ASSESSMENT OF HETEROLOGOUS MEMBRANE-PROTEIN POLARITY IN TRANSIENTLY TRANSFECTED MDCK CELLS

We have evaluated transient transfection of MDCK cells by the DEAE-dex tran/chloroquine method as a rapid method for study of heterologous pl asma membrane protein polarity. Transiently transfected cells reseeded onto permeable supports formed confluent monolayers with normal tight junctions and normal distribution of endogenous apical and basolatera l surface markers. Transfected monolayers reseeded onto opaque polycar bonate filters attained cell heights 3 times greater than on transpare nt filters. Conventional and confocal immunofluorescence microscopy we re used to assess polarity of transient expression of heterologous pro teins previously defined in stably transfected cell lines as apical (D AF-CD55), basolateral (VSV-G), and nonpolarized (CD7) in distribution. Through each transiently expressed protein exhibited a polarity pheno type in most cells which resembled the stable phenotype, consistency o f polarized localization was less than in stably transfected cells. Si milar results were obtained by lipofection. We conclude that transient transfection of MDCK cells may be useful as a rapid screen, but is no t sufficiently reliable for definitive assessment of heterologous memb rane proein polarity.