IDENTIFICATION OF THE BASOLATERAL TARGETING DETERMINANT OF A PERIPHERAL MEMBRANE-PROTEIN, MACMARCKS, IN POLARIZED CELLS

Background: Although the molecular determinants that specify the targe ting of transmembrane proteins to the apical or basolateral membrane d omains within polarized epithelial cells have been well characterized, very little is known about the targeting of peripheral membrane prote ins within these cells. MacMARCKS is a member of the MARCKS family of protein kinase C (PKC) substrates. This myristoylated protein regulate s actin structure at cell membranes and is essential for the morphogen ic movement of neuroepithelial cells during the formation of the neura l tube, Results: MacMARCKS was specifically targeted to sites of cell- cell contact in the basolateral domain of polarized Madin-Darby canine kidney (MDCK) epithelial cells and was displaced from this location u pon activation of PKC. We defined the basolateral targeting determinan t of MacMARCKS to be the effector domain, a basic region spanning 24 a mino acids and containing the PKC phosphorylation sites as well as bin ding sites for calmodulin and actin. This domain, in conjunction with a myristoyl moiety, was sufficient to target a non-membrane-associated protein-green fluorescent protein-specifically to the basolateral sur face of polarized MDCK cells. Conclusions: This is the first descripti on of a specific amino acid sequence that specifies targeting of a per ipheral membrane protein to the basolateral membrane in polarized epit helial cells.