 4-6 begins just past the start of the horizontal axis at the horizontal axis and rises to peak at almost one-quarter of the horizontal axis and three-quarters of the height of the vertical axis in G 1, then decreases almost vertically to end at the horizontal axis at just past one-quarter of its length at the beginning of the S stage. The blue curve for cyclin E- C D K 2 begins at the horizontal axis at about halfway through the G 1 stage and almost immediately begins to rise rapidly to peak at the boundary between the G 1 and S stages just below the peak for cyclin D- C D K 4-6 before dropping symmetrically to end at the horizontal axis at about one-third of its length in the first half of the S stage. The green curve for cyclin A- C D K 2 begins at the horizontal axis just before the start of the S stage and rises relatively slowly as almost a diagonal line to half the height of the vertical axis at the border between the S and G 2 stages at the midpoint of the horizontal axis, after which it rises very slowly to peak at just past three-quarters of the length of the horizontal axis and just below the peak of cyclin E- C D K 2 at the beginning of the M stage. The red curve for cyclin B- C D K 1 begins at the horizontal axis halfway through the S stage and rises very slowly to about one-quarter of the way through the G 2 stage, after which it rises more rapidly to peak just below the peak for cyclin A- C D K 2 at about half the height of the vertical axis just before the start of the right-hand region representing the G 1 stage. It then drops rapidly to reach the horizontal axis at the boundary between the M stage and the right-hand region representing the G 1 stage. All data are approximate.

CDKs Are Regulated by Phosphorylation, Cyclin Degradation, Growth Factors, and Specific Inhibitors

The activity of a CDK is strikingly affected by phosphorylation and dephosphorylation of two specific residues in the protein (Fig. 12-37). Phosphorylation of ${Thr}^{160}$ $Thr Superscript 160$ of CDK2 stabilizes a conformation in which an autoinhibitory “T loop” is moved away from the substrate-binding cleft in the kinase, opening it to bind protein substrates. Dephosphorylation of - ${Tyr}^{15}$ $Tyr Superscript 15$ of CDK2 removes a negative charge that blocks ATP from approaching its binding site. This mechanism for activating a CDK is self-reinforcing; the enzyme (PTP) that dephosphorylates - ${Tyr}^{15}$ $Tyr Superscript 15$ is itself a substrate for the CDK and is activated by phosphorylation. The combination of these factors activates the CDK manyfold, allowing it to phosphorylate downstream protein targets required for progression of the cell cycle (Fig. 12-38a).

A three-part figure shows the activation of cyclin-dependent protein kinases (C D Ks) by showing the crystal structure of a inactive C D K 2 without the cyclin in part a, the changed conformation of the crystal structure of inactive C D K 2 with bound cyclin in part b, and the effects of phosphorylation of a T h r residue in the T loop to produce active C D K 2 in part c. — FIGURE 12-37 Activation of cyclin-dependent protein kinases (CDKs) by cyclin and phosphorylation. CDKs are active only when associated with a cyclin. The crystal structure of CDK2 with and without a cyclin reveals the basis for this activation. (a) Without the cyclin, CDK2 folds so that one segment, the T loop, obstructs the binding site for protein substrates. The binding site for ATP is also near the T loop and is blocked when ${Tyr}^{15}$ $Tyr Superscript 15$ is phosphorylated (not shown). (b) When the cyclin binds, it forces conformational changes that move the T loop away from the active site and reorient an amino-terminal helix, bringing a residue critical to catalysis $({Glu}^{51})$ $left-parenthesis Glu Superscript 51 Baseline right-parenthesis$ into the active site. **(c)** When a Thr residue in the T loop is phosphorylated, its negative charges are stabilized by interaction with three Arg residues, holding the T loop away from the substrate-binding site. Removal of the phosphoryl group on ${Tyr}^{15}$ $Tyr Superscript 15$ gives ATP access to its binding site, fully activating CDK2 (see Fig. 12-38). [Data from (a) PDB ID 1HCK, U. Schulze-Gahmen et al., *J. Med. Chem.* 39:4540, 1996; (b) PDB ID 1FIN, P. D. Jeffrey et al., *Nature* 376:313, 1995; (c) PDB ID 1JST, A. A. Russo et al., *Nature Struct. Biol.* 3:696, 1996.]

