Phosphatidylethanolamine-binding Protein 4 Research Articles

Knockout of phosphatidylethanolamine binding protein4 (PEBP4) promotes chronic non-bacterial prostatitis by mediating the activation of NF-κB signaling.

The etiology of chronic prostatitis remains unclear; consequently, this disease is associated with recurrence and ineffective clinical therapy. Therefore, there is an urgent need to investigate the underlying pathogenesis of chronic prostatitis in order to develop more efficacious treatments. The previous study found that knocking out of PEBP4 leads to chronic prostatitis in the male mice. This research aimed to identify the role of PEBP4 in prostatitis, determine the molecular pathogenic mechanisms associated with chronic prostatitis, and provide guidelines for the development of new treatment strategies for chronic prostatitis. A PEBP4 exon knockout strain (PEBP4-/-) was established in C57BL/6 mice via the Cre-loxP system. Hematoxylin-eosin (H&E) staining was used to investigate histological changes. RNA-sequencing was used to investigate the gene expression signature of the prostate and the levels of inflammatory cytokines were determined by real-time polymerase chain reaction (RT-PCR). The expression of PEBP4 protein in prostate tissue was determined by immunohistochemistry in specimens from patients with BPH and BPH combined with chronic prostatitis. Finally, we used a CRISPR-Cas9 plasmid to knockout PEBP4 in RWPE-1 cells; western blotting was subsequently used to measure the level of activation in the NF-κB signaling pathway after activating with TNF-α. Hemorrhage and inflammatory cell infiltration were incidentally observed in the seminal vesicles and prostate glands of PEBP4-/- mice after being fed with a normal diet for 1 year. In addition, we found significantly lower (p<0.001) expression levels of PEBP4 protein in prostate tissues from patients with benign prostate hyperplasia (BPH) and chronic and non-bacterial prostatitis (CNP)when compared to those with BPH only. The reduced expression of PEBP4 led to a higher risk of prostatitis recurrence in patients after 2 years of follow-up. Increased levels of NF-κB and IκB phosphorylation were observed in PEBP4-knockout RWPE-1 cells and prostate glands from PEBP4-/- mice. The knockout of PEBP4 in experimental mice led to chronic prostatitis and the reduced expression of PEBP4 in patients with higher risk of chronic and non-bacterial prostatitis suggested that PEBP4 might act as a protective factor against chronic prostatitis. The knockout of PEBP4 in RWPE-1 cells led to the increased activation of NF-κB and IκB, thus indicating that inhibition of PEBP4 faciliated the NF-κB signaling cascade. Our findings provide a new etiology and therapeutic target for chronic prostatitis.

PEBP4 deficiency aggravates LPS-induced acute lung injury and alveolar fluid clearance impairment via modulating PI3K/AKT signaling pathway

Acute lung injury (ALI) is a common clinical syndrome, which often results in pulmonary edema and respiratory distress. It has been recently reported that phosphatidylethanolamine binding protein 4 (PEBP4), a basic cytoplasmic protein, has anti-inflammatory and hepatoprotective effects, but its relationship with ALI remains undefined so far. In this study, we generated PEBP4 knockout (KO) mice to investigate the potential function of PEBP4, as well as to evaluate the capacity of alveolar fluid clearance (AFC) and the activity of phosphatidylinositide 3-kinases (PI3K)/serine-theronine protein kinase B (PKB, also known as AKT) signaling pathway in lipopolysaccharide (LPS)-induced ALI mice models. We found that PEBP4 deficiency exacerbated lung pathological damage and edema, and increased the wet/dry weight ratio and total protein concentration of bronchoalveolar lavage fluid (BALF) in LPS-treated mice. Meanwhile, PEBP4 KO promoted an LPS-induced rise in the pulmonary myeloperoxidase (MPO) activity, serum interleuin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α levels, and pulmonary cyclooxygenase-2 (COX-2) expression. Mechanically, PEBP4 deletion further reduced the protein expression of Na+ transport markers, including epithelial sodium channel (ENaC)-α, ENaC-γ, Na,K-ATPase α1, and Na,K-ATPase β1, and strengthened the inhibition of PI3K/AKT signaling in LPS-challenged mice. Furthermore, we demonstrated that selective activation of PI3K/AKT with 740YP or SC79 partially reversed all of the above effects caused by PEBP4 KO in LPS-treated mice. Altogether, our results indicated the PEBP4 deletion has a deterioration effect on LPS-induced ALI by impairing the capacity of AFC, which may be achieved through modulating the PI3K/AKT pathway.

