Beginning in mid-February 2008, the 1997-2007 online version of the Science Watch® newsletter, ESI-Topics.com, and in-cites.com, will all be featured together on the redesigned ScienceWatch.com. All previous content from the three sites will be permanently archived, and remain accessible from any existing bookmarks to the archived pages. No new content will be added to this site. Updates and new content (updated biweekly) are available at ScienceWatch.com now.
The Thomson Corporation inin-cites logoites
ScientistsPapersInstitutionsJournalsCountriesH O M ERSS feeds


S E A R C H
incites



SCIENTISTS
Scientists
• Papers
• Institutions
• Journals
• Countries
 

• The Top 10...
• Analysis of...
• Site Map by Fields
• Overview Menu of all Interviews
• Podcasts
• Hot Papers published within the last 2 years
• Current Classics
• SCI-BYTES - What's New in Research
What's New in Research
•

in-cites, May 2003
Citing URL: http://www.in-cites.com/scientists/ProfSirPhilipCohen.htm

Scientists

             
An essay by:
Professor Sir Philip Cohen
           

n this in-cites essay, Professor Sir Philip Cohen of the University of Dundee talks about his highly cited investigations of one of the major pathways of insulin signal transduction. Professor Cohen is currently the second most-cited scientist in the field of Biology & Biochemistry in the ISI Essential Science Indicators Web product, with 192 papers cited a total of 14,186 times to date. Professor Cohen also has 44 papers cited a total of 5,434 times to date in the field of Molecular Biology & Genetics. At the University of Dundee, Professor Cohen is the head of the MRC Protein Phosphorylation Unit in the Division of Cell Signalling, School of Life Sciences.

Dissection of one of the major pathways of insulin signal transduction

The discovery of insulin in 1921 and its spectacular effects on Type-1 diabetics must rank as one of the greatest medical advances of the 20th century. However, understanding the way in which insulin signals to the cell interior proved a far more difficult nut to crack and, even 60 years later, was still a complete mystery. Then in 1982, Ron Kahn showed that the insulin receptor was a protein tyrosine kinase which becomes active and phosphorylates itself upon binding to insulin. Later, he and Morris White established that receptor autophosphorylation is followed by the recruitment and phosphorylation of insulin receptor substrate 1 (IRS1) which, in turn, recruits phosphatidylinositol 3-kinase (PI 3-kinase) to the plasma membrane. This enzyme, which converts PI(4,5)P2 (a minor inositol phospholipid in the plasma membrane) to PI(3,4,5)P3, was discovered in the late 1980s by Lew Cantley in a completely different context. Following the identification of a relatively specific inhibitor of PI 3-kinase (wortmannin) by Michio Ui and his colleagues in the early 1990s, this compound was found to block nearly all the metabolic actions of insulin, indicating that PI(3,4,5)P3 was likely to be the long sought after "second messenger" for insulin. However, how PI(3,4,5)P3 mediated the intracellular actions of insulin was unclear.

My own research on insulin signalling began in late 1973, when I decided to follow up seminal findings made by Joseph Larner about a decade earlier. His studies had revealed that, within minutes, insulin triggers the activation of glycogen synthase, an enzyme which catalyses the last step in the synthesis of glycogen. This facilitates the conversion of glucose to glycogen, one of the major metabolic actions of insulin. Larner also showed that activation results from the conversion of glycogen synthase to a less highly phosphorylated state. Thus, it was already clear that insulin must induce the inhibition of a protein kinase and/or the activation of a protein phosphatase acting on glycogen synthase. My first postdoctoral fellow, Hugh Nimmo, and third graduate student, Chris Proud, were put to work on this problem and their studies soon revealed that the regulation of glycogen synthase was surprisingly complex, involving several phosphorylation sites and at least two protein kinases. Then, in 1979, my student Noor Embi identified a novel protein kinase capable of phosphorylating and inactivating glycogen synthase in vitro, and which we termed glycogen synthase kinase 3 (GSK3) [1]. This enzyme was purified to homogeneity and characterised by Brian Hemmings, a postdoctoral fellow, and by Jim Woodgett, a graduate student. Peter Parker, another postdoctoral fellow, then made the key demonstration that most of the phosphate released from glycogen synthase in vivo in response to insulin is removed from the serine residues that are targeted by GSK3 in vitro [2]. This result implied that insulin must exert its effect on glycogen synthase by inducing the inhibition of GSK3 and/or by activating a protein phosphatase. The relevant phosphatase was later identified as a glycogen-associated form of protein phosphatase-1 by Peter Stralfors, a postdoctoral fellow ([3], reviewed in [4]).

