scholarly journals Smoothened Stabilizes and Protects TRAF6 from Proteosomal Degradation: A Novel Non-Canonical Role of Smoothened with Implications in Lymphoma Biology

Blood ◽  
2017 ◽  
Vol 130 (Suppl_1) ◽  
pp. 646-646
Author(s):  
Changju QU ◽  
Amineh Vaghefi ◽  
Kranthi Kunkalla ◽  
Jennifer R Chapman ◽  
Yadong Liu ◽  
...  
Keyword(s):  
Cell Survival ◽  
Cell Lines ◽  
E3 Ligase ◽  
B Cell Lymphoma ◽  
Mrna Levels ◽  
Protein Levels ◽  
Multiple Cancers ◽  
Proteosomal Degradation ◽  
Hh Signaling

Abstract Tumor necrosis factor receptor-associated factor 6 (TRAF6), an (K63) E3-ligase, plays a crucial role in many biological processes and its activity is relevant in the biology of multiple cancers including diffuse large B cell lymphoma (DLBCL). Although molecules that trigger TRAF6 activation have been defined, those that stabilize TRAF6 levels and/or enhance TRAF6 function remain largely unclear. Previously, we found that activation of smoothened (SMO) with recombinant Hedgehog (Hh) ligand increased the binding between SMO with TRAF6, as well as TRAF6 protein levels (Blood 2013; 121:4718-28). In addition, transient overexpression of SMO resulted in increased K63-Ub of both TRAF6 and NEMO indicating stabilization of these proteins resulting in NF-kB activation. This is relevant, as more recently we found that TRAF6 amplifies pAKT signaling in DLBCL and that TRAF6 is the dominant E3 ligase for the K63-Ub of AKT in DLBCL. Moreover, TRAF6 recruitment to the cell membrane, and stabilization of its ubiquitination profile are facilitated by SMO. SMO is a member of the Frizzled-class G-protein-coupled receptor (GPCRs) and is traditionally known for its role as signal transducer in canonical Hedgehog (Hh) signaling. These observations prompted us to investigate whether the ability of SMO to increase TRAF6 levels is limited to ligand induced signaling, whether it contributes to chemoresistance in DLBCL cells, and whether SMO directly participates in controlling TRAF6 levels. To confirm the regulatory role of SMO in the TRAF6/AKT axis in DLBCL cells (HBL1 and HT) and further outline the nature of the underlying regulation, we measured the impact of activation of the Hh pathway with recombinant Shh ligand on TRAF6 levels, with and without SMO knockdown or recombinant SMO overexpression. Canonical Hh signaling results in the activation of the GLI1 transcription factor and the subsequent elevation of GLI1 mRNA levels is an established indicator of activation of the Hh pathway. However, neither SMO activation nor the knockdown of GLI1 had a significant impact on TRAF6 mRNA levels. These findings indicate that TRAF6 is not transcriptionally regulated by SMO signaling through GLI1 (canonical Hh signaling). In contrast, overexpression of SMO or siRNA knockdown of SMO resulted in an increase or decrease of TRAF6 protein levels, respectively. Consistent with the decrease of AKT activation (pAKT T308 and S473) after TRAF6 knockdown, the increase in TRAF6 levels that follows SMO overexpression resulted in an increase in the levels of AKT phosphorylation. Altogether, these observations suggest a post-translational regulation of TRAF6 by SMO. Indeed, stable knockdown of SMO dramatically reduces the half-life of TRAF6 in both HBL1 and HT cells in the presence of cyclohexamide. Furthermore, overexpression of SMO increases K63-Ub of both TRAF6 and AKT. In contrast, the SMO induced decrease in K48-Ub occurred only for TRAF6 but not for AKT. These data link the SMO-stimulated activation of TRAF6 to the enhancement of AKT signaling and protection of TRAF6 from proteasomal degradation. Mechanistically, we found that SMO, through its C-terminal tail, stabilizes TRAF6 and protects TRAF6 from proteosomal degradation, an effect mediated by ubiquitin-specific protease-8 (USP8). Importantly, this functional link between SMO and TRAF6 is reflected in DLBCL patient samples where high expression of both molecules correlates with poor prognosis. Resistance to DXR is a serious challenge in the treatment of DLBCL, and activated AKT is known to contribute to DXR resistance in multiple cancers including DLBCL. We evaluated whether SMO and TRAF6 support resistance to DXR in DLBCL cell lines. We exposed HT and HBL1 cells as well as their counterparts with stable knockdown of TRAF6 or SMO to DXR for 96hrs. Cell viability after exposure to DXR was determined by an Annexin V and PI staining assay. Silencing SMO or TRAF6 dramatically decreased cell survival after treatment with DXR. In summary, we report that SMO is needed to facilitate and maintain TRAF6-dependent elevated pAKT levels in DLBCL cell lines of germinal (GC) and non-GC subtypes, and that the SMO/TRAF6 axis contributes to DXR resistance in DLBCL. Our study reveals a novel and potential central cell survival signaling mechanism in which SMO stabilizes and protects TRAF6 from proteosomal degradation. Disclosures Lossos: Affimed: Research Funding.

