Prenatal interleukin 6 elevation increases glutamatergic synapse density and disrupts hippocampal connectivity in offspring.

Early prenatal inflammatory conditions are thought to be a risk factor for different neurodevelopmental disorders. Maternal interleukin-6 (IL-6) elevation during pregnancy causes abnormal behavior in offspring, but whether these defects result from altered synaptic developmental trajectories remains unclear. Here we showed that transient IL-6 elevation via injection into pregnant mice or developing embryos enhanced glutamatergic synapses and led to overall brain hyperconnectivity in offspring into adulthood. IL-6 activated synaptogenesis gene programs in glutamatergic neurons and required the transcription factor STAT3 and expression of the RGS4 gene. The STAT3-RGS4 pathway was also activated in neonatal brains during poly(I:C)-induced maternal immune activation, which mimics viral infection during pregnancy. These findings indicate that IL-6 elevation at early developmental stages is sufficient to exert a long-lasting effect on glutamatergic synaptogenesis and brain connectivity, providing a mechanistic framework for the association between prenatal inflammatory events and brain neurodevelopmental disorders.

MGST3 regulates BACE1 protein translation and amyloidogenesis by controlling the RGS4-mediated AKT signaling pathway.

Microsomal glutathione transferase 3 (MGST3) regulates eicosanoid and glutathione metabolism. These processes are associated with oxidative stress and apoptosis, suggesting that MGST3 might play a role in the pathophysiology of Alzheimer's disease. Here, we report that knockdown (KD) of MGST3 in cell lines reduced the protein level of beta-site amyloid precursor protein cleaving enzyme 1 (BACE1) and the resulting amyloidogenesis. Interestingly, MGST3 KD did not alter intracellular reactive oxygen species level but selectively reduced the expression of apoptosis indicators which could be associated with the receptor of cysteinyl leukotrienes, the downstream metabolites of MGST3 in arachidonic acid pathway. We then showed that the effect of MGST3 on BACE1 was independent of cysteinyl leukotrienes but involved a translational mechanism. Further RNA-seq analysis identified that regulator of G-protein signaling 4 (RGS4) was a target gene of MGST3. Silencing of RGS4 inhibited BACE1 translation and prevented MGST3 KD-mediated reduction of BACE1. The potential mechanism was related to AKT activity, as the protein level of phosphorylated AKT was significantly reduced by silencing of MGST3 and RGS4, and the AKT inhibitor abolished the effect of MGST3/RGS4 on phosphorylated AKT and BACE1. Together, MGST3 regulated amyloidogenesis by controlling BACE1 protein expression, which was mediated by RGS4 and downstream AKT signaling pathway.

Ophiopogonin D' inhibited tumour growth and metastasis of anaplastic thyroid cancer by modulating JUN/RGS4 signalling.

Anaplastic thyroid cancer (ATC), an aggressive malignancy with virtually 100% disease-specific mortality, has long posed a formidable challenge in oncology due to its resistance to conventional treatments and the severe side effects associated with current regimens such as doxorubicin chemotherapy. Consequently, there was urgent need to identify novel candidate compounds that could provide innovative therapeutic strategies for ATC. Ophiopogonin D' (OPD'), a triterpenoid saponin extracted, yet its roles in ATC has not been reported. Our data demonstrated that OPD' potently inhibited proliferation and metastasis of ATC cells, promoting cell cycle arrest and apoptosis. Remarkably, OPD' impeded growth and metastasis of ATC in vitro and in vivo, displaying an encouraging safety profile. Regulator of G-protein signalling 4 (RGS4) expression was significantly up-regulated in ATC compared to normal tissues, and this upregulation was suppressed by OPD' treatment. Mechanistically, we elucidated that the transcription factor JUN bound to the RGS4 promoter, driving its transactivation. However, OPD' interacted with JUN, attenuating its transcriptional activity and thereby disrupting RGS4 overexpression. In summary, our research revealed that OPD' bound with JUN, which in turn resulted in the suppression of transcriptional activation of RGS4, thereby eliciting cell cycle arrest and apoptosis in ATC cells. These findings could offer promise in the development of high-quality candidate compounds for treatment in ATC.

RGS4 controls secretion of von Willebrand factor to the subendothelial matrix.

