Inhibition of epidermal growth factor‐stimulated DNA synthesis by a bovine sialoglycopeptide inhibitor occurs at an intracellular level

Abstract

The control of cell proliferation involves the complex interaction between growth factors and growth inhibitors. We have examined this interaction with the mitogen epidermal growth factor (EGF) and a recently purified 18 kD, pI 3, sialoglycopeptide that reversibly inhibits cellular metabolism of a variety of cells. The sialoglycopeptide was a very potent inhibitor of EGF action; 0.22 nM of the inhibitor completely blocked the mitogenic effect of 1.60 nM of EGF. The sialoglycopeptide, however, did not affect the binding of EGF to 3T3 cells. Neither the mixed affinities (0.11-1.9 nM) of binding nor the total number of receptors (50,000 receptors/cell) for EGF were altered by the addition of the sialoglycopeptide. In addition, competitive binding experiments demonstrated the specificity of inhibitor binding to 3T3 cells and also showed that EGF and the sialoglycopeptide did not share the same receptor, suggesting that the inhibitor blocked EGF action at a postreceptor, intracellular event in the signal cascade. We further demonstrated that the sialoglycopeptide had to be added within 2.5 hr after EGF to block effectively the stimulation of DNA synthesis by the growth factor, suggesting that the inhibitor blocked EGF stimulation at a relatively early step in the signal transduction mechanism.

Role of calcium in growth inhibition induced by a novel cell surface sialoglycopeptide

“…These studies must be interpreted with caution, however, since the compounds utilized to block various intracellular calcium changes may have other, as of yet unknown, effects on cellular constituents. Previous studies in our laboratory have demonstrated the ability of the CeReS-18 growth inhibitor to inhibit the induction of DNA synthesis mediated by such growth factors (Bascom et al, 1987;Johnson and Sharifi, 1989). In addition, reports by Takuwa et al (1992Takuwa et al ( , 1993 demonstrated that stimulation of the phosphorylation of the retinoblastoma gene product (Rb) and activation of p34cdc2 kinase are calciumicalmodulin-dependent processes.…”

“…CeReS-18 binds to a specific cell surface receptor (Sharifi and Johnson, 1987), and binding to this receptor correlates with protein synthesis inhibition (Bascom et al, 1986).…”

“…CeReS-18 appears to block most cell types in the late GI phase of the cell cycle, and results in cell cycle synchronization upon 0 1995 WILEY-LISS, INC. removal (Fattaey et al, 1989Edson et al, 1991). CeReS-18 binds to a specific cell surface receptor (Sharifi and Johnson, 1987), and binding to this receptor correlates with protein synthesis inhibition (Bascom et al, 1986).…”