scholarly journals Assessment of TSPAN Expression Profile and Their Role in the VSCC Prognosis

2021 ◽  
Vol 22 (9) ◽  
pp. 5015
Author(s):  
Kelly Pedrozo Ferreira ◽  
Bruna Cristine de Almeida ◽  
Laura Gonzalez dos Anjos ◽  
Glauco Baiocchi ◽  
Fernando Augusto Soares ◽  
...  
Keyword(s):  
Cell Lines ◽  
Expression Profile ◽  
Tissue Level ◽  
Cellular Level ◽  
Metastasis Suppressor ◽  
Mrna Levels ◽  
Vulvar Squamous Cell Carcinoma ◽  
Protein Levels ◽  
Worse Prognosis

The role and prognostic value of tetraspanins (TSPANs) in vulvar squamous cell carcinoma (VSCC) remain poorly understood. We sought to primarily determine, at both the molecular and tissue level, the expression profile of the TSPANs CD9, CD63, CD81, and CD82 in archived VSCC samples (n = 117) and further investigate their clinical relevance as prognostic markers. Our studies led us to identify CD63 as the most highly expressed TSPAN, at the gene and protein levels. Multicomparison studies also revealed that the expression of CD9 was associated with tumor size, whereas CD63 upregulation was associated with histological diagnosis and vascular invasion. Moreover, low expression of CD81 and CD82 was associated with worse prognosis. To determine the role of TSPANs in VSCC at the cellular level, we assessed the mRNA levels of CD63 and CD82 in established metastatic (SW962) and non-metastatic (SW954) VSCC human cell lines. CD82 was found to be downregulated in SW962 cells, thus supporting its metastasis suppressor role. However, CD63 was significantly upregulated in both cell lines. Silencing of CD63 by siRNA led to a significant decrease in proliferation of both SW954 and SW962. Furthermore, in SW962 particularly, CD63-siRNA also remarkably inhibited cell migration. Altogether, our data suggest that the differential expression of TSPANs represents an important feature for prognosis of VSCC patients and indicates that CD63 and CD82 are likely potential therapeutic targets in VSCC.

Expression of Lipoprotein Lipase In Chronic Lymphocytic Leukemia Cells: Is There a Biological Function?

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 3580-3580
Author(s):  
Edit anna Porpaczy ◽  
Stefanie Tauber ◽  
Martin Bilban ◽  
Gerhard Kostner ◽  
Michaela Gruber ◽  
...  
Keyword(s):  
Enzymatic Activity ◽  
Cell Survival ◽  
Mrna Expression ◽  
Lipoprotein Lipase ◽  
Cell Lines ◽  
Knock Down ◽  
Protein Levels ◽  
Healthy Donors