The haemostatic protein von Willebrand factor (VWF) exists in plasma and subendothelial pools. The plasma pools are secreted from endothelial storage granules, Weibel-Palade bodies (WPBs), by basal secretion with a contribution from agonist-stimulated secretion, and the subendothelial pool is secreted into the subendothelial matrix by a constitutive pathway not involving WPBs. We set out to determine whether the constitutive release of subendothelial VWF is actually regulated and, if so, what functional consequences this might have. Constitutive VWF secretion can be increased by a range of factors, including changes in VWF expression, levels of TNF and other environmental cues. An RNA-seq analysis revealed that expression of regulator of G protein signalling 4 (RGS4) was reduced in endothelial cells (HUVECs) grown under these conditions. siRNA RGS4 treatment of HUVECs increased constitutive basolateral secretion of VWF, probably by affecting the anterograde secretory pathway. In a simple model of endothelial damage, we show that RGS4-silenced cells increased platelet recruitment onto the subendothelial matrix under flow. These results show that changes in RGS4 expression alter levels of subendothelial VWF, affecting platelet recruitment. This introduces a novel control over VWF function.

RGS4 controls Gαi3-mediated regulation of Bcl-2 phosphorylation on TGN38-containing intracellular membranes.

Intracellular pools of the heterotrimeric G-protein α-subunit Gαi3 (encoded by GNAI3) have been shown to promote growth factor signaling, while at the same time inhibiting the activation of JNK and autophagic signaling following nutrient starvation. The precise molecular mechanisms linking Gαi3 to both stress and growth factor signaling remain poorly understood. Importantly, JNK-mediated phosphorylation of Bcl-2 was previously found to activate autophagic signaling following nutrient deprivation. Our data shows that activated Gαi3 decreases Bcl-2 phosphorylation, whereas inhibitors of Gαi3, such as RGS4 and AGS3 (also known as GPSM1), markedly increase the levels of phosphorylated Bcl-2. Manipulation of the palmitoylation status and intracellular localization of RGS4 suggests that Gαi3 modulates phosphorylated Bcl-2 levels and autophagic signaling from discreet TGN38 (also known as TGOLN2)-labeled vesicle pools. Consistent with an important role for these molecules in normal tissue responses to nutrient deprivation, increased Gαi signaling within nutrient-starved adrenal glands from RGS4-knockout mice resulted in a dramatic abrogation of autophagic flux, compared to wild-type tissues. Together, these data suggest that the activity of Gαi3 and RGS4 from discreet TGN38-labeled vesicle pools are critical regulators of autophagic signaling that act via their ability to modulate phosphorylation of Bcl-2.

Identification of a five-gene signature of the RGS gene family with prognostic value in ovarian cancer.

The RGS (regulator of G protein signaling) gene family, which includes negative regulators of G protein-coupled receptors, comprises important drug targets for malignant tumors. It is thus of great significance to explore the value of RGS family genes for diagnostic and prognostic prediction in ovarian cancer. The RNA-seq, immunophenotype, and stem cell index data of pan-cancer, The Cancer Genome Atlas (TCGA) data, and GTEx data of ovarian cancer were downloaded from the UCSC Xena database. In the pan-cancer database, the expression level of RGS1, RGS18, RGS19, and RGS13 was positively correlated with stromal and immune cell scores. Cancer patients with high RGS18 expression were more sensitive to cyclophosphamide and nelarabine, whereas those with high RGS19 expression were more sensitive to cladribine and nelarabine. The relationship between RGS family gene expression and overall survival (OS) and progression-free survival (PFS) of ovarian cancer patients was analyzed using the KM-plotter database, RGS17, RGS16, RGS1, and RGS8 could be used as diagnostic biomarkers of the immune subtype of ovarian cancer, and RGS10 and RGS16 could be used as biomarkers to predict the clinical stage of this disease. Further, Lasso cox analysis identified a five-gene risk score (RGS11, RGS10, RGS13, RGS4, and RGS3). Multivariate COX analysis showed that the risk score was an independent prognostic factor for patients with ovarian cancer. Immunohistochemistry and the HPA protein database confirmed that the five-gene signature is overexpressed in ovarian cancer. GSEA showed that it is mainly involved in the ECM-receptor interaction, TGF-beta signaling pathway, Wnt signaling pathway, and chemokine signaling pathway, which promote the occurrence and development of ovarian cancer. The prediction model of ovarian cancer constructed using RGS family genes is of great significance for clinical decision making and the personalized treatment of patients with ovarian cancer.

Polyphenon E Effects on Gene Expression in PC-3 Prostate Cancer Cells.

