Brain areas interconnected to ventral pathway circuits are independently able to induce enhancement in object recognition memory and cause reversal in object recognition memory deficit.

AIMS: Ventral pathway circuits are constituted by the interconnected brain areas that are distributed throughout the brain. These brain circuits are primarily involved in processing of object related information in brain. However, their role in object recognition memory (ORM) enhancement remains unknown. Here, we have studied on the implication of these circuits in ORM enhancement and in reversal of ORM deficit in aging. METHODS: The brain areas interconnected to ventral pathway circuits in rat brain were activated by an expression of a protein called regulator of G-protein signaling 14 of 414 amino acids (RGS14414). RGS14414 is an ORM enhancer and therefore used here as a gain-in-function tool. ORM test and immunohistochemistry, lesions, neuronal arborization, and knockdown studies were performed to uncover the novel function of ventral pathway circuits. RESULTS: An activation of each of the brain areas interconnected to ventral pathway circuits individually induced enhancement in ORM; however, same treatment in brain areas not interconnected to ventral pathway circuits produced no effect. Further study in perirhinal cortex (PRh), area V2 of visual cortex and frontal cortex (FrC), which are brain areas that have been shown to be involved in ORM and are interconnected to ventral pathway circuits, revealed that ORM enhancement seen after the activation of any one of the three brain areas was unaffected by the lesions in other two brain areas either individually in each area or even concurrently in both areas. This ORM enhancement in all three brain areas was associated to increase in structural plasticity of pyramidal neurons where more than 2-fold higher dendritic spines were observed. Additionally, we found that an activation of either PRh, area V2, or FrC not only was adequate but also was sufficient for the reversal of ORM deficit in aging rats, and the blockade of RGS14414 activity led to loss in increase in dendritic spine density and failure in reversal of ORM deficit. CONCLUSIONS: These results suggest that brain areas interconnected to ventral pathway circuits facilitate ORM enhancement by an increase in synaptic connectivity between the local brain area circuits and the passing by ventral pathway circuits and an upregulation in activity of ventral pathway circuits. In addition, the finding of the reversal of ORM deficit through activation of an interconnected brain area might serve as a platform for developing not only therapy against memory deficits but also strategies for other brain diseases in which neuronal circuits are compromised.

Age-Dependent RGS5 Loss in Pericytes Induces Cardiac Dysfunction and Fibrosis.

BACKGROUND: Pericytes are capillary-associated mural cells involved in the maintenance and stability of the vascular network. Although aging is one of the main risk factors for cardiovascular disease, the consequences of aging on cardiac pericytes are unknown. METHODS: In this study, we have combined single-nucleus RNA sequencing and histological analysis to determine the effects of aging on cardiac pericytes. Furthermore, we have conducted in vivo and in vitro analysis of RGS5 (regulator of G-protein signaling 5) loss of function and finally have performed pericytes-fibroblasts coculture studies to understand the effect of RGS5 deletion in pericytes on the neighboring fibroblasts. RESULTS: Aging reduced the pericyte area and capillary coverage in the murine heart. Single-nucleus RNA sequencing analysis further revealed that the expression of Rgs5 was reduced in cardiac pericytes from aged mice. In vivo and in vitro studies showed that the deletion of RGS5 impaired cardiac function, induced fibrosis, and morphological changes in pericytes characterized by a profibrotic gene expression signature and the expression of different ECM (extracellular matrix) components and growth factors, for example, TGFB2 and PDGFB. Indeed, culturing fibroblasts with the supernatant of RGS5-deficient pericytes induced their activation as evidenced by the increased expression of αSMA (alpha smooth muscle actin) in a TGFß (transforming growth factor beta)2-dependent mechanism. CONCLUSIONS: Our results have identified RGS5 as a crucial regulator of pericyte function during cardiac aging. The deletion of RGS5 causes cardiac dysfunction and induces myocardial fibrosis, one of the hallmarks of cardiac aging.

RGS4 controls airway hyperresponsiveness through GAP-independent mechanisms.

