Encyclopedia of Signaling Molecules

2018 Edition
| Editors: Sangdun Choi

Pin1

Reference work entry
DOI: https://doi.org/10.1007/978-3-319-67199-4_581

Synonyms

Historical Background

Peptidyl-prolyl cis-trans isomerase NIMA-interacting 1 (PIn1) is a phosphorylation-dependent peptidyl-prolyl isomerase that was first isolated by a yeast two-hybrid screen designed to identify human proteins which interact with the product of the “never in mitosis” gene A (NIMA) (Lu et al. 1996). Sequence analysis revealed that human Pin1 exhibits approximately 45% amino acid sequence similarity with the product of the ESS1 gene that was previously identified as essential for growth in the budding yeast Saccharomyces cerevisiae (Hanes et al. 1989). It has subsequently been found that Pin1-like proteins are highly conserved and found in both eukaryotes and prokaryotes (Maruyama et al. 2004). While it initially implicated as a regulator of mitosis, it is evident that Pin1 has roles in a number of biological processes.

Pin1 is classified as a peptidyl-prolyl isomerase (PPIase), catalyzing the cis-trans conversion of the peptide bond between a proline and the preceding amino acid (Fig. 1). It is part of a large superfamily of PPIases, which is divided into three families: the cyclophilins, the FK506-binding proteins (FKBPs), and the parvulins (Gothel and Marahiel 1999). One distinguishing characteristic among the three families of PPIases is their substrate specificities, particularly concerning the residue directly preceding the proline. Like all members of the parvulin family, Pin1 has a preference for hydrophobic residues. However, unique to Pin1 is its phosphorylation dependence, requiring that the preceding residue be a phosphorylated serine or phosphorylated threonine (Yaffe et al. 1997). With phosphorylation playing a pivotal role in cell signaling, one can infer that an isomerase with phosphorylated residues in its specificity determinants would add an additional layer of complexity to signaling pathways. This has been shown in multiple cellular processes where Pin1-catalyzed isomerization regulates the conformation of key cellular proteins (Lu and Zhou 2007; Nakamura et al. 2012; Zhou and Lu 2016).
Pin1, Fig. 1

Peptidyl-prolyl isomerization. Cis-trans isomerization of the peptide bond (arrow) preceding the proline. Xxx represents any amino acid

Structural and Enzymatic Features of Pin1

High-resolution structures of Pin1 determined by x-ray crystallography revealed that it consists of two structural domains connected by a relatively short linker, which had originally been predicted from its primary sequence (Fig. 2). The N-terminal WW domain is named for two conserved tryptophan residues and comprises residues 1–39. It consists of a triple stranded antiparallel β-sheet, with a hydrophobic patch in the surface (Zhou et al. 1999). Generally described as a protein-protein interaction domain, the WW domain of Pin1 binds pSer/Thr-Pro motifs, thus facilitating interactions between Pin1 and its substrates (Lu et al. 1999). The 118 amino acid catalytic PPIase domain (residues 45–163) is found on the C-terminal end of the protein and consists of four antiparallel β-sheets and four α-helices. Within this domain are two relatively well-described regions, the proline-binding pocket and the phosphate-binding loop, which lie on opposite sides of the active site. The hydrophobic proline-binding pocket contains three highly conserved residues, Leu122, Met130, and Phe134, which are thought to be responsible for holding the proline in place during catalysis (Ranganathan et al. 1997). The phosphate-binding loop contains two positively charged arginine residues at positions 68 and 69, as well as another positively charged amino acid, lysine at position 63, conferring upon Pin1 its preference for phosphorylated residues preceding the proline (Zhou et al. 1999). A short linker connects the WW and PPIase domains, whose flexibility may contribute to the broad substrate specificity of Pin1 (Li et al. 2005). Interestingly, although both domains of Pin1 bind the pSer/Thr-Pro motif, it appears that they may bind differently since the WW domain typically has a higher binding affinity for peptides than the PPIase domain (Lu et al. 1999).
Pin1, Fig. 2

(a) Linear representation of Pin1. Phosphorylation sites and protein kinases responsible for phosphorylation of Pin1. (b and c) High-resolution structure of Pin1 determined by x-ray crystallography (PDB: 1NMV). (b) The WW domain is shown in blue, the linker region is yellow, and the PPIase domain is shown in red. (c) Key residues of the PPIase domain are highlighted (See text for details)

In spite of evidence confirming the ability of Pin1 to catalyze cis-trans isomerization (Stukenberg and Kirschner 2001; Nakamura et al. 2012), there are still questions concerning its precise catalytic mechanism. On the basis of a crystal structure of Pin1, Ranganathan et al. (1997) initially proposed a mechanism that involved the formation of a covalent enzyme-substrate intermediate with Cys113, His59, and His157 being key residues involved in catalysis. Since then, however, additional evidence has argued instead for a non-covalent mechanism. In this respect, Lippens et al. (2007) proposed that the role of Cys113 is to destabilize the peptide prolyl bond to allow for its rotation (Lippens et al. 2007). This hypothesis is supported by data from Behrsin et al. (2007) showing that the Cys113/Asp substitution did not abolish Pin1 function. Additionally, with regard to the histidine residues, it has been shown that they do not directly participate in catalysis, suggesting they instead act structurally to support the integrity of the active site (Bailey et al. 2008).

