RECOMBINANT EXPRESSION OF THE BRUSH-BORDER MYOSIN-I HEAVY-CHAIN

Although the specific functions of myosin I motors are not known, thei r localization to membrane structures suggests a function in membrane motility. Different myosin I isoforms in the same cell or in different cells can possess different localizations. To determine if the locali zation and biochemical activity of the best-characterized mammalian my osin I, chicken intestinal epithelium brush border myosin I, was depen dent on determinants of the membrane or actin cytoskeleton specific to epithelial cells, we transfected the cDNA for the heavy chain of this myosin into COS cells. Transient transfection of COS cells with the c hicken brush border myosin I heavy chain resulted in the production of recombinant myosin I. Recombinant brush border myosin I localized to protrusions of the plasma membrane, particularly at spreading cell edg es, and also to unknown cytoplasmic structures. Some cells expressing particularly high levels of brush border myosin I possessed a highly i rregular surface. Recombinant brush border myosin I purified from COS cells bound to actin filaments in an ATP-dependent manner and decorate d actin filaments to form a characteristic appearance. The recombinant myosin also catalyzed calcium-sensitive, actin-activated MgATPase act ivity similar to that of the native enzyme. Thus, any cellular factor required for the general membrane localization or biochemical activity of brush border myosin I is present in COS cells as well as intestina l epithelium. (C) 1995 Wiley-Liss, Inc.