Phosphatidylethanolamine-binding protein 4 deficiency exacerbates carbon tetrachloride-induced liver fibrosis by regulating the NF-κB signaling pathway.

Liver fibrosis is a pathological process which can progress to hepatocirrhosis, even hepatocellular carcinoma. Phosphatidylethanolamine-binding protein 4 (PEBP4) is a secreted protein involved in regulating many molecular pathways, whereas its roles in diseases including hepatic fibrosis remain undefined. The nuclear factor-κappa B (NF-κB) signaling pathway has been found to be involved in the development of liver fibrosis. In this study, we generated a hepatocyte-conditional knockout (CKO) mouse model of PEBP4, and explored the potential functions of PEBP4 on liver fibrosis and the NF-κB signaling pathway in a mouse model of carbon tetrachloride (CCl4)-induced liver fibrosis. We demonstrated that PEBP4 CKO aggravated CCl4-triggered liver fibrosis, as evidenced by altered histopathology, an increase in the activities of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and hydroxyproline (HYP) levels, and more collagen deposition, as well as by enhanced expression of fibrotic markers including α-smooth muscle actin (α-SMA), collagen I and collagen III. Mechanistically, PEBP4 deficiency activated the NF-κB signaling pathway, as indicated by increased phosphorylation of NF-κB p65 and inhibitor protein κB inhibitor-α (IκB-α), and nuclear NF-κB p65 expression in the fibrotic liver. Notably, the NF-κB inhibitor pyrrolidine dithiocarbamate (PDTC) partially blocked the activation of the NF-κB pathway, and reversed the pro-fibrotic effect of PEBP4 deletion in CCl4-treated mice. Together, these results suggest that PEBP4 deficiency results in aggravation of liver fibrosis and activation of the NF-κB signaling pathway, supporting a novel concept that PEBP4 is a crucial player in hepatic fibrosis, but also might be a negative regulator of the NF-κB signaling in liver fibrosis.

PEBP4 Directs the Malignant Behavior of Hepatocellular Carcinoma Cells via Regulating mTORC1 and mTORC2.

Phosphatidylethanolamine binding protein 4 (PEBP4) is an understudied multifunctional small protein. Previous studies have shown that the expression of PEBP4 is increased in many cancer specimens, which correlates to cancer progression. The present study explored the mechanism by which PEBP4 regulates the growth and progression of hepatocellular carcinoma cells. Thus, we showed that knockdown of PEBP4 in MHCC97H cells, where its expression was relatively high, diminished activities of serine/threonine protein kinase B (PKB, also known as Akt), mammalian target of rapamycin complex 1(mTORC1), and mTORC2, events that were not restored by insulin-like growth factor 1 (IGF-1). Conversely, overexpression of PEBP4 in MHCC97L cells with the low endogenous level yielded opposite effects. Furthermore, physical association of PEBP4 with Akt, mTORC1, and mTORC2 was observed. Interestingly, introduction of AktS473D mutant, bypassing phosphorylation by mTORC2, rescued mTORC1 activity, but without effects on mTORC2 signaling. In contrast, the effect of PEBP4 overexpression on the activity of mTORC1 but not that of mTORC2 was suppressed by MK2206, a specific inhibitor of Akt. In conjunction, PEBP4 knockdown-engendered reduction of cell proliferation, migration and invasion was partially rescued by Akt S473D while increases in these parameters induced by overexpression of PEBP4 were completely abolished by MK2206, although the expression of epithelial mesenchymal transition (EMT) markers appeared to be fully regulated by the active mutant of Akt. Finally, knockdown of PEBP4 diminished the growth of tumor and metastasis, whereas they were enhanced by overexpression of PEBP4. Altogether, our study suggests that increased expression of PEBP4 exacerbates malignant behaviors of hepatocellular cancer cells through cooperative participation of mTORC1 and mTORC2.

