Domains of type 1 protein phosphatase inhibitor-2 required for nuclear and cytoplasmic localization in response to cell-cell contact.
Inhibitor-2 of type 1 protein phosphatase is a phosphoprotein conserved among all eukaryotes, and it appears in both the nucleus and cytoplasm of tissue culture cells. We discovered that endogenous inhibitor-2 is concentrated in the nucleus of cells cultured at low density, whereas cells growing at high density excluded inhibitor-2 from the nucleus. There was rapid redistribution of inhibitor-2 when cells were replated at low or high density. Localization of myc-tagged forms of inhibitor-2 showed that residues 119-197 were required for nuclear accumulation in low-density cells and residues 78-119 were required for cytoplasmic localization in high-density cells. Fusion of inhibitor-2 residues 78-119 to green fluorescent protein was sufficient to produce cytoplasmic retention. Inhibitor-2 fused to triple tandem green fluorescent protein (100 kDa) was imported into the nucleus of low-density cells but was not excluded from the nucleus when cells reached high density, implying that inhibitor-2 was actively imported into the nucleus but exited by passive diffusion instead of active export. We conclude that inhibitor-2 contains two separate domains that control its localization in the nucleus or cytoplasm. This change in inhibitor-2 localization may direct inhibitor-2 to different forms of protein phosphatase 1 or change the localization of protein phosphatase, as part of the cellular response to cell-cell contacts at high density.
Leach
Craig
C
Center for Cell Signaling, University of Virginia School of Medicine, Box 800577-MSB7225, Charlottesville VA 22908, USA. cal4a@virginia.edu
Eto
Masumi
M
Brautigan
David L
DL
D021581:Active Transport, Cell Nucleus / Q000187:drug effects / Q000502:physiology; D000595:Amino Acid Sequence / Q000502:physiology; D020134:Catalytic Domain / Q000502:physiology; D002450:Cell Communication / Q000502:physiology*; D002451:Cell Compartmentation / Q000502:physiology; D002467:Cell Nucleus / Q000201:enzymology*; D003593:Cytoplasm / Q000201:enzymology*; D005057:Eukaryotic Cells / Q000201:enzymology*; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D008164:Luminescent Proteins; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D054645:Protein Phosphatase 1; D017434:Protein Structure, Tertiary / Q000502:physiology; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D000097847:Protein Phosphatase Inhibitory Proteins
D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; D011506:Proteins; D011993:Recombinant Fusion Proteins; C024833:protein phosphatase inhibitor-2; D000097847:Protein Phosphatase Inhibitory Proteins