Abstract Abstract 3580 The expression of lipoprotein lipase (LPL) in CLL cells is an established mRNA surrogate marker for immunoglobulin heavy chain (IgVH) mutational status. High expression of LPL correlates with poor prognosis. However, the possible functional role of LPL in CLL is still unclear. LPL is normally expressed in muscle cells, adipose tissue and macrophages, transported to the luminal surface of endothelial cells where it is bound heparan sulfate-proteoglycans (HSPG). Heparin competes with HSPG for the binding sites and intravenous injection leads to elevated plasma LPL protein levels and enzymatic activity (“heparin release test”). LPL mRNA levels correlate with intracellular protein expression (Heintel et al. Leukemia. 2005; Mansouri et al. Leuk Res. 2010). Moreover cellular lysates from CLL patients contain elevated LPL enzymatic activity compared to healthy donors (Pallasch et al. Leukemia. 2008.). In this study, we investigated the basal (pre-heparin) LPL protein levels by enzyme-linked immunosorbent assay in the serum of 42 CLL patients, 14 non-CLL patients (lymphoma in remission), and 4 healthy donors (HD): Median pre-heparin LPL protein levels were 40.10 ng/ml (range: 5.66–108.44), 44.11 ng/ml (18.26-84.08), and 68.14 ng/ml (33.28-174.38), respectively. Among CLL patients there were no significant differences between those with high (N=16; median LPL protein in serum: 38.10 ng/ml (8.72-73.49)) and low (N=26; 43.12 ng/ml (5.66-108.44) (p=0.354) LPL mRNA expression. Thirteen patients with known LPL mRNA expression were investigated for LPL protein “release” after heparin injection. Ten and twenty minutes after 50 U/kg heparin injection, the elevation of both parameters, LPL protein amount in serum and enzymatic activity in plasma, was similar to those of HD normal values. In detail, medium serum protein levels in samples with high LPL mRNA (N=5) increased from 16.11 to 214.33 and 332.78 ng/ml and in the samples with low mRNA (N=8) from 13.08 to 219.68 and 386.65 ng/ml, respectively. The corresponding median values of the LPL enzymatic activities in high vs. low expressors were: 7.25/15.52/20.01 and 7.45/19.13/20.57 μ M/ml/h. In addition, release of LPL from peripheral mononuclear cells (PBMC) of CLL patients (N=3) by heparin in vitro was absent. Cell viability and LPL mRNA expression remained unaffected in both in vivo and in vitro samples after heparin addition. In order to assess the impact of LPL on cell survival, CLL cells were cultured (N=3) for up to 72 hours with different doses of purified LPL protein. There was no positive effect on cell survival irrespective of primary LPL mRNA expression or culture conditions (with or without FCS). Since these results point to an intracellular effect of LPL, we aimed to identify downstream targets by knock down with siRNA against LPL in 7 CLL samples and 5 cell lines (hepatocellular carcinoma, cervix carcinoma, colon carcinoma, multiple myeloma and acute monocytic leukemia) with high LPL mRNA expression. Gene expression changes were analyzed by microarrays (GeneChip® Human Gene 1.0 ST Array, Affymetrix). Fifteen genes were up- (N=4) or downregulated (N=11) in at least 3 of 5 cell lines by more than 1.5-fold (e.g. GSTP1, COROC1). Nine genes were at least 1.5-fold downregulated in parallel with LPL in the CLL samples only. These genes belong to various pathways (e.g. cell cycle, signaling in immune system, metabolism of carbohydrates) and seem to be specific for CLL. Cross-validation of individual genes is under way. Our data suggest that (1) neither basal serum LPL protein levels nor heparin-induced LPL release in CLL patients are suitable clinical prognostic markers; (2) Stimulation with external LPL protein does not affect CLL cell survival; (3) siRNA knock-down of LPL induces changes in various functional pathways. We conclude that the key role of LPL expression in high-risk CLL is related to its (intra)cellular expression. Disclosures: No relevant conflicts of interest to declare.

GLI1 Directly Regulates the Transcription of AKT Genes in Diffuse Large B-Cell Lymphoma.

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 2399-2399
Author(s):  
Nitin K Agarwal ◽  
Changju Qu ◽  
Kranthi Kunkalla ◽  
Yadong Liu ◽  
Francisco Vega
Keyword(s):  
Cell Viability ◽  
Cell Survival ◽  
Mrna Expression ◽  
Real Time ◽  
Cell Lines ◽  
Real Time Pcr ◽  
Luciferase Reporter ◽  
Knock Down ◽  
Hh Signaling

