This paper investigates the mechanism of action of heavy ion radiation (HIR) on mouse testes. The testes of male mice subjected to whole body irradiation with carbon ion beam (0.5 and 4Gy) were analyzed at 7days after irradiation. A two-dimensional gel electrophoresis approach was employed to investigate the alteration of protein expression in the testes. Spot detection and matching were performed using the PDQuest 8.0 software. A difference of more than threefold in protein quantity (normalized spot volume) is the standard for detecting differentially expressed protein spots. A total of 11 differentially expressed proteins were found. Protein identification was performed using matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry (MALDI-TOF-TOF). Nine specific proteins were identified by searching the protein sequence database of the National Center for Biotechnology Information. These proteins were found involved in molecular chaperones, metabolic enzymes, oxidative stress, sperm function, and spermatogenic cell proliferation. HIR decreased glutathione activity and increased malondialdehyde content in the testes. Given that Pin1 is related to the cell cycle and that proliferation is affected by spermatogenesis, we analyzed testicular histological changes and Pin1 protein expression through immunoblotting and immunofluorescence. Alterations of multiple pathways may be associated with HIR toxicity to the testes. Our findings are essential for studies on the development, biology, and pathology of mouse testes after HIR in space or radiotherapy.

Microtubule assembly and disassembly is required for the maintenance of cell structure, mobility, and division. However, the cellular and biochemical implications of microtubule disruption are not fully understood. Using a proteomic approach, we found that the peptidyl-prolyl isomerase, cyclophilin A, was increased in plasma membrane extracts from chronic myeloid leukemia cells after microtubule disruption. In addition, we found that two peptidyl-prolyl isomerases, cyclophilin A and pin1, are overexpressed up to 10-fold in hematological malignancies compared with normal peripheral blood mononuclear cells. Although previous reports suggest that cyclophilin A is localized to the cytosol of mammalian cells, we found that cyclophilin A and pin1 are both localized to the nucleus and nuclear domains in hematopoietic cells. Microtubule disruption of hematopoietic cells caused a dramatic subcellular redistribution of cyclophilin A and pin1 from the nucleus to the cytosol and plasma membrane. We suggest that this accounts for the increased cyclophilin A at the plasma membrane of chronic myeloid leukemia cells after microtubule disruption. The subcellular redistribution of cyclophilin A and pin1 occurred in a c-Jun NH(2)-terminal kinase- and serine protease-dependent manner. Moreover, the altered subcellular localization of the peptidyl-prolyl isomerases occurred in a dose- and time-dependent manner after microtubule disruption and was found to correlate with G(2)/M arrest and precede induced cell death. These results suggest that the function of peptidyl-prolyl isomerases may be influenced by microtubule dynamics throughout the cell cycle, and their altered localization may be an important part of the mechanism by which microtubule-disrupting agents exert their cytostatic effects.

Pin1 is a peptidylprolyl cis/trans isomerase and it has a unique enzymatic activity of catalyzing isomerization of the peptide bond between phospho-serine/threonine and proline. Through the conformational change of its substrates, Pin1 regulates diverse biological processes including adipogenesis. In mouse embryonic fibroblasts and 3T3-L1 preadipocytes, overexpression of Pin1 enhances adipocyte differentiation whereas inhibition of Pin1 activity suppresses it. However, the precise functions of Pin1 during adipogenesis are not clear. In the present study, we investigated the potential targets of Pin1 during adipogenesis. We found that Pin1 interacts directly with and regulates the transcriptional activity of PPARγ, a key regulator of adipogenesis. In addition, ERK activity and Ser273 of PPARγ, a potential ERK phosphorylation target site, are important for the regulation of PPARγ function by Pin1 in 3T3-L1 cells. Taken together our results suggest a novel regulatory mechanism of Pin1 during adipogenesis, in which Pin1 enhances adipocyte differentiation by regulating the function of PPARγ.

