Role of Protein Phosphatase‐1 Inhibitor‐3 in the Isoform‐Selective Expression, Subcellular Targeting and Nuclear/Nucleolar Transport of Protein Phosphatase‐1

Abstract

Inhibitor-3 (Inh3) an inhibitor protein of protein phosphatase-1 (PP1) which is localized to the nucleolus and to the centrosomes. Three closely related isoforms of PP1, α, β, and γ1, are ubiquitously expressed in mammalian tissue. Our data show that Inh3 is co-localized with PP1α in the centrosome and with PP1γ1 in nucleoli, i.e., Inh3 exhibits a specificity for these two isoforms. The localization of Inh3 to the nucleoli is dependent on two discrete localization signals, one for nuclear targeting and one for nucleolar targeting. Attempts to study the localization of PP1 isoforms by overexpression of red fluorescent protein (DsRed)-tagged PP1 isoforms in HEK 293 cells revealed they could not be overexpressed. However, DsRed-tagged isoforms α and γ1, but not isoform β, are expressed when green fluorescent protein (GFP)-Inh3 is co-expressed. The co-expression of PP1 can be disrupted by the mutation of two PP1 binding regions in Inh3. Furthermore, isoform α is co-localized with Inh3 to the nuclei and isoform γ1 is co-localized with Inh3 to the nuclei and nucleoli. DsRed-PP1α did not exhibit any fluorescence in the centrosomes even when co-expressed with GFP-Inh3. These findings provide evidence that 1) Inh3 may function as a chaperone protein for PP1α and γ1, 2) Inh-3 is required for their nuclear/nucleolar transport. (Supported by Philip Morris USA Inc. and Philip Morris International and by NIH DK 18512.)