1. 8. Lecture WS 2010/11Cellular Programs1 Already simple genetic circuits can give rise to oscillations. E.g., a negative feedback loop X  R ─┤ X can yield oscillations (X activates R, which inhibits X, so that R goes down, so that X goes back up...). Such a circuit requires significant non- linearity or a time delay to keep from rapidly settling to a constant steady state. An oscillator of this sort is thought to be the core of many eukaryotic cell cycles. V8: Cell cycle control Oikonomou & Cross, Curr. Opin. Genet Devel. 20, 605 (2010) FrederickCatherine Cross,Oikonomou Rockefeller University Oscillatory networks underlie the - circadian clock, - the beating of our hearts, and - the cycle of cell division, which creates two cells from one, driving the reproduction and development of living systems.

2. 8. Lecture WS 2010/11Cellular Programs2 Cell cycle control system Tyson et al., Curr.Opin.Cell.Biol. 15, 221 (2003) APC: anaphase-promoting complex, Cdk1, Wee1, Cdc25:kinases CKI: cyclin-dependent kinase inhibitor

3. 8. Lecture WS 2010/11Cellular Programs3 cell-cycle machinery Central components of the cell-cycle machinery are cyclin-dependent kinases (such as CDK1/ CDC2). Their sequential activation and inactivation govern cell- cycle transitions. The activity of CDK1/CDC2 is low (off) in the G1 phase and has to be high (on) for entry into mitosis (M phase). Tyson et al., Curr.Opin.Cell.Biol. 15, 221 (2003)

4. 8. Lecture WS 2010/11Cellular Programs4 A negative feedback loop can give rise to oscillations. Here, such an oscillator forms the core of eukaryotic cell cycles. Cyclin–CDK acts as activator, and APC- Cdc20 acts as repressor. Non-linearity in APC-Cdc20 activation prevents the system from settling into a steady state. Positive and negative feedback loops in the cyclin– CDK oscillator of eukaryotic cells Oikonomou & Cross, Curr. Opin. Genet Devel. 20, 605 (2010) - CDKs require the binding of a cyclin subunit for activity. These cyclin partners can also determine the localization of the complex and its specificity for targets. - At the beginning of the cell cycle, cyclin–CDK activity is low, and ramps up over most of the cycle. Early cyclins trigger production of later cyclins and these later cyclins then turn off the earlier cyclins, so that control is passed from one set of cyclin–CDKs to the next. - The last set of cyclins to be activated, the G2/M-phase cyclins, initiate mitosis, and also initiate their own destruction by activating the APC-Cdc20 negative feedback loop. APC-Cdc20 targets the G2/M-phase cyclins for destruction, resetting the cell to a low-CDK activity state, ready for the next cycle.

5. 8. Lecture WS 2010/11Cellular Programs5 mutual inhibition: toggle switch Tyson et al., Curr.Opin.Cell.Biol. 15, 221 (2003) S: signalE: enzyme R: responseEP: phosphorylated form of enzyme This bifurcation is called toggle switch („Kippschalter“): if S is decreased enough, the switch will go back to the off-state. For intermediate stimulus strengh (S crit1 < S < S crit2 ), the response of the system can be either small or large, depending on how S was changed. This is often called „hysteresis“.

6. 8. Lecture WS 2010/11Cellular Programs6 Cell cycle control system Tyson et al., Curr.Pin.Cell.Biol. 15, 221 (2003) signal: concentration of Cdk1:CycB response: free Cdk1/CycB The G1/S module is a toggle switch, based on mutual inhibition between Cdk1-cyclin B and CKI, a stoichiometric cyclin-dependent kinase inhibitor.

7. 8. Lecture WS 2010/11Cellular Programs7 Cell cycle control system Tyson et al., Curr.Pin.Cell.Biol. 15, 221 (2003) The G2/M module is a second toggle switch, based on mutual activation between Cdk1-cyclinB and Cdc25 (a phosphotase that activates the dimer) and mutual inhibition between Cdk1-cyclin B and Wee1 (a kinase that inactivates the dimer).

8. 8. Lecture WS 2010/11Cellular Programs8 Cell cycle control system Tyson et al., Curr.Pin.Cell.Biol. 15, 221 (2003) The M/G1 module is an oscillator, based on a negative-feedback loop: Cdk1-cyclin B activates the anaphase-promoting complex (APC) by phosphorylating it. This activates Cdc20, which degrades cyclin B. The „signal“ that drives cell proliferation is cell growth: a newborn cell cannot leave G1 and enter the DNA synthesis/division process (S/G2/M) until it grows to a critical size.