Transcription factor LSF is essential for cell cycle progression, being required for activating expression of the thymidylate synthase (Tyms) gene at the G1/S transition. We previously established that phosphorylation of LSF in early G1 at Ser-291 and Ser-309 inhibits its transcriptional activity and that dephosphorylation later in G1 is required for its reactivation. Here we reveal the role of prolyl cis-trans isomerase Pin1 in activating LSF, by facilitating dephosphorylation at both Ser-291 and Ser-309. We demonstrate that Pin1 binds LSF both in vitro and in vivo. Using coimmunoprecipitation assays, we identify three SP/TP motifs in LSF (at residues Ser-291, Ser-309, and Thr-329) that are required and sufficient for association with Pin1. Co-expression of Pin1 enhances LSF transactivation potential in reporter assays. The Pin1-dependent enhancement of LSF activity requires residue Thr-329 in LSF, requires both the WW and PPiase domains of Pin1, and correlates with hypophosphorylation of LSF at Ser-291 and Ser-309. These findings support a model in which the binding of Pin1 at the Thr-329-Pro-330 motif in LSF permits isomerization by Pin1 of the peptide bonds at the nearby phosphorylated SP motifs (Ser-291 and Ser-309) to the trans configuration, thereby facilitating their dephosphorylation.

Abnormal cell cycle events are increasingly becoming important attributes of neurodegenerative pathology. Pin1 is a crucial target of neurodegeneration in relation to its functions regarding these abnormal cell cycle events in neurons. Pin1 is majorly involved in many aspects of cell cycle regulation and it has also been suggested to have a neuroprotective function against neurodegenerative pathologies. Oxidative dysregulation of Pin1 affects not only normal tau regulation, eventually causing tangle formation, but also cell cycle regulation in neurons. Presence of cell cycle proteins has been shown in many neurodegenerative diseases. Importantly, many of these proteins have physical interactions with Pin1. Hence, understanding Pin1's role in abnormal cell cycle re-entry is critical in terms of finding new approaches for the future therapeutic options treating neurodegenerative pathologies. Here, we show that inhibition of Pin1 by its selective inhibitor juglone leads to up-regulation of cyclinD1, phospho-tau, and caspase 3, producing apoptosis in cultured rat hippocampal neurons. We also observed axonal retraction with a change in sub-cellular localizations of cyclins. Therefore, Pin1 dysregulation, in relation to its role in cell cycle regulation in neurons, may have profound effects in the progression of neurodegenerative pathology, making it a possible crucial target behind many neurodegenerative diseases.

The canonical Wnt signaling pathway, in which β-catenin nuclear localization is a crucial step, plays an important role in osteoblast differentiation. Pin1, a prolyl isomerase, is also known as a key enzyme in osteogenesis. However, the role of Pin1 in canonical Wnt signal-induced osteoblast differentiation is poorly understood. We found that Pin1 deficiency caused osteopenia and reduction of β-catenin in bone lining cells. Similarly, Pin1 knockdown or treatment with Pin1 inhibitors strongly decreased the nuclear β-catenin level, TOP flash activity, and expression of bone marker genes induced by canonical Wnt activation and vice versa in Pin1 overexpression. Pin1 interacts directly with and isomerizes β-catenin in the nucleus. The isomerized β-catenin could not bind to nuclear adenomatous polyposis coli, which drives β-catenin out of the nucleus for proteasomal degradation, which consequently increases the retention of β-catenin in the nucleus and might explain the decrease of β-catenin ubiquitination. These results indicate that Pin1 could be a critical target to modulate β-catenin-mediated osteogenesis.

Parvulins belong to a family of peptidyl-prolyl cis/trans isomerases (PPIases) that catalyze the cis/trans conformations of prolyl-peptidyl bonds. Herein, we characterized two novel parvulins, TbPIN1 and TbPAR42, in Trypanosoma brucei. TbPIN1, a 115 amino-acid protein, contains a single PPIase domain but lacks the N-terminal WW domain. Using NMR spectroscopy, TbPIN1 was found to exhibit PPIase activity toward a phosphorylated substrate. Overexpression of TbPIN1 can rescue the impaired temperature-sensitive phenotype in a mutant yeast strain. TbPAR42, containing 383 amino acids, comprises a novel FHA domain at its N terminus and a C-terminal PPIase domain but is a non-Pin1-type PPIase. Functionally, a knockdown of TbPAR42 in its procyclic form results in reduced proliferation rates suggesting an important role in cell growth.