Polyphenon E (Poly E) is a standardized, caffeine-free green tea extract with defined polyphenol content. Poly E is reported to confer chemoprotective activity against prostate cancer (PCa) progression in the TRAMP model of human PCa, and has shown limited activity against human PCa in human trials. The molecular mechanisms of the observed Poly E chemopreventive activity against PCa are not fully understood. We hypothesized that Poly E treatment of PCa cells induces gene expression changes, which could underpin the molecular mechanisms of the limited Poly E chemoprevention activity against PCa. PC-3 cells were cultured in complete growth media supplemented with varied Poly E concentrations for 24 h, then RNA was isolated for comparative DNA microarray (0 vs. 200 mg/L Poly E) and subsequent TaqMan qRT-PCR analyses. Microarray data for 54,613 genes were filtered for >2-fold expression level changes, with 8319 genes increased and 6176 genes decreased. Eight genes involved in key signaling or regulatory pathways were selected for qRT-PCR. Two genes increased expression significantly, MXD1 (13.98-fold; p = 0.0003) and RGS4 (21.98-fold; p = 0.0011), by qRT-PCR. MXD1 and RGS4 significantly increased gene expression in Poly E-treated PC-3 cells, and the MXD1 gene expression increases were Poly E dose-dependent.

Regulator of G-Protein Signaling-4 Attenuates Cardiac Adverse Remodeling and Neuronal Norepinephrine Release-Promoting Free Fatty Acid Receptor FFAR3 Signaling.

Propionic acid is a cell nutrient but also a stimulus for cellular signaling. Free fatty acid receptor (FFAR)-3, also known as GPR41, is a Gi/o protein-coupled receptor (GPCR) that mediates some of the propionate's actions in cells, such as inflammation, fibrosis, and increased firing/norepinephrine release from peripheral sympathetic neurons. The regulator of G-protein Signaling (RGS)-4 inactivates (terminates) both Gi/o- and Gq-protein signaling and, in the heart, protects against atrial fibrillation via calcium signaling attenuation. RGS4 activity is stimulated by ß-adrenergic receptors (ARs) via protein kinase A (PKA)-dependent phosphorylation. Herein, we examined whether RGS4 modulates cardiac FFAR3 signaling/function. We report that RGS4 is essential for dampening of FFAR3 signaling in H9c2 cardiomyocytes, since siRNA-mediated RGS4 depletion significantly enhanced propionate-dependent cAMP lowering, Gi/o activation, p38 MAPK activation, pro-inflammatory interleukin (IL)-1ß and IL-6 production, and pro-fibrotic transforming growth factor (TGF)-ß synthesis. Additionally, catecholamine pretreatment blocked propionic acid/FFAR3 signaling via PKA-dependent activation of RGS4 in H9c2 cardiomyocytes. Finally, RGS4 opposes FFAR3-dependent norepinephrine release from sympathetic-like neurons (differentiated Neuro-2a cells) co-cultured with H9c2 cardiomyocytes, thereby preserving the functional ßAR number of the cardiomyocytes. In conclusion, RGS4 appears essential for propionate/FFAR3 signaling attenuation in both cardiomyocytes and sympathetic neurons, leading to cardioprotection against inflammation/adverse remodeling and to sympatholysis, respectively.

The association between RGS4 and choline in cardiac fibrosis.

BACKGROUND: Myocardial fibrosis is caused by the adverse and powerful remodeling of the heart secondary to the death of cardiomyocytes after myocardial infarction. Regulators of G protein Signaling (RGS) 4 is involved in cardiac diseases through regulating G protein-coupled receptors (GPCRs). METHODS: Cardiac fibrosis models were established through cardiac fibroblasts (CFs) treatment with transforming growth factor (TGF)-ß1 in vitro and mice subjected to myocardial infarction in vivo. The mRNA expression of RGS4, collagen I/III and α-SMA detected by qRT-PCR. Protein level of RGS4, collagen I, CTGF and α-SMA detected by Western blot. The ejection fraction (EF%) and fractional shortening (FS%) of mice were measured by echocardiography. Collagen deposition of mice was tested by Masson staining. RESULTS: The expression of RGS4 increased in CFs treatment with TGF-ß1 and in MI mice. The model of cardiac fibrosis detected by qRT-PCR and Western blot. It was demonstrated that inhibition of RGS4 expression improved cardiac fibrosis by transfection with small interfering RNA in CFs and injection with lentivirus shRNA in mice. The protective effect of choline against cardiac fibrosis was counteracted by overexpression of RGS4 in vitro and in vivo. Moreover, choline inhibited the protein level of TGF-ß1, p-Smad2/3, p-p38 and p-ERK1/2 in CFs treated with TGF-ß1, which were restored by RGS4 overexpression. CONCLUSION: This study demonstrated that RGS4 promoted cardiac fibrosis and attenuated the anti-cardiac fibrosis of choline. RGS4 may weaken anti-cardiac fibrosis of choline through TGF-ß1/Smad and MAPK signaling pathways. Video Abstract: Video Byte of this article.