Abstract Abstract 2399 Activation of the Hedgehog (Hh)/glioma-associated oncogene (GLI) pathway has been found in a growing number of malignancies. We have provided evidence that canonical Hh signaling is required for cell survival and proliferation of DLBCL cell lines. To confirm the pathogenic role of GLI1 in DLBCL, we established GLI1 knock down DLBCL cell lines (OCI-Ly19, HBL-1 and BJAB) using a lentiviral shRNA system and performed cell viability and apoptosis assays. Cell viability assays demonstrated that GLI1 knockdown DLBCL cells experienced a statistically significantly decrease in the number of viable cells in comparison with control cells harboring scramble shRNA. To examine whether decreases number of cell viability in GLI1 knock down cells were due to apoptosis, we performed annexin V and PI assays. We observed marked increase of apoptosis in GLI1 knock down DLBCL cells versus controls (2.5 fold increase for OCI-Ly10, and 5 fold for HBL1 and BJAB). To investigate the mechanism by which GLI1 regulates tumorigenesis and cell survival, we searched for whole genome GLI1-target genes in DLBCL cells using CHIP sequencing technique and identified AKT genes as potential targets of GLI1. Using pharmacological and silencing approaches, we observed that Hh signaling modulates the expression of AKT genes in DLBCL cells. We further identified two putative binding sites for GLI1 in the AKT1 promoter region and confirmed their functionality using chromatin immunoprecipitation, luciferase reporter and site-directed mutagenesis assays. To investigate whether there is any correlation between AKT1 and GLI1 mRNA expression in human DLBCL tumors, we performed quantitative real-time PCR analyses in 17 frozen DLBCL specimens including apharesis samples from pleural effusions. The real time PCR analysis revealed a strong Spearmen correlation coefficient (R2=0.9) between GLI1 and AKT1 mRNA expression. In summary, we provide evidence of the role of GLI1 in the pathobiology of DLBCL and demonstrated a cross talk, at the transcriptional level, between Hh signaling and AKT in DLBCL. A link between these 2 pathways at the trasncriptional level was not previoulsy documented. This finding is of clinical interest as AKT has a key role in lymphoma cell survival and constitutive activation of AKT has been described in DLBCL. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Novel Role of Raft-Associated Smoothened (SMO) in AKT Signal Regulation in Diffuse Large B Cell Lymphoma

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 3972-3972
Author(s):  
Nitin Agarwal ◽  
Kranthi Kunkalla ◽  
Daniel Bilbao ◽  
Ralf Landgraf ◽  
Francisco Vega
Keyword(s):  
Lipid Rafts ◽  
Lipid Raft ◽  
B Cell Lymphoma ◽  
Signaling Proteins ◽  
Mrna Levels ◽  
Surface Receptors ◽  
Akt Activation ◽  
Hh Signaling ◽  
Signal Regulation

