Peptidyl-prolyl Isomerase Pin1 Research Articles

The essential mitotic peptidyl-prolyl isomerase Pin1 binds and regulates mitosis-specific phosphoproteins.

Phosphorylation of mitotic proteins on the Ser/Thr-Pro motifs has been shown to play an important role in regulating mitotic progression. Pin1 is a novel essential peptidyl-prolyl isomerase (PPIase) that inhibits entry into mitosis and is also required for proper progression through mitosis, but its substrate(s) and function(s) remain to be determined. Here we report that in both human cells and Xenopus extracts, Pin1 interacts directly with a subset of mitotic phosphoproteins on phosphorylated Ser/Thr-Pro motifs in a phosphorylation-dependent and mitosis-specific manner. Many of these Pin1-binding proteins are also recognized by the monoclonal antibody MPM-2, and they include the important mitotic regulators Cdc25, Myt1, Wee1, Plk1, and Cdc27. The importance of this Pin1 interaction was tested by constructing two Pin1 active site point mutants that fail to bind a phosphorylated Ser/Thr-Pro motif in mitotic phosphoproteins. Wild-type, but not mutant, Pin1 inhibits both mitotic division in Xenopus embryos and entry into mitosis in Xenopus extracts. We have examined the interaction between Pin1 and Cdc25 in detail. Pin1 not only binds the mitotic form of Cdc25 on the phosphorylation sites important for its activity in vitro and in vivo, but it also inhibits its activity, offering one explanation for the ability of Pin1 to inhibit mitotic entry. In a separate paper, we have shown that Pin1 is a phosphorylation-dependent PPIase that can recognize specifically the phosphorylated Ser/Thr-Pro bonds present in mitotic phosphoproteins. Thus, Pin1 likely acts as a general regulator of mitotic proteins that have been phosphorylated by Cdc2 and other mitotic kinases.

Structural and Functional Analysis of the Mitotic Rotamase Pin1 Suggests Substrate Recognition Is Phosphorylation Dependent

The human rotamase or peptidyl-prolyl cis-trans isomerase Pin1 is a conserved mitotic regulator essential for the G2/M transition of the eukaryotic cell cycle. We report the 1.35 Å crystal structure of Pin1 complexed with an AlaPro dipeptide and the initial characterization of Pin1's functional properties. The crystallographic structure as well as pH titration studies and mutagenesis of an active site cysteine suggest a catalytic mechanism that includes general acid-base and covalent catalysis during peptide bond isomerization. Pin1 displays a preference for an acidic residue N-terminal to the isomerized proline bond due to interaction of this acidic side chain with a basic cluster. This raises the possibility of phosphorylation-mediated control of Pin1–substrate interactions in cell cycle regulation.