Hepatocyte-Conditional Knockout of Phosphatidylethanolamine Binding Protein 4 Aggravated LPS/D-GalN-Induced Acute Liver Injury via the TLR4/NF-κB Pathway.

Acute liver injury (ALI) is a disease that seriously threatens human health and life, and a dysregulated inflammation response is one of the main mechanisms of ALI induced by various factors. Phosphatidylethanolamine binding protein 4 (PEBP4) is a secreted protein with multiple biological functions. At present, studies on PEBP4 exist mainly in the field of tumors and rarely in inflammation. This study aimed to explore the potential roles and mechanisms of PEBP4 on lipopolysaccharide (LPS)/D-galactosamine (D-GalN)-induced ALI. PEBP4 was downregulated after treatment with LPS/D-GalN in wild-type mice. PEBP4 hepatocyte-conditional knockout (CKO) aggravated liver damage and repressed liver functions, including hepatocellular edema, red blood cell infiltration, and increased aspartate aminotransferase (AST)/alanine aminotrans-ferase (ALT) activities. The inflammatory response was promoted through increased neutrophil infiltration, myeloperoxidase (MPO) activities, and cytokine secretions (interleukin-1β, IL-1β; tumor necrosis factor alpha, TNF-α; and cyclooxygenase-2, COX-2) in PEBP4 CKO mice. PEBP4 CKO also induced an apoptotic effect, including increasing the degree of apoptotic hepatocytes, the expressions and activities of caspases, and pro-apoptotic factor Bax while decreasing anti-apoptotic factor Bcl-2. Furthermore, the data demonstrated the levels of Toll-like receptor 4 (TLR4), phosphorylation-inhibitor of nuclear factor kappaB Alpha (p-IκB-α), and nuclear factor kappaB (NF-κB) p65 were upregulated, while the expressions of cytoplasmic IκB-α and NF-κB p65 were downregulated after PEBP4 CKO. More importantly, both the NF-κB inhibitor (Ammonium pyrrolidinedithiocarbamate, PDTC) and a small-molecule inhibitor of TLR4 (TAK-242) could inhibit TLR4/NF-κB signaling activation and reverse the effects of PEBP4 CKO. In summary, the data suggested that hepatocyte-conditional knockout of PEBP4 aggravated LPS/D-GalN-induced ALI, and the effect is partly mediated by activation of the TLR4/NF-κB signaling pathway.

Overexpression of phosphatidylethanolamine-binding protein 4 (PEBP4) associates with recurrence of meningiomas

Background and purposeAbnormal expression of phosphatidylethanolamine-binding protein 4 (PEBP4) has been identified in various types of malignant tumors. In the present study, we investigated the expression of PEBP4 in meningioma cases and examined whether PEBP4 expression was correlated with outcomes among these patients. Materials and methodsThe expression levels of PEBP4 and Ki-67 in human meningioma tissues from 65 patients were evaluated by immunohistochemical staining. The correlation between PEBP4 immunoreactivity in meningioma samples and patients’ clinical outcomes was examined using the Kruskal–Wallis correlation test. The prognostic value of PEBP4 expression in meningiomas patients also was investigated. ResultsImmunohistochemical analysis revealed up-regulated PEBP4 expression in both atypical and anaplastic meningiomas compared with classical meningiomas (13.38 ± 4.19% vs. 3.64 ± 2.04%, P < 0.001). PEBP4 immunoreactivity in meningioma samples was closely correlated with that for Ki-67 (Spearman r = 0.7922, P < 0.0001). PEBP4 expression was also associated with tumor differentiation grade and clinical recurrence (P < 0.05). Multivariate regression analysis showed with high PEBP4 expression was associated with a longer recurrence-free survival (hazard ratio=0.252, 95% confidence interval: 0.067–0.940, P = 0.040). ConclusionPEBP4 may play an important role in the progression of meningioma, as high PEBP4 expression was associated with a higher pathological grade of meningioma. Moreover, PEBP4 expression may be a meaningful prognostic biomarker in meningioma.