FIGURE 12-37 Activation of cyclin-dependent protein kinases (CDKs) by cyclin and phosphorylation. CDKs are active only when associated with a cyclin. The crystal structure of CDK2 with and without a cyclin reveals the basis for this activation. (a) Without the cyclin, CDK2 folds so that one segment, the T loop, obstructs the binding site for protein substrates. The binding site for ATP is also near the T loop and is blocked when ${Tyr}^{15}$ $Tyr Superscript 15$ is phosphorylated (not shown). (b) When the cyclin binds, it forces conformational changes that move the T loop away from the active site and reorient an amino-terminal helix, bringing a residue critical to catalysis $({Glu}^{51})$ $left-parenthesis Glu Superscript 51 Baseline right-parenthesis$ into the active site. **(c)** When a Thr residue in the T loop is phosphorylated, its negative charges are stabilized by interaction with three Arg residues, holding the T loop away from the substrate-binding site. Removal of the phosphoryl group on ${Tyr}^{15}$ $Tyr Superscript 15$ gives ATP access to its binding site, fully activating CDK2 (see Fig. 12-38). [Data from (a) PDB ID 1HCK, U. Schulze-Gahmen et al., *J. Med. Chem.* 39:4540, 1996; (b) PDB ID 1FIN, P. D. Jeffrey et al., *Nature* 376:313, 1995; (c) PDB ID 1JST, A. A. Russo et al., *Nature Struct. Biol.* 3:696, 1996.]

Part a, labeled C D K 2 (inactive) shows the surface contour of a white protein that resembles two roughly spherical pieces joined by a slightly narrower center. Along the upper right of the left-hand sphere, there is a green helix that runs at a slight diagonal from the upper left of the right side of the sphere to its lower right, near the upper center of the sphere. This green helix is labeled amino-terminal helix and has a green wireframe model representing G l u superscript 51 that extends from its right center into the opening between the two halves above the center of the protein. Beneath the right end of the green helix, there is a light blue wireframe model containing a sphere that is labeled T y r superscript 15. White threads extend to the lower left and lower right from the left end of T y r superscript 15. A red thread begins above the red piece at the right end of T y r superscript 15, loops over the red piece at the right end of T y r superscript 16, loops to the left next to the left end of T y r superscript 15, and then runs back to the right in front of the upper part of a roughly spherical yellow space-filling model of A T P before looping down slightly to form a T loop just to the right of the center of the molecule before ending. Part b shows the surface contour of a protein with a roughly oval cyclin subunit on top and a roughly oval C D K (inactive) below. The two pieces are joined at the left and center, leaving a small space between the left and center pieces and an opening between them on the right side. A green helix is present in the opening between the upper left and center pieces of the subunits and is angled slightly down to the right as it extends away from the viewer. A green wireframe piece labeled G l u superscript 51 extends slightly to the left below it, where it is near the red end of a light blue wireframe structure labeled T y r superscript 15 that has a white piece extending down almost to a roughly spherical yellow space-filling model of A T P below. A red strand begins in the upper right center of the bottom subunit and runs up to just to the right of the green helix, at which point it forms a loop that runs up from the bottom subunit toward the upper right and into the top subunit. Part c, labeled C D K 2 (active) has a similar structure to the protein in part b except that the T y r superscript 15 and the T loop have changed orientation and a phosphate is attached to the T loop. A close-up of this region shows the green helix on the left with a purple wireframe model of A r g superscript 50 extending up to the right to end close to a phosphorylated T h r superscript 160. The green helix also has a green wireframe model labeled G l u superscript 51 extending down past a red piece at the end of the light blue T y r and behind the yellow spheres of A T P below. The red strand begins to the right of the yellow spheres of A T P, loops slightly toward the G l u superscript 51, then runs up behind the phosphorylated T h r superscript 160, then loops up over a blue helix of the upper subunit that runs from the lower left to upper right, then curves almost vertically along a vertical white helix with the phosphorylated T h r superscript 160 shown as a wireframe with a phosphate that extends from this vertical portion toward the left. The top diagonal helix of the upper subunit has the beginning of another diagonal subunit near its upper left side and the white vertical helix has another vertical helix to its right.