9. 8. Lecture WS 2010/11Cellular Programs9 Positive feedback is added to the oscillator in multiple ways. A highly conserved but non-essential mechanism consists of ‘handoff’ of cyclin proteolysis from APC- Cdc20 to APC-Cdh1. Cdh1 is a relative of Cdc20 which activates APC late in mitosis and into the ensuing G1. Cdh1 is inhibited by cyclin–CDK activity, resulting in mutual inhibition (which is logically equivalent to positive feedback). Positive feedback in the cyclin–CDK oscillator Oikonomou & Cross, Curr. Opin. Genet Devel. 20, 605 (2010)

10. 8. Lecture WS 2010/11Cellular Programs10 Size control in S. pombe in G2 phase Oikonomou & Cross, Curr. Opin. Genet Devel. 20, 605 (2010) Pom1, localized to cell poles, indirectly inhibits CDK activity (through inhibition of Cdr2, which inhibits Wee1, which in turn inhibits CDK). As the cell elongates, the concentration of Pom1 at the center of the cell (where the nucleus is located) drops, allowing CDK activation leading to mitosis.

11. 8. Lecture WS 2010/11Cellular Programs11 Coupling of multiple cellular oscillators Oikonomou & Cross, Curr. Opin. Genet Devel. 20, 605 (2010) Schematic of multiple peripheral oscillators coupled to the CDK oscillator in budding yeast. Coupling entrains such peripheral oscillators to cell cycle progression; peripheral oscillators also feed back on the cyclin–CDK oscillator itself. E.g. major genes in the periodic transcription program include most cyclins, CDC20, and CDC5. Cdc14 directly promotes establishment of the low-cyclin–CDK positive feedback loop by activating Cdh1 and Sic1 as well as more indirectly antagonizing cyclin– CDK activity by dephosphorylating cyclin–CDK targets. The centrosome and budding cycles could communicate with the cyclin–CDK cycle via the spindle integrity and morphogenesis checkpoints.

12. 8. Lecture WS 2010/11Cellular Programs12 Phase locking of cellular oscillators Oikonomou & Cross, Curr. Opin. Genet Devel. 20, 605 (2010)

13. 8. Lecture WS 2010/11Bioinformatics III13 Role of protein complexes Cell cycle proteins that are part of complexes or other physical interactions are shown within the circle. For the dynamic proteins, the time of peak expression is shown by the node color; static proteins are represented as white nodes. Outside the circle, the dynamic proteins without interactions are positioned and colored according to their peak time. Lichtenberg et al. Science 307, 724 (2005)

14. 8. Lecture WS 2010/11Bioinformatics III14 Conditional gene expression c, Standard statistics (global topological measures and local network motifs) describing network structures. These vary between endogenous and exogenous conditions; those that are high compared with other conditions are shaded. (Note, the graph for the static state displays only sections that are active in at least one condition, but the table provides statistics for the entire network including inactive regions.) Luscombe, Babu, … Teichmann, Gerstein, Nature 431, 308 (2004) a, Schematics and summary of properties for the endogenous and exogenous sub-networks. b, Graphs of the static and condition-specific networks. Transcription factors and target genes are shown as nodes in the upper and lower sections of each graph respectively, and regulatory interactions are drawn as edges; they are coloured by the number of conditions in which they are active. Different conditions use distinct sections of the network.

15. 8. Lecture WS 2010/11Bioinformatics III15 Forward-directed TF-network Luscombe, Babu, … Teichmann, Gerstein, Nature 431, 308 (2004) a, The 70 TFs active in the cell cycle. The diagram shades each cell by the normalized number of genes targeted by each TF in a phase. Five clusters represent phase-specific TF and one cluster is for ubiquitously active TFs. Note, both hub and non-hub TF are included. b, Serial inter-regulation between phase- specific TFs. Network diagrams show TFs that are active in one phase regulate TFs in subsequent phases. In the late phases, TFs apparently regulate those in the next cycle. c, Parallel inter-regulation between phase- specific and ubiquitous TFs in a two-tiered hierarchy. Serial and parallel inter-regulation operate in tandem to drive the cell cycle while balancing it with basic house-keeping processes.