Physiological Regulation of Pin1

It appears that Pin1 is subject to regulation at a number of levels (Lu and Zhou 2007). Its expression is upregulated in response to growth factors through E2F-mediated transcription, an observation consistent with its role in the cell cycle (Lu et al. 2002). Pin1 is also regulated through posttranslational modifications including phosphorylation and oxidation that have been shown to modulate its activity or functional properties (Litchfield et al. 2015; Innes et al. 2015). For example, phosphorylation on Ser16 and Ser65 has opposing effects: the former prevents interactions with substrates, while the latter reduces ubiquitylation, thus increasing stability of Pin1. As illustrated in Fig. 2, Pin1 has been shown to be a substrate for a number of distinct protein kinases including PKA, COT, Aurora A, PLK1, and DAPK (reviewed in Litchfield et al. 2015). Oxidation of Pin1 may have a relationship to pathologies such as Alzheimer’s disease, rather than as a part of normal cell regulation (Sultana et al. 2006; Innes et al. 2015; Chen et al. 2015). In addition to these modifications, analysis of databases such as PhosphoSitePlus (Hornbeck et al. 2014) reveals that Pin1 is also potentially regulated by modification of numerous other sites.

Cellular Functions of Pin1

Pin1 is primarily localized in the nucleus; however, it can also be detected in the cytoplasm. This pervasive distribution of Pin1 is consistent with its extensive list of target proteins that are localized throughout the cell (Lu et al. 2002; Zhou and Lu 2016). As previously noted, Pin1 was first identified due to its interaction with NIMA, a protein kinase involved in mitotic regulation. This relationship was the first of many which suggested that Pin1 plays an integral role in regulation of the cell cycle and growth. Since then, Pin1 has been shown to be involved in a variety of additional cellular processes, emphasizing its diversity and importance (Fig. 3). Loss of function mutations, deletions, or knockdowns of Pin1 in yeast and mammalian cells provides striking evidence for its role in the cell cycle, as these cells undergo mitotic arrest and apoptosis (Lu et al. 2002). Furthermore, Pin1 has a lengthy list of substrates that are known to be involved in the cell cycle, including a number of mitotic regulatory proteins (e.g., CDC25 and WEE1) that are targets of proline-directed protein kinases, such as CDKs and MAPKs (Lu and Zhou 2007; Litchfield et al. 2015; Zhou and Lu 2016). Pin1-catalyzed isomerization of these phosphorylated sites may be responsible for coordinating the activity of mitotic proteins, thus allowing for progression through the cell cycle (Lu and Zhou 2007). Pin1 has also been shown to coordinate duplication of centrosomes, DNA synthesis (Suizu et al. 2006), and to assist in chromosome condensation (Xu and Manley 2007) further emphasizing its role in the cell cycle.
Pin1, Fig. 3

Selected Pin1 substrates and consequences of interaction

Similar to its actions in the cell cycle, Pin1 has been shown to interact with proteins involved in cell signaling events and pathways involving proline-directed protein kinases (Litchfield et al. 2015). One such example is the MAPK pathway, where, following proline-directed phosphorylation by MAPK, the proteins c-Jun and c-Fos are acted upon by Pin1 (Lu et al. 2007).

Adding to the growing list of functions, Pin1 has also been shown to regulate expression of some genes through regulation of their transcription factors (Lu et al. 2007), to assist in the maintenance of telomeres through interactions with TRF1 (Lee et al. 2009), to facilitate DNA repair through interactions with p53 (Takahashi et al. 2008), and finally to support breast development (Wulf et al. 2003; Rustighi et al. 2017).

Additionally, Pin1 has been shown to have specific roles in the immune and nervous systems. These additional functions provide links to the implication of Pin1 in various pathogenic conditions, which will be discussed in the following section. In short however, it has been shown that Pin1 is important for regulating transcription of cytokines in T cells, as well as for survival of eosinophils (Lu et al. 2007). The importance of Pin1 in the brain is evident in Pin1 knockout mice, which have progressive and age-related neurodegeneration. This is directly related to the ability of Pin1 to promote normal neuronal cell functioning and survival through the interaction with proteins such as Tau and amyloid precursor protein (APP) (Lu and Zhou 2007; Driver et al. 2015).

