THE ANNEXIN II(2)P11(2) COMPLEX IS THE MAJOR PROTEIN-COMPONENT OF THETRITON X-100-INSOLUBLE LOW-DENSITY FRACTION PREPARED FROM MDCK CELLS IN THE PRESENCE OF CA2+

Annexin II(2)p11(2) is present in the submembranous region of cells ex pressing both subunits of the complex. Most probably, this subcellular distribution is maintained through the interaction of annexin II(2)p1 1(2) with membrane phospholipids and/or elements of the cortical cytos keleton known to occur in vitro in a Ca2+-dependent manner. To determi ne whether membrane or cytoskeleton interactions are primarily respons ible for anchoring annexin II(2)p11(2) in the cell cortex, we subjecte d Madin-Darby canine kidney (MDCK) cells to serial extractions using d ifferent detergents and identified annexin II and p11 in the different fractions employing specific antibodies. The complex but not monomeri c annexin II remains insoluble when the cells are extracted with Trito n X-100 in the presence of Ca2+. However, treatment of the Triton X-10 0-insoluble cell remnants with a series of other detergents known to s olubilize GPI-anchored proteins leads to a partial extraction of annex in II(2)p11(2) even in the presence of Ca2+. Using sucrose density gra dient analysis in the presence of Ca2+ as a different means of fractio nating the Triton X-100-insoluble cell remnants we show that the major ity of annexin II(2)p11(2) copurifies with a low-density fraction whic h has been reported to contain GPI-anchored proteins, certain glycolip ids, and VIP21/caveolin. Annexin II(2)p11(2) is by far the most abunda nt protein component in this fraction indicating that its association with the low-density material occurs via lipid binding and is not due to the interaction with a certain protein.