Regulators of G protein signaling (RGS) proteins constrain G protein-coupled receptor (GPCR)-mediated and other responses throughout the body primarily, but not exclusively, through their GTPase-activating protein activity. Asthma is a highly prevalent condition characterized by airway hyper-responsiveness (AHR) to environmental stimuli resulting in part from amplified GPCR-mediated airway smooth muscle contraction. Rgs2 or Rgs5 gene deletion in mice enhances AHR and airway smooth muscle contraction, whereas RGS4 KO mice unexpectedly have decreased AHR because of increased production of the bronchodilator prostaglandin E2 (PGE2) by lung epithelial cells. Here, we found that knockin mice harboring Rgs4 alleles encoding a point mutation (N128A) that sharply curtails RGS4 GTPase-activating protein activity had increased AHR, reduced airway PGE2 levels, and augmented GPCR-induced bronchoconstriction compared with either RGS4 KO mice or WT controls. RGS4 interacted with the p85α subunit of PI3K and inhibited PI3K-dependent PGE2 secretion elicited by transforming growth factor beta in airway epithelial cells. Together, these findings suggest that RGS4 affects asthma severity in part by regulating the airway inflammatory milieu in a G protein-independent manner.

A splice acceptor variant in RGS6 associated with intellectual disability, microcephaly, and cataracts disproportionately promotes expression of a subset of RGS6 isoforms.

Intellectual disability (ID) is associated with an increased risk of developing psychiatric disorders, suggesting a common underlying genetic factor. Importantly, altered signaling and/or expression of regulator of G protein signaling 6 (RGS6) is associated with ID and numerous psychiatric disorders. RGS6 is highly conserved and undergoes complex alternative mRNA splicing producing ~36 protein isoforms with high sequence similarity historically necessitating a global approach in functional studies. However, our recent analysis in mice revealed RGS6 is most highly expressed in CNS with RGS6L(+GGL) isoforms predominating. A previously reported genetic variant in intron 17 of RGS6 (c.1369-1G>C), associated with ID, may provide further clues into RGS6L(+GGL) isoform functional delineation. This variant was predicted to alter a highly conserved canonical 3' acceptor site creating an alternative branch point within exon 18 (included in a subset of RGS6L(+GGL) transcripts) and a frameshift forming an early stop codon. We previously identified this alternative splice site and demonstrated its use generates RGS6Lζ(+GGL) isoforms. Here, we show that the c.1369-1G>C variant disrupts the canonical, preferred (>90%) intron 17 splice site and leads to the exclusive use of the alternate exon 18 splice site, inducing disproportionate expression of a subset of isoforms, particularly RGS6Lζ(+GGL). Furthermore, RGS6 global knockout mice do not exhibit ID. Thus, ID caused by the c.1369-1G>C variant likely results from altered RGS6 isoform expression, rather than RGS6 isoform loss. In summary, these studies highlight the importance of proper RGS6 splicing and identify a previously unrecognized role of G protein signaling in ID.

RGS4 Actions in Mouse Prefrontal Cortex Modulate Behavioral and Transcriptomic Responses to Chronic Stress and Ketamine.

The signal transduction protein, regulator of G protein signaling 4 (RGS4), plays a prominent role in physiologic and pharmacological responses by controlling multiple intracellular pathways. Our earlier work identified the dynamic but distinct roles of RGS4 in the efficacy of monoamine-targeting versus fast-acting antidepressants. Using a modified chronic variable stress (CVS) paradigm in mice, we demonstrate that stress-induced behavioral abnormalities are associated with the downregulation of RGS4 in the medial prefrontal cortex (mPFC). Knockout of RGS4 (RGS4KO) increases susceptibility to CVS, as mutant mice develop behavioral abnormalities as early as 2 weeks after CVS resting-state functional magnetic resonance imaging I (rs-fMRI) experiments indicate that stress susceptibility in RGS4KO mice is associated with changes in connectivity between the mediodorsal thalamus (MD-THL) and the mPFC. Notably, RGS4KO also paradoxically enhances the antidepressant efficacy of ketamine in the CVS paradigm. RNA-sequencing analysis of naive and CVS samples obtained from mPFC reveals that RGS4KO triggers unique gene expression signatures and affects several intracellular pathways associated with human major depressive disorder. Our analysis suggests that ketamine treatment in the RGS4KO group triggers changes in pathways implicated in synaptic activity and responses to stress, including pathways associated with axonal guidance and myelination. Overall, we show that reducing RGS4 activity triggers unique gene expression adaptations that contribute to chronic stress disorders and that RGS4 is a negative modulator of ketamine actions. SIGNIFICANCE STATEMENT: Chronic stress promotes robust maladaptation in the brain, but the exact intracellular pathways contributing to stress vulnerability and mood disorders have not been thoroughly investigated. In this study, the authors used murine models of chronic stress and multiple methodologies to demonstrate the critical role of the signal transduction modulator regulator of G protein signaling 4 in the medial prefrontal cortex in vulnerability to chronic stress and the efficacy of the fast-acting antidepressant ketamine.