Synergistic regulation of Rgs4 mRNA by HuR and miR-26/RISC in neurons.

The negative regulator of G-protein signalling 4 (Rgs4) is linked to several neurologic diseases, e.g. schizophrenia, addiction, seizure and pain perception. Consequently, Rgs4 expression is tightly regulated, resulting in high mRNA and protein turnover. The post-transcriptional control of gene expression is mediated via RNA-binding proteins (RBPs) that interact with mRNAs in a combinatorial fashion. Here, we show that in neurons the RBP HuR reduces endogenous Rgs4 expression by destabilizing Rgs4 mRNA. Interestingly, in smooth muscle cells, Rgs4 is stabilized by HuR, indicating tissue-dependent differences in HuR function. Using in vitro RNA-based pulldown experiments, we identify the functional AU-rich element (ARE) within the Rgs4 3'-UTR that is recognized and bound by HuR. Bioinformatic analysis uncovered that this ARE lies within a highly conserved area next to a miR-26 binding site. We find that the neuronal-enriched miR-26 negatively influences Rgs4 expression in neurons. Further, HuR and miR-26 act synergistically in fluorescent reporter assays. Together, our data suggest a regulatory mechanism, in which an RBP selectively destabilizes a target mRNA in cooperation with a miRNA and the RISC machinery.

TRPM2-AS inhibits the growth, migration, and invasion of gliomas through JNK, c-Jun, and RGS4.

Gliomas are a group of brain cancers with high mortality and morbidity. Understanding the molecular mechanisms is important for the prevention or treatment of gliomas. The present study was to investigate the effects and mechanisms of long noncoding RNA TRPM2-AS in gliomas proliferation, migration, and invasion. We first compared the levels of TRPM2-AS in 111 patients with glioma to that of the normal control group by a quantitative polymerase chain reaction. The results indicated a significant increase of TRPM2-AS in patients with glioma (2.43 folds of control, p = .0135). MTT methods, wound healing assays, transwell analysis, and clone formation analysis indicated the overexpression of TRPM2-AS promoted the proliferation, migration, and invasion of U251 and U87 cells, while downregulation of TRPM2-AS inhibited the cell proliferation, migration, and invasion significantly (p < .05). To further uncover the mechanisms, bioinformatics analysis was conducted on the expression profiles, GSE40687 and GSE4290, from the Gene Expression Omnibus database. One hundred fifty-six genes were differentially expressed in both datasets (FC > 2.0; p = .05). Among these differentially expressed genes, the level of RGS4 messenger RNA was drastically regulated by TRPM2-AS. Further western-blot analysis indicated the increase of RGS4 protein expression and decrease of p-JNK/JNK and p-c-Jun/c-Jun ratio after TRPM2-AS overexpression. On the other hand, inhibition of TRPM2-AS by small interfering RNA suppressed the expression of RGS4 and promoted the ratios of p-JNK/JNK and p-c-Jun/c-Jun. The present work indicated the mechanisms of the participation of TRPM2-AS in the progression of gliomas might, at least partly, be related to JNK, c-Jun, and RGS4. Our work provided new insights into the underlying mechanisms of glioma cellular functions.

lncRNA RPL34-AS1 inhibits cell proliferation and invasion while promoting apoptosis by competitively binding miR-3663-3p/RGS4 in papillary thyroid cancer.