Diagnostic value of phosphatidylethanolamine binding protein 4 levels in patients receiving nursing interventions for advanced chronic kidney disease.

ObjectiveTo explore the diagnostic role of phosphatidylethanolamine binding protein 4 (PEBP4) in patients with chronic kidney disease (CKD) receiving nursing interventions.MethodsELISA was used to evaluate serum PEBP4 levels. Receiver-operating characteristic curve analysis was used to assess diagnostic accuracy. Spearman correlation analysis was used to assess the relationships between PEBP4 levels and biochemical indexes.ResultsSerum PEBP4 was high in CKD patients compared with healthy individuals. PEBP4 levels were positively correlated with pathological stage in CKD patients. PEBP4 had higher sensitivity for diagnosis of CKD than common indexes including blood urea nitrogen, creatinine and C-reactive protein. Among CKD patients treated with calcium channel blockers, serum PEBP4 levels declined notably and were associated with concentrations of K+, Na+, Cl− and Ca2+. Nursing interventions significantly decreased serum PEBP4 levels. A significant association between serum PEBP4 level and ionic concentration was observed in CKD patients receiving nursing interventions.ConclusionsThis prospective study demonstrated that PEBP4 level might represent an effective diagnostic biomarker in CKD patients. PEBP4 also acted as a valuable care compliance factor for determining the necessity for nursing interventions. Nursing interventions restored ion channel function and subsequently resulted in decreased PEBP4 levels and proteinuria.

Immuno-histological mapping and functional association of seminal proteins in testis and excurrent ducts with sperm function in buffalo.

The region-specific expression of seminal proteins in testis and excurrent duct system determines the quality and function of the spermatozoa. In the present study, localization and expression of some of the seminal proteins such as insulin-like growth factor receptor 1β (IGF-1Rβ), phosphatidylethanolamine-binding protein 4 (PEBP4), α-tubulin and tissue factor pathway inhibitor 2 (TFPI2) were carried out in testis, excurrent duct system and spermatozoa of buffalo. IGF-1Rβ was localized in the cells of the seminiferous tubules of the testis, except in primary spermatocytes. The PEBP4 was localized only in the elongated spermatid, whereas α-tubulin and TFPI2 proteins were localized in all cells of the seminiferous tubule including spermatocyte. In the buffalo spermatozoa, IGF-1Rβ, PEBP4, α-tubulin and TFPI2 were localized in the acrosome region, the post-acrosomal region till the tail end, post-acrosome to the entire tail region and the equatorial region, respectively. The study indicates that IGF-1R, α-tubulin and PEBP4 proteins regulate spermatogenesis, whereas TFPI2 may be involved during the zona binding process of the buffalo spermatozoa.

PEBP4 gene expression in lung squamous cell carcinoma: A meta-analysis-based study of the molecular pathways involved.

Previous studies have suggested increased activity of phosphatidylethanolamine binding protein 4 (PEBP4) may be associated with the prognosis of non-small cell lung cancer. However, to the best of our knowledge, no direct association between PEBP4 and lung squamous cell carcinoma (LSCC) has been reported. In the present study, a systematic review and meta-analysis was performed to examine the gene expression activity of PEBP4 in LSCC. A total of 10 out of 131 gene expression datasets from the Gene Expression Omnibus (GEO) were selected, including 574 samples (319 patients with LSCC and 255 healthy controls). Subsequently, multiple linear regression (MLR) was employed to study three potential influencing factors: Sample size, population region and study date. A literature-based pathway analysis was then conducted to examine the potential mechanisms through which PEBP4 may exert influence on LSCC. The results of a meta-analysis indicated that, in LSCC, PEBP4 exhibited significantly low expression levels (P<0.033), with mildly increased gene expression levels observed in three studies (log fold-change: 0.072–2.13). However, a significant between-study variance was observed from the heterogeneity analysis. MLR indicated that population region was a significant factor (P<0.0065), whereas sample size and study age were not (P>0.46). Eight functional pathways were subsequently identified, through which PEBP4 may influence the prognosis of LSCC and its response to treatment. The results of the present study suggested that the effects of PEBP4 on LSCC can be neglected in most cases of LSCC, where PEBP4 demonstrated decreased expression levels. However, in the case of PEBP4 overexpression, it may contribute to the progression of LSCC and lead to the development of drug resistance.

