Objectives of Research:

A highly investigated field for several decades now relates to insulin-dependent signal transmission networks. A key question having clinical significance as well is whether insulin effects can be manifested via alternative (insulin-independent) signaling pathways. Our long-term studies on the insulino-mimetic actions of vanadium [i.e. Karlish and Shechter, (1980) Nature 284, 556; Degani et al. (1981) Biochem-istry 20, 5795; Shechter and Ron (1986) J. Biol. Chem. 261, 14951; Shechter et al. (1992) Biochem-istry 30, 2063, reviewed in Shechter Y. (1990) Diabetes 39, 1] revealed that the metabolic effects of insulin can be fully manifested through insulin-receptor independent pathways. The identification and characterization of the 'key players' in this backup system is the main objectives of our research.

Recent Findings:

Vanadium salts that mimic virtually all the metabolic effects of insulin, have no or minor effect in activating the insulin-receptor tyrosine kinase (InsRTK) in intact cellular systems (reviewed in Shechter et al., 1995). This vanadate bypassing receptor activation was fully validated, using specific cell permeable blockers of InsRTK [Shisheva and Shechter (1992) Biochemistry 31, 2063]. Endogenous tyrosine-phosphorylation, however, is a prerequisite condition for manifesting the metabolic effects of insulin. We therefore searched for additional vanadium-activatable non-receptor protein tyrosine kinase. Such a protein has been identified in the cytosolic fraction of rat adipocytes (CytPTK, Shisheva and Shechter, 1993).

The relevant features of CytPTK, and the basic differences between CytPTK and the insulin-receptor are summarized in Table I.

CytPTK is activated several fold by vanadium salts and participates in several of the insulin-like effects manifested by vanadium salts.

Establishment of a cell-free system: The lack of a cell-free experimental system for studying insulin-dependent signaling pathways was the predominant factor in slowing down this field of research. Recently, however, we have managed to establish such a cell-free system for the investigation of vanadium-dependent mechanism(s) of action (Elberg et al., 1994). This experimental system allowed us to determine that CytPTK activation is preceded and dependent on inhibiting vanadium-sensitive protein phosphotyrosine phosphatases (PTPases, Elberg et al. , 1994; Elberg et al. , 1995, Li et al., 1995a, b). Most of the vanadate-sensitive PTPases are intrinsic plasma membrane proteins. The more relevant PTPases to manifest the bioresponses of insulin are now identified (in preparation). An additional non-receptor protein-tyrosine kinase that is activated by vanadate was identified as well (submitted). In summary, the key players of the backup system are PTPases, and nonreceptor protein tyrosine kinases. The 'cross-talk' relationships between these components is the main interest of our laboratory.

Table I. Comparison between CytPTK and InsRTK.
estimated molecular-weight 53 kDa 350-400 kDa
requirement for bivalent anion Co2+ Mn2+
good exogenous substrate PolyGlu4Tyr PolyGlu4Tyr
sensitivity to N-ethylmaleimide no effect inactivation
inhibition by staurosporine (IC50) 2 uM 8 uM

Recent Publications:

Shisheva, A. and Shechter, Y. (1993) Role of cytosolic tyrosine kinase in mediating insulin-like actions of vanadate in rat adipocytes. J. Biol. Chem. 268: 6463-6469.

Shisheva, A. and Shechter, Y. (1993) Mechanism of pervanadate stimulation and potentiation of insulin-activated glucose transport in rat adipocytes; dissociation from vanadate effect. Endocrinology 133: 1562-1568.

Elberg, G., Li, J. and Shechter, Y. (1994) Vanadium activates or inhibits receptor and non-receptor protein tyrosine kinases in cell-free experiments, dependent on its oxidation state. Possible role of endogenous vanadium in controlling cellular protein tyrosine kinase activity. J. Biol. Chem. 269: 9521-9527.

Li, J., Elberg, G., Gefel, D. and Shechter, Y. (1995a) Permolybdate and pertungstate-potent stimulators of insulin effects in rat adipocytes; mechanism of action. Biochemistry 34: 6216-6225.

Li, J., Elberg, G., Libman, J., Shanzer, A., Gefel, D. and Shechter, Y. (1995b). Insulin-like effects of tungstate and molybdate: mediation through insulin receptor independent pathways. Endocrine 3: 631-637.

Elberg, G., Li, J., Leibovitch, A. and Shechter, Y. (1995) Non-receptor cytosolic protein tyrosine kinases from various rat tissues. Biochim. Biophys. Acta 1296: 299-306.

Shechter, Y., Li, J., Elberg, G., Gefel, D., Meyerovitch, J. and Shisheva, A. (1995) J. Cell Biochem. 153: 39-47.

Shechter, Y., Li J., Meyerovitch J., Gefel D., Bruck R., Elberg G., Miller D.S., Shisheva A. (1995) Insulin-like actions of vanadate are mediated in an insulin-receptor-independent manner via non-receptor protein tyrosine kinases and protein phosphotyrosine phosphatases, Mol. and Cel. Biochem. 153: 39-47.

Goldwaser, I., Gefel, D., Gershonov, E., Fridkin, M. and Shechter, Y. (2000) Insulin-like effects of vanadium; basic and clinical implications. J. Inorganic Biochem. 80:21-25.

Sun, Q., Sekar, N., Goldwaseer, I., Gershonov, E., Fridkin, M. and Shechter, Y. (2000) Vanadate restores glucose-6-phosphate in diabetic rats; A mechanism to enhance glucose metabolism. Am. J. Physiol. Endocrinol. Metab. 279:E403-E410.

Gershonov, E., Goldwaser, I., Fridkin, M. and Shechter, Y. (2000) A novel approach for a water-soluble long-acting insulin; Design, preparation and analysis of [2-sulfo]-9-fluorenylmethoxycarbonyl-insulin (FMS3-insulin). J. Med. Chem. 43:2530-2537.

Goldwaser, Y., Qian, S., Gershonov, E., Fridkin, M. and Shechter, Y. (2000) Organic vanadium chelators potentiate vanadium-evoked glucose metabolism in vitro and in vivo. Establishing criteria for optimal chelators. Mol. Pharmacol. 58:738-746.

Yoram Shechter, Liana Preciado-Patt, Gideon Schreiber, and Mati Fridkin (2001) Prolonging the half-life of human interferon-a2 in circulation: Design, preparation, and analysis of (2-sulfo-9-fluorenylmthoxycarbonyl)7-interferon-a2. Proc. Natl. Acad. Sci. USA 98: 1212-1217.

Yoram Shechter, Itzhak Goldwaser, Iris Lavon, Eytan Gershonov, Brenda Mester, Marina Mironchik, Lianna Preciado-Patt and Mati Fridkin (2001) A new approach for prolonging the half-life of peptides, proteins and low-molecular-weight drugs in vivo. Drugs of the Future 26: 472-480.

Yoram Shechter, Itzhak Goldwaser and Mati Fridkin (2001) Historic perspective and recent developments on the insulin-like actions of vanadium: Toward developing vanadium based drugs for diabetes. J. Am. Chem. Soc., in press.