lncRNA LINC00960 promotes apoptosis by sponging ubiquitin ligase Nrdp1-targeting miR-183-5p.

The ubiquitin ligase Nrdp1/RNF41 promotes the ubiquitin-dependent degradation of multiple important substrates, including BRUCE/BIRC6, a giant ubiquitin-conjugating enzyme inhibiting both apoptosis and autophagy. miR-183-5p is associated with various malignancies potentially by targeting dozens of genes. Here, we show that the lncRNA LINC00960 binds to the Nrdp1-targeting miR-183-5p and promotes apoptosis. Compared to other known miR-183-5p targets, Nrdp1 mRNA is among the few with top scores to complement miR-183-5p. miR-183-5p binds to the 3'UTR of Nrdp1 mRNA and downregulates Nrdp1 at both the mRNA and protein levels. The miR-183-5p mimics inhibit DNA damage-induced apoptosis probably by upregulating BRUCE level, whereas the miR-183-5p inhibitor suppresses the effects of miR-183-5p. LINC00960 is the noncoding RNA with the highest score to complement miR-183-5p. LINC00960 overexpression reduces, but its knockdown increases, the level of miR-183-5p, whereas LINC00960 overexpression increases, but its knockdown decreases, the level of Nrdp1 and apoptosis. Importantly, the expression of LINC00960, which is associated with multiple types of tumors, positively correlates with that of Nrdp1 in several tumors but inversely correlates with that of miR-183-5p in multiple human tumor cell lines, as analysed by quantitative PCR. Thus, miR-183-5p downregulates Nrdp1 expression and inhibits apoptosis, whereas LINC00960 upregulates Nrdp1 and promotes apoptosis by inhibiting miR-183-5p. These results may provide new ideas for the prevention, diagnosis and treatment of apoptosis-related diseases, such as tumors and neurodegenerative diseases.

A20/Nrdp1 interaction alters the inflammatory signaling profile by mediating K48- and K63-linked polyubiquitination of effectors MyD88 and TBK1.

A20 is a potent anti-inflammatory protein that mediates both inflammation and ubiquitination in mammals, but the related mechanisms are not clear. In this study, we performed mass spectrometry (MS) screening, gene ontology (GO) analysis, and coimmunoprecipitation (co-IP) in a lipopolysaccharide (LPS)-induced inflammatory cell model to identify novel A20-interacting proteins. We confirmed that the E3 ubiquitin ligase Nrdp1, also known as ring finger protein 41 (RNF41), interacted with A20 in LPS-stimulated cells. Further co-IP analysis demonstrated that when A20 was knocked out, degradation-inducing K48-linked ubiquitination of inflammatory effector MyD88 was decreased, but protein interaction-mediating K63-linked ubiquitination of another inflammatory effector TBK1 was increased. Moreover, western blot experiments showed that A20 inhibition induced an increase in levels of MyD88 and phosphorylation of downstream effector proteins as well as of TBK1 and a downstream effector, while Nrdp1 inhibition induced an increase in MyD88 but a decrease in TBK1 levels. When A20 and Nrdp1 were coinhibited, no further change in MyD88 was observed, but TBK1 levels were significantly decreased compared with those upon A20 inhibition alone. Gain- and loss-of-function analyses revealed that the ZnF4 domain of A20 is required for Nrdp1 polyubiquitination. Upon LPS stimulation, the inhibition of Nrdp1 alone increased the secretion of IL-6 and TNF-α but decreased IFN-ß secretion, as observed in other studies, suggesting that Nrdp1 preferentially promotes the production of IFN-ß. Taken together, these results demonstrated that A20/Nrdp1 interaction is important for A20 anti-inflammation, thus revealing a novel mechanism for the anti-inflammatory effects of A20.

Cellular and molecular mechanisms underlying planar cell polarity pathway contributions to cancer malignancy.