The Roles And Signaling Pathways Of Phosphatidylethanolamine-Binding Protein 4 In Tumors.

Phosphatidylethanolamine-binding protein 4 (PEBP4) has been found to be highly expressed in many tumors and to be closely related to the proliferation, differentiation, and metastasis of tumors. PEBP4 has also been found to be involved in many cancer-activated signaling pathways and to cause therapeutic resistance. In this study, we first reviewed the morphological structure and expression of PEBP4, then discussed the roles of PEBP4 in individualized treatment of some cancers, and finally explored the possibilities of cultivating PEBP4 as a therapeutic target.We also identified the main signaling pathways in which PEBP4 affects different cancers. It is here concluded that over-expression of PEBP4 can enhance the proliferation and metastasis of the cancer cells and the resistance to radiotherapy/chemotherapy in cancers.

Phosphatidylethanolamine binding protein-4 (PEBP4) is increased in IgA nephropathy and is associated with IgA-positive B-cells in affected kidneys

IgA nephropathy (IgAN) is the most common glomerulonephritis worldwide and a major cause of chronic kidney disease and failure. IgAN is driven by an autoimmune reaction against galactose-deficient IgA1 that results in the generation of autoantibodies and large IgG-IgA immune complexes. Immune complexes accumulate in the glomerular mesangium causing chronic inflammation and renal scarring. A significant proportion of IgAN patients develop end-stage kidney disease and require dialysis or transplantation. Currently, there are no approved specific therapies that can ameliorate the systemic autoimmune reaction in IgAN and no biomarkers that can predict renal inflammation and scarring. In this study, we used shotgun LC-MS/MS proteomics to compare small volumes of urine from healthy subjects and IgAN patients. We identified multiple urine proteins with unknown renal or IgAN function. Our attention was captured by the increase of phosphatidylethanolamine binding protein-4 (PEBP4) in IgAN urine. The function of PEBP4 in IgAN or renal disease is unknown. Increased levels of urine and serum PEBP4 were subsequently validated in different cohorts of IgAN patients and PEBP4 was linked to declining kidney function in IgAN. Strong PEBP4 staining was sporadically seen in IgAN kidney biopsies, colocalising with IgA in glomeruli and in the lumen of kidney tubules. In a small number of IgAN biopsies, PEBP4 colocalised with IgA and CD19 while the increased excretion of PEBP4 in IgAN urine was accompanied by increased excretion of classic B-cell factors BAFF, BCMA and TACI as well as IgA and IgG. PEBP4 is a new IgAN-related protein with unknown function and a likely renal disease marker in urine and serum.

Phosphatidylethanolamine-binding protein 4 promotes the epithelial-to-mesenchymal transition in non-small cell lung cancer cells by activating the sonic hedgehog signaling pathway.

Phosphatidylethanolamine-binding protein 4 (PEBP4), a member of the PEBP family, has been reported to play a pivotal role in tumor progression. However, its role in epithelial-to-mesenchymal transition (EMT) in non-small cell lung cancer (NSCLC) cells remains unclear. Here, we investigated the effects and underlying mechanism of PEBP4 in NSCLC EMT. Three human NSCLC cell lines (A549, H1299, and H460) were transfected with pcDNA3.1-PEBP4 or PEBP4-targeting small interfering RNA. Then, cell proliferation was analyzed by the MTT assay, and cell migration and invasion were analyzed by the transwell chamber assay. Protein and messenger RNA expression of the related genes and proteins were assessed by Western blot analysis and quantitative real-time polymerase chain reaction, respectively. Results showed that PEBP4 was highly expressed in the human lung cancer tissues and three human NSCLC cell lines. Pretreatment with pcDNA3.1-PEBP4 promoted the proliferation, invasion, and migration of NSCLC cells and increased EMT in vitro and lung tumor metastasis in vivo. Whereas knockdown of PEBP4 suppressed NSCLC cell migration, PEBP4, and invasion with prevented EMT. Furthermore, PEBP4 overexpression significantly promoted the transcriptional activity of sonic hedgehog (Shh) signaling in NSCLC cells. Further analysis showed that using cyclopamine to inhibit Shh signaling significantly ameliorated the effect on cell proliferation, invasion, and migration, as well as EMT triggered by PEBP4 overexpression. Together, these results suggest that PEBP4 may promote tumorigenesis in NSCLC by regulating cell proliferation and EMT via activation of the Shh signaling pathway.