In the early 1990s Jim Woodgett and Chris Proud, who had by now set up their own research groups, independently demonstrated that GSK3 was indeed inhibited acutely by insulin [5, 6]. Chris Proud [6] found that the insulin-induced inhibition of GSK3 could be reversed by treatment with a serine/threonine-specific protein phosphatase. My student Darren Cross then made the key observation that the PI 3-kinase inhibitor wortmannin prevented the insulin-induced inhibition of GSK3 [7]. Thus it started to become clear that PI(3,4,5)P3 must trigger the activation of a protein kinase, which then phosphorylated and inhibited GSK3.

Calum Sutherland, another graduate student, found that two insulin-stimulated protein kinases, termed MAPKAP-K1 and p70 S6 kinase both inhibited GSK3 in vitro by phosphorylating a specific serine residue [8, 9]. However, surprisingly, Darren Cross then showed that two different drugs, PD 98059 and rapamycin, which prevent the activation of MAPKAP-K1 and p70 S6 kinase, respectively, had no effect on the insulin-induced inhibition of GSK3 [10]. By incubating cells in the presence of both PD 98059 and rapamycin, Darren was then able to reveal the presence of a further insulin-stimulated protein kinase capable of inactivating GSK3.

At this juncture Brian Hemmings, who had set up his own laboratory at the Friedrich-Miescher Institute in Basel, Switzerland, returned to Dundee to examine the thesis of a graduate student. After Brian was shown our data, he suggested that the novel insulin-stimulated protein kinase might be protein kinase B (PKB, also called AKT), because he was aware of work to be published shortly which would demonstrate that PKB was activated by insulin or growth factors via a PI(3,4,5)P3-dependent pathway [11, 12]. Fortunately, Brian was also able to provide an anti-PKB antibody to test this idea. This enabled Darren to rapidly establish that the new activity could indeed be immunoprecipitated by this antibody [10].

The next step was clearly to discover how PKB was activated. Dario Alessi, a postdoctoral fellow in the lab, found that the insulin-induced activation of PKB was accompanied by its phosphorylation at two sites, Thr308 and Ser473. Moreover activation and phosphorylation were both prevented by wortmannin and reversed by treatment with PP2A [13]. Dario was then able to detect and purify a further protein kinase capable of activating PKB in vitro. Excitingly, this protein kinase was only able to activate PKB in the presence of lipid vesicles containing PI(3,4,5)P3 and we therefore called it 3-phosphoinositide-dependent protein kinase-1 (PDK1) [14]. PDK1 was the missing link in the chain of events by which insulin, acting via PI(3,4,5)P3, activates glycogen synthase, and its discovery launched Dario's independent research career. Dario went on to clone PDK1 showing that, like PKB, it contains a pleckstrin homology (PH) domain [15]. We now know that PI(3,4,5)P3 interacts with the PH domains of PKB and PDK1, co-localising these enzymes at the plasma membrane and allowing PDK1 to activate PKB [16, 17]. Dario's group also showed that the activation of PKB does not occur in cells that lack PDK1, providing genetic evidence for the essential role of PDK1 in the pathway [18].

We demonstrated that PKB has a restricted specificity, only phosphorylating serine and threonine residues that lie in Arg-Xaa-Arg-Xaa-Xaa-Ser/Thr- sequences (where Xaa is any amino acid) [19]. This facilitated the identification of further key physiological substrates of PKB, such as FKHR (recently renamed FOXO-1) [20, 21] and related transcription factors. We, and others, also found that PDK1 activates a number of other protein kinases of the "AGC subfamily," such as isoforms of protein kinase C (PKC) [22] and the serum and glucocorticoid-regulated protein kinase (SGK) [23, 24]. The finding that PDK1 switches on a number of protein kinases, each of which have multiple substrates, helps to explain how so many actions of insulin can be mediated by PI(3,4,5)P3.