RGS6 drives cardiomyocyte death following nucleolar stress by suppressing Nucleolin/miRNA-21.

BACKGROUND: Prior evidence demonstrated that Regulator of G protein Signaling 6 (RGS6) translocates to the nucleolus in response to cytotoxic stress though the functional significance of this phenomenon remains unknown. METHODS: Utilizing in vivo gene manipulations in mice, primary murine cardiac cells, human cell lines and human patient samples we dissect the participation of a RGS6-nucleolin complex in chemotherapy-dependent cardiotoxicity. RESULTS: Here we demonstrate that RGS6 binds to a key nucleolar protein, Nucleolin, and controls its expression and activity in cardiomyocytes. In the human myocyte AC-16 cell line, induced pluripotent stem cell derived cardiomyocytes, primary murine cardiomyocytes, and the intact murine myocardium tuning RGS6 levels via overexpression or knockdown resulted in diametrically opposed impacts on Nucleolin mRNA, protein, and phosphorylation.RGS6 depletion provided marked protection against nucleolar stress-mediated cell death in vitro, and, conversely, RGS6 overexpression suppressed ribosomal RNA production, a key output of the nucleolus, and triggered death of myocytes. Importantly, overexpression of either Nucleolin or Nucleolin effector miRNA-21 counteracted the pro-apoptotic effects of RGS6. In both human and murine heart tissue, exposure to the genotoxic stressor doxorubicin was associated with an increase in the ratio of RGS6/Nucleolin. Preventing RGS6 induction via introduction of RGS6-directed shRNA via intracardiac injection proved cardioprotective in mice and was accompanied by restored Nucleolin/miRNA-21 expression, decreased nucleolar stress, and decreased expression of pro-apoptotic, hypertrophy, and oxidative stress markers in heart. CONCLUSION: Together, these data implicate RGS6 as a driver of nucleolar stress-dependent cell death in cardiomyocytes via its ability to modulate Nucleolin. This work represents the first demonstration of a functional role for an RGS protein in the nucleolus and identifies the RGS6/Nucleolin interaction as a possible new therapeutic target in the prevention of cardiotoxicity.

Epigenetic insights into Familial Mediterranean Fever: Increased RGS10 expression and histone modifications accompanies inflammation in familial Mediterranean fever disease.