Knockdown of PEBP4 inhibits human glioma cell growth and invasive potential via ERK1/2 signaling pathway.

Phosphatidylethanolamine (PE)-binding protein 4 (PEBP4) is an antiapoptotic protein that is aberrantly expressed in various malignancies. We previously demonstrated that PEBP4 expression is dramatically induced in human gliomas and positively correlated with tumor grade and patient survival. However, the function of PEBP4 in human glioma development and underlying mechanisms remain largely unknown. By stable lentiviral vector-mediated silencing of PEBP4, we examined the effects of PEBP4 knockdown on the growth, apoptosis, and invasion of U251 and U373 human glioma cell lines using MTT, Transwell, colony formation, and flow cytometric assays. We examined the in vivo role of PEBP4 in tumor growth by inoculation of BALB/c nu/nu male mice with PEBP4-deficient U251 and U373 cells. The expression of cell cycle- and apoptosis-related proteins was analyzed by Western blotting and immunostaining. Knockdown of PEBP4 significantly reduced the proliferation and invasion of human glioma cells while inducing cell apoptosis by altering the expression of cell cycle- and apoptosis-related proteins. Mechanistically, PEBP4 knockdown led to activation of the extracellular signal-regulated kinases 1/2 (ERK1/2) pathway, an effect that could be reversed by U0126, a selective inhibitor of MEK1/2 (upstream of ERK1/2), suggesting involvement of ERK1/2 signaling in the regulation of glioma development and progression by PEBP4. We identified PEBP4 as a novel regulator mediating human glioma cell proliferation, invasion, and apoptosis as well as tumor formation and growth. Therefore, PEBP4 may be a potential therapeutic target in human glioma treatment.

Knockdown of PEBP4 suppresses proliferation, migration and invasion of human breast cancer cells.

Phosphatidylethanolamine-binding protein 4 (PEBP4), a member of the PEBP family, plays a pivotal role in tumor progression. However, the roles of PEBP4 in breast cancer remain unclear. Therefore, in the present study, we investigated the effects of PEBP4 on breast cancer cell proliferation, migration and invasion, and the underlying mechanism was also explored. Our results showed that the expression of PEBP4 was significantly up-regulated in breast cancer cell lines. Knockdown of PEBP4 inhibited breast cancer cell proliferation in vitro and tumor growth in vivo. Furthermore, knockdown of PEBP4 suppressed breast cancer cell migration and invasion with prevented EMT. Mechanistically, knockdown of PEBP4 inhibited breast cancer cell proliferation and migration through the inactivation of PI3K/Akt signaling pathway. In conclusion, the present study demonstrated for the first time that knockdown of PEBP4 inhibited the proliferation, invasion and tumorigenesis in breast cancer cells. Thus, PEBP4 may serve as a potential therapeutic target for the treatment of breast cancer.

Role of the PEBP4 protein in the development and metastasis of gastric cancer.

Phosphatidylethanolamine-binding protein 4 (PEBP4) has previously been reported to be upregulated in various cancers. However, the physiological functions of PEBP4 in gastric cancer are still unknown. Aiming to clarify the properties and role of PEBP4 in the development and invasion of gastric cancer, we performed several biological assays and a knockdown assay. The expression level of PEBP4 was shown to be significantly upregulated in gastric cancer tissue samples, and knockdown of the expression of PEBP4 induced significant inhibitory effects on cell proliferation, migration and invasiveness. In addition, it was demonstrated that PEBP4 was associated with the development and invasion of gastric cancer cells through activation of the PI3K/Akt signaling pathway. Our findings supported the hypothesis that PEBP4 might be a novel potential drug target for the treatment of gastric cancer.

