The Cell Cycle: Regulation and Division CSC 858 3/17/05 Dr. Andrew Bieberich Dr. Michael Goldman Reading: Tozeren and Byers Chapter 7 For April 7, read Chapter 3 Copyright Andrew Bieberich 2005

Ultimately, cells decide whether to divide or not based upon input of information from outside the cell. single cells (yeast, bacteria) Are there enough nutrients? Are toxic waste molecules too concentrated to proceed without cell damage? cells within multi-cell organisms Is the cell attached to others? Is it too crowded? Are the correct growth factors present?

Regulation of the multi-celled eukaryotic cell cycle 1. Semi-modular control system. 2. Five major checkpoints that act as switches in the system. 3. Cyclin and Cyclin and Cyclin Growth factors coordinate cell cycles across multiple cells. 5. G 0, when time stands still. 6. Cancer: when switches malfunction. G1 START M S DNA replication checkpoint DNA damage checkpoint G2 metaphase checkpoint degredation of metaphase cyclins

Across cell types, the cell cycle may take minutes, months, or arrest indefinitely. Therefore, we know that something sophisticated must be controlling it. A “clock” is not flexible. Phase triggering is only slightly more so. The system required is one that uses switches controlled by subsystems with feedback. G1 START M S DNA replication checkpoint DNA damage checkpoint G2 metaphase checkpoint degredation of metaphase cyclins

G1 Chromosomes become uncondensed. Also called ‘G0’ if cell arrests in this state. START M daughter cells S DNA replication checkpoint DNA damage checkpoint DNA is replicated. G2 Chromosomes condense, Topoisomerase II helps to untangle them. metaphase checkpoint degredation of metaphase cyclins (check before entering G1)

Growth factors and integrin-mediated singalling pathways both affect production of cyclin D in the nucleus. cell surface receptor nuclear membrane nuclear pore growth factor integrin bound to ECM Rb E2F signal transduction pathways with many phosphorylation steps

Growth factors and integrin-mediated singalling pathways both affect production of cyclin D in the nucleus. cell surface receptor nuclear membrane nuclear pore growth factor integrin bound to ECM signal transduction pathways with many phosphorylation steps Rb E2F CDK4 cyclin D cyclin D gene expression

Rb CDK4 cyclin D E2F Turn on genes for DNA replication in S phase. Turn on gene for cyclin E and cyclin A Rb CDK2 cyclin E / A Progression past START is reached when the feedback loop increases the rate at which Rb is phosphorylated and DNA replication genes are turned on by E2F.

Rb CDK2 cyclin A Cyclin A-CDK2, in addition to increasing the rate at which Rb is phosphorylated, also begins phosphorylating (and thus activating) the DNA Replication Complex.

CDK2 cyclin A Activated DNA RC binds to replication fork and recruits DNA Pol III When activated DNA RC begins to bind replication forks, the G1 - S phase transition is complete.

S phase: all DNA in all chromosomes must be replicated and checked for damage. DNA Pol epsilon detects presence of replication forks- check before exiting S phase. Many proteins (and thus many genes) are involved in checking for damaged DNA at this point. Apoptosis may occur if damage is too great. p53 BRCA1 regulatory cross-talk

Apoptosis can be thought of as “programmed cell death.” A set of molecular machinery kept in reserve for this purpose is turned on, and the cell auto-destructs by digesting all of its components. Loss of function of p53, or other genes that encode damage checking proteins, can be a “pre-cancerous” condition. Cells that divide without checking for DNA damage may replicate mutant forms of cell cycle regulating genes. These cells may in turn become the progenitors of tumors.

ribosomes mitochondria histones centrosomes/centrioles Cell components other than DNA must be replicated also during S phase.

John Kyrk’s webpage has an illustration of primary DNA packaging with histones, and this serves as a powerful suplement to our discussion of G2 phase. You may see this by clicking this link: As is mentioned in Tozeren and Byers, not much is known about the molecular signalling that controls timing of G2. We know that S phase ends when the DNA replication and DNA damage checkpoints are passed, and mitosis (M phase) is considered to have begun when chromosomes are fully condensed. During G2, in between S and M, DNA topoisomerase II works to untangle the uncondensed chromosomes so that they may condense separately.

Mitosis: Tozeren and Byers explain the separate phases of mitosis completely, so it is not necessary to simply repeat what they said. However, you should make sure that you are familiar with what happens during each of these phases: Prophase Metaphase Anaphase Telophase You will notice that not everyone describes these in exactly the same way. (For instance, John Kyrk lists ‘Prometaphase’ as an extra step. I’ve never seen that before.) The basic events that take place are as described in Chapter 7 of Tozeren and Byers, so you may go by what they have written. To see John Kyrk’s mitosis animation:

centrosome microtubule paired sister chromatids aligned along metaphase plate There is a checkpoint at metaphase of mitosis. The sensor proteins detect whether tension is exerted on centromeres of all chromatid pairs. centromere

After the metaphase checkpoint sensory system detects perfect alignment, anaphase begins. The MT motors pull each chromatid along its microtubule, and the microtubule disassembles as it is pulled toward the kinetochore.

Growing animal cells under artificial culture conditions: Normal cells will usually not divide more than ~10 times under culture conditions, so primary cultures of recently isolated cells do not last long. Alternatively, cell lines that are immortal may be used- these are often cells containing genetic mutations that cause cancer-like growth, and so are less accurate model systems for “normal” cell activity.