Role Of Prolidase Research Articles

Prolidase Deficiency Causes Spontaneous T Cell Activation and Lupus-like Autoimmunity.

Prolidase deficiency (PD) is a multisystem disorder caused by mutations in the PEPD gene, which encodes a ubiquitously expressed metallopeptidase essential for the hydrolysis of dipeptides containing C-terminal proline or hydroxyproline. PD typically presents in childhood with developmental delay, skin ulcers, recurrent infections, and, in some patients, autoimmune features that can mimic systemic lupus erythematosus. The basis for the autoimmune association is uncertain, but might be due to self-antigen exposure with tissue damage, or indirectly driven by chronic infection and microbial burden. In this study, we address the question of causation and show that Pepd-null mice have increased antinuclear autoantibodies and raised serum IgA, accompanied by kidney immune complex deposition, consistent with a systemic lupus erythematosus-like disease. These features are associated with an accumulation of CD4 and CD8 effector T cells in the spleen and liver. Pepd deficiency leads to spontaneous T cell activation and proliferation into the effector subset, which is cell intrinsic and independent of Ag receptor specificity or antigenic stimulation. However, an increase in KLRG1+ effector CD8 cells is not observed in mixed chimeras, in which the autoimmune phenotype is also absent. Our findings link autoimmune susceptibility in PD to spontaneous T cell dysfunction, likely to be acting in combination with immune activators that lie outside the hemopoietic system but result from the abnormal metabolism or loss of nonenzymatic prolidase function. This knowledge provides insight into the role of prolidase in the maintenance of self-tolerance and highlights the importance of treatment to control T cell activation.

Extracellular Prolidase (PEPD) Induces Anabolic Processes through EGFR, β1-integrin, and IGF-1R Signaling Pathways in an Experimental Model of Wounded Fibroblasts.

The role of prolidase (PEPD) as a ligand of the epidermal growth factor receptor (EGFR) was studied in an experimental model of wound healing in cultured fibroblasts. The cells were treated with PEPD (1–100 nM) and analysis of cell viability, proliferation, migration, collagen biosynthesis, PEPD activity, and the expressions of EGFR, insulin-like growth factor 1 (IGF-1), and β1-integrin receptor including downstream signaling proteins were performed. It has been found that PEPD stimulated proliferation and migration of fibroblasts via activation of the EGFR-downstream PI3K/Akt/mTOR signaling pathway. Simultaneously, PEPD stimulated the expression of β1-integrin and IGF-1 receptors and proteins downstream to these receptors such as FAK, Grb2, and ERK1/2. Collagen biosynthesis was increased in control and “wounded” fibroblasts under PEPD treatment. The data suggest that PEPD-induced EGFR signaling may serve as a new attempt to therapy wound healing.

Platelet-Rich Plasma Promotes the Proliferation of Human Keratinocytes via a Progression of the Cell Cycle. A Role of Prolidase.

Although the role of platelet-rich plasma (PRP) in tissue regeneration has been confirmed in many studies, the mechanism of this process is still not fully understood. Human keratinocytes (HaCaT) cells were used as an experimental model for studies on the effects of PRP on cell proliferation, migration, collagen biosynthesis, prolidase activity, and its expression and anabolic signaling. The activation of epidermal growth factor receptor (EGFR), β1-integrin, and insulin-like growth factor-1 receptor (IGF-1R) by PRP were investigated by western blot and immunocytochemistry. It has been found that PRP induced keratinocytes migration and proliferation through activation of cell cycle progression and EGFR downstream signaling. Similar biological effects were achieved by an addition to the culture medium of prolidase (PEPD), a ligand of EGFR (PRP is a rich source of PEPD–2 ng/mL). PRP-dependent stimulation of collagen biosynthesis was accompanied by an increase in the expression of NF-κβ, IGF-1R-downstream signaling proteins, and PEPD activity. The data suggest that PRP activates a complex of growth factors and adhesion receptors that stimulate cell proliferation, migration, and collagen biosynthesis. PRP induces PEPD-dependent human keratinocyte proliferation through activation of the EGFR receptor. Our study provides a novel mechanism of PRP-dependent wound healing.

High prolidase levels in patients with Familial Mediterranean Fever (FMF).

Familial Mediterranean Fever (FMF) is an autoinflammatory disease. Prolidase is a specific imidodipeptidase that plays a role in collagen degradation, and an important role in inflammation and wound healing. Hypoxia-inducible factor-1α (HIF-1) is an important protein in the regulation of immunological response, hemostasis, vascularization. The aim of the study was to compare serum prolidase and HIF-1α levels in patients with FMF in attack-free period and healthy control group. Between August 2017 and December 2017, sixty patients diagnosed with FMF according to the criteria of the Tel-hashomer and admitted to Sivas Cumhuriyet University Medical Faculty, Internal Medicine Rheumatology Department and sixty healthy volunteers were enrolled in the study. Median serum prolidase levels were 72.1 (25.1-114.9) ng/ml in FMF group and 30.7 (21.3-86.2) ng/mL in healthy control (HC) group (p = 0.018). ROC analysis showed that the sensitivity was 65% and the specificity was 68.3% at serum prolidase levels 54.03 ng/mL (p < 0.05). The median serum levels of HIF-1α in the FMF group was 482.0 (292.0-3967.0) pg/mL and 632.0 (362.0-927.0) pg/mL in the HC group (p > 0.05). There was no significant correlation between laboratory findings, sex, age, and prolidase (p > 0.05). Serum prolidase enzyme levels in FMF patients with attack-free period were significantly higher than in the HC group. However, the role of prolidase and HIF1-α in the FMF disease needs to be clarified with more extensive and comprehensive studies.