A two-part figure shows the regulation of C D K by phosphorylation in eight steps with activation shown in part a as steps 1 through 5 and inactivation shown in part b as steps 6 through 8. — FIGURE 12-38 Regulation of CDK by phosphorylation and proteolysis. **(a)** The series of events that leads to activation of a cyclin-dependent protein kinase. **(b)** The periodic proteolytic degradation of cyclin, inactivating the cyclin-dependent protein kinase. The steps are described in the text.

The steps are shown in a cycle with the top half labeled part a and the bottom half labeled part b. Part a begins with a blue rectangle labeled C D K at the boundary between the top and bottom halves. Step 1: No cyclin present; C D K is inactive. A light red structure that resembles an oval with flat sides labeled cyclin is added to an arrow pointing up and then right. Step 2: Cyclin synthesis leads to its accumulation. A blue rectangle labeled C D K at the top center of the cycle has cyclin fitted into a concavity on its lower side and a wedge labeled T y r at its upper left corner that has a line extending to a red circle labeled P to its upper left. Step 3: Cyclin-C D K complex forms, but phosphorylated T y r superscript 15 blocks A T P-binding site; still inactive. An arrow points to a similar molecule with a second wedge labeled T h r in the upper right corner with a line connecting it to a green circle labeled P to the right. Step 4: Phosphorylation of T h r superscript 160 in T loop and dephosphorylation of T y r superscript 15 activates cyclin-C D K manyfold. An arrow points down with an arrow branching off to show the loss of P subscript i. This produces a blue rectangle labeled C D K at the boundary between the upper and lower halves with the left-hand wedge labeled T h r gone, the right-hand wedge labeled T h r attached to a green circle labeled P that is now to its upper right instead of directly to its right, and short lines radiating from its top and sides. A dashed arrow points to the left to an arrow pointing from a green oval labeled phosphatase to a green oval labeled phosphatase with a line extending down to a green circle labeled P at the lower right and with short radiating lines extending from all sides. Step 5: C D K phosphorylates phosphatase, which activates more C D K. A dashed arrow points back to the start of the arrow between C D K with T y r bonded to a red circle labeled P and T h r bonded to a green circle labeled P down, with a loss of P subscript 1, to activated C D K without T y r. Part b begins with two arrows pointing from activated C D K. A dashed horizontal arrow points to the arrow from a purple oval labeled D B R P to a similar purple oval with a line to a green circle labeled P to the lower right and with short radiating lines all around it. Step 6: C D K phosphorylates D B R P, activating it. A dashed arrow points from activated D B R P to the second arrow from activated C D K. This arrow is joined by a line from U, showing its addition, and points to a blue rectangle labeled C D K at the bottom center of the figure that no longer has T h r, still has short radiating lines from its upper half, and has a chain of four U attached to the cyclin bound to its bottom side. Step 7: D B R P triggers addition of ubiquitin molecules to cyclin by ubiquitin ligase. An arrow points to the left and then up to the starting C D K at the boundary between the top and bottom halves. Before it reaches C D K, this arrow has an arrow pointing to the left to a dashed red outline of the shape of cyclin. Step 8: Cyclin is degraded by proteasome, leaving C D K inactive.

The presence of a single-strand break in DNA signals arrest of the cell cycle in G2 by activating two proteins (ATM and ATR; see Fig. 12-40). These proteins trigger a cascade of responses that include inactivation of the PTP that dephosphorylates ${Tyr}^{15}$ $Tyr Superscript 15$ of the CDK. With the CDK inactivated, the cell is arrested in G2, unable to divide until the DNA is repaired and the effects of the cascade are reversed.

Highly specific and precisely timed proteolytic breakdown of mitotic cyclins regulates CDK activity throughout the cell cycle (Fig. 12-38b). How is the timing of cyclin breakdown controlled? A feedback loop occurs in the overall process shown in Figure 12-38. As a cell enters mitosis, the M-phase CDK is inactive (step ). As cyclin is synthesized (step ), the cyclin-CDK complex forms (step ). The T loop lies in the substrate-binding site of CDK, and - ${Tyr}^{15}$ $Tyr Superscript 15$ blocks its ATP-binding site, keeping the complex inactive. When ${Thr}^{160}$ $Thr Superscript 160$ in the T loop is phosphorylated, the loop moves out of the substrate-binding site, and when ${Tyr}^{15}$ $Tyr Superscript 15$ is dephosphorylated, ATP can bind. These two changes make the cyclin-CDK complex many times more active (step ). Further activation is achieved as CDK also phosphorylates and activates the enzyme that dephosphorylates - ${Tyr}^{15}$ $Tyr Superscript 15$ (step ). The active cyclin-CDK complex triggers its own inactivation by phosphorylation of DBRP (destruction box recognizing protein; step ). DBRP and ubiquitin ligase then attach several molecules of ubiquitin (U) to the cyclin (step ), targeting it for destruction by proteolytic enzyme complexes called proteasomes (step ). The role of ubiquitin and proteasomes is not limited to the regulation of cyclins; as we shall see in Chapter 27, both also take part in the turnover of cellular proteins, a process fundamental to cellular housekeeping.

