DIFFERENTIAL COLOCALIZATION OF PROFILIN WITH MICROFILAMENTS IN PTK2 CELLS

Profilins are thought to be involved in the control of actin dynamics in eukaryotic cells. in accordance with this concept, profilin was fou nd to be colocalized with the cortical microfilament webs in leading l amellae of locomoting and spreading fibroblasts. However, so far, ther e is little information on the distribution of profilin in other cell types. In this study, we report on the colocalization of profilin with various microfilament suprastructures in the epithelial cell line PtK 2. This cell line, which is derived fr-um rat kangaroo, contains a pro filin sharing an N-terminal epitope with bovine and human profilin I, as seen by immunoblotting with monoclonal antibodies. By using immunof luorescence in conjunction with conventional fluorescence microscopy a nd confocal laser-scanning microscopy, we found profilin in ruffling a reas of the peripheral lamellae and nascent stress fibres of spreading cells, whereas the peripheral belts of stationary cells growing in ep ithelioid sheets lacked profilin staining. In these cells. profilin wa s primarily distributed in a fine reticular or vesicular network that was not related to the microfilament system. Conspicuously low levels of profilins were observed in the contractile ring of mitotic cells. T his was found for different fixation protocols and antibodies of the I gG and IgM type, respectively. indicating that lack of staining of the cleavage furrow was not due to antibody penetration problems. Dependi ng on the fixation protocol, the nuclear matrix appeared strongly posi tive or negative for profilin. Cells microinjected with birch pollen p rofilin and labeled with a birch profilin-specific monoclonal antibody corroborated the results obtained with the endogeneous protein: The i njected profilin was targeted to the cortical web and to nascent stres s fibers of spreading cells but not to thee cleavage ring: of mitotic cells. These results suggest that high concentrations of a profilin 1 homologue are preferentially located with those microfilament suprastr uctures in PtK2 cells that are subject to rapid modulation by external signals. (C) 1997 Wiley-Liss, Inc.