PI 3-kinase, PKB, protein synthesis and cellular survival

1. PI 3-kinase, PKB, protein synthesis and cellular survival IJsbrand Kramer i.kramer@iecb.u-bordeaux.fr

3. Modifications of phosphoinositol lipids by: a kinase (PI 3-kinase), two phosphatases (PTEN, SHIP) and one lipase (phospholipase C). All products are signaling molecules (second messengers).

4. extracellular PDGF-R PI 3-kinase acts downstream of a number of tyrosine-kinase containing receptors. Here we show the example of the PDGF-R. Importanlty, insulin signals through PI 3-kinase and this is essential for the regulation of the glucose metabolism (together with the regulation by PKA). PDGF A/B ATP ATP y751 Y741 intracellular

5. PI 3-kinase adaptor protein p85 PDGF-R P85 SH2 domain (C-terminal) pTyr751 PDGF A/B Met755 PDGF-R (aa751-755) ATP ATP P110 PI 3-kinase y751 Y741 p85 The adoptor/regulatory protein p85 recruits the catalytic subunit p110 (a,b or d) to the membrane. This way it places the enzyme in the vincinity of its substrate (inositol phospholipids). Compare this with the role of Grb for the recruitment of Sos). Binding of p85 to the receptor may also have an allosteric effect that promotes kinase activity of the catalytic subunit.

6. PI 3-kinase p110g can also be recruited to the membrane by an interaction with RasGTP. Both Ras and p85 have a membrane recruitment and an allosteric role in the regulation of activity of the catalytic subunit (p110)

7. Activation of PI 3-kinase downstream of the insulin receptor

8. Numerous PI 3-kinases, classified as type I, II or II, and their regulators.

9. Numerous interactions between PI 3-kinase and other signalling components. PI 3-kinase may control Cdc42 and Rac activity through its BCR/GAP domain.

10. The core of type 1g PI 3-kinase has a typical kinase fold; with its small (N-terminal) and big (C-terminal) lobes separated by the catalytic cleft that binds ATP (in red) and substrate (inositol-1,4,5-trisphosphate). • The helix domain forms a scaffold to which RBD, the catalytic and the C2-domain are firmly attached.

11. Wortmannin and Lilly294002 are PI 3-kinase inhibitors that occupy the catalytic cleft.

12. Domain architecture of the family of protein kinase B and its upstream kinase PDK1

13. Molecular structure of inactive and active PKB (panel a and b). The position of the HM domain in panel (a) is hypothetical. Note the subtle cascade of events that render PKB catalytically competent (panel c)

14. Activation of PKB by membrane attachment followed by two phosphorylations • Members of the AGC family of protein kinases that are activated in a similar manner

15. PKB controls protein synthesis through activation of mTOR. This leads to activation of S6K1 as well as assocation of initiation factors with mRNA

16. mTOR regulates protein synthesis by phosphorylating and removing BP. This signals the assembly of the ribosomal initiation complex. S6K1, primed by mTOR, is activated by PDK1 and this too contributes to the process of initiation of protein synthesis

17. Dual role of Ras and PI 3-K/PKB in the activation of protein synthesis

18. Adhesion of cells to matrix contributes to cellular survival

19. PKB protects against apoptosis in a number of ways

20. Initiation of apoptosis by leakage of cytochrome c into the cytoplasm: a process facilitated by the pro-apoptotic mediator Bad

21. PKB inhibits activity of GSK3b, thereby preventing the breakdown of cyclin D and thus stimulating the entry into the G1 phase of the cell cycle

22. Downystream of the Insulin receptor; PKB plays a role in the metabolism of glucose; activation of glycogen synthase b An essential task of PKB is to inactivate GSK-3b.

23. PTEN classifies as a tyrosine phosphatase but it prefers inositol substrates. It opposes the action of PI 3-kinase and its loss is associated with cell transformation, hence its label « tumour suppressor ».

24. PTEN is inhibited by the action of growth factors. This occurs through the activation of Src which phosphorylates PTEN on two tyrosine residues. This reduces its half-live and inhibits membrane localisation (no access to substrate)

25. The tumour suppresive effect of PTEN is not limited to counteracting PI 3-kinase. It also plays a role in the nucleus, independent of its phosphatase action, where it associates with proteins that warrant « chromosome integrity ».