While the mutational activation of oncogenes drives tumor initiation and growth by promoting cellular transformation and proliferation, increasing evidence suggests that the subsequent re-engagement of largely dormant developmental pathways contributes to cellular phenotypes associated with the malignancy of solid tumors. Genetic studies from a variety of model organisms have defined many of the components that maintain epithelial planar cell polarity (PCP), or cellular polarity in the axis orthogonal to the apical-basal axis. These same components comprise an arm of non-canonical Wnt signaling that mediates cell motility events such as convergent extension movements essential to proper development. In this review, we summarize the increasing evidence that the Wnt/PCP signaling pathway plays active roles in promoting the proliferative and migratory properties of tumor cells, emphasizing the importance of subcellular localization of PCP components and protein-protein interactions in regulating cellullar properties associated with malignancy. Specifically, we discuss the increased expression of Wnt/PCP pathway components in cancer and the functional consequences of aberrant pathway activation, focusing on Wnt ligands, Frizzled (Fzd) receptors, the tetraspanin-like proteins Vangl1 and Vangl2, and the Prickle1 (Pk1) scaffold protein. In addition, we discuss negative regulation of the Wnt/PCP pathway, with particular emphasis on the Nrdp1 E3 ubiquitin ligase. We hypothesize that engagement of the Wnt/PCP pathway after tumor initiation drives malignancy by promoting cellular proliferation and invasiveness, and that the ability of Wnt/PCP signaling to supplant oncogene addiction may contribute to tumor resistance to oncogenic pathway-directed therapeutic agents.

Clinicopathological and prognostic correlations of HER3 expression and its degradation regulators, NEDD4-1 and NRDP1, in primary breast cancer.

BACKGROUND: Human epidermal growth factor receptor HER3 (ErbB3), especially in association with its relative HER2 (ErbB2), is known as a key oncogene in breast tumour biology. Nonetheless, the prognostic relevance of HER3 remains controversial. NEDD4-1 and NRDP1 are signalling molecules closely related to the degradation of HER3 via ubiquitination. NEDD4-1 and NRDP1 have been reported to contribute to HER3-mediated signalling by regulating its localization and cell membrane retention. We studied correlations between HER3, NEDD4-1, and NRDP1 protein expression and their association with tumour histopathological characteristics and clinical outcomes. METHODS: The prevalence of immunohistochemically detectable expression profiles of HER3 (n = 177), NEDD4-1 (n = 145), and NRDP1 (n = 145) proteins was studied in primary breast carcinomas on archival formalin-fixed paraffin-embedded (FFPE) samples. Clinicopathological correlations were determined statistically using Pearson's Chi-Square test. The Kaplan-Meier method, log-rank test (Mantel-Cox), and Cox regression analysis were utilized for survival analysis. RESULTS: HER3 protein was expressed in breast carcinomas without association with HER2 gene amplification status. Absence or low HER3 expression correlated with clinically aggressive features, such as triple-negative breast cancer (TNBC) phenotype, basal cell origin (cytokeratin 5/14 expression combined with ER negativity), large tumour size, and positive lymph node status. Low total HER3 expression was prognostic for shorter recurrence-free survival time in HER2-amplified breast cancer (p = 0.004, p = 0.020 in univariate and multivariate analyses, respectively). The majority (82.8%) of breast cancers demonstrated NEDD4-1 protein expression - while only a minor proportion (8.3%) of carcinomas expressed NRDP1. NEDD4-1 and NRDP1 expression were not associated with clinical outcomes in HER2-amplified breast cancer, irrespective of adjuvant trastuzumab therapy. CONCLUSIONS: Low HER3 expression is suggested to be a valuable prognostic biomarker to predict recurrence in HER2-amplified breast cancer. Neither NEDD4-1 nor NRDP1 demonstrated relevance in prognostics or in the subclassification of HER2-amplified breast carcinomas.

CgNrdp1, a conserved negative regulating factor of MyD88-dependent Toll like receptor signaling in oyster Crassostrea gigas.

Toll like receptor (TLR) signaling cascades are under precise regulations to ensure the proper immune responses during various pathogen invasions. The neuregulin receptor degradation protein-1 (Nrdp1) has been demonstrated to be a novel negative regulator of TLR signaling by targeting MyD88 to induce degradation in mammals. In the present study, an Nrdp1 homologue, CgNrdp1, was identified from the genome of Pacific oyster Crassostrea gigas. It contained an open reading frame encoding a polypeptide of 315 amino acids which shared high identities with other homologues from different species. There was a conserved RING domain in CgNrdp1, indicating the functional E3 ubiquitin ligase activity. The bacterially expressed recombinant CgNrdp1 and CgMyD88 showed much stronger affinity compared to control groups in the ELISA assay, showing the interacting ability between CgNrdp1 and CgMyD88. When CgMyD88 or HsMyD88 was co-transfected with CgNrdp1 into HEK293T cells, the luciferase activities of NF-κB were significantly decreased compared to those in MyD88 single-transfection groups, indicating the conserved negative regulating function of CgNrdp1 on the MyD88 induced TLR signaling. These results indicated that CgNrdp1 was a negative regulator of TLR signaling in oyster and the Nrdp1-MyD88 axis was functional and highly conserved from mollusks to mammals in the negative regulation of TLR signaling.