The identification of PDK1 in early 1997, more than 23 years after I first started to work on insulin signalling, was a momentous event for my laboratory. The delineation, at least in outline, of one of the major pathways of insulin signal transduction offers new opportunities to overcome the resistance to insulin that can lead to Type-II diabetes, and intense efforts to develop such drugs are underway in many pharmaceutical companies. However, the PDK1-PKB-GSK3 "cascade" is switched on by growth factors, as well as by insulin, delivering an anti-apoptotic signal that is crucial for the survival of many cancer cells. For this reason there is also enormous interest in developing inhibitors of this pathway as anti-cancer drugs.

Of course, many unsolved problems remain. For example, PDK1 only phosphorylates PKB at Thr308, and the protein kinase that phosphorylates PKB at Ser473 (provisionally termed PDK2) has yet to be identified. Many of the key substrates of PKB and the other protein kinases that are activated by PDK1 have yet to be identified. Finally, it remains entirely possible that the stimulation of glycogen synthase by insulin involves the activation of a phosphatase that dephosphorylates glycogen synthase, as well as the inhibition of GSK3. Many laboratories are now working on these and other outstanding questions and an even clearer picture will undoubtedly emerge over the next few years.

References.
  1. Embi, N., Rylatt, D.B. and Cohen, P. (1980) Eur.J.Biochem.107, 519-527."Glycogen synthase kinase-3 from rabbit skeletal muscle and phosphorylation of glycogen synthase by three different protein kinases".
  2. Parker, P.J., Caudwell, F.B. and Cohen, P. (1983) Eur. J. Biochem. 130, 227-234. "Glycogen synthase from rabbit skeletal muscle: effects of insulin on the state of phosphorylation of the seven phosphoserine residues in vivo'.
  3. Strålfors,P., Hiraga, A. and Cohen P. (1985) Eur.J.Biochem.149, 295-303. "The protein phosphatases involved in cellular regulation: Purification and characterisation of the glycogen-bound form of protein phosphatase-1 from rabbit skeletal muscle
  4. Hubbard, M. J. and Cohen, P (1993) TIBS. 18, 172-177. "The regulation of protein phosphatases and protein kinases by targeting subunits".
  5. Hughes, K., Ramakrishna, S., Benjamin, W.B. and Woodgett, J.R. (1992) Biochem. J. 288, 309-314. Identification of multifunctional ATP-citrate lyase kinase as the α-isoform of glycogen synthase kinase-3."
  6. Welsh, G.I. and Proud, C.G. (1993) Biochem J. 294, 625-629. "Glycogen synthase kinase-3 is rapidly inactivated in response to insulin and phosphorylates eukaryotic initiation factor eIF2B."
  7. Cross, D.A.E., Alessi, D.R., Vandenheede, J.R., McDowell, H., Harinder, S. and Cohen, P. (1994) Biochem. J. 303, 21-26. "The inhibition of glycogen synthase kinase-3 by insulin or IGF-1 in the rat skeletal muscle cell line L6 is blocked by wortmannin, but not by rapamycin: evidence that wortmannin blocks activation of the MAP kinase pathway in L6 cells between Ras and Raf."
  8. Sutherland, C., Leighton, I. and Cohen, P. (1993) Biochem. J. 296, 15 - 19. "Inactivation of glycogen synthase kinase-3 by phosphorylation; new kinase connections in insulin and growth factor signalling".
  9. Sutherland, C. and Cohen, P. (1994) FEBS Lett. 338, 37-42 " The α-isoform of glycogen synthase kinase-3 from rabbit skeletal muscle is inactivated by p70 S6 kinase or MAP kinase-activated protein kinase-1 in vitro".
  10. Cross, D.A.E., Alessi, D.R., Cohen, P., Andjelkovic, M. and Hemmings, B. (1995) Nature 378, 785-789 "Inhibition of glycogen synthase kinase-3 by insulin is mediated by Akt/PKB."
