The recycling pathway of protein ERGIC-53 and dynamics of the ER-Golgi intermediate compartment

To establish recycling routes in the early secretory pathway we have studie d the recycling of the ER-Golgi intermediate compartment (ERGIC) marker ERG IC-53 in HepG2 cells. Immunofluorescence microscopy showed progressive conc entration of ERGIC-53 in the Golgi area at 15 degrees C. Upon rewarming to 37 degrees C ERGIC-53 redistributed into the cell periphery often via tubul ar processes that largely excluded anterograde transported albumin. Immunog old labeling of cells cultured at 37 degrees C revealed ERGIC-53 predominan tly in characteristic P-COP-positive tubulo-vesicular clusters both near th e Golgi apparatus and in the cell periphery. Concentration of ERGIC-53 at 1 5 degrees C resulted from both accumulation of ERGIC-53 in the ERGIC and mo vement of ERGIC membranes closer to the Golgi apparatus. Upon rewarming to 37 degrees C the labeling of ERGIC-53 in the ERGIC rapidly returned to norm al levels whereas ERGIC-53's labeling in the cis-Golgi was unchanged. Tempe rature manipulations had no effect on the average number of ERGIC-53 cluste rs. Density gradient centrifugation indicated that the surplus ERGIC-53 acc umulating in the ERGIC at 15 degrees C was rapidly transported to the ER up on rewarming. These results suggest that the ERGIC is a dynamic membrane sy stem composed of a constant average number of clusters and that the major r ecycling pathway of ERGIC-53 bypasses the Golgi apparatus.