Cytoplasmic polyadenylation is a conserved mechanism that controls mRNA translation and stability. A key protein that promotes polyadenylation-induced translation of mRNAs in maturing Xenopus oocytes is the cytoplasmic polyadenylation element binding protein (CPEB). During this meiotic transition, CPEB is subjected to phosphorylation-dependent ubiquitination and partial destruction, which is necessary for successive waves of polyadenylation of distinct mRNAs. Here we identify the peptidyl-prolyl cis-trans isomerase Pin1 as an important factor mediating CPEB destruction. Pin1 interacts with CPEB in an unusual manner in which it occurs prior to CPEB phosphorylation and prior to Pin1 activation by serine 71 dephosphorylation. Upon induction of maturation, CPEB becomes phosphorylated, which occurs simultaneously with Pin1 dephosphorylation. At this time, the CPEB-Pin1 interaction requires cdk1-catalyzed CPEB phosphorylation on S/T-P motifs. Subsequent CPEB ubiquitination and destruction are mediated by a conformational change induced by Pin1 isomerization of CPEB. Similar to M phase progression in maturing Xenopus oocytes, the destruction of CPEB during the mammalian cell cycle requires Pin1 as well. These data identify Pin1 as a new and essential factor regulating CPEB degradation.

Atomic-level molecular dynamic simulations are capable of fully folding structurally diverse proteins; however, they are limited in their ability to accurately represent electrostatic interactions. Here we have experimentally tested the role of charged residues on stability and folding kinetics of one of the most widely simulated β-proteins, the WW domain. The folding of wild type Pin1 WW domain, which has two positively charged residues in the first turn, was compared to the fast folding mutant FiP35 Pin1, which introduces a negative charge into the first turn. A combination of FTIR spectroscopy and laser-induced temperature-jump coupled with infrared spectroscopy was used to probe changes in the amide I region. The relaxation dynamics of the peptide backbone, β-sheets and β-turns, and negatively charged aspartic acid side chain of FiP35 were measured independently by probing the corresponding bands assigned in the amide I region. Folding is initiated in the turns and the β-sheets form last. While the global folding mechanism is in good agreement with simulation predictions, we observe changes in the protonation state of aspartic acid during folding that have not been captured by simulation methods. The protonation state of aspartic acid is coupled to protein folding; the apparent pKa of aspartic acid in the folded protein is 6.4. The dynamics of the aspartic acid follow the dynamics of the intermediate phase, supporting assignment of this phase to formation of the first hairpin. These results demonstrate the importance of electrostatic interactions in turn stability and formation of extended β-sheet structures.

The peptidyl-prolyl cis-trans isomerases (PPIases) that include immunophilins (cyclophilins and FKBPs) and parvulins (Pin1, Par14, Par17) participate in cell signaling, transcription, pre-mRNA processing and mRNA decay. The human genome encodes 19 cyclophilins, 18 FKBPs and three parvulins. Immunophilins are receptors for the immunosuppressive drugs cyclosporin A, FK506, and rapamycin that are used in organ transplantation. Pin1 has also been targeted in the treatment of Alzheimer's disease, asthma, and a number of cancers. While these PPIases are characterized as molecular chaperones, they also act in a nonchaperone manner to promote protein-protein interactions using surfaces outside their active sites. The immunosuppressive drugs act by a gain-of-function mechanism by promoting protein-protein interactions in vivo. Several immunophilins have been identified as components of the spliceosome and are essential for alternative splicing. Pin1 plays roles in transcription and RNA processing by catalyzing conformational changes in the RNA Pol II C-terminal domain. Pin1 also binds several RNA binding proteins such as AUF1, KSRP, HuR, and SLBP that regulate mRNA decay by remodeling mRNP complexes. The functions of ribonucleoprotein associated PPIases are largely unknown. This review highlights PPIases that play roles in RNA-mediated gene expression, providing insight into their structures, functions and mechanisms of action in mRNP remodeling in vivo.