Nrdp1S, short variant of Nrdp1, inhibits human glioma progression by increasing Nrdp1-mediated ErbB3 ubiquitination and degradation.

The ubiquitin ligase neuregulin receptor degradation protein 1 (Nrdp1) is involved in the induction of apoptosis and suppression of tumour formation. We previously showed that it was expressed at lower levels in human glioma tissues compared with normal brain tissues. However, the mechanism underlying this is unclear. Here, we reported that a novel short variant (Nrdp1S), lacking 71 amino acids at the N-terminal, was expressed in normal human brain tissue, but absent from glioma tissues. Similar to Nrdp1, Nrdp1S could be degraded by the proteasomal pathway, but exhibited an even longer half-life than Nrdp1. Nrdp1S was also shown to form a heterodimer with Nrdp1, which increased its stability, thereby augmenting the Nrdp1-mediated ubiquitination and degradation of ErbB3. EdU incorporation, MTT assay and in vitro colony formation demonstrated that Nrdp1S significantly inhibited the cell tumourigenicity. These results together suggest that Nrdp1S is a tumour suppressor that which potentiates the Nrdp1-mediated ubiquitination and degradation of ErbB3. An Nrdp1S deficiency may also be an important factor in the loss of Nrdp1.

Phosphorylation of the E3 ubiquitin ligase RNF41 by the kinase Par-1b is required for epithelial cell polarity.

The establishment and maintenance of cell polarity is an essential property governing organismal homeostasis, and loss of polarity is a common feature of cancer cells. The ability of epithelial cells to establish apical-basal polarity depends on intracellular signals generated from polarity proteins, such as the Par-1 family of proteins, as well as extracellular signals generated through cell contacts with the extracellular matrix (ECM). The Par-1 family has a well-established role in regulating cell-cell contacts in the form of tight junctions by phosphorylating Par-3. In addition, Par-1 has been shown to impact on cell-ECM interactions by regulating laminin receptor localization and laminin deposition on the basal surface of epithelial cells. Laminins are major structural and signaling components of basement membrane (BM), a sheet of specialized ECM underlying epithelia. In this study, we identify RNF41, an E3 ubiquitin ligase, as a novel Par-1b (also known as MARK2) effector in the cell-ECM pathway. Par-1b binds to and phosphorylates RNF41 on serine 254. Phosphorylation of RNF41 by Par-1b is required for epithelial cells to localize laminin-111 receptors to their basolateral surfaces and to properly anchor to laminin-111. In addition, phosphorylation of RNF41 is required for epithelial cells to establish apical-basal polarity. Our data suggests that phosphorylation of RNF41 by Par-1b regulates basolateral membrane targeting of laminin-111 receptors, thereby facilitating cell anchorage to laminin-111 and ultimately forming the cell-ECM contacts required for epithelial cells to establish apical-basal cell polarity.

Hepatitis B virus X protein reduces the stability of Nrdp1 to up-regulate ErbB3 in hepatocellular carcinoma cells.

Hepatitis B virus (HBV)-associated hepatocellular carcinoma (HCC) is the most widespread type of liver cancer. However, the underlying mechanism of HCC tumorigenesis is very intricate and HBV-encoded X protein (HBx) has been reported to play a key role in this process. It has been reported that HBx up-regulates the transcription of ErbB3. However, it remains unclear whether HBx can regulate ErbB3 expression at post-translational modification level. In this study, we showed that HBx interacts with ubiquitin ligase Nrdp1 (neuregulin receptor degradation protein 1) and decreases its stability, which results in the up-regulation of ErbB3 and promotion of HCC cells. Moreover, the expression of ErbB3 was almost undetectable in normal liver tissues but was relative abundant in HCC tissues, and the level of ErbB3 and Nrdp1 significantly showed a negative correlation in HCC tissues. Taken together, these findings suggest that HBx promotes the progression of HCC by decreasing the stability of Nrdp1, which results in up-regulation of ErbB3, suggesting that ErbB3 may be a target for HCC therapy.

Regulation of ubiquitin-specific processing protease 8 suppresses neuroinflammation.