BACKGROUND: Familial Mediterranean fever (FMF) is an autosomal recessive autoinflammatory disease characterized by recurring fever, erythema, joint pain, and abdominal discomfort during acute episodes. While FMF patients typically share MEFV gene mutations, they display varying clinical manifestations, suggesting the involvement of modifying genes, epigenetic mechanisms, or environmental factors. G protein regulator signal 10 (RGS10), a member of the RGS protein family, exhibits anti-inflammatory effects in autoinflammatory diseases. There are no studies on the role of plays in FMF pathogenesis or histone modification in FMF. AIMS: This study aimed to shed light on the epigenetic regulation of FMF from several perspectives. The relationship between RGS10 DNA hypermethylation in FMF clinical parameters and the regulation of 22 histone modifications were examined in FMF attack patients and the control group. METHODS: Sixty FMF (remission/attack) and thirty healthy individuals were included in the study. First, RNA was isolated from the blood of patients/controls, and the expression of RGS10 was examined. Then, DNA was isolated from the patients, and gene-specific hypermethylation was investigated using the bisulfite conversion method. Finally, histone extraction was performed for FMF patients and controls and 22 histone H3 modifications were determined. In addition, using ADEX bioinformatics tools, RGS10 expression and methylation profiles were detected in different autoinflammatory diseases. RESULTS: This study indicate that RGS10 expression decreased in attack-free/attack patients than control, attributed to DNA methylation. In addition, there were a positive correlation between FMF patients and attack, WBC, neutrophil, MCHC and MPV. Moreover, higher H3K4 me3, H3K9 me2, and H3K14ac levels were observed in patients with FMF attacks. This research also showed a consistent decrease in RGS10 expression in patients with SjS, SSc, and T1D compared with controls. I also obtained five prognosis-related CpGs (cg17527393, cg19653161, cg20445950, cg18938673 and cg13975098) of RGS10 in patients with SjS, RA, SSc, SLE and T1D. CONCLUSION: The present study provides insights into the complex relationship between RGS10, epigenetic modifications, and immune responses in FMF. While RGS10 may initially enhance immune responses, genetic mutations and epigenetic changes associated with FMF acute episode may override this regulatory effect, resulting in increased inflammation and clinical symptoms. Moreover, our study revealed elevated levels of specific histone modifications in the context of FMF, suggesting significant epigenetic changes that could contribute to the disease pathogenesis. Understanding these associations opens new avenues for research and potential therapeutic interventions, potentially involving epigenetic therapies targeting histone modifications.

Regulator of G protein signalling 18 promotes osteocyte proliferation by activating the extracellular signal­regulated kinase signalling pathway.

Osteocyte function is critical for metabolism, remodelling and regeneration of bone tissue. In the present study, the roles of regulator of G protein signalling 18 (RGS18) were assessed in the regulation of osteocyte proliferation and bone formation. Target genes and signalling pathways were screened using the Gene Expression Omnibus (GEO) database and Gene Set Enrichment Analysis (GSEA). The function of RGS18 and the associated mechanisms were analysed by Cell Counting Kit 8 assay, 5­ethynyl­2'­deoxyuridine assay, flow cytometry, reverse transcription­quantitative PCR, western blotting and immunostaining. Overlap analysis of acutely injured subjects (AIS) and healthy volunteers (HVs) from the GSE93138 and GSE93215 datasets of the GEO database identified four genes: KIAA0825, ANXA3, RGS18 and LIPN. Notably, RGS18 was more highly expressed in peripheral blood samples from AIS than in those from HVs. Furthermore, RGS18 overexpression promoted MLO­Y4 and MC3T3­E1 cell viability, proliferation and S­phase arrest, but inhibited apoptosis by suppressing caspase­3/9 cleavage. Silencing RGS18 exerted the opposite effects. GSEA of GSE93138 revealed that RGS18 has the ability to regulate MAPK signalling. Treatment with the MEK1/2 inhibitor PD98059 reversed the RGS18 overexpression­induced osteocyte proliferation, and treatment with the ERK1/2 activator 12­O­tetradecanoylphorbol­13­acetate reversed the effects of RGS18 silencing on osteocyte proliferation. In conclusion, RGS18 may contribute to osteocyte proliferation and bone fracture healing via activation of ERK signalling.

Phosphorylation Dynamics in a flg22-Induced, G Protein-Dependent Network Reveals the AtRGS1 Phosphatase.