Plant cells experience a tremendous amount of mechanical stress caused by turgor pressure. Because cells are glued to their neighbors by the middle lamella, supracellular patterns of physical forces are emerging during growth, usually leading to tension in the epidermis. Cortical microtubules have been shown to reorient in response to these mechanical stresses, and to resist them, indirectly via their impact on the anisotropic structure of the cell wall. In a recent study, we show that the polar localization of the auxin efflux carrier PIN1 can also be under the control of physical forces, thus linking cell growth rate and anisotropy by a common mechanical signal. Because of the known impact of auxin on the stiffness of the cell wall, this suggests that the mechanical properties of the extracellular matrix play a crucial signaling role in morphogenesis, notably controlling the polarity of the cell, as observed in animal systems.

The novel 3H-spiro[1-benzofuran-2-cyclopentan]-3-one skeleton has been made accessible by different routes. One- and two-step protocols lead to tricyclic and tetracyclic benzofuranones 2 and 3, respectively. A four-step synthesis to spirocompound 4 is described. The novel spirocyclic benzofuranones display modest to no inhibition of the human peptidyl prolyl cis/trans isomerase Pin1.

The binding of epigallocatechin-3-gallate (EGCG) to wild type Pin1 in solution was studied by spectroscopic methods and molecular dynamics simulations in this research to explore the binding mode and inhibition mechanism. The binding constants and number of binding sites per Pin1 for EGCG were calculated through the Stern-Volmer equation. The values of binding free energy and thermodynamic parameters were calculated and indicated that hydrogen bonds, electrostatic interaction and Van der Waals interaction played the major role in the binding process. The alterations of Pin1 secondary structure in the presence of EGCG were confirmed by far-UV circular dichroism spectra. The binding model at atomic-level revealed that EGCG was bound to the Glu12, Lys13, Arg14, Met15 and Arg17 in WW domain. Furthermore, EGCG could also interact with Arg69, Asp112, Cys113 and Ser114 in PPIase domain.

Pin1 isomerizes the phosphorylated Ser/Thr-Pro peptide bonds and regulates the functions of its binding proteins by inducing conformational changes. Involvement of Pin1 in the aging process has been suggested based on the phenotype of Pin1-knockout mice and its interaction with lifespan regulator protein, p66 (Shc) . In this study, we utilize a proteomic approach and identify peroxiredoxin 1 (PRDX1), another regulator of aging, as a novel Pin1 binding protein. Pin1 binds to PRDX1 through interacting with the phospho-Thr ( 90) -Pro ( 91) motif of PRDX1, and this interaction is abolished when the Thr ( 90) of PRDX1 is mutated. The Pin1 binding motif, Thr-Pro, is conserved in the 2-Cys PRDXs, PRDX1-4 and the interactions between Pin1 and PRDX2-4 are also demonstrated. An increase in hydrogen peroxide buildup and a decrease in the peroxidase activity of 2-Cys PRDXs were observed in Pin1 (-/-) mouse embryonic fibroblasts (MEFs), with the activity of PRDXs restored when Pin1 was re-introduced into the cells. Phosphorylation of PRDX1 at Thr ( 90) has been shown to inhibit its peroxidase activity; however, how exactly the activity of PRDX1 is regulated by phosphorylation still remains unknown. Here, we demonstrate that Pin1 facilitates the protein phosphatase 2A-mediated dephosphorylation of PRDX1, which helps to explain the accumulation of the inactive phosphorylated form of PRDX1 in Pin1 (-/-) MEFs. Collectively, we identify Pin1 as a novel PRDX1 binding protein and propose a mechanism for Pin1 in regulating the metabolism of reactive oxygen species in cells.

Neurodegenerative diseases termed Tauopathies, including Alzheimer disease, are characterized by the presence of intraneuronal neurofibrillary tangles (NFTs), composed by hyperphosphorylated protein Tau. Peptidyl-prolyl cis/trans isomerase Pin1 plays a pivotal role in the regulation of Tau phosphorylation/dephosphorylation state. Indeed, Pin1 specifically recognizes pThr231-Pro232 motif of Tau, catalyzes its isomerisation and, in dependence of the cellular environment, promotes its dephosphorylation by PP2A phosphatase: in the dephosphorylated state Tau is able to exert its physiological activity, promoting microtubules polymerization. However, Pin1 activity in Tauopathies in which Tau is mutated can be harmful, because the isomerase can accelerate progression of the disease. Taking into account the complexity of pathways in which Pin1, under a strict regulation, exerts its biological functions, this isomerase can be consider a promising target in the improvement and design of new therapies against Tauopathies.