In our previous study, we reported that luteolin might exert neuroprotective functions by inhibiting the production of proinflammatory mediators, thereby suppressing microglial activation. In this study, we used two-dimensional gel electrophoresis (2-DE) and mass spectrometry (MS) to study the effect of ubiquitin-specific processing protease 8 (USP8) in luteolin-treated microglia. Western blot analysis verified that USP8 expression is upregulated by luteolin. Researchers have found that USP8 markedly enhanced the stability of neuregulin receptor degradation protein-1 (Nrdp1), which in turn inhibited the production of proinflammatory cytokines in toll-like receptor-triggered macrophages. We next hypothesized that luteolin inhibits microglial inflammation by regulating USP8 gene expression. After transfecting BV2-immortalized murine microglial cells with USP8, a significant reduction in the degradation of Nrdp1 was observed. USP8 overexpression also reduced the production of lipopolysaccharide (LPS)-induced proinflammatory mediators such as inducible nitric oxide synthase (iNOS), nitric oxide (NO), cyclooxygenase-2 (COX-2), and prostaglandin E2 (PGE2). We also found that USP8 siRNA blocked luteolin inhibition of pro-inflammatory gene expression such as iNOS, NO, COX-2, and PGE2. Taken together, our findings suggested that luteolin inhibits microglial inflammation by enhancing USP8 protein production. We concluded that in addition to anti-inflammatory luteolin, USP8 might represent a novel mechanism for the treatment of neuroinflammation and neurodegeneration.

Reciprocal cross-regulation between RNF41 and USP8 controls cytokine receptor sorting and processing.

The mechanisms controlling the steady-state cell surface levels of cytokine receptors, and consequently the cellular response to cytokines, remain poorly understood. The number of surface-exposed receptors is a dynamic balance of de novo synthesis, transport to the plasma membrane, internalization, recycling, degradation and ectodomain shedding. We previously reported that the E3 ubiquitin ligase RING finger protein 41 (RNF41) inhibits basal lysosomal degradation and enhances ectodomain shedding of JAK2-associated cytokine receptors. Ubiquitin-specific protease 8 (USP8), an RNF41-interacting deubiquitylating enzyme (DUB) stabilizes RNF41 and is involved in trafficking of various transmembrane proteins. The present study identifies USP8 as a substrate of RNF41 and reveals that loss of USP8 explains the aforementioned RNF41 effects. RNF41 redistributes and ubiquitylates USP8, and reduces USP8 levels. In addition, USP8 knockdown functionally matches the effects of RNF41 ectopic expression on the model leptin and leukemia inhibitory factor (LIF) receptors. Moreover, RNF41 indirectly destabilizes the ESCRT-0 complex through suppression of USP8. Collectively, our findings demonstrate that RNF41 controls JAK2-associated cytokine receptor trafficking by acting as a key regulator of USP8 and ESCRT-0 stability. Balanced reciprocal cross-regulation of RNF41 and USP8 thus determines whether receptors are sorted for lysosomal degradation or recycling, this way regulating basal cytokine receptor levels.

MiR-497 promotes metastasis of colorectal cancer cells through Nrdp1 inhibition.

We have recently shown that Nrdp1 inhibits phosphorylation of ErB3 in colorectal cancer (CRC) cells, to suppress epidermal growth factor receptor (EGFR) signaling-stimulated MMP7 activation for CRC metastasis. In this study, we examined the control of Nrdp1 in CRC cells. We detected significant increases in miR-497 in CRC specimen, compared to paired normal colorectal tissue. Moreover, we detected a strong positive correlation between miR-497 levels and Nrdp1 levels, and a strong inverse correlation between miR-497 levels and MMP7 levels. In vitro, overexpression of miR-497 in human CRC cells significantly decreased Nrdp1 transcripts and protein, and vice versa. Moreover, overexpression of miR-497 in human CRC cells also significantly increased MMP7 transcripts, cellular protein, and secreted protein, resulting in increases in cell invasiveness in a transwell cell migration assay. Furthermore, we found that MiR-497 directly targeted 3'-UTR of Nrdp1 mRNA to inhibit its translation. Together, our data suggest that the regulation of MMP7 by Nrdp1 in CRC cells could be inhibited by miR-497 through suppressing Nrdp1 translation. Our work thus highlights a novel molecular regulatory machinery that regulates metastasis of CRC.

Nrdp1-mediated ErbB3 degradation inhibits glioma cell migration and invasion by reducing cytoplasmic localization of p27(Kip1).