The microbe-associated molecular pattern flg22 is recognized in a flagellin-sensitive 2-dependent manner in root tip cells. Here, we show a rapid and massive change in protein abundance and phosphorylation state of the Arabidopsis root cell proteome in WT and a mutant deficient in heterotrimeric G-protein-coupled signaling. flg22-induced changes fall on proteins comprising a subset of this proteome, the heterotrimeric G protein interactome, and on highly-populated hubs of the immunity network. Approximately 95% of the phosphorylation changes in the heterotrimeric G-protein interactome depend, at least partially, on a functional G protein complex. One member of this interactome is ATBα, a substrate-recognition subunit of a protein phosphatase 2A complex and an interactor to Arabidopsis thaliana Regulator of G Signaling 1 protein (AtRGS1), a flg22-phosphorylated, 7-transmembrane spanning modulator of the nucleotide-binding state of the core G-protein complex. A null mutation of ATBα strongly increases basal endocytosis of AtRGS1. AtRGS1 steady-state protein level is lower in the atbα mutant in a proteasome-dependent manner. We propose that phosphorylation-dependent endocytosis of AtRGS1 is part of the mechanism to degrade AtRGS1, thus sustaining activation of the heterotrimeric G protein complex required for the regulation of system dynamics in innate immunity. The PP2A(ATBα) complex is a critical regulator of this signaling pathway.

RGS proteins and cardiovascular Angiotensin II Signaling: Novel opportunities for therapeutic targeting.

Angiotensin II (AngII), as an octapeptide hormone normally ionized at physiological pH, cannot cross cell membranes and thus, relies on, two (mainly) G protein-coupled receptor (GPCR) types, AT1R and AT2R, to exert its intracellular effects in various organ systems including the cardiovascular one. Although a lot remains to be elucidated about the signaling of the AT2R, AT1R signaling is known to be remarkably versatile, mobilizing a variety of G protein-dependent and independent signal transduction pathways inside cells to produce a biological outcome. Cardiac AT1R signaling leads to hypertrophy, adverse remodeling, fibrosis, while vascular AT1R signaling raises blood pressure via vasoconstriction, but also elicits hypertrophic, vascular growth/proliferation, and pathological remodeling sets of events. In addition, adrenal AT1R is the major physiological stimulus (alongside hyperkalemia) for secretion of aldosterone, a mineralocorticoid hormone that contributes to hypertension, electrolyte abnormalities, and to pathological remodeling of the failing heart. Regulator of G protein Signaling (RGS) proteins, discovered about 25 years ago as GTPase-activating proteins (GAPs) for the Gα subunits of heterotrimeric G proteins, play a central role in silencing G protein signaling from a plethora of GPCRs, including the AngII receptors. Given the importance of AngII and its receptors, but also of several RGS proteins, in cardiovascular homeostasis, the physiological and pathological significance of RGS protein-mediated modulation of cardiovascular AngII signaling comes as no surprise. In the present review, we provide an overview of the current literature on the involvement of RGS proteins in cardiovascular AngII signaling, by discussing their roles in cardiac (cardiomyocyte and cardiofibroblast), vascular (smooth muscle and endothelial cell), and adrenal (medulla and cortex) AngII signaling, separately. Along the way, we also highlight the therapeutic potential of enhancement of, or, in some cases, inhibition of each RGS protein involved in AngII signaling in each one of these cell types.

RGS5 maintaining vascular homeostasis is altered by the tumor microenvironment.

BACKGROUND: Regulator of G protein signaling 5 (RGS5), as a negative regulator of G protein-coupled receptor (GPCR) signaling, is highly expressed in arterial VSMCs and pericytes, which is involved in VSMC phenotypic heterogeneity and vascular remodeling in tumors. However, its role in normal and tumor vascular remodeling is controversial. METHODS: RGS5 knockout (Rgs5-KO) mice and RGS5 overexpression or knockdown in VSMCs in vivo by adeno-associated virus type 9 (AAV) carrying RGS5 cDNA or small hairpin RNA (shRNA) targeting RGS5 were used to determine the functional significance of RGS5 in vascular inflammation. RGS5 expression in the triple-negative (TNBCs) and non-triple-negative breast cancers (Non-TNBCs) was determined by immunofluorescent and immunohistochemical staining. The effect of breast cancer cell-conditioned media (BC-CM) on the pro-inflammatory phenotype of VSMCs was measured by phagocytic activity assays, adhesion assay and Western blot. RESULTS: We identified that knockout and VSMC-specific knockdown of RGS5 exacerbated accumulation and pyroptosis of pro-inflammatory VSMCs, resulting in vascular remodeling, which was negated by VSMC-specific RGS5 overexpression. In contrast, in the context of breast cancer tissues, the role of RGS5 was completely disrupted. RGS5 expression was increased in the triple-negative breast cancer (TNBC) tissues and in the tumor blood vessels, accompanied with an extensive vascular network. VSMCs treated with BC-CM displayed enhanced pro-inflammatory phenotype and higher adherent with macrophages. Furthermore, tumor-derived RGS5 could be transferred into VSMCs. CONCLUSIONS: These findings suggest that tumor microenvironment shifts the function of RGS5 from anti-inflammation to pro-inflammation and induces the pro-inflammatory phenotype of VSMCs that is favorable for tumor metastasis.