Pin1 in Pathogenesis of Human Disease

Considering the diversity of its roles and importance as a key regulator of many cellular and biological processes, it is not unexpected that Pin1 appears to be involved in various pathological conditions, including cancer, Alzheimer’s disease, and asthma. In this respect, Pin1 has been implicated in a variety of cancers, including breast, lung, colon, and prostate cancer (Lu and Zhou 2007; Driver et al. 2015; Zhou and Lu 2016). This is not surprising given its role as a regulator of the cell cycle. However, the precise role Pin1 plays in cancer is controversial, as levels of Pin1 have been shown to be either positively or negatively related to cancer (Yeh and Means 2007). One of the better understood pathways in which overexpression of Pin1 appears to participate in cancer involves cyclin D1. Not only can Pin1 increase expression of cyclin D1, Pin1 can also directly bind and stabilize cyclin D1 to enhance cyclin D1/CDK activity (Lu et al. 2006). Conversely, loss of Pin1 can suppress transformation by Neu or Ras (Ryo et al. 2002). Additionally, Pin1 has been shown to stabilize p53, an important tumor suppressor which promotes apoptosis in response to genotoxic stresses (Yeh and Means 2007). With regard to Alzheimer’s disease, the precise role of Pin1 in pathogenesis remains uncertain, although evidence suggests that various mechanisms in Alzheimer’s disease downregulate and/or inactivate Pin1 (e.g., through oxidation), suggesting it has a neuroprotective role (Lu and Zhou 2007). The loss of Pin1 function has impacts on two proteins, namely, APP and Tau, both found in senile plaques and neurofibrillary tangles. One model suggests that without Pin1, the pThr668-Pro motif of APP remains in the cis form and accumulates in plaques. Similarly, the Tau pThr231-Pro motif is also found mostly in the cis form, leading to its hyperphosphorylation and subsequent accumulation (Lu and Zhou 2007). The association between Pin1 and asthma relates to the role Pin1 plays in immune cell function. By regulating the release of cytokines from eosinophils, and participating in the apoptotic decision of both T cells and eosinophils, activated Pin1 modulates the allergic inflammatory response in the lungs associated with asthma (Esnault et al. 2008).

Emergence of Pin1 as a Candidate for Molecular-Targeted Therapy

The involvement of Pin1 in various human diseases, including cancer, makes it an obvious candidate for therapies (Zhou and Lu 2016). Additionally, the fact that other PPIase proteins, including cyclophilin and FKBP, have been shown to be targets of therapeutically effective agents lends support to attempts to achieve the same success with Pin1. The first general inhibitor of parvulins was juglone, and although it has the ability to irreversibly inhibit Pin1, its use as an anticancer therapy is limited by its nonspecificity (Wang and Etzkorn 2006). More recently, work has been focused on structure-based design of Pin1 inhibitors. Features that have been targeted by these rationally designed Pin1 inhibitors include its hydrophobic binding pocket, the phosphate-binding loop (Potter et al. 2010), or Cys113 within its active site (Dong et al. 2010). Thus far, these Pin1 inhibitors have had varying degrees of specificity, as well as issues with potency, degradation, and cell permeability. In addition to these inhibitors, there have also been efforts to isolate inhibitors in the form of cyclic peptides which are less likely to be subject to proteolysis and may bind Pin1 with a higher affinity due to their reduced flexibility (Liu et al. 2010; Duncan et al. 2011). Although some inhibitors are able to inhibit Pin1 at nanomolar concentrations, their usage currently appears to be more appropriate for further investigations regarding the cellular functions of Pin1, rather than as therapeutic agents. However, this does not preclude the notion of using them as models to guide the design of novel, potentially therapeutic inhibitors of Pin1 (Zhou and Lu 2016).

Summary

Since its discovery in 1996, much has been learned regarding the structure, function, and regulation of Pin1. In comparison to other PPIases, one particularly intriguing feature of Pin1 is its phosphorylation dependence which enables Pin1 to introduce an additional level of control in pathways involving proline-directed protein kinases such as CDKs that are central drivers of cell cycle progression. While Pin1 was initially implicated as a key regulator of mitosis, it has subsequently been shown to be important in a diverse array of cellular processes. In concert with its participation in a broad spectrum of biological events, it is noteworthy that Pin1 has been implicated in a variety of diseases including cancer and neurological disorders such as Alzheimer’s disease and asthma. Pin1 has thus emerged as a potential candidate for molecular-targeted therapy. Consequently, it can be anticipated that ongoing efforts to understand its regulation and functions and to elucidate its precise catalytic mechanism will foster efforts to develop new approaches that will harness its promise as a therapeutic target.

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Copyright information

© Springer International Publishing AG 2018

Authors and Affiliations

  1. 1.Department of BiochemistrySchulich School of Medicine and Dentistry, The University of Western OntarioLondonCanada
  2. 2.Department of OncologySchulich School of Medicine and Dentistry, The University of Western OntarioLondonCanada