We previously reported that loss of Nrdp1 contributes to human glioma progression by reducing apoptosis. However, the role of Nrdp1 in glioma migration and invasion has not been investigated. Here, we report that ErbB3, a substrate of Nrdp1, is undetectable in normal brain tissues and grade II/III glioma tissues, but is abundant in a certain percentage of grade IV glioma tissues and is associated with the loss of Nrdp1. This suggests that Nrdp1 may be involved in glioma migration and invasion by regulating ErbB3. Thus, the role of Nrdp1/ErbB3 signaling in glioma cell migration and invasion was investigated using Nrdp1 loss- and gain-of-function. The results show that down-regulation of Nrdp1 by use of short hairpin RNA promoted glioma cell migration and invasion. In contrast, overexpression of Nrdp1 significantly inhibited glioma cell migration and invasion. Further investigation on molecular targets revealed that Nrdp1 decreased the level of ErbB3, which resulted in decreasing p-AKT thereby reducing cytoplasmic p27(Kip1). Taken together, these findings suggest that Nrdp1-mediated ErbB3 degradation suppresses glioma migration and invasion and that loss of Nrdp1 may amplify ErbB3 signaling to contribute to glioma migration and invasion. These findings suggest that Nrdp1 may be a target for glioma therapy.

Expression profiles and interaction suggest TBK1 can be regulated by Nrdp1 in response to immune stimulation in large yellow croaker Larimichthys crocea.

TBK1 has been extensively studied in mammals because of its important roles as a molecular bridge, linking the TLRs (TLR3 and TLR4) and RLRs signals to activate transcriptional factors IRF3 and IRF7 for IFN-I production. However, the information on molecular and functional characteristics of TBK1 in teleosts is limited. In this study, the molecular characterization and immune response of TBK1 in Larimichthys crocea (named as LcTBK1) as well as its interaction with Nrdp1 were investigated. Sequence analysis demonstrated that LcTBK1 included four functional motifs, the N-terminal protein kinase domain and ATP-binding site, middle ULD and C-terminal coiled-coil domain. The tissue expression profiles indicated that LcTBK1 gene was constitutively expressed in the twelve tissues examined, with high expression in brain. Temporal expression analysis showed that LcTBK1 mRNA was obviously increased in the liver after injection of LPS, Poly I:C and inactive Vibrio parahaemolyticus, however, declined at some time points in spleen and head-kidney. Furthermore, we found that LcTBK1 can interact with LcNrdp1, an E3 ubiquitin ligase that involved in immune response to Cryptocaryon irritans infection in L. crocea. The qPCR showed that LcNrdp1 was also significantly up-regulated in liver, down-regualted at some time points in spleen and head-kidney after LPS, Poly I:C and inactive V. parahaemolyticus injection, although the expression patterns of the two genes after the three treatments were different in change magnitude and up-regulation timespan. These results suggested that LcTBK1 was involved in L. crocea defense against the pathogen infection and can be regulated by Nrdp1 in PPRs signaling pathway of fishes.

Lower expression of Nrdp1 in human glioma contributes tumor progression by reducing apoptosis.

Ubiquitin ligase Nrdp1 (neuregulin receptor degradation protein 1) plays important roles in multiple physiological process because it can ubiquitinate various substrates such as ErbB3, BRUCE, MyD88, C/EBPß, and Parkin, and so forth. In addition to the physiological function, it was also found to be involved in tumor progression. It has been shown that loss of Nrdp1 enhances breast cancer cell growth. Up to now, the role of Nrdp1 in glioma has not been elucidated. Here, we reported that Nrdp1 as well as cleaved caspase 3 was lower expressed in human glioma tissues comparing with the nontumorous. And then we found that the expression of Nrdp1 and cleaved caspase 3 was increased in the treatment of Temozolomide (TMZ), a drug for glioma chemotherapy. Further investigation indicated that transient transfection of Nrdp1 significantly promoted cell apoptosis by aggravating the degradation of BRUCE and activation of caspase 3. In addition, overexpression of Nrdp1 augmented TMZ induced apoptosis by evaluating the degradation of BRUCE and the activation of caspase 3, while silencing of Nrdp1 reduced the sensitivity to the TMZ by inhibiting the degradation of BRUCE and the activation of caspase 3 in human glioma cells. These observations show that Nrdp1 is a pro-apoptotic protein in human glioma and lower expression of Nrdp1 in human glioma may promote tumor progression by reducing apoptosis, suggesting that Nrdp1 may be an important regulator in the development of human glioma.