RGS1 Modulates Autophagic and Metabolic Programs and Is a Critical Mediator of Human Regulatory T Cell Function.

Regulatory T cells (Tregs) are critical mediators of immune tolerance and play a diametric role in cancer and autoimmunity. Tumor-infiltrating Tregs are often associated with poor prognosis in solid tumors because their enrichment in the tumor microenvironment contributes to immunosuppression. Conversely, dysregulation in the Treg compartment can disrupt self-tolerance, leading to autoimmunity. In the present study, we describe what is, to our knowledge, a novel regulator of Tregs, the GTPase activator regulator of G protein 1 (RGS1), demonstrating that RGS1-deficient human Tregs show downregulation of Treg-associated genes and are less immunosuppressive. These RGS1-deficient Tregs exhibit perturbations to the FOXP3-c-MYC transcriptional axis and downstream metabolic and autophagy programs by shifting their energy demands toward glycolysis and rendering them less autophagic. Taken together, RGS1 may serve as an apical node of Treg function by regulating the FOXP3-c-MYC transcriptional axis, thereby providing a therapeutic rationale for targeting RGS1 for treatment of cancer and autoimmune diseases.

Listeria-based vaccination against the pericyte antigen RGS5 elicits anti-vascular effects and colon cancer protection.

Colorectal cancer (CRC) remains a leading cause of cancer-related mortality despite efforts to improve standard interventions. As CRC patients can benefit from immunotherapeutic strategies that incite effector T cell action, cancer vaccines represent a safe and promising therapeutic approach to elicit protective and durable immune responses against components of the tumor microenvironment (TME). In this study, we investigate the pre-clinical potential of a Listeria monocytogenes (Lm)-based vaccine targeting the CRC-associated vasculature. CRC survival and progression are reliant on functioning blood vessels to effectively mediate various metabolic processes and oxygenate underlying tissues. We, therefore, advance the strategy of initiating immunity in syngeneic mouse models against the endogenous pericyte antigen RGS5, which is a critical mediator of pathological vascularization. Overall, Lm-based vaccination safely induced potent anti-tumor effects that consisted of recruiting functional Type-1-associated T cells into the TME and reducing tumor blood vessel content. This study underscores the promising clinical potential of targeting RGS5 against vascularized tumors like CRC.

RGS5 as a Biomarker of Pericytes, Involvement in Vascular Remodeling and Pulmonary Arterial Hypertension.

Introduction: Pulmonary arterial hypertension (PAH) is a life-threatening disease characterized by a sustained rise in mean pulmonary artery pressure. Pulmonary vascular remodeling serves an important role in PAH. Identifying a key driver gene to regulate vascular remodeling of the pulmonary microvasculature is critical for PAH management. Methods: Differentially expressed genes were identified using the Gene Expression Omnibus (GEO) GSE117261, GSE48149, GSE113439, GSE53408 and GSE16947 datasets. A co-expression network was constructed using weighted gene co-expression network analysis. Novel and key signatures of PAH were screened using four algorithms, including weighted gene co-expression network analysis, GEO2R analysis, support vector machines recursive feature elimination and robust rank aggregation rank analysis. Regulator of G-protein signaling 5 (RGS5), a pro-apoptotic/anti-proliferative protein, which regulate arterial tone and blood pressure in vascular smooth muscle cells. The expression of RGS5 was determined using reverse transcription-quantitative PCR (RT-qPCR) in PAH and normal mice. The location of RGS5 and pericytes was detected using immunofluorescence. Results: Compared with that in the normal group, RGS5 expression was upregulated in the PAH group based on GEO and RT-qPCR analyses. RGS5 expression in single cells was enriched in pericytes in single-cell RNA sequencing analysis. RGS5 co-localization with pericytes was detected in the pulmonary microvasculature of PAH. Conclusion: RGS5 regulates vascular remodeling of the pulmonary microvasculature and the occurrence of PAH through pericytes, which has provided novel ideas and strategies regarding the occurrence and innovative treatment of PAH.