Papillary thyroid cancer (PTC) accounts for 80% of all thyroid cancers and seriously impacts the quality of people's lives. Long noncoding RNAs (lncRNAs) play an important role in PTC. In previous studies, thousands of lncRNAs were screened to study their potential relationships with PTC. The aim of this study was to investigate the effect of RPL34-AS1 in PTC and to explore its potential mechanisms. Bioinformatic analyses were performed to characterize the possible function and biological features of RPL34-AS1. Apoptosis, proliferation, and invasion were detected to assess the effect of RPL34-AS1. Cell proliferation was measured using a Cell Counting Kit-8 assay. Western blot analysis was used to assess the apoptosis proteins Bax and Bcl-2. Cell invasion was measured using a Transwell assay. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis was performed to examine RPL34-AS1, miR-3663-3P, and RGS4 expression. Dual-luciferase assay was performed to assess the binding of miR-3663-3P by RPL34-AS1. RIP experiment was used to verify the combination between miR-3663-3p and RGS4. We found that overexpression of RPL34-AS1 could inhibit proliferation and invasion while promoting apoptosis in PTC cell lines. Moreover, RPL34-AS1 could also competitively bind miR-3663-3p and exert its function by regulating the miR-3663-3p/RGS4 in PTC cell lines. We found a previously uncharacterized lncRNA, RPL34-AS1, and studied its function and mechanism in PTC. Our research will provide new insights into PTC and new clues for its clinical treatment.

miR-874-3p inhibits cell migration through targeting RGS4 in osteosarcoma.

BACKGROUND: The present study explored the role and mechanism of microRNA-874-3p (miR-874-3p) in the migration of the osteosarcoma cell line, U-2 OS. METHODS: The expression profile of osteosarcoma (OS) microRNA (GSE65071) datasets was downloaded from the Gene Expression Omnibus (GEO) database (https://www.ncbi.nlm.nih.gov/geo) to identify differentially expressed miRNAs in OS and its biological functions. A quantitative reverse transcription-polymerase chain reaction was performed to detect the expression of miR-874-3p and its target gene regulator of G protein 4 (RGS4) in human osteosarcoma cells U-2 OS and normal osteoblast hFOB1.19. Plasmid overexpression miR-874-3p and pcDNA-RGS4 were transfected into U-2 OS using Lipofectamine 2000 (Thermo Fisher, Waltham, MA, USA). Cell migration was measured using Transwell migration assays. Bioinformatic analysis and luciferase reporter assay were conducted to search for the target gene of miR-874-3p. RESULTS: In total, 167 differentially expressed miRNAs were detected after the analysis of GSE65071; of which 78 were up-regulated genes and 89 were down-regulated. miR-874-3p was down-regulated and selected for further analysis. The expression level of miR-874-3p in U-2 OS cells was significantly decreased compared to the hFOB1.19 cell line (p < 0.05). Overexpression of miR-874-3p significantly inhibited the proliferation and migration of U-2 OS cells and overexpression of RGS4 reversed the inhibitory effect of miR-874-3p on U-2 OS cells. Through luciferase report analyses and bioinformatic analysis, RGS4 may be the candidate target gene of miR-874-3p. CONCLUSIONS: In conclusion, overexpression of miR-874-3p suppressed OS cell proliferation and migration. Thus, miR-874-3p might present a therapeutic agent for the treatment of OS.

Targeting RGS4 Ablates Glioblastoma Proliferation.

Glioblastoma (GBM) is the most common type of adult primary brain tumor with a median survival rate of less than 15 months, regardless of the current standard of care. Cellular heterogeneity, self-renewal ability and tumorigenic glioma cancer stem cell (GSC) populations contribute to the difficulty in treating GBM. G-protein-coupled receptors (GPCRs) are the largest group of membrane proteins and mediate many cellular responses. Regulators of G-protein signaling 4 (RGS4) are negative regulators of G-protein signaling, and elevated levels of RGS4 are reportedly linked with several human diseases, including cancer. This study investigates the effect of silencing RGS4, resulting in inhibition of GSC growth, invasion and migration. Data obtained from The Cancer Genome Atlas (TCGA) demonstrated poor patient survival with high expression of RGS4. Immunohistochemistry and immunoblot analysis conducted on GBM patient biopsy specimens demonstrated increased RGS4 expression correlative with the TCGA data. RNA sequencing confirmed a significant decrease in the expression of markers involved in GSC invasion and migration, particularly matrix metalloproteinase-2 (MMP2) in knockout of RGS4 using CRISPR plasmid (ko-RGS4)-treated samples compared to parental controls. Gelatin zymography confirmed the reduced activity of MMP2 in ko-RGS4-treated samples. Silencing RGS4 further reduced the invasive and migratory abilities and induction of apoptosis of GSCs as evidenced by Matrigel plug assay, wound healing assay and human apoptosis array. Collectively, our results showed that the silencing of RGS4 plays an important role in regulating multiple cellular functions, and is an important therapeutic target in GBM.

