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

Abstract

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.

The Xenopus Suc1/Cks Protein Promotes the Phosphorylation of G2/M Regulators

“…2B). Previously, the endogenous concentration of Pin1 in Xenopus egg extracts was estimated to be 0.5 M or 9 g/ml (20). Furthermore, recombinant Pin1 at a final concentration of 10 M (180 g/ml) completely blocked the entry of egg extracts into mitosis (20).…”

“…Overexpression of Pin1 in human cells (18) and Xenopus egg extracts inhibits the entry into mitosis (20,21). Depletion of Pin1 from human cells by expression of an antisense construct and deletion of the gene encoding the budding yeast Pin1 homologue Ess1 both result in a defect in the progression through mitosis (18).…”

“…Depletion of Pin1 from HeLa cells results in a mitotic arrest, whereas overexpression of Pin1 in HeLa cells causes a G 2 arrest (18). Similarly, the addition of excess recombinant human His6-Pin1 protein to cycling Xenopus egg extracts can block the entry of these extracts into mitosis (20,21). These studies indicated that Pin1 can impinge upon the Cdc2/cyclin B-dependent pathway of mitotic regulation.…”

Gephyrin: a central GABAergic synapse organizer

“…88,89 These conformational changes profoundly affect on the function of Pin1 substrates by modulating their catalytic activity, phosphorylation status, protein interactions and/or protein stability. 90 Gephyrin undergoes proline-directed phosphorylation and interacts with Pin1 via a large region that encompasses the E domain and part of the C domain, eliciting conformational changes in gephyrin.…”

Pin1 overexpression is associated with poor differentiation and survival in oral squamous cell carcinoma

“…The PPIase domain has catalytic activity that specifically isomerizes Ser/Thr-Pro peptide bonds (4). This novel post-phosphorylation isomerase regulates the conformation of a subset of phosphoproteins such as p53, Cdc25C and the microtubule-associated protein tau, thereby affecting their activity and/or protein-protein interactions (1,(9)(10)(11)(12)(13)(14). Pin1 has been shown to interact with a number of cancer-related phosphoproteins, which suggested Pin1 might link signal transduction to pathogenesis of cancer (1,15,16).…”