RGS10 inhibits proliferation and migration of pulmonary arterial smooth muscle cell in pulmonary hypertension via AKT/mTORC1 signaling.

Objective: Excessive proliferation and migration of pulmonary arterial smooth muscle cell (PASMC) is a core event of pulmonary hypertension (PH). Regulators of G protein signaling 10 (RGS10) can regulate cellular proliferation and cardiopulmonary diseases. We demonstrate whether RGS10 also serves as a regulator of PH.Methods: PASMC was challenged by hypoxia to induce proliferation and migration. Adenovirus carrying Rgs10 gene (Ad-Rgs10) was used for external expression of Rgs10. Hypoxia/SU5416 or MCT was used to induce PH. Right ventricular systolic pressure (RVSP) and right ventricular hypertrophy index (RVHI) were used to validate the establishment of PH model.Results: RGS10 was downregulated in hypoxia-challenged PASMC. Ad-Rgs10 significantly suppressed proliferation and migration of PASMC after hypoxia stimulus, while silencing RGS10 showed contrary effect. Mechanistically, we observed that phosphorylation of S6 and 4E-Binding Protein 1 (4EBP1), the main downstream effectors of mammalian target of rapamycin complex 1 (mTORC1) as well as phosphorylation of AKT, the canonical upstream of mTORC1 in hypoxia-induced PASMC were negatively modulated by RGS10. Both recovering mTORC1 activity and restoring AKT activity abolished these effects of RGS10 on PASMC. More importantly, AKT activation also abolished the inhibitory role of RGS10 in mTORC1 activity in hypoxia-challenged PASMC. Finally, we also observed that overexpression of RGS10 in vivo ameliorated pulmonary vascular wall thickening and reducing RVSP and RVHI in mouse PH model.Conclusion: Our findings reveal the modulatory role of RGS10 in PASMC and PH via AKT/mTORC1 axis. Therefore, targeting RGS10 may serve as a novel potent method for the prevention against PH."

Identification of Breast Cancer Metastasis Markers from Gene Expression Profiles Using Machine Learning Approaches.

Cancer metastasis accounts for approximately 90% of cancer deaths, and elucidating markers in metastasis is the first step in its prevention. To characterize metastasis marker genes (MGs) of breast cancer, XGBoost models that classify metastasis status were trained with gene expression profiles from TCGA. Then, a metastasis score (MS) was assigned to each gene by calculating the inner product between the feature importance and the AUC performance of the models. As a result, 54, 202, and 357 genes with the highest MS were characterized as MGs by empirical p-value cutoffs of 0.001, 0.005, and 0.01, respectively. The three sets of MGs were compared with those from existing metastasis marker databases, which provided significant results in most comparisons (p-value < 0.05). They were also significantly enriched in biological processes associated with breast cancer metastasis. The three MGs, SPPL2C, KRT23, and RGS7, showed highly significant results (p-value < 0.01) in the survival analysis. The MGs that could not be identified by statistical analysis (e.g., GOLM1, ELAVL1, UBP1, and AZGP1), as well as the MGs with the highest MS (e.g., ZNF676, FAM163B, LDOC2, IRF1, and STK40), were verified via the literature. Additionally, we checked how close the MGs were to each other in the protein-protein interaction networks. We expect that the characterized markers will help understand and prevent breast cancer metastasis.

The therapeutic potential of targeting cardiac RGS4.

