Development of the Na +-dependent hexose carrier in LLC-PK 1 cells is dependent on microtubules

Abstract

The Na +-dependent hexose carrier, an endogenous apical marker, develops during differentiation of LLC-PK 1, an established cell line with characteristics of the proximal tubule. This development was inhibited by the microtubule-disrupting drugs, colchicine and nocodazole, while it was insensitive to lumicolchicine. This strongly suggests that microtubules are involved in the plasma membrane expression of the Na +-dependent hexose carrier. We also analyzed the increase in activity of endogenous apical and basolateral membrane proteins during the polarization process. The development of three apical (Na +-dependent hexose carrier, γ-glutamyltransferase and alkaline phosphatase) and one basolateral membrane protein (Na + / K +-ATPase) was studied during the reorganization of LLC-PK 1 cells into a polarized epithelium. Colchicine inhibited the rapid, transient increase in the expression of the Na +-dependent hexose carrier during this polarization process. A similar result was observed for the development of the other apical proteins, while the development of Na + / K +-ATPase seemed to be largely insensitive to colchicine. Our results are in agreement with the model that the vesicles containing the apical membrane proteins use microtubules as tracks to reach the plasma membrane. The transport of vesicles containing basolateral membrane proteins clearly occurs by a different pathway which is independent on an intact microtubular network. Since the inhibition by the microtubule-disrupting drugs was complete, it can be concluded that after disruption of microtubules, the apical vesicles do not use the basolateral pathway by default.