Ubiquitin Ligase Nrdp1 Controls Autophagy-Associated Acrosome Biogenesis and Mitochondrial Arrangement during Spermiogenesis.

Autophagy is critical to acrosome biogenesis and mitochondrial quality control, but the underlying mechanisms remain unclear. The ubiquitin ligase Nrdp1/RNF41 promotes ubiquitination of the mitophagy-associated Parkin and interacts with the pro-autophagic protein SIP/CacyBP. Here, we report that global deletion of Nrdp1 leads to formation of the round-headed sperm and male infertility by disrupting autophagy. Quantitative proteome analyses demonstrated that the expression of many proteins associated with mitochondria, lysosomes, and acrosomes was dysregulated in either spermatids or sperm of the Nrdp1-deficient mice. Deletion of Nrdp1 increased the levels of Parkin but decreased the levels of SIP, the mitochondrial fission protein Drp1 and the mitochondrial protein Tim23 in sperm, accompanied by the inhibition of autophagy, the impairment of acrosome biogenesis and the disruption of mitochondrial arrangement in sperm. Thus, our results uncover an essential role of Nrdp1 in spermiogenesis and male fertility by promoting autophagy, providing important clues to cope with the related male reproductive diseases.

Androgen Receptor-Mediated Nuclear Transport of NRDP1 in Prostate Cancer Cells Is Associated with Worse Patient Outcomes.

To our knowledge, our group is the first to demonstrate that NRDP1 is located in the nucleus as well as the cytoplasm of CaP cells. Subcellular fractionation, immunohistochemistry, and immunofluorescence analysis combined with confocal microscopy were used to validate this finding. Subcellular fractionation followed by western blot analysis revealed a strong association between AR and NRDP1 localization when AR expression and/or cellular localization was manipulated via treatment with R1881, AR-specific siRNA, or enzalutamide. Transfection of LNCaP with various NRDP1 and AR constructs followed by immunoprecipitation confirmed binding of NRDP1 to AR is possible and determined that binding requires the hinge region of AR. Co-transfection with NRDP1 constructs and HA-ubiquitin followed by subcellular fractionation confirmed that nuclear NRDP1 retains its ubiquitin ligase activity. We also show that increased nuclear NRDP1 is associated with PSA recurrence in CaP patients (n = 162, odds ratio; 1.238, p = 0.007) and that higher levels of nuclear NRDP1 are found in castration resistant cell lines (CWR22Rv1 and PC3) compared to androgen sensitive cell lines (LNCaP and MDA-PCa-3B). The combined data indicate that NRDP1 plays a role in mediating CaP progression and supports further investigation of both the mechanism by which nuclear transport occurs and the identification of specific nuclear targets.

MiRNA-494 enhances M1 macrophage polarization via Nrdp1 in ICH mice model.

BACKGROUND: Ubiquitination-mediated M1/M2 macrophage polarization plays important roles in the pathogenesis of immune disease. However, the regulatory mechanism of ubiquitination during M1/M2 macrophage polarization following intracerebral hemorrhage (ICH) has not been well studied. METHODS: In the experiment, macrophages were administered with erythrocyte lysates, and then miR-494-, Nrdp1-, and M1/M2-related markers were analyzed. Brain inflammatory response, brain edema, and neurological functions of ICH mice were also assessed. RESULTS: We found that miR-494 levels increased while Nrdp1 levels decreased in macrophages after ICH. We also demonstrated that miR-494 inhibited Nrdp1 expression by directly binding its 3'-untranslated region. MiR-494 attenuated C/EBP-ß activation and downstream proinflammatory factor production. Upregulation of Nrdp1 in macrophages significantly promoted M2 macrophage polarization via ubiquitinating and activating C/EBP-ß. Moreover, the results indicated that miR-494 could enhance M1 macrophage polarization, promote brain edema, and impair neurological functions in ICH mice. CONCLUSIONS: Taken together, our results demonstrated that Nrdp1 contributed to M1/M2 macrophage polarization and neuroinflammation via ubiquitination and activation of C/EBP-ß in ICH. miR-494 may provide a promising therapeutic clue for ICH.

Role of the Nrdp1 in Brain Injury Induced by Chronic Intermittent Hypoxia in Rats via Regulating the Protein Levels of ErbB3.