The third mechanism for changing CDK activity is regulation of the rate of synthesis of the cyclin or CDK or both. Extracellular signals such as growth factors and cytokines (developmental signals that trigger cell division) activate, by phosphorylation, the nuclear transcription factors Jun and Fos, which promote the synthesis of many gene products, including cyclins, CDKs, and the transcription factor E2F. In turn, E2F stimulates production of several enzymes essential for the synthesis of deoxynucleotides and DNA, and the CDK and cyclin allow the cell to enter the S phase (Fig. 12-39).

A figure shows how cell division is regulated by growth factors. — FIGURE 12-39 Regulation of cell division by growth factors.

Finally, specific protein inhibitors bind to and inactivate specific CDKs. One such protein is p21, which we discuss below.

These four control mechanisms modulate the activity of specific CDKs that, in turn, control whether a cell will divide, differentiate, become permanently quiescent, or begin a new cycle of division after a period of quiescence. The details of cell cycle regulation, such as the number of different cyclins and kinases and the combinations in which they act, differ from species to species, but the basic mechanism has been conserved in the evolution of all eukaryotic cells.

CDKs Regulate Cell Division by Phosphorylating Critical Proteins

We have examined how cells maintain close control of CDK activity, but how does the activity of CDKs control the cell cycle? There are scores of known CDK targets, and much remains to be learned. But we can see a general pattern behind CDK regulation by inspecting the effect of CDKs on the structures of lamin and myosin and on the activity of retinoblastoma protein.

The structure of the nuclear envelope is maintained in part by highly organized meshworks of intermediate filaments composed of the protein lamin. Breakdown of the nuclear envelope before segregation of the sister chromatids in mitosis is partly due to the phosphorylation of lamin by a CDK, which causes lamin filaments to depolymerize.

A second kinase target is the ATP-driven contractile machinery (actin and myosin) that pinches a dividing cell into two equal parts during cytokinesis. After the division, a CDK phosphorylates a small regulatory subunit of myosin, causing dissociation of myosin from actin filaments and inactivating the contractile machinery. Subsequent dephosphorylation allows reassembly of the contractile apparatus for the next round of cytokinesis.

A third and very important CDK substrate is the retinoblastoma protein, pRb; when DNA damage is detected, this protein participates in a mechanism that arrests cell division in G1 (Fig. 12-40). Named for the retinal tumor cell line in which it was discovered, pRb functions in most, perhaps all, cell types to regulate cell division in response to a variety of stimuli. Unphosphorylated pRb binds the transcription factor E2F; while bound to pRb, E2F cannot promote transcription of a group of genes necessary for DNA synthesis (the genes for DNA polymerase α, ribonucleotide reductase, and other proteins; see Chapter 25). In this state, the cell cycle cannot proceed from the G1 phase to the S phase, the step that commits a cell to mitosis and cell division. The pRb-E2F blocking mechanism is relieved when pRb is phosphorylated by cyclin E–CDK2, which occurs in response to a signal for cell division to proceed.

A figure shows how the phosphorylation of p R b, the retinoblastoma protein, regulates the passage of cells from G 1 to S. — FIGURE 12-40 Regulation of passage from G1 to S by phosphorylation of pRb. Transcription factor E2F promotes transcription of genes for certain enzymes essential to DNA synthesis. The retinoblastoma protein, pRb, can bind E2F (lower left), inactivating it and preventing transcription of these genes. Phosphorylation of pRb by CDK2 prevents it from binding and inactivating E2F, and the genes are transcribed, allowing cell division. Damage to the cell’s DNA (upper left) triggers a series of events that inactivate CDK2, blocking cell division. When the protein MRN detects damage to the DNA, it activates two protein kinases, ATM and ATR, and they phosphorylate and activate the transcription factor p53. Active p53 promotes the synthesis of another protein, p21, an inhibitor of CDK2. Inhibition of CDK2 stops the phosphorylation of pRb, which therefore continues to bind and inhibit E2F. With E2F inactivated, genes essential to cell division are not transcribed and cell division is blocked. When DNA has been repaired, this inhibition is released, and the cell divides.