Constitutive PI3K/AKT activation is relevant to multiple aspects of tumor growth and survival in numerous cancers including diffuse large B cell lymphoma (DLBCL). For example, PTEN loss is one of the mechanisms leading to constitutive PI3K/AKT activation in a subset of DLBCL. Smoothened (SMO) is a seven transmembrane spanning and Frizzled-class G-protein coupled receptor that functions as a Hedgehog (Hh) signal transducer. SMO is overexpressed in DLBCL cell lines and tumors. While canonical Hh signaling culminates in the activation of GLI transcription factors and is best understood in the context of cilia, "noncanonical" Hh signaling does not involve GLI transcriptional activity and remains less well characterized. Here, we found that SMO is not only an integral component of lipid rafts but also plays an unexpected central role in the organization of raft microdomains (specialized glycolipid-enriched microdomains known to serve as a highly dynamic signaling platform for cell surface receptors and signaling proteins) and in the sorting of lipid raft-associated proteins. To address whether SMO co-localizes to lipid rafts in the context of DLBCL, HBL1 cells were engineered to stably overexpress a C-terminal SMO-mCherry-fusion protein and were incubated with FITC-Cholera toxin. Within 15 min, a large fraction of SMO-mCherry was co-localized with FITC-Cholera toxin in lipid raft clusters. We also performed immunostaining of endogenous SMO and CD59 in HBL1 cells. CD59 is a glycol-phosphatidyl inositol-anchored lipid raft protein. Immunostaining of live HBL1 cells revealed the colocalization of SMO and CD59 on the extracellular surface of HBL1 cells. No colocalization was found between SMO and the transferrin receptor (TF-R), a plasma membrane protein not associated with lipid rafts. Finally, immunoblotting analysis of detergent-free fractionation further corroborated SMO as a bona fide component of lipid rafts. We then tried to explore the functional relevance of SMO association to the lipid compartment. First, we examined the effects of SMO stable knockdown in DLBCL. We observed a marked decrease in the expression of raft-associated receptors and signaling proteins (e.g. IGFR1, EGFR, IRS1) while caveolin and flotillin, two functional components of lipid rafts, remained unaltered in their levels and distribution. Although only a portion of the overall pool of AKT and pAKT were localized to lipid rafts, SMO loss significantly reduced raft-localized total AKT and pAKT (T308/S473). Consistent with the well-established role of AKT in cell survival, SMO silencing also resulted in reduced DLBCL cell viability. To evaluate whether SMO regulated IGF1R expression at the transcriptional level, we analyzed the mRNA levels of GLI1, the immediate transcriptional target of canonical SMO signaling, and IGF1R in SMO-/- MEFs. Even if GLI1 transcript levels were reduced, consistent with the established mode of GLI1 regulation by SMO through the GLI2 transcription factor, IGF1R mRNA levels remained unchanged. Using GLI1-/- MEFs, we could confirm that the effect of SMO on surface receptors was independent of canonical Hh signaling. We also determined that the rate of IGFR degradation was comparable in the presence and absence of SMO. Total IGF1R receptor levels at steady state represent the balance of total protein synthesis and the fraction of existing receptors that is directed towards lysosomal degradation. To test whether the absence of SMO results in increased degradation, we inhibited lysosomal function with chloroquine (CHLQ). CHLQ had a far more pronounced impact on restoring IGF1R protein levels in SMO deficient cells than it did in control cells. These last results suggested that in the absence of SMO, a larger fraction of this receptor is directed towards lysosomal degradation, and thus resulting in lower steady state levels. In summary, our data confirm that SMO is localized to raft microdomains in lymphoma cells and play a novel role in the sorting of surface proteins for degradation or recycling. In particular, SMO increases the levels of raft resident receptors and facilitates the assembly of an AKT activating machinery to enhance lymphoma cell survival. This novel role of SMO in signal regulation at the level of lipid rafts has broad implications for cancer biology. Disclosures Vega: National Cancer Institute, national Institutes of Health: Other: Grant Funding-R01CA222918.

scholarly journals TRIM27 interacts with Iκbα to promote the growth of human renal cancer cells through regulating the NF-κB pathway

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Chengwu Xiao ◽  
Wei Zhang ◽  
Meimian Hua ◽  
Huan Chen ◽  
Bin Yang ◽  
...  
Keyword(s):  
Cell Lines ◽  
Prognostic Indicator ◽  
The Cancer Genome Atlas ◽  
Mrna Levels ◽  
Loss Of Function ◽  
Protein Levels ◽  
Kaplan Meier ◽  
Cancer Genome Atlas

Abstract Background The tripartite motif (TRIM) family proteins exhibit oncogenic roles in various cancers. The roles of TRIM27, a member of the TRIM super family, in renal cell carcinoma (RCC) remained unexplored. In the current study, we aimed to investigate the clinical impact and roles of TRIM27 in the development of RCC. Methods The mRNA levels of TRIM27 and Kaplan–Meier survival of RCC were analyzed from The Cancer Genome Atlas database. Real-time PCR and Western blotting were used to measure the mRNA and protein levels of TRIM27 both in vivo and in vitro. siRNA and TRIM27 were exogenously overexpressed in RCC cell lines to manipulate TRIM27 expression. Results We discovered that TRIM27 was elevated in RCC patients, and the expression of TRIM27 was closely correlated with poor prognosis. The loss of function and gain of function results illustrated that TRIM27 promotes cell proliferation and inhibits apoptosis in RCC cell lines. Furthermore, TRIM27 expression was positively associated with NF-κB expression in patients with RCC. Blocking the activity of NF-κB attenuated the TRIM27-mediated enhancement of proliferation and inhibition of apoptosis. TRIM27 directly interacted with Iκbα, an inhibitor of NF-κB, to promote its ubiquitination, and the inhibitory effects of TRIM27 on Iκbα led to NF-κB activation. Conclusions Our results suggest that TRIM27 exhibits an oncogenic role in RCC by regulating NF-κB signaling. TRIM27 serves as a specific prognostic indicator for RCC, and strategies targeting the suppression of TRIM27 function may shed light on future therapeutic approaches.