PEBP4 silencing inhibits hypoxia-induced epithelial-to-mesenchymal transition in prostate cancer cells.

Hypoxia induced epithelial-to-mesenchymal transition (EMT) to facilitate the tumor biology. Phosphatidylethanolamine-binding protein 4 (PEBP4) is a member of the PEBP family and has been reported to be upregulated in various cancer types. The definite function of PEBP4 in regulating the EMT of prostate cancer, however, is still unclear. Here, we examined the functional role of PEBP4 and the underlying molecular mechanisms in hypoxia-induced EMT in prostate cancer cells. Our results showed that PEBP4 mRNA and protein expression was markedly increased in the human prostate cancer tissues and cell lines. Knockdown of PEBP4 significantly inhibited hypoxia-induced migration/invasion and EMT program. Furthermore, knockdown of PEBP4 prevented hypoxia-induced the expression of p-Akt and p-mTOR in prostate cancer cells. Taken together, this study reported here provided evidence that knockdown of PEBP4 inhibited hypoxia-induced EMT in prostate cancer cells. Our study uncovered a novel role for PEBP4 in prostate cancer progression, which might support the potential for PEBP4 targeting in prostate cancer therapy.

Phosphatidylethanolamine binding protein 4 (PEBP4) is a secreted protein and has multiple functions

Phosphatidylethanolamine binding proteins (PEBP) represent a superfamily of proteins that are conserved from bacteria to humans. In mammals, four members have been identified, PEBP1–4. To determine the functional differences among PEBP1–4 and the underlying mechanism for their actions, we performed a sequence alignment and found that PEBP4 contains a signal peptide and potential glycosylation sites, whereas PEBP1–3 are intracellular proteins. To test if PEBP4 is secreted, we made constructs with Myc epitope at the amino (N) terminus or carboxyl (C) terminus to mask the signal sequence or keep it free, respectively. Our data revealed that both mouse and human PEBP4 were secreted when the epitope was tagged at their C-terminus. To our surprise, secretion was dependent upon the C-terminal conserved domain in addition to the N-terminal signal sequence. When the epitope was placed to the N-terminus, the recombinant protein failed to secrete and instead, was retained in the cytoplasm. Mass spectrometry detected asparagine (N)-glycosylation on the secreted PEBP4. Although overexpression of N-terminal tagged PEBP4 resulted in an inhibition of ERK activation by EGF, that with a C-terminal epitope tag did not have such an effect. Likewise, transfection of PEBP4 shRNA did not appear to affect ERK activation, suggesting that PEBP4 does not participate in the regulation of this pathway. In contrast, PEBP4 siRNA suppressed phosphorylation of Act at S473. Therefore, our results suggest that PEBP4 is a multifunctional protein and can be secreted. It will be important to investigate the mechanism by which PEBP4 is secreted and regulates cellular events.

Increased expression of phosphatidylethanolamine-binding protein 4 (PEBP4) strongly associates with human gliomas grade.

Abnormal expression of phosphatidylethanolamine-binding protein 4 (PEBP4) has been found in various types of malignancies. However, the PEBP4 expression in human gliomas is still unclear. In this study, we aim to compare the expression of PEBP4 in tumor samples derived from 58 patients with different grades of gliomas with that in 5 non-neoplastic brain samples and to investigate the clinical significance of PEBP4 expression in gliomas. The mRNA and protein expressions of PEBP4 were measured by real-time quantitative polymerase chain reaction and western blot, respectively. The intracellular expressions of PEBP4 in samples were examined by immunohistochemistry. The association between PEBP4 expression and the clinicopathologic characteristics of gliomas patients were analyzed. Our results demonstrated that the mRNA and protein levels of PEBP4 were upregulated in gliomas tissues, especially in high-grade (World Health Organization Grades III and IV) gliomas, when compared to normal control (p < 0.01). Immunohistochemical analysis indicated that PEBP4 was highly expressed in 82.4% (28/34) of high-grade gliomas, when compared to 41.7% (10/24) of high expression in low-grade gliomas and 20.0% (1/5) in non-neoplastic brain samples (p = 0.001). Multivariate Cox regression analysis revealed that increased PEBP4 expression was an independent prognostic factor for gliomas. Patients with low level of PEBP4 had longer survival time compared to those with high PEBP4 expression (p = 0.003). These data indicate a clinical significance of PEBP4 for predicting the tumor grade and the prognosis in patients with gliomas.

