CAVEOLAE AND ENDOPLASMIC-RETICULUM - IMMUNOFLUORESCENCE MICROSCOPY AND TIME-LAPSE ANALYSIS

Caveolae have been hypothesized to be involved in Ca2+ signaling. By e lectron microscopy, caveolae were observed to be apposed to the endopl asmic reticulum (ER), which is a major intracellular Ca2+ pool. In the present study, we examined the relationship between caveolae and the ER when the distribution of the latter was changed by depolymerization of microtubules. Double immunofluorescence microscopy for detection o f caveolin and the ER antigens, and time-lapse observation of green fl uorescent protein (GFP)-tagged caveolin were employed. in normal human fibroblasts and PtK2 cells, the ER was seen as a network extending th roughout the cytoplasm, and most caveolin occurred in patches along th e cell edge. When microtubules were depolymerized by Colcemid or nocod azole, the ER became retracted from the cell periphery and aggregated around the nucleus; in the same cells, caveolin was not seen along the cell edge, but was aligned along the edge of the retracted ER. By tim e-lapse analysis, GFP-caveolin expressed in PtK2 cells was observed to move from the cell periphery toward the cell center in Colcemid-treat ed cells. The result shows that the apposition of caveolae and the ER is maintained even after the gross distributional change, and suggests a mechanical linkage between the two organelles.