Differences in plasma membrane organization of neuroblastoma cells grown in the differentiated and undifferentiated states

Abstract

Two methods have been developed for the isolation of plasma membranes from the neuro-2A clone of murine neuroblastoma grown in the differentiated (monolayer) and undifferentiated (spinner) states. In the first method, the membranes were isolated in 40 to 45% yield by fractionation in a two-phase polymer system and a linear sucrose gradient (primary plasma membranes). In the second method, those areas of the plasma membrane internalized during phagocytosis of polystyrene latex beads were isolated by fractionation in two discontinuous sucrose gradients. Both membrane preparations were free of any significant contamination by mitochondria, microsomes, and lysosomes. The primary plasma membranes from monolayer cells were richer in Na + + K +-activated ATPase, acetylcholinesterase, and choline acetyltransferase activities than those from spinner cells, but the polypeptide composition of these membranes was very similar in both types of culture as determined by polyacrylamide gel electrophoresis in buffers containing sodium dodecyl sulfate and urea. In contrast to these results, the internalized plasma membrane fraction from monolayer cells differed significantly from that isolated from spinner cells in its enzymatic properties and polypeptide composition. Furthermore, the effects of dibutyryl adenosine 3′,5′-monophosphate and sodium butyrate on these membrane properties were different in the two types of culture. With either monolayer or spinner cells, the internalized plasma membrane fraction differed in its enzymatic and polypeptide composition from the primary plasma membranes. These results support the conclusion that differentiation of the neuro-2A clone is accompanied by extensive changes in the organization and dynamics of the plasma membrane.