What is the role of prolidase in pathogenesis of primary varicose veins?

What is the role of prolidase in pathogenesis of primary varicose veins?

Phosphoenolpyruvate-dependent inhibition of collagen biosynthesis, α2β1 integrin and IGF-I receptor signaling in cultured fibroblasts

The mechanism of collagen biosynthesis regulation is not fully understood. The finding that prolidase plays an important role in collagen biosynthesis and phosphoenolpyruvate inhibits prolidase activity "in vitro" led to evaluate its effect on collagen biosynthesis in cultured human skin fibroblasts. Confluent fibroblasts were treated with millimolar concentrations (1-4 mM) of phosphoenolpyruvate monopotassium salt (PEP) for 24 h. It was found that PEP-dependent decrease in prolidase activity and expression was accompanied by parallel decrease in collagen biosynthesis. However, the experiments with inhibitor of PEP production, 3-mercaptopicolinate revealed no direct correlation between collagen biosynthesis and prolidase activity and expression. Since insulin-like growth factor (IGF-I) is the most potent stimulator of both collagen biosynthesis and prolidase activity, and prolidase is regulated by beta(1) integrin signaling, the effect of PEP on IGF-I receptor (IGF-IR) and beta(1) integrin receptor expressions were evaluated. It was found that the exposure of the cells to 4 mM PEP contributed to a decrease in IGF-IR and beta(1) integrin receptor expressions. The data suggest that PEP-dependent decrease of collagen biosynthesis in cultured human skin fibroblasts may undergo through depression of alpha(2)beta(1) integrin and IGF-IR signaling. The hypothetical mechanism of the role of prolidase in IGF-IR, beta(1) integrin receptor expressions, and clinical significance of the process are discussed.

Introduction to second proline symposium

The idea that proline, the only proteogenic secondary amino acid, has special metabolic and regulatory features has increasingly found adherents. Because its alpha-nitrogen is within the pyrrolidine ring, proline cannot be substrate for generic amino acid-metabolizing enzymes. Instead, a paradigm of enzymes with their special tissue and subcellular localizations and regulatory mechanisms participate in a metabolically sequestered system. Increasingly, the regulatory functions of this system are being discovered. However, as recently as the early 1980s, the proposal that metabolism of proline could serve as a regulatory system, even under special conditions, met with considerable resistance. In fact, to paraphrase critics, “if there are unusual features to proline metabolism lending to its regulatory function, certainly they would have been discovered by now (i.e. pre-1980).” That such was not the case was affirmed recently. Special features of proline metabolism lending to its role in regulating general metabolism and cell behavior were explored at the 1st International Symposium on Proline Metabolism held October 2004 at Frederick, Maryland. That meeting focused on findings in two specific areas. First, genetic epidemiology linked schizophrenia with mutations in proline oxidase/proline dehydrogenase (POX/PRODH). The neurophysiologic function of proline as a neurotransmitter suppressing glutamergic neurons was presented as a potential pathophysiologic link between proline and the neuropsychiatgric disorder in humans. The finding that proline oxidase/proline dehydrogenase is a p53-induced gene (PIG6) led to the documentation of POX/PRODH as a ROS generating enzyme capable of inducing apoptosis independent of p53, thereby establishing the importance of POX/PRODH in carcinogenesis.. Seminal advances had been made in the structural biology of PRODH from E. coli. The response of the participants at this meeting was enthusiastic, and the concensus was to have a meeting on proline metabolism triennially. We thank the Center for Cancer Research, NCI, the Office of Dietary Supplements, NIH and the Office of Rare Diseases, NIH for sponsoring this 2nd International Symposium on Proline Metabolism and Human Disease. In planning this meeting, the organizers responded to comments made by the attendants at the last meeting and included several areas in addition to those focusing on human disease. The new invitees and their subject area are as follows: 1) functions in prokaryotic regulation of proline including proline transporters (Janet Wood, Donald Becker); 2) plant biochemistry especially proline as an osmoprotectant (Nathalie Verbruggen) and the role of proline in regulating plant biochemistry and metabolism (Kalidas Shetty); 3) collagen metabolism (Steve Krane) and the role of prolidase in collagen metabolism (Jerzy Palka); 4) the importance of metabolism in cancer (Chi Dang), and 5) the role of TOR in the response to nutrient stress (Alan Kimmel). As a basis for understanding the role of proline in neurophysiologic function, the function of glutamate as a neurotransmitter was reviewed (Joseph Cable). Significant advances have been made in the areas previously explored in 2004. In the area of neuropsychiatriac disorders, David Valle reviewed the work characterizing biochemical phenotypes expressed by specific mutations identified in patients with early schizophrenia. In the cancer area, Yongmin Liu presented his work on POX induction of apoptosis by both intrinsic and extrinsic pathways and Jim Phang presented the work on responses of POX/PRODH to stress signaling. Guoyao Wu presented his work on proline as a source of arginine for in utero development of pigs. Especially revealing was the new structural biological findings that substantiates PRODH as a source of superoxide (Jack Tanner) and the possibility that the electrons from proline transferred by PRODH can be used either for electron transport (ATP generation) or for reducing oxygen (production of superoxide). Dave Valle summarized insights gained from characterization of the inborn errors of proline and ornithine metabolism, Antonella Forlino presented the clinical and genetic characterization of prolidase deficiency. C. Andy Hu, the Editor-in-Chief of this volume, reviewed the knowledge of pyrroline-5-carboxylate synthase. It is encouraging to find advances in the understanding of proline metabolism on a broad front. The impact of these discoveries is beginning to be felt. In fact, if you googled either “proline oxidase” or “proline dehydrogenase” AND cancer, you would get more than 4,000 hits. Interestingly, the antibody for POX/PRODH is now commercially available from at least 2 sources, and perhaps more importantly, siRNA for studying the effects of POX/PRODH knockdown has recently become available. It is evident that momentum has been generated to establish this symposium as a continuing event. The organizing committee will be expanded, and an enlarged steering committee will be responsible for future meetings.

