Cell surface changes in transformed human lymphocytes : I. ConA and E-PHA induced unique changes in surface topography

Binding studies with six purified plant lectins were used to investigate membrane alterations that occur in lymphocyte transformation. Normal human peripheral blood lymphocytes transformed with E-Phytohemagglutinin (E-PHA) or concanavalin-A (Con-A) generally possessed increased numbers of lectin receptors. When this increase was corrected for the expanded surface area of transformed lymphocytes, it appeared that E-PHA and ConA each produced a unique and complex reorganization of cell surface topography. Surface alterations occurred independently of DNA synthesis, cell proliferation, and microtubule or microfilament function. Puromycin inhibited emergence of new lectin receptors on cells transformed with E-PHA, but not with ConA. Lymphocytes incubated with either lectin showed increased incorporation of [¹⁴C]galactose into trypsin-sensitive cell surface glycoproteins. This incorporation was abolished by puromycin in cells stimulated by E-PHA but not by ConA. These studies demonstrate that although both lectins induce similar morphological alterations in human lymphocytes, at the molecular level the structural changes induced in the cell membrane by these two lectins differ considerably. Furthermore, these structural alterations are mediated via different mechanisms in the two groups of cells. De novo protein synthesis is required for cell surface reorganization in PHA-stimulated cells, but not in cells stimulated by ConA. The effect of ConA appears to be to enhance attachment of saccharide structures to pre-synthesized membrane proteins.     

Receptors for pea and lentil lectins on human lymphoid cells: demonstration of distinct lectin-defined cell subclasses

Lectins are useful probes for studying cell surface glycoconjugates. Pea (PL) and lentil (LL) lectin each requires for binding a fucosyl- and two alpha-mannosyl residues in core regions of glycopeptides, but differences in outer chain carbohydrates may alter their relative binding affinities. We used binding studies with [125I]-PL and LL and flow cytometry with fluorescein-conjugated (FITC)-PL and -LL to study their interactions with peripheral lymphocytes. Binding of both lectins to lymphocytes was saturable, reversible, and inhibited by alpha-methyl mannose. Scatchard analyses were consistent with two classes of receptors for each lectin. Flow cytometric analyses demonstrated that cell to cell receptor densities varied. Sixty-five percent of lymphocytes bound PL (mean 2 X 10(6) receptors/cell) and 45% bound LL (mean 3 X 10(6) receptors/cell). Competition studies demonstrated mutual inhibition, but flow cytometry revealed persistent FITC-PL or -LL binding depsite 20-fold molar excess of the other lectin. Distributions of receptors for PL and LL on lymphocytes were as follows: 45% of lymphocytes bound both PL and LL; 20% of lymphocytes bound PL alone; 35% of lymphocytes bound neither PL nor LL. Despite similar binding requirements for PL and LL and overlap between their receptors on lymphocytes, there appear to be subsets of receptors specific for each lectin. These results may reflect abilities of PL and LL to discriminate subtle carbohydrate differences on lymphocyte surfaces.