MiR-21-3p modulates lipopolysaccharide-induced inflammation and apoptosis via targeting TGS4 in retinal pigment epithelial cells.

Age-related macular degeneration (AMD) is a major reason of blindness in the elderly. MicroRNAs are implicated in various pathological processes, including inflammation and apoptosis. In this study, we aim to investigate the biological functions of miR-21-3p in inflammation and apoptosis caused by lipopolysaccharide (LPS) in human retinal pigment epithelial (ARPE-19) cells. The miR-21-3p inhibitor and mimic were transfected into ARPE-19 cells for 48 hours, followed by exposed to LPS (10 µg/mL) for 24 hours. The mRNA and protein expression of IL-6 and MCP-1 were measured using real-time PCR (RT-PCR) and enzyme-linked immunosorbent assays. Cell viability, apoptosis, caspase 3 activity, cleaved caspase-3 and cleaved-PARP protein levels were detected to evaluate the effects of miR-21-3p on apoptosis. Additionally, the target relationship between miR-21-3p and regulator of G-protein signalling 4 (RGS4) was verified by dual luciferase reporter assay. RT-PCR analysis demonstrated that LPS induced miR-21-3p expression. Inhibition of miR-21-3p reduced the mRNA and protein levels of IL-6 and MCP-1. Apoptosis, caspase-3 activity, and cleaved-caspase 3 and cleaved PARP protein levels were repressed by the miR-21-3p inhibitor. However, overexpression of miR-21-3p showed the opposite results. Furthermore, we identified that miR-21-3p directly targeted the 3' untranslated region of RGS4. MiR-21-3p negatively regulated the expression of RGS4 both in mRNA and protein levels. Silencing RGS4 reduced the anti-inflammatory and anti-apoptotic effects of miR-21-3p inhibitor. Our results revealed that miR-21-3p inhibition targeted RGS4 to attenuate inflammatory responses and apoptosis caused by LPS in ARPE-19 cells.

A gene-based review of RGS4 as a putative risk gene for psychiatric illness.

Considerable efforts have been made to characterize RGS4 as a potential candidate gene for schizophrenia. Investigations span across numerous modalities and include explorations of genetic risk associations, mRNA and protein levels in the brain, and functionally relevant interactions with other candidate genes as well as links to schizophrenia relevant neural phenotypes. While these lines of investigations have yielded partially inconsistent findings, they provide a perspective on RGS4 as an important part of a larger biological system contributing to schizophrenia risk. This gene-based review aims to provide a comprehensive overview of published data from different experimental modalities and discusses the current knowledge of RGS4's systems-biological impact on the schizophrenia pathology.

Rosiglitazone Inhibits Angiotensin II-Induced Proliferation of Glomerular Mesangial Cells via the Gαq/Plcß4/TRPC Signaling Pathway.

BACKGROUND/AIMS: Mesangial cell proliferation and extracellular matrix accumulation (ECM) deposition play an important role in the pathogenesis of glomerulosclerosis. TRPC and PPAR-γ can regulate cell proliferation. Angiotensin II (AngII) can induce mesangial cell proliferation and affect TRPC expression. However, the mechanism has not been fully elucidated. This study was designed to investigate the role of TRPC and the effect of rosiglitazone (RSG) in the proliferation of rat glomerular mesangial cells (HBZY-1) that were stimulated by AngII and the underlying mechanisms. METHODS: Immunofluorescence staining and qRT-PCR were performed to examine the expression levels of TRPCs in HBZY-1. Gene expression levels of TRPC, PPAR-γ, RGS4 (regulators of G protein signaling), the GPCR/Gαq/PLCß4/TRPC signaling pathway and major downstream molecules (PCNA, SKP2, P21 and P27) were detected by qRT-PCR and western blotting. Additionally, changes in intracellular Ca2+ levels were determined through Fluo-4 Ca2+ imaging, and the cell cycle was analyzed by flow cytometry. RESULTS: Our results found that TRPC1 and 6 were at higher expression levels in HBZY-1 cells. Following AngII stimulation, there were increased levels of TRPC1 and 6, Ca2+ entry, PCNA and SKP2, decreased expression levels of P21 and P27 and a reduced G0/G1 percentage. Silencing TRPC1 and 6 by siRNAs led to decrease in Ca2+ influx, G0/G1 cell cycle arrest and cell proliferation. Notably, PPAR-γ activation by RSG upregulated RGS4 expression, which can interact with the Gαq family to inhibit the Gαq-mediated signaling cascade. The results were similar to silencing TRPC1 and 6 by siRNAs. CONCLUSION: All these results indicate that RSG could inhibit HBZY-1 cell proliferation via the Gαq/PLCß4/TRPC signaling pathway.