Sulfonation by cytosolic sulfotransferases plays an important role in the metabolism of both endogenous and exogenous compounds. Sulfotransferase 4A1 (SULT4A1) is a novel sulfotransferase found primarily in neurons in the brain. It is highly conserved between species, but no substantial enzyme activity has been identified for the protein. Consequently, little is known about the role of this enzyme in the brain. We performed a yeast two-hybrid screen of a human brain library to isolate potential SULT4A1-interacting proteins that might identify the role or regulation of the sulfotransferase in humans. The screen isolated the peptidyl-prolyl cis-trans isomerase Pin1. Its interaction with SULT4A1 was confirmed by coimmunoprecipitation studies in HeLa cells and by in vitro pull-down of expressed proteins. Moreover, Pin1 binding was dependent on phosphorylation of the SULT4A1 protein. Pin1 destabilized SULT4A1, decreasing its half-life from more than 8 h to approximately 4.5 h. This effect was dependent on the isomerase activity of Pin1 and was inhibited by okadaic acid, suggesting a role for the phosphatase PP2A. Pin1-mediated SULT4A1 degradation did not involve the proteosomes or macroautophagy, but it was inhibited by the calpain antagonists N-acetyl-Leu-Leu-Nle-CHO and Z-Val-Phe-CHO. Finally, Pin1 binding was mapped to two threonine-proline motifs (Thr(8) and Thr(11)) that are not present in any of the other human cytosolic sulfotransferases. Our findings suggest that SULT4A1 is subject to post-translational modification that alters its stability in the cell. These modifications may also be important for enzyme activity, which explains why specific substrates for SULT4A1 have not yet been identified.

Parvulins and FKBPs are members of the peptidyl-prolyl cis/trans isomerases (PPIase) enzyme family whose role is to catalyze the interconversion between the cis trans forms of a peptide bond preceding internal proline residues in a polypeptide substrate. Members of the parvulin subfamily have been found to be involved in a variety of diseases, including Alzheimer's disease and cancer and are also considered possible antiparasitic targets. Genes Y110A2AL.13 (pin-1) and Y48C3A.16 (pin-4) were found in the worm's genome, possibly encoding parvulins. One is homologous to human and fly PIN1 whereas the other is homologous to human and fly PIN4. Both were expressed in Escherichia coli, purified and found to have in vitro PPIase activity. Expression levels of both genes, as well as the fkb genes (that encode FK506-binding proteins) were measured during development and under cold or heat stress conditions. The results revealed a potential role for these genes under temperature-related stress. RNAi silencing was performed for wild type and mutant strain worms under normal and cold or heat stress conditions. A reduced lifespan was observed when pin-4 dsRNA was fed to the fkb-5 deficient worms. Our work presents a first attempt to characterize the Caenorhabditis elegans parvulins and may present an interesting starting point for further experimentation concerning their role, along with the FKBP subfamily, in nematode physiology and their possible use as antiparasitic targets.

Tau pathology is the main pathological characteristic of mild cognitive impairment (MCI) and Alzheimer disease (AD), and tau-based therapeutic strategies have great implications in the prevention of MCI and AD. The phosphorylation of threonine 231 preceding proline 232 (pThr231-Pro232) triggers tau hyperphosphorylation, tau aggregation, and tau pathology. Interestingly, the pThr231-Pro232 motif may be in a cis or trans configuration, but several recent studies have firstly indicated that cis, but not trans, pThr231-Pro232 tau is a striking therapeutic target for MCI and AD. Cis pThr231-Pro232 tau appears firstly in MCI and accumulates exclusively in the development of AD. Moreover, cis pThr231-Pro232 tau has low affinity to microtubules, high resistance to dephosphorylation and degradation, and a potent tendency to aggregate. On the contrary, trans pThr231-Pro232 tau has normal physiological activity in vivo. Fortunately, Pin1 is the only known isomerase that catalyzes pThr231-Pro232 tau from the neurotoxic cis to nontoxic trans configuration, which prevents MCI and AD. Nonetheless, as we have mentioned before, Pin1 is frequently inactivated under abnormal physiological conditions in vivo. Therefore, it is necessary to clear cis pThr231-Pro232 tau by immunotherapy when Pin1 is insufficient, in order to avoid the occurrence of MCI and AD.

