Fig. 9

An Arf6-DN mutant blocks is localized at the cell surface and on lysosomes. SN56 cells were transfected with HA-βAPP-CFP, LAMP1-mRFP, and Arf6 bearing either a dominant negative GFP-Arf6-T27N-DN or constitutively active GFP-Arf6-Q67L-CA mutation. Cells were surface labeled Alexa Fluor 647 labeled anti-HA antibodies on ice and then allowed to internalize at 37 °C and imaged by confocal microscopy. a Images show the individual color channels of LAMP1, HA (surface-labeled APP), and CFP (APP) along the top row. On the bottom row is the distribution of GFP-Arf6-DN by itself, and merged each of the above channeled. Arf6-DN protein appears on the cell surface and in large intracellular organelles, colocalized with LAMP1 (arrowheads), surface-labeled HA-APP (arrowheads) and with the CFP tag on APP (arrowheads). b The constitutively active GFP-Arf6-Q67L-CA, is also present at the cell surface, the cytosol, and on many intracellular organelles. It demonstrates on a much less localization with LAMP1 (arrowheads), surface-labeled HA-APP (arrowheads) or the CFP tag on APP (arrowheads)