scholarly journals Topoisomerase II is regulated by translationally controlled tumor protein for cell survival during organ growth in Drosophila

2021 ◽  
Vol 12 (9) ◽  
Author(s):  
Dae-Wook Yang ◽  
Jung-Wan Mok ◽  
Stephanie B. Telerman ◽  
Robert Amson ◽  
Adam Telerman ◽  
...  
Keyword(s):  
Cell Survival ◽  
Topoisomerase Ii ◽  
Wing Disc ◽  
Organ Development ◽  
Translationally Controlled Tumor Protein ◽  
Protein Levels ◽  
Eye Disc ◽  
Mutual Regulation

AbstractRegulation of cell survival is critical for organ development. Translationally controlled tumor protein (TCTP) is a conserved protein family implicated in the control of cell survival during normal development and tumorigenesis. Previously, we have identified a human Topoisomerase II (TOP2) as a TCTP partner, but its role in vivo has been unknown. To determine the significance of this interaction, we examined their roles in developing Drosophila organs. Top2 RNAi in the wing disc leads to tissue reduction and caspase activation, indicating the essential role of Top2 for cell survival. Top2 RNAi in the eye disc also causes loss of eye and head tissues. Tctp RNAi enhances the phenotypes of Top2 RNAi. The depletion of Tctp reduces Top2 levels in the wing disc and vice versa. Wing size is reduced by Top2 overexpression, implying that proper regulation of Top2 level is important for normal organ development. The wing phenotype of Tctp RNAi is partially suppressed by Top2 overexpression. This study suggests that mutual regulation of Tctp and Top2 protein levels is critical for cell survival during organ development.

scholarly journals Unlike its Paralog LEDGF/p75, HRP-2 Is Dispensable for MLL-R Leukemogenesis but Important for Leukemic Cell Survival

Cells ◽  
2021 ◽  
Vol 10 (1) ◽  
pp. 192
Author(s):  
Siska Van Belle ◽  
Sara El Ashkar ◽  
Kateřina Čermáková ◽  
Filip Matthijssens ◽  
Steven Goossens ◽  
...  
Keyword(s):  
Growth Factor ◽  
Cell Survival ◽  
Cell Lines ◽  
Protein Interactions ◽  
Leukemic Cell ◽  
Related Protein ◽  
Histone Tails ◽  
Knockout Mouse Model ◽  
Leukemic Cell Lines

HDGF-related protein 2 (HRP-2) is a member of the Hepatoma-Derived Growth Factor-related protein family that harbors the structured PWWP and Integrase Binding Domain, known to associate with methylated histone tails or cellular and viral proteins, respectively. Interestingly, HRP-2 is a paralog of Lens Epithelium Derived Growth Factor p75 (LEDGF/p75), which is essential for MLL-rearranged (MLL-r) leukemia but dispensable for hematopoiesis. Sequel to these findings, we investigated the role of HRP-2 in hematopoiesis and MLL-r leukemia. Protein interactions were investigated by co-immunoprecipitation and validated using recombinant proteins in NMR. A systemic knockout mouse model was used to study normal hematopoiesis and MLL-ENL transformation upon the different HRP-2 genotypes. The role of HRP-2 in MLL-r and other leukemic, human cell lines was evaluated by lentiviral-mediated miRNA targeting HRP-2. We demonstrate that MLL and HRP-2 interact through a conserved interface, although this interaction proved less dependent on menin than the MLL-LEDGF/p75 interaction. The systemic HRP-2 knockout mice only revealed an increase in neutrophils in the peripheral blood, whereas the depletion of HRP-2 in leukemic cell lines and transformed primary murine cells resulted in reduced colony formation independently of MLL-rearrangements. In contrast, primary murine HRP-2 knockout cells were efficiently transformed by the MLL-ENL fusion, indicating that HRP-2, unlike LEDGF/p75, is dispensable for the transformation of MLL-ENL leukemogenesis but important for leukemic cell survival.

scholarly journals IL6 sensitizes prostate cancer to the antiproliferative effect of IFNα2 through IRF9