We have shown that Sp1 phosphorylation at Thr739 decreases its DNA-binding activity. In this study, we found that phosphorylation of Sp1 at Thr739 alone is necessary, but not sufficient for the inhibition of its DNA-binding activity during mitosis. We demonstrated that Pin1 could be recruited to the Thr739(p)-Pro motif of Sp1 to modulate the interaction between phospho-Sp1 and CDK1, thereby facilitating CDK1-mediated phosphorylation of Sp1 at Ser720, Thr723 and Thr737 during mitosis. Loss of the C-terminal end of Sp1 (amino acids 741-785) significantly increased Sp1 phosphorylation, implying that the C-terminus inhibits CDK1-mediated Sp1 phosphorylation. Binding analysis of Sp1 peptides to Pin1 by isothermal titration calorimetry indicated that Pin1 interacts with Thr739(p)-Sp1 peptide but not with Thr739-Sp1 peptide. X-ray crystallography data showed that the Thr739(p)-Sp1 peptide occupies the active site of Pin1. Increased Sp1 phosphorylation by CDK1 during mitosis not only stabilized Sp1 levels by decreasing interaction with ubiquitin E3-ligase RNF4 but also caused Sp1 to move out of the chromosomes completely by decreasing its DNA-binding activity, thereby facilitating cell cycle progression. Thus, Pin1-mediated conformational changes in the C-terminal region of Sp1 are critical for increased CDK1-mediated Sp1 phosphorylation to facilitate cell cycle progression during mitosis.

Electric pulses with high field strength and durations in the nanosecond range (nsPEFs) are of considerable interest for biotechnological and medical applications. However, their actual cellular site of action is still under debate--due to their extremely short rise times, nsPEFs are thought to act mainly in the cell interior rather than at the plasma membrane. On the other hand, nsPEFs can induce membrane permeability. We have revisited this issue using plant cells as a model. By mapping the cellular responses to nsPEFs of different field strength and duration in the tobacco BY-2 cell line, we could define a treatment that does not impinge on short-term viability, such that the physiological responses to the treatment can be followed. We observe, for these conditions, a mild disintegration of the cytoskeleton, impaired membrane localization of the PIN1 auxin-efflux transporter and a delayed premitotic nuclear positioning followed by a transient mitotic arrest. To address the target site of nsPEFs, we made use of the plant-specific KCH kinesin, which can assume two different states with different localization (either near the nucleus or at the cell membrane) driving different cellular functions. We show that nsPEFs reduce cell expansion in nontransformed cells but promote expansion in a line overexpressing KCH. Since cell elongation and cell widening are linked to the KCH localized at the cell membrane, the inverted response in the KCH overexpressor provides evidence for a direct action of nsPEFs, also at the cell membrane.

L-Cysteine plays a prominent role in sulfur metabolism of plants. However, its role in root development is largely unknown. Here, we report that L-cysteine reduces primary root growth in a dosage-dependent manner. Elevating cellular L-cysteine level by exposing Arabidopsis thaliana seedlings to high L-cysteine, buthionine sulphoximine, or O-acetylserine leads to altered auxin maximum in root tips, the expression of quiescent center cell marker as well as the decrease of the auxin carriers PIN1, PIN2, PIN3, and PIN7 of primary roots. We also show that high L-cysteine significantly reduces the protein level of two sets of stem cell specific transcription factors PLETHORA1/2 and SCR/SHR. However, L-cysteine does not downregulate the transcript level of PINs, PLTs, or SCR/SHR, suggesting that an uncharacterized post-transcriptional mechanism may regulate the accumulation of PIN, PLT, and SCR/SHR proteins and auxin transport in the root tips. These results suggest that endogenous L-cysteine level acts to maintain root stem cell niche by regulating basal- and auxin-induced expression of PLT1/2 and SCR/SHR. L-Cysteine may serve as a link between sulfate assimilation and auxin in regulating root growth.