Obstructive sleep apnea syndrome (OSAS) is known as a repeated obstruction of the upper airway during sleep, leading to generalized hypoxia episodes and associated with cardiovascular and cerebrovascular diseases. We mainly explored the role of neuregulin receptor degradation protein-1 (Nrdp1, also known as FLRF) in brain injury induced by chronic intermittent hypoxia (CIH) in rats. Wistar rats were randomly divided into 4 groups (n = 12 per group), including the sham + adeno-associated virus-NC (AAV-NC) group, the sham + AAV-siNrdp1 group, the IH-4w (intermittent hypoxia for 4 weeks) + AAV-NC group, and the IH-4w + AAV-siNrdp1 group. Morphologic changes in brain tissue were observed by hematoxylin and eosin (HE) staining. Apoptosis in the hippocampus was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) staining. Spatial learning and memory were assessed by the Morris water maze test. The expression of Nrdp1 mRNA and protein in the hippocampus was detected by qualitative real-time polymerase chain reaction (qRT-PCR) and Western blotting. The concentration of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in serum was detected via enzyme-linked immunosorbent assay (ELISA) kits. Nrdp1 expression was increased after intermittent hypoxia exposure over time. Western blotting and H&E results showed that pathological changes of hippocampus neurons in chronic intermittent hypoxia rat were diminished by shNrdp1. Western blotting and TUNEL staining showed that apoptotic cells in the hippocampus of CIH rats were decreased by shNrdp1. The Morris water maze results proved that shNrdp1 improved spatial learning performance of chronic intermittent hypoxia rats. ELISA kits results showed that CIH-induced inflammatory response was decreased by shNrdp1. Western blotting and qRT-PCR results showed protein expression of ErbB3 in the hippocampus of CIH rats. Nrdp1 could regulate ErbB3 protein levels in brain-injured rats with CIH, which demonstrates that Nrdp1 is a potential therapeutic target in the cognition deficits associated with OSAS.

Prediction of Ubiquitin Ligase Nrdp1-Associated Proteins in Glioma Database.

The ubiquitin proteasome pathway is conserved from yeast to mammals and is necessary for the targeted degradation of most short-lived proteins in eukaryotic cells. Its protein substrates include cell cycle regulatory proteins and proteins that are not properly folded in the endoplasmic reticulum. Owing to the ubiquity of its protein substrates, ubiquitination regulates a variety of cellular activities, including cell proliferation, apoptosis, autophagy, endocytosis, DNA damage repair, and immune response. With new genomic data continuously being obtained, ubiquitination through genomic data analysis will be an effective method. We obtained 83 overlapping genes from four glioma databases, which differed from ubiquitin ligase Nrdp1 expression, including 36 downregulated and 47 upregulated genes. The KEGG pathways, molecular functions, cellular components, and biological processes potentially associated with Nrdp1 were obtained using GSEA and Cytoscape. In human gliomas, differences in the expression of Nrdp1 were identified between nontumor brain tissue and different glioma tissues, but no difference in expression was found between low­grade glioma (LGG) and anaplastic glioma (AG). In survival analysis, we found no significant association between Nrdp1 expression level and patient prognosis.

Nrdp1 increases neuron apoptosis via downregulation of Bruce following intracerebral haemorrhage.

BACKGROUND: Neuregulin receptor degradation protein-1 (Nrdp1) is an E3 ubiquitin ligase that plays an important role in regulating cell growth, apoptosis and oxidative stress. However, the data regarding its expression and exact mechanism in neuronal injury following ICH has not been well identified. METHODS: In this study, primary cortical neurons from C57BL/6 mice were subjected to erythrocyte lysates. Nrdp1 expression, cell apoptosis, caspase-3 and BRUCE levels were detected. In addition, inflammatory response, brain edema, and neurological injury in ICH mice were also assessed. RESULTS: We found that the expression of Nrdp1 was significantly increased in neuron cells accompanied by up-regulation of active caspase-3 and decreased expression of BRUCE (an inhibitor of apoptosis protein). However, inhibiting Nrdp1 levels of neurons reduced caspase-3 activity but induced up-regulation of BRUCE. In vivo, inhibiting Nrdp1 levels increased pro-inflammatory cytokines, brain edema, and neurological injury following ICH. CONCLUSIONS: Taken together, the data suggested that Nrdp1 might play a crucial role in neuronal apoptosis via inhibiting BRUCE following ICH.