G protein-coupled receptors (GPCRs) play pivotal roles in regulation of cardiac function and homeostasis. To function properly, every cell needs these receptors to be stimulated only when a specific extracellular stimulus is present, and to be silenced the moment that stimulus is removed. The regulator of G protein signaling (RGS) proteins are crucial for the latter to occur at the cell membrane, where the GPCR normally resides. Perturbations in both activation and termination of G protein signaling underlie numerous heart pathologies. Although more than 30 mammalian RGS proteins have been identified, each RGS protein seems to interact only with a specific set of G protein subunits and GPCR types/subtypes in any given tissue or cell type, and this applies to the myocardium as well. A large number of studies have provided substantial evidence for the roles various RGS proteins expressed in cardiomyocytes play in cardiac physiology and heart disease pathophysiology. This review summarizes the current understanding of the functional roles of cardiac RGS proteins and their implications for the treatment of specific heart diseases, such as heart failure and atrial fibrillation. We focus on cardiac RGS4 in particular, since this isoform appears to be selectively (among the RGS protein family) upregulated in human heart failure and is also the target of ongoing drug discovery efforts for the treatment of a variety of diseases.

RGS1 mediates renal interstitial fibrosis through activation of the inflammatory response.

Renal interstitial fibrosis (RIF) is considered as a common pathway for all patients with chronic kidney disease (CKD) to progress to end-stage kidney disease (ESRD). The basic pathological manifestation is the increase of matrix component in the tubular interstitium, while the injury of tubular epithelial cells in the renal interstitium and the excessive accumulation of matrix will eventually lead to tubular atrophy and obstruction, loss of effective renal units, and finally impaired renal filtration function. The relevant mechanism of RIF remains unclear. The present study will investigate the function and relevant mechanism of RGS1 in RIF. The RIF-related microarrays GSE22459 and GSE76882 were downloaded and analyzed. Renal parenchymal atrophic calyx tissues were collected from clinical RIF patients. Cellular inflammation, fibrosis and animal RIF models were constructed using Lipopolysaccharide (LPS), TGF-ß1 and unilateral ureteral occlusion (UUO). HE and Masson staining were performed to detect morphological alterations of renal tissue samples. qRT-PCR, Western blot and ELISA were carried out to detect the expression of relevant genes/proteins. RGS1 is a gene co-differentially expressed by GSE22459 and GSE76882. RGS1 expression was elevated in renal tissues of RIF patients, cells and animal RIF models. Knockdown of RGS1 inhibited renal cell inflammatory response, fibrosis and renal fibrosis in RIF mice. Overexpression of RGS1 plays the opposite role. Knockdown of RGS1 inhibited the inflammatory response in the RIF cell and mouse model. Targeting RGS1 might be a potential therapeutic strategy for RIF treatment.

A regulatory node involving Gαq, PLCß, and RGS proteins modulates platelet reactivity to critical agonists.

BACKGROUND: Most platelet agonists work through G protein-coupled receptors, activating pathways that involve members of the Gq, Gi, and G12/G13 families of heterotrimeric G proteins. Gq signaling has been shown to be critical for efficient platelet activation. Growing evidence suggests that regulatory mechanisms converge on G protein-coupled receptors and Gq to prevent overly robust platelet reactivity. OBJECTIVES: To identify and characterize mechanisms by which Gq signaling is regulated in platelets. METHODS: Based on our prior experience with a Gαi2 variant that escapes regulation by regulator of G protein signaling (RGS) proteins, a Gαq variant was designed with glycine 188 replaced with serine (G188S) and then incorporated into a mouse line so that its effects on platelet activation and thrombus formation could be studied in vitro and in vivo. RESULTS AND CONCLUSIONS: As predicted, the G188S substitution in Gαq disrupted its interaction with RGS18. Unexpectedly, it also uncoupled PLCß-3 from activation by platelet agonists as evidenced by a loss rather than a gain of platelet function in vitro and in vivo. Binding studies showed that in addition to preventing the binding of RGS18 to Gαq, the G188S substitution also prevented the binding of PLCß-3 to Gαq. Structural analysis revealed that G188 resides in the region that is also important for Gαq binding to PLCß-3 in platelets. We conclude that the Gαq signaling node is more complex than that has been previously understood, suggesting that there is cross-talk between RGS proteins and PLCß-3 in the context of Gαq signaling.