  11. Franke, T.F., Yang, S.I., Chan, T.O., Datta, K., Kazlauskas, A., Morrison, D.K., Kaplan, D.R. and Tsichlis, P.N. (1995) Cell 81, 727-736. "The protein kinase encoded by the AKT proto-oncogene is a target of the PDGF-activated phosphatidylinositol 3-kinase."
  12. Burgering, B.M.Th. and Coffer, P.J. (1995) Nature 376, 599-602. "Protein kinase B (c-AKT) in phosphatidylinositol-3-OH kinase signal transduction."
  13. Alessi, D.R., Andjelkovic, M., Caudwell, B., Cron, P., Morrice, N., Cohen, P. and Hemmings, B. (1996) EMBO J. 15, 6541-6551. "Mechanism of activation of protein kinase B by insulin and IGF-1".
  14. Alessi, D.R., James, S.R., Downes, C.P., Holmes, A.B., Gaffney, P.R.J., Reece, C.B. and Cohen, P. (1997) Curr. Biol. 7, 261-269. "Characterization of a 3-phosphoinositide-dependent protein kinase which phosphorylates and activates protein kinase Bα".
  15. Alessi, D.R., Deak, M., Casamayor, A., Caudwell, F.B., Morrice, N., Norman, D.G., Gaffney, P., Reese, C.B., MacDougall, C.N., Harbison, D., Ashworth, A. and Bownes, M. (1997) Curr. Biol. 7, 776-789. "3-phosphoinositide-dependent protein kinase-1 (PDK1): structural and functional homology with the Drosophila DSTPK61 kinase."
  16. Andjelkovic, M., Alessi, D.R., Meier, R., Fernandez, A., Lamb, N.J.C., Frech, M., Cron, P., Cohen, P., Lucocq, J.M. and Hemmings, B.A. (1997) J. Biol. Chem. 272, 31515-31524. "Role of Translocation in the Activation and Function of Protein Kinase B".
  17. Currie, R.A., Walker, K. S., Gray, A., Deak, M., Casamayor, A., Downes, C.P., Cohen, P., Alessi, D.R. and Lucocq, J. (1999) Biochem. J. 337, 575-583. "The role of phosphatidylinositol 3,4,5-trisphosphate in regulating the activity and localisation of 3-phosphoinositide-dependent protein kinase-1".
  18. Williams, M.R., Arthur, J.S.C., Balendran, A., van der Kaay, J., Poli, V., Cohen, P. and Alessi, D.R. (2000) Curr. Biol. 10, 439-448. "The role of 3-phosphoinositide-dependent protein kinase 1 in activating AGC kinases defined in embryonic stem cells".
  19. Alessi, D.R., Caudwell, F.B., Andjelkovic, M., Hemmings, B. and Cohen, P. (1996) FEBS Lett. 399, 333-338. "Molecular basis for the substrate specificity of protein kinase B; comparison with MAPKAP kinase-1 and p70 S6 kinase".
  20. Rena, G., Guo, S., Cichy, S., Unterman, T.G. and Cohen, P. (1999) J. Biol. Chem. 274, 17179-17183. "Phosphorylation of the transcription factor forkhead family member FKHR by protein kinase B."
  21. Guo, S., Rena, G., Cichy, S., He, X., Cohen, P and Unterman, T.G. (1999) J. Biol. Chem. 274, 17184-17192. "Phosphorylation of serine 256 by protein kinase B disrupts transactivation by FKHR and mediates effects of insulin on IGF binding protein-1 promoter activity through a conserved insulin response sequence".
  22. Le Good, J.A., Ziegler, H., Parekh, D.B., Alessi, D.R., Cohen, P. and Parker, P.J. (1998) Science 281, 2042-2045. "Protein kinase C isotypes are controlled by phosphoinositide 3-kinase through the upstream kinase PDK1".
  23. Kobayashi, T. and Cohen, P. (1999) Biochem J. 339, 319-328. "Activation of SGK by agonists that activate phosphatidylinositide 3-kinase is mediated by PDK1 and PDK2".
  24. Kobayashi, T., Deak, M., Morrice, N. and Cohen, P. (1999) Biochem. J. 344, 189-197. "Characterisation of the structure and regulation of two novel isoforms of serum- and glucocorticoid-induced protein kinase".End

Professor Sir Philip Cohen, FRS, FRSE
MRC Protein Phosphorylation Unit
School of Life Sciences
MSI/WTB Complex
University of Dundee
Dundee, Scotland 
  

in-cites, May 2003
Citing URL: http://www.in-cites.com/scientists/ProfSirPhilipCohen.htm

ScienceWatch.com - Tracking Trends and Perfomance in Basic Research
Go to the new ScienceWatch.com
Home | Search | Disclaimer | Terms of Use | Privacy Policy | Copyright
Contact Webmaster with questions/comments |
(c) 2008 The Thomson Corporation.