Prolidase Dependent Inhibition of Collagen Biosynthesis in Chinese Hamster Ovary Cells

Collagen is responsible for maintenance of connective tissue integrity, and through interaction with integrin receptors may participate in regulation of numerous physiological and pathological processes. An important role in collagen biosynthesis plays prolidase. It was previously found that nickel chloride inhibited prolidase activity in Chinese hamster ovary cells (CHO-C9). The cells lack any detectable ornithine aminotransferase and P5C synthase activities, and therefore require addition of free proline or glicyl-proline (converted to glycine and proline) for growth. We have found that Ni(II) contributed to decrease in collagen and hydroxyproline content in CHO cells incubated with Gly-Pro, whereas it had no effect on hydroxyproline content in the cells incubated with proline. Decrease in collagen content was not related to decrease in type I collagen mRNA level suggesting regulation of this process at post-transcriptional level. However decrease in expression of Sos and phosphorylated MAP-kinases were found in the cells growing in the presence of Gly-Pro and Ni(II). Decrease in the expression of these proteins was not related to inhibition of signalling induced by growth factors, since no changes were observed in expression of AKT in CHO cells incubated with Ni(II). The results presented provide evidence for important role of prolidase in collagen biosynthesis.

Extracellular matrix and HIF‐1 signaling: The role of prolidase

Hypoxia-inducible factor-1 (HIF-1) plays an important role in stress-responsive gene expression. Although primarily sensitive to hypoxia, HIF-1 signaling can be regulated by a number of stress factors including metabolic stress, growth factors and molecules present in the extracellular matrix (ECM). Degradation of ECM by metalloproteinases (MMP) is important for tumor progression, invasion and metastasis. ECM is predominantly collagen, and the imino acids (Pro and HyPro) comprise 25% of collagen residues. The final step in collagen degradation is catalyzed by prolidase, the obligate peptidase for imidodipeptides with Pro and HyPro in the carboxyl terminus. Defective wound healing in patients with inherited prolidase deficiency is associated with histologic features of angiopathy suggesting that prolidase may play a role in angiogenesis. Because HIF-1 alpha is central to angiogenesis, we considered that prolidase may modulate this pathway. To test this hypothesis, we made expression constructs of human prolidase and obtained stable transfectants in colorectal cancer cells (RKO). Overexpression of prolidase resulted in increased nuclear hypoxia inducible factor (HIF-1 alpha) levels and elevated expression of HIF-1-dependent gene products, vascular endothelial growth factor (VEGF) and glucose transporter-1 (Glut-1). The activation of HIF-1-dependent transcription was shown by prolidase-dependent activation of hypoxia response element (HRE)-luciferase expression. We used an oxygen-dependent degradation domain (ODD)-luciferase reporter construct as a surrogate for HIF-1 alpha as an in situ prolyl-hydroxylase assay. Since this reporter is degraded by VHL-dependent mechanisms, the increased levels of luciferase observed with prolidase expression reflected the decreased HIF-1 alpha prolyl hydroxylase activity. Additionally, the differential expression of prolidase in 2 breast cancer cell lines showed prolidase-dependent differences in HIF-1 alpha levels. These findings show that metabolism of imidodipeptides by prolidase plays a previously unrecognized role in angiogenic signaling.