2013 ◽  
Vol 20 (5) ◽  
pp. 677-689 ◽  
Author(s):  
Holger H H Erb ◽  
Regina V Langlechner ◽  
Patrizia L Moser ◽  
Florian Handle ◽  
Tineke Casneuf ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Cell Lines ◽  
Cellular Proliferation ◽  
Tissue Expression ◽  
Type I ◽  
Regulatory Factor ◽  
Quantitative Real Time Pcr ◽  
Antiproliferative Effects ◽  
Protein Levels

Development and progression of prostate cancer (PCa) are associated with chronic inflammation. The cytokine interleukin 6 (IL6) can influence progression, differentiation, survival, and angiogenesis of PCa. To identify novel pathways that are triggered by IL6, we performed a gene expression profiling of two PCa cell lines, LNCaP and MDA PCa 2b, treated with 5 ng/ml IL6. Interferon (IFN) regulatory factor 9 (IRF9) was identified as one of the most prevalent IL6-regulated genes in both cell lines. IRF9 is a mediator of type I IFN signaling and acts together with STAT1 and 2 to activate transcription of IFN-responsive genes. The IL6 regulation of IRF9 was confirmed at mRNA and protein levels by quantitative real-time PCR and western blot respectively in both cell lines and could be blocked by the anti-IL6 antibody Siltuximab. Three PCa cell lines, PC3, Du-145, and LNCaP-IL6+, with an autocrine IL6 loop displayed high expression of IRF9. A tissue microarray with 36 PCa tissues showed that IRF9 protein expression is moderately elevated in malignant areas and positively correlates with the tissue expression of IL6. Downregulation and overexpression of IRF9 provided evidence for an IFN-independent role of IRF9 in cellular proliferation of different PCa cell lines. Furthermore, expression of IRF9 was essential to mediate the antiproliferative effects of IFNα2. We concluded that IL6 is an inducer of IRF9 expression in PCa and a sensitizer for the antiproliferative effects of IFNα2.

scholarly journals The Role of the FOXO1/β2-AR/p-NF-κB p65 Pathway in the Development of Endometrial Stromal Cells in Pregnant Mice under Restraint Stress

2021 ◽  
Vol 22 (3) ◽  
pp. 1478
Author(s):  
Jiayin Lu ◽  
Yaoxing Chen ◽  
Zixu Wang ◽  
Jing Cao ◽  
Yulan Dong
Keyword(s):  
Oxidative Stress ◽  
Stromal Cells ◽  
Restraint Stress ◽  
Target Genes ◽  
Mrna Levels ◽  
Endometrial Stromal Cells ◽  
Protein Levels ◽  
Pregnant Mice

Restraint stress causes various maternal diseases during pregnancy. β2-Adrenergic receptor (β2-AR) and Forkhead transcription factor class O 1 (FOXO1) are critical factors not only in stress, but also in reproduction. However, the role of FOXO1 in restraint stress, causing changes in the β2-AR pathway in pregnant mice, has been unclear. The aim of this research was to investigate the β2-AR pathway of restraint stress and its impact on the oxidative stress of the maternal uterus. In the study, maternal mice were treated with restraint stress by being restrained in a transparent and ventilated device before sacrifice on Pregnancy Day 5 (P5), Pregnancy Day 10 (P10), Pregnancy Day 15 (P15), and Pregnancy Day 20 (P20) as well as on Non-Pregnancy Day 5 (NP5). Restraint stress augmented blood corticosterone (CORT), norepinephrine (NE), and blood glucose levels, while oestradiol (E2) levels decreased. Moreover, restraint stress increased the mRNA levels of the FOXO family, β2-AR, and even the protein levels of FOXO1 and β2-AR in the uterus and ovaries. Furthermore, restraint stress increased uterine oxidative stress level. In vitro, the protein levels of FOXO1 were also obviously increased when β2-AR was activated in endometrial stromal cells (ESCs). In addition, phosphorylated-nuclear factor kappa-B p65 (p-NF-κB p65) and its target genes decreased significantly when FOXO1 was inhibited. Overall, it can be said that the β2-AR/FOXO1/p-NF-κB p65 pathway was activated when pregnant mice were under restraint stress. This study provides a scientific basis for the origin of psychological stress in pregnant women.

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