Vincristine-Induced Overexpression of P-Glycoprotein in L1210 Cells Is Associated with Remodeling of Cell Surface Saccharides

Abstract

Multidrug resistance of murine leukemic cell line L1210/VCR (R), obtained by adaptation of parental L1210 cells (S) on vincristine, is associated with overexpression of P glycoprotein (P-gp, the ATP-dependent drug efflux pump). Previously, we found that cytochemical staining of negatively charged cell surface binding sites (probably sialic acid) by ruthenium red (RR) revealed a compact layer of RR bound to the external coat of S cells. This is in contrast to R cells and L1210/VCR cells cultured in the presence of vincristine during the last cultivation prior to the experiment (V cells), where the RR layer was either reduced or absent. In the current paper, we observed differences in the interactions of S, R and V cells with Concanavalin A (ConA) and tomato lectin (lycopersicum esculentum agglutinin, LEA). ConA bound and induced cell damage more effectively in S cells than in R or V cells. Both of these effects could be prevented by methyl-manopyranose, but not by N-acetylglucosamine. In contrast, LEA lectin preferentially bound to R and V cells. While LEA agglutinated cells more effectively than ConA, it did not cause cell damage comparable to ConA. Binding of LEA to the cell surface could be prevented by chitooligosaccharides. Both LEA and ConA failed to identify P-gp in lectin blots. Thus, changes in ConA and LEA interactions are not caused by massive expression of P-gp in the plasma membrane and the consequent exposure of the inner saccharides to the external side of the plasma membrane.Taken together, the above facts suggest that S cells differ from R and V cells in the composition of cell surface glycosides not directly linked to P-gp.

The expression of P-gp in leukemia cells is associated with cross-resistance to protein N-glycosylation inhibitor tunicamycin

“…This leads to differences in protein glycosylation between P-gp-positive and -negative cells (reviewed in Breier et al 2005Breier et al , 2013. Consistent with this, overexpression of P-gp in leukemia cells leads to large-scale remodeling of the cell surface glycosides that can be detected by several lectins (Molnar et al 2009;Sulova et al 2009Sulova et al , 2010Bubencikova et al 2012). Moreover, aberrant cell surface sialylation was described in MDR HL60 cells (Ma et al 2015).…”

“…Changes in cell surface glycoproteins are considered responsible for the ConA resistance of non-differentiating L6 myoblasts selected for resistance to this lectin (Parfett et al 1983). Both P-gppositive variants of L1210 cells (L1210/R and L1210/T) are less sensitive to ConA than the parental L1210 cells (Fiala et al 2003;Sulova et al 2009Sulova et al , 2010). This resistance is based on a lower binding capacity of the cell surface of resistant cells to ConA than sensitive cells.…”

“…This resistance is based on a lower binding capacity of the cell surface of resistant cells to ConA than sensitive cells. Alterations of the cell surface binding capacity of the tomato lectin (Lycopersicum esculentum agglutinin, Sulova et al 2009), SNA and WGA (Bubencikova et al 2012) on the cell surface of P-gp-positive L1210 cells are associated with altered cell death effects induced by these lectins. These facts indicate that expression of P-gp in L1210 cells alters the cell surface sugar composition, and the composition of cell surface sugars is associated with changes to the binding and cell death effect of lectins.…”

Tunicamycin Depresses P-Glycoprotein Glycosylation Without an Effect on Its Membrane Localization and Drug Efflux Activity in L1210 Cells

“…The MDR phenotype of R cells was associated with an alteration in transglycosylation reactions linked with decreases in UDP-sugars, glycogen and cell surface sialic acid [16]. In addition, R cells differ from S cells in the composition of the cell surface saccharides that are ligands of concanavalin A and the tomato lectin Lycopersicum esculentum agglutinin [17]. While concanavalin A interacts more potently with S cells than with R cells, the tomato lectin exhibited the opposite behavior.…”

“…While concanavalin A interacts more potently with S cells than with R cells, the tomato lectin exhibited the opposite behavior. Both lectins were shown to interact with glycosylated proteins in the plasma membrane other than P-gp [17]. The weak interaction of concanavalin A with P-gp-positive cells is directly related to presence of P-gp in L1210 cells because T cells displayed a similarly reduced interaction compared to S cells [15].…”