Association between RGS4 variants and psychotic-like experiences in nonclinical individuals.

The psychosis phenotype is expressed across a continuum known as schizotypy, which ranges from personality variation through subclinical symptoms to severe psychopathology. The study of subclinical manifestations in non-affected individuals minimizes confounding factors associated with the clinical phenotype and facilitates the differentiation of dimension-specific etiological mechanisms. The aim of the present study was to investigate the association between the variation in the regulator of G-protein signaling 4 (RGS4) gene, a putative candidate gene for psychosis previously associated with schizophrenia endophenotypes, and psychotic-like experiences (PLEs). In total, 808 healthy individuals completed the community assessment of psychic experiences (CAPE) to measure positive and negative PLEs and provided a DNA sample. Two RGS4 single-nucleotide polymorphisms (SNPs) (rs951436 [SNP4] and rs2661319 [SNP18]) were genotyped. Analyses of covariance (ANCOVA) were used to explore the association of positive and negative PLEs with RGS4 variation. Our results showed associations of positive and negative PLEs with the two polymorphisms studied: subjects with the T allele (SNP4) and the A allele (SNP18) had higher scores on both the positive and the negative dimensions. Haplotypic analyses supported these results, showing the highest scores in those with the TA haplotype (SNP4-SNP18). The RGS4 variants might exert gene-specific modulating effects on psychosis proneness.

Expression regulation and functional analysis of RGS2 and RGS4 in adipogenic and osteogenic differentiation of human mesenchymal stem cells.

BACKGROUND: Understanding the molecular basis underlying the formation of bone-forming osteocytes and lipid-storing adipocytes will help provide insights into the cause of disorders originating in stem/progenitor cells and develop therapeutic treatments for bone- or adipose-related diseases. In this study, the role of RGS2 and RGS4, two members of the regulators of G protein signaling (RGS) family, was investigated during adipogenenic and osteogenenic differentiation of human mesenchymal stem cells (hMSCs). RESULTS: Expression of RGS2 and RGS4 were found to be inversely regulated during adipogenesis induced by dexamethasone (DEX) and 3-isobutyl-methylxanthine, regardless if insulin was present, with RGS2 up-regulated and RGS4 down-regulated in response to adipogenic induction. RGS2 expression was also up-regulated during osteogenesis at a level similar to that induced by treatment of DEX alone, a shared component of adipogenic and osteogenic differentiation inducing media, but significantly lower than the level induced by adipogenic inducing media. RGS4 expression was down-regulated during the first 48 h of osteogenesis but up-regulated afterwards, in both cases at levels similar to that induced by DEX alone. Expression knock-down using small interfering RNA against RGS2 resulted in decreased differentiation efficiency during both adipogenesis and osteogenesis. On the other hand, expression knock-down of RGS4 also resulted in decreased adipogenic differentiation but increased osteogenic differentiation. CONCLUSIONS: RGS2 and RGS4 are differentially regulated during adipogenic and osteogenic differentiation of hMSCs. In addition, both RGS2 and RGS4 play positive roles during adipogenesis but opposing roles during osteogenesis, with RGS2 as a positive regulator and RGS4 as a negative regulator. These results imply that members of RGS proteins may play multifaceted roles during human adipogenesis and osteogenesis to balance or counterbalance each other's function during those processes.

Decreased levels of RGS4 in the paraventricular nucleus facilitate GABAergic inhibition during the acute stress response.

A healthy acute stress response requires both rapid increase and rapid clearance of blood corticosteroids. We previously showed that regulators of G-protein signaling 4 (RGS4), which decreases in the paraventricular nucleus (PVN) during acute stress, forms a complex with the GABAB receptor. In the present study, we show that this decrease in RGS4 levels in the PVN during an acute stress response facilitates the return of blood corticosteroids to basal levels. Moreover, the effect of RGS4 decrease is attenuated by a GABAB receptor antagonist. These results suggest that RGS4 in the PVN regulates blood corticosteroid-related GABAB receptor signaling during the acute stress response.