Pin1 is prevalently overexpressed in human cancers and implicated to regulate cell growth and apoptosis. Thus far, however, no role for Pin1 has been described in modulating vascular smooth muscle cell (VSMC) senescence.

Immunohistochemistry and Western blotting were used to assess Pin1 protein level in human normal and atherosclerotic tissues. β-galactosidase staining, cumulative population doubling level, telomerase activity, and relative telomere length measurement were used to confirm VSMC senescence. The expressions of Pin1 and other genes involved in this research were analyzed by quantitative reverse-transcription polymerase chain reaction and Western blotting in VSMCs. Apolipoprotein E gene-deleted mice (ApoE-/-) fed a high-fat diet were treated with juglone or 10% ethanol, respectively, for 3 weeks. The extent of atherosclerosis was evaluated by Oil Red O, Masson trichrome staining, and immunohistology.

Pin1 protein level decreased in human atherosclerotic tissues and VSMCs, synchronously with increased VSMC senescence. Adenoviral-mediated Pin1 overexpression rescued cellular senescence in atherosclerotic VSMCs, with concurrent down-regulation of P53, p21, growth arrest and DNA-damage-inducible protein 45-alpha (Gadd45a), phosphorylated retinoblastoma (p-pRb), p65 and upregulation of cyclin subfamilies (cyclin B, D, and E), and cyclin-dependent kinase subfamilies (2, 4, and 6), whereas Pin1 knockdown resulted in the converse effects, indicating that VSMC senescence mediated by Pin1 is an integrated response to diverse signals. In vivo data from ApoE-/- mice showed that treatment of juglone led to accelerated atherosclerosis development.

Altogether this work supports a role for Pin1 as a vital modulator of VSMC senescence, thereby providing a novel target for regulation and control of atherosclerosis.

Aging of tendon stem/progenitor cells (TSPCs) can lead to tissue degeneration and subsequent injury. However, the molecular mechanisms controlling TSPC aging are not completely understood. In the present study, we investigated the role of Pin1 in aging of human TSPCs. Pin1 mRNA and protein expression levels were significantly decreased during prolonged in vitro culture of human TSPCs. Furthermore, overexpression of Pin1 delayed the progression of cellular senescence, as confirmed by downregulation of senescence-associated β-galactosidase, increased telomerase activity and decreased levels of the senescence marker, p16(INK4A). Conversely, Pin1 siRNA transfection promoted senescence in TSPCs. In addition, miR-140-5p regulated Pin1 expression at the translational level via directly targeting its 3'UTR. Our results collectively demonstrate that Pin1 acts as an important regulator of TSPC aging.

The p53 protein averts tumor formation by preventing the proliferation of damaged cells. The presence of functional p53 is critical for efficient and proper cellular responses to a variety of stress conditions. Interestingly, p63 and p73, which are the homologous ancestors of p53, retain a broader set of activities than their progeny, particularly during early embryonic development. The link of these homologues to cancer and their effect on p53 tumor suppression is only beginning to be unravelled. The tight regulation of p53 is governed by the Mdm2 E3 ligase, but also by at least two other E3 ligases. Recent findings suggest fine-tuning of p53 regulation through changes in the ratio of p53 and Mdm2. This regulation of p53 is modulated by the Mdm2 homologue, Mdmx. Genetic studies reveal the critical role Mdmx plays in p53 regulation, although the mode of action is yet to be fully explored. The relief of p53 from this tight regulation is imperative in order for it to respond to stress signals. An intriguing player in this process is the prolyl isomerase Pin1, which induces a conformational change in p53, and more recently identified, also in p73, in response to DNA damage. This complex network of regulation emerges as a family affair. This wealth of knowledge has been translated into the development of novel anti-cancer strategies based on the p53 status in the cancer cell.