Disruption of the in vivo distribution of the intermediate filaments in fibroblasts through the microinjection of a specific monoclonal antibody

Abstract

Monoclonal antibodies (JLB1 and JLB7) that recognize minor components of the intermediate filament system of cultured cells were introduced into living fibroblasts by microinjection. Several minutes after injection of the JLB7 antibody virtually all of the intermediate filaments of the cells were found to be aggregated into tight bundles near or around the nucleus. In contrast, injection of the JLB1 antibody caused little or no aggregation of the intermediate filaments. Electron microscopy showed that the perinuclear bundles that formed after injection of the JLB7 antibody each consisted of ten or more filaments apparently crosslinked together. Double-label immunofluorescence microscopy showed that virtually all of the vimentin-containing intermediate filaments in the JLB7 antibody-injected cells were redistributed to the perinuclear region and remained there for at least 24 hr. The distributions of actin microfilaments and microtubules were seemingly undisturbed following microinjection. No obvious changes in cell morphology or behavior were apparent in the cells injected with JLB7 antibody; the cells displayed a flat appearance, showed a polarity, were able to ruffle and bleb and even appeared to show the normal saltatory movements of intracellular vesicles, granules and mitochondria, suggesting that intermediate filaments are not involved in these activities. The microinjection of highly specific monoclonal antibodies that recognize and alter components of the cell provides an additional approach to determine the in vivo functions of intracellular elements.