THE EFFECT OF APICAL AND BASOLATERAL LIPIDS ON THE FUNCTION OF THE BAND-3 ANION-EXCHANGE PROTEIN

Although many polarized proteins are sorted to the same membrane domai n in all epithelial tissues, there are some that exhibit a cell type-s pecific polarity. We recently found that band 3 (the anion exchanger A E1) was present in the apical membrane of a renal intercalated cell li ne when these cells were seeded at low density, but its targeting was reversed to the basolateral membrane under the influence of an extrace llular matrix protein secreted when the cells were seeded at high dens ity. Because apical and basolateral lipids differ in epithelia, we ask ed what effect might these lipids have on band 3 function. This questi on is especially interesting since apical anion exchange in these cell s is resistant to disulfonic stilbene inhibitors while basolateral ani on exchange is quite sensitive. Furthermore, the apical anion exchange r cannot be stained by antibodies that readily identify the basolatera l protein. We used short chain sphingolipid analogues and found that s phingomyelin was preferentially targeted to the basolateral domain in the intercalated cell line. The ganglioside GM(1) (Gal1 beta 1,3GalNAc beta 1, 4Gal-NeuAc alpha 2, 3Gal beta 1, 4Glc ceramide) was confined to the apical membrane as visualized by confocal microscopy after addi tion of fluorescent cholera toxin to filter grown cells, We reconstitu ted erythrocyte band 3 into liposomes using apical and basolateral typ es of lipids and examined the inhibitory potency of 4,4'-dinitorsostil bene-2,2'-disulfonic acid (DNDS; a reversible stilbene) on (SO4)-S-35/ SO4 exchange. Although anion exchange in sphingomyelin liposomes was s ensitive to inhibition, the addition of increasing amounts of the gang lioside GM(1) reduced the potency of the inhibitor drastically. Becaus e these polarized lipids are present in the exofacial surface of the b ilayer, we propose that the lipid structure might influence the packin g of the transmembrane domains of band 3 in that region, altering the binding of the stilbenes to these chains. These results highlight the role of polarized lipids in changing the function of unpolarized prote ins or of proteins whose locations differ in different epithelia.