The figure has two halves. The left-hand side is labeled cell division blocked by p 53 and the right-hand side is labeled cell division occurs normally. The left-hand side begins with a double-stranded piece of D N A that is broken in the middle, where a yellow oval labeled M R N is attached. M R N is below the space between the broken pieces and attached to the lower left and lower right ends of D N A on either side of the break. The right-hand side begins with an intact double helix of D N A. On the left side, an arrow points down from M R N to a gray oval labeled A T M, A T R, from with an arrow points to a red circle labeled p 53 that has short radiating lines around it. Accompanying text reads, active. An arrow below p 53 is accompanied by text reading, transcriptional regulation leads to upward arrow symbol [p 21]. The arrow points to a half-circle with a rectangular protrusion from its lower left. An arrow points down from p 21 to join an arrow pointing from active C D K 2 on the right-hand side to inactive C D K 2 on the left-hand side. Active C D K 2 is a blue rectangle labeled C D K 2 with short radiating lines on the left, top, and right and a light red partial oval with flat ends labeled cyclin E. Inactive C D K 2 has p 21 fitted into onto its right side and no short radiating lines. Below inactive C D K 2, there is an orange half-circle labeled p R b with an irregular shape to its flat side. Two arrows point from p R b. One arrow points to the right and is joined on the right-hand side by a dashed arrow from active C D K 2 to point to a smooth orange half-circle labeled p R b with a bond to a red circle labeled P to its upper right. The second arrow points down to join an arrow from active E 2 F on the right-hand side to inactive E 2 F on the left-hand side. Inactive E 2 F is a purple rectangle with an irregular shape to its top side that fits into the irregular bottom of p R b, which is above it. E 2 F active is a similar purple rectangle with short radiating lines extending from all sides. An arrow labeled transcriptional regulation points right from active E 2 F to text reading, enzymes for D N A synthesis, from which an arrow points down to text reading, passage from G 1 to S.

When the protein kinases ATM and ATR detect damage to DNA (signaled by the presence of the protein MRN at a double-strand break site), they phosphorylate p53, activating it to serve as a transcription factor that stimulates the synthesis of the protein p21 (Fig. 12-40). This protein inhibits the protein kinase activity of cyclin E–CDK2. In the presence of p21, pRb remains unphosphorylated and bound to E2F, blocking the activity of this transcription factor, and the cell cycle is arrested in G1. This gives the cell time to repair its DNA before entering the S phase, thereby avoiding the potentially disastrous transfer of a defective genome to one or both daughter cells. When the damage is too severe to allow effective repair, this same machinery triggers apoptosis (described below), a process that leads to the death of the cell, preventing the possible development of a cancer.

SUMMARY 12.8 Regulation of the Cell Cycle by Protein Kinases

Progression through the cell cycle is regulated by the cyclin-dependent protein kinases (CDKs), which act at specific points in the cycle, phosphorylating key proteins and modulating their activities. The catalytic subunit of CDKs is inactive unless associated with the regulatory cyclin subunit.
The activity of a cyclin-CDK complex changes during the cell cycle through differential synthesis of CDKs, specific degradation of the cyclin, phosphorylation and dephosphorylation of critical residues in CDKs, and binding of inhibitory proteins to specific cyclin-CDKs.
A cyclin sequence (the destruction box) marks cyclin for tagging with ubiquitin and degradation in proteasomes. The rise of cyclin concentration by its synthesis ultimately triggers its degradation, yielding oscillations in cyclin level keyed to the cell cycle.
Cells receive extracellular signals that determine the timing of their division. Scores of proteins are known targets of CDKs, many with unknown functions. Among the targets phosphorylated by cyclin-CDKs are proteins of the nuclear envelope and proteins required for cytokinesis and DNA repair.