Vimentin dephosphorylation by protein phosphatase 2A is modulated by the targeting subunit B55.
The intermediate filament protein vimentin is a major phosphoprotein in mammalian fibroblasts, and reversible phosphorylation plays a key role in its dynamic rearrangement. Selective inhibition of type 2A but not type 1 protein phosphatases led to hyperphosphorylation and concomitant disassembly of vimentin, characterized by a collapse into bundles around the nucleus. We have analyzed the potential role of one of the major protein phosphatase 2A (PP2A) regulatory subunits, B55, in vimentin dephosphorylation. In mammalian fibroblasts, B55 protein was distributed ubiquitously throughout the cytoplasm with a fraction associated to vimentin. Specific depletion of B55 in living cells by antisense B55 RNA was accompanied by disassembly and increased phosphorylation of vimentin, as when type 2A phosphatases were inhibited using okadaic acid. The presence of B55 was a prerequisite for PP2A to efficiently dephosphorylate vimentin in vitro or to induce filament reassembly in situ. Both biochemical fractionation and immunofluorescence analysis of detergent-extracted cells revealed that fractions of PP2Ac, PR65, and B55 were tightly associated with vimentin. Furthermore, vimentin-associated PP2A catalytic subunit was displaced in B55-depleted cells. Taken together these data show that, in mammalian fibroblasts, the intermediate filament protein vimentin is dephosphorylated by PP2A, an event targeted by B55.
Turowski
P
P
Cell Biology Unit, Institut de Genetique Humaine, Centre National de la Recherche Scientifique UPR 1142, F-34396 Montpellier Cedex 5, France.
Myles
T
T
Hemmings
B A
BA
Fernandez
A
A
Lamb
N J
NJ
D000935:Antifungal Agents / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D007382:Intermediate Filaments / Q000378:metabolism* / Q000648:ultrastructure; D007399:Interphase / Q000502:physiology; D019319:Okadaic Acid / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D054648:Protein Phosphatase 2; D011714:Pyrans; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013141:Spiro Compounds; D014746:Vimentin / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure
D000935:Antifungal Agents; D004791:Enzyme Inhibitors; D011714:Pyrans; D011994:Recombinant Proteins; D013141:Spiro Compounds; D014746:Vimentin; C053079:tautomycin; D019319:Okadaic Acid; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2