Phosphatidylethanolamine-binding protein 4 promotes lung cancer cells proliferation and invasion via PI3K/Akt/mTOR axis.

While phosphatidylethanolamine-binding protein 4 (PEBP4) is a key factor in the malignant proliferation and metastasis of tumor cells, the exact regulatory network governing its roles remains unclear. This study was designed to investigate the effect of PEBP4 on PI3K/Akt/mTOR pathway and explore its molecular network that governs the proliferation and metastasis of tumor cells. After the recombinant plasmid pcDNA3.1-PEBP4 was constructed, the recombinant plasmid pcDNA3.1-PEBP4 and PEBP4-targeting siRNA were transfected into lung cancer HCC827 cell line. The expressions of PI3K/Akt/mTOR pathway components in HCC827 cells in each group were determined using Western blotting. In the HCC827 cells, the effect of PI3K pathway inhibitor LY294002 on the expressions of PI3K/Akt/mTOR pathway components under the effect of PEBP4 was determined using Western blotting, and the effects of LY294002 on the cell viability, proliferation, and migration capabilities under the overexpression of PEBP4 were determined using MTT method, flow cytometry, and Transwell migration assay. Furthermore, the effect of mTOR inhibitor rapamycin (RAPA) on the expressions of PI3K/Akt/mTOR pathway components under the effect of PEBP4 was determined using Western blotting, and the effects of RAPA on the cell viability, proliferation, and migration capabilities under the overexpression of PEBP4 were determined using MTT method, flow cytometry, and Transwell migration assay. As shown by Western blotting, the protein expressions of p-Akt and phosphorylated mTOR (p-mTOR) were significantly higher in the pcDNA3.1-PEBP4-transfected group than in the normal control group and PEBP4 siRNA group (P<0.05); furthermore, the protein expressions of p-Akt and p-mTOR significantly decreased in the PEBP4 targeting siRNA-transfected group (P<0.05). Treatment with LY294002 significantly inhibited the protein expressions of p-Akt and p-mTOR in HCC827 cells (P<0.05). In contrast, treatment with RAPA only significantly inhibited the protein expression of p-mTOR (P<0.05). As shown by MTT, flow cytometry, and Transwell migration assay, both LY294002 and RAPA could significantly lower the viability of HCC827 cells and inhibit their proliferation and invasion (P<0.05); meanwhile, they could reverse the effect of PEBP4 in promoting the proliferation and migration of HCC827 cells (P<0.05). The overexpression of PEBP4 increases the phosphorylation levels of Akt and mTOR in lung cancer cells. The PI3K/Akt/mTOR signaling axis may be a key molecular pathway via which PEBP4 promotes the proliferation and invasion of non-small cell lung cancer (NSCLC) cells; also, it may serve as a potential therapeutic target.

PEBP4 promoted the growth and migration of cancer cells in pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most common malignancies in the world. Numerous studies have linked the activation of AKT to the progression of PDAC. Phosphatidylethanolamine-binding protein 4 (PEBP4) has been reported to be upregulated in various cancer types. However, its expression pattern and biological functions in PDAC are unknown. In this study, it was found that the messenger RNA (mRNA) and protein level of PEBP4 was elevated in PDAC samples. Forced expression of PEBP4 in PDAC cell lines promoted cell growth and migration, while downregulation of PEBP4 in PDAC cells by RNA interference (RNAi) inhibited the growth, migration, and metastasis of the cancer cells. PEBP4 interacted with AKT and promoted the phosphorylation of serine 473 in AKT. Collectively, this study suggested that PEBP4 might promote the progression of PDAC through activating AKT signaling and PEBP4 might be a promising therapeutic target for PDAC treatment.