Rearrangement of the keratin cytoskeleton after combined treatment with microtubule and microfilament inhibitors.

Abstract

In addition to containing microtubule and microfilament systems, vertebrate epithelial cells contain an elaborate keratin intermediate-filament cytoskeleton . Little is known about its structural organization or function . Using indirect immunofluorescence microscopy with an antikeratin antiserum probe, we found that destabilization of microtubules and microfilaments with cytostatic drugs induces significant alterations in the cytoskeletal organization of keratin filaments in HeLa and fetal mouse epidermal cells . Keratin filament organization was observed to undergo a rapid (1-2 h) transition from a uniform distribution to an open lattice of keratin fibers stabilized by membrane-associated focal centers . Since addition of any one drug alone did not elicit significant organizational change in the keratin cytoskeleton, we suggest that microfilaments and microtubules have a combined role in maintaining the arrangement of keratin in these cells . Vimentin filaments, the only other intermediatesized filaments found in HeLa cells, did not co-distribute with keratin in untreated or drugtreated cells . These findings offer a new way to approach the study of the dynamics and functional roles of the keratin cytoskeleton in epithelial cells .

De novo formation of cytokeratin filament networks originates from the cell cortex in A‐431 cells

“…Our finding that filament network dynamics are controlled from the cell cortex by affecting successful ligation of rodlets into networks and by determining CKF extension vs. retraction, identifies the cell cortex as an organizing center where, depending on the mitotic stage, differential decisions are made. This contrasts with previous studies that implicated the nucleus and cytoplasmic foci, or specialized membrane sites acting as initiation sites for IF network formation [Eckert et al, 1982;Knapp et al, 1983;Kreis et al, 1983;Celis et al, 1984;Bologna et al, 1986;Albers and Fuchs, 1989;Magin et al, 1990;Raats et al, 1990;Sarria et al, 1990;Djabali, 1999]. By immunoelectronmicroscopy, we found strong labeling of a circumferential layer of non-filamentous material just below the actin cortical mesh after complete filament breakdown (not shown).…”

“…Focal organizing centers have been proposed to act as initiation sites for IF re-formation, among them the nucleus [Eckert et al, 1982;Albers and Fuchs, 1989], certain Contract grant sponsor: Stiftung Rheinland-Pfalz für Innovation; Contract grant sponsor: German Research Council; Contract grant number: LE 566/7-1. cytoplasmic sites [Kreis et al, 1983;Celis et al, 1984;Magin et al, 1990;Raats et al, 1990;Sarria et al, 1990], or plasmamembrane domains including desmosomes and hemidesmosomes in epithelial cells [Knapp et al, 1983;Bologna et al, 1986]. There is, however, no satisfactory explanation as to how IF formation may be regulated from any of these focal points, as there is common agreement that, due to the absence of defined ends, filament growth is facilitated by lateral exchange of subunits [Vikstrom et al, 1989;Ngai et al, 1990;Miller et al, 1993;Herrmann and Aebi, 1998;Julien, 1999] and not by end-on elongation and shortening as is the case for actin filaments and microtubules.…”

Disorganization of microfilaments and intermediate filaments interferes with the assembly and stability of desmosomes in MDCK epithelial cells

“…These associations between cytokeratin and actin filaments were only observed in drug-treated cultures and were not apparent in control cells suggesting that the proper arrangement of the microfilament network is required to support the organization of cytokeratin filaments in MDCK cells. These observations are consistent with previous studies which have suggested the possibility of an interaction between intermediate filaments and microfilaments that has yet to be characterized [Green et al, 1986[Green et al, , 1987Kitajima et al, 1986;Knapp et al, 1983;Roscioli and Mullins, 19891.…”

Intermediate filaments.

“…The collapse of vimentin IF into the nuclear region on treatment of mesenchymal cells with colchicine suggests close association vith MT . In epithelial cells, Ca" ions, injection of antikeratin antibodies, and combinations of MT and MF inhibitors (7,27,29) can cause the reversible collapse of the IF networks . The common phenomenen ofIF collapse into the perinuclear region caused by various factors has suggested the existence of IF-organizing centers that direct or regulate the cytoplasmic distribution and even synthesis of IF (7).…”