Mesenchymal stem cell-derived exosomes overexpressing SRC-3 protect mice from cerebral ischemia by inhibiting ferroptosis.

BACKGROUND: The treatment for cerebral ischemia remains limited, and new therapeutic strategies are urgently needed. Exosome has shown great promise for the treatment of cerebral ischemia. Steroid receptor coactivator-3 (SRC-3) was reported to be involved in neurological performances. In this study, we aimed to investigate the protective effects of mesenchymal stem cell (MSC)-derived exosomes overexpressing SRC-3 on cerebral ischemia in mice. METHODS: The mice were treated with an intracerebroventricular injection of GFP-overexpressed exosomes (GFP-exo) and SRC-3-overexpressed exosomes (SRC3-exo) in a middle cerebral artery occlusion (MCAO) model of cerebral ischemia. RESULTS: The results showed that SRC3-exo treatment significantly inhibited lipid peroxidation and ferroptosis of the neurons subjected to oxygen-glucose deprivation. It further suppressed the activation of microglia and astrocytes, and decreased the production of pro-inflammatory cytokines in the brains of MCAO mice. Furthermore, SRC3-exo treatment reduced the water content of brain tissue and infarct size, which alleviated the neurological damage and improved neurological performances in the MCAO mice. CONCLUSIONS: Our results suggest that MSC-derived exosomes expressing SRC3 can be a therapeutic strategy for cerebral ischemia by inhibiting ferroptosis.

Lead Compound Development of SRC-3 Inhibitors with Improved Pharmacokinetic Properties and Anticancer Efficacy.

Steroid receptor coactivator 3 (SRC-3) is a critical mediator of many intracellular signaling pathways that are crucial for cancer proliferation and metastasis. In this study, we performed structure-activity relationship exploration and drug-like optimization of the hit compound SI-2, guided by in vitro/in vivo metabolism studies and cytotoxicity assays. Our efforts led to the discovery of two lead compounds, SI-10 and SI-12. Both compounds exhibit potent cytotoxicity against a panel of human cancer cell lines and demonstrate acceptable pharmacokinetic properties. A biotinylated estrogen response element pull-down assay demonstrated that SI-12 could disrupt the recruitment of SRC-3 and p300 in the estrogen receptor complex. Importantly, SI-10 and SI-12 significantly inhibited tumor growth and metastasis in vivo without appreciable acute toxicity. These results demonstrate the potential of SI-10 and SI-12 as drug candidates for cancer therapy, given their potent SRC-3 inhibition and promising pharmacokinetic and toxicity profiles.

Deficiency of Nuclear Receptor Coactivator 3 Aggravates Diabetic Kidney Disease by Impairing Podocyte Autophagy.

Nuclear receptors (NRs) are important transcriptional factors that mediate autophagy, preventing podocyte injury and the progression of diabetic kidney disease (DKD). However, the role of nuclear receptor coactivators that are powerful enhancers for the transcriptional activity of NRs in DKD remains unclear. In this study, a significant decrease in Nuclear Receptor Coactivator 3 (NCOA3) is observed in injured podocytes caused by high glucose treatment. Additionally, NCOA3 overexpression counteracts podocyte damage by improving autophagy. Further, Src family member, Fyn is identified to be the target of NCOA3 that mediates the podocyte autophagy process. Mechanistically, NCOA3 regulates the transcription of Fyn in a nuclear receptor, PPAR-γ dependent way. Podocyte-specific NCOA3 knockout aggravates albuminuria, glomerular sclerosis, podocyte injury, and autophagy in DKD mice. However, the Fyn inhibitor, AZD0530, rescues podocyte injury of NCOA3 knockout DKD mice. Renal NCOA3 overexpression with lentivirus can ameliorate podocyte damage and improve podocyte autophagy in DKD mice. Taken together, the findings highlight a novel target, NCOA3, that protects podocytes from high glucose injury by maintaining autophagy.

USP24-dependent stabilization of Runx2 recruits a p300/NCOA3 complex to transactivate ADAMTS genes and promote degeneration of intervertebral disc in chronic inflammation mice.

BACKGROUND: Intervertebral disc degeneration (IDD) naturally occurs during the aging process. Its occurrence is closely related to chronic inflammation; however, the causal relationship between them is controversial. This study aimed to investigate if inflammation would promote IDD incidence and explore the underlying mechanism. METHODS: A chronic inflammation mouse model was established by intraperitoneal injection of lipopolysaccharide (LPS). Enzyme-linked immunosorbent assay was performed to determine proinflammatory cytokines in serum. Histological staining was used to evaluate the degeneration of IVDs. Immunoblots and RT-qPCR analyses were performed to measure protein and mRNA expression levels. Immunoprecipitation, mass spectrometry, and co-immunoprecipitation assays were used to determine the assembly of protein complex. RESULTS: We found that an inflammatory microenvironment activated p38 kinase, which phosphorylated the Runx2 transcription factor at the Ser28 site. The phosphorylated Runx2 (pRunx2) then recruited a deubiquitinase, ubiquitin-specific peptidase 24 (USP24), which stabilized pRunx2 and protected it from ubiquitin-dependent proteasomal degradation. The stabilized pRunx2 recruited histone acetyltransferase p300 and nuclear receptor coactivator 3 (NCOA3) to assemble a complex. This NCOA3-p300-pRunx2 complex then transactivated the expression of 13 ADAMTS (a disintegrin and metalloproteinase with thrombospondin motif) genes, thereby promoting the degradation of extracellular matrix (ECM) in intervertebral discs (IVDs) and causing IDD. Administration of either a p38 inhibitor (doramapimod), a NCOA3 inhibitor (bufalin), or a p300 inhibitor (EML425) significantly decreased the expression of the 13 ADAMTS genes and slowed the degeneration of IVDs. CONCLUSION: In summary, our results demonstrate that USP24 protects pRunx2 from proteasomal degradation under chronic inflammation conditions, enabling pRunx2 to transactivate ADAMTS genes and degrade ECM. Our findings provide direct evidence that chronic inflammation triggers IDD and offer a therapeutic strategy for retarding IDD in patients with chronic inflammation.

[The relationship between the expression of amplified in breast cancer 1、androgen receptor and tamoxifen resistance in breast cancer].

Objective: To discuss the value of amplified in breast cancer 1(AIB1) and androgen receptor (AR) for the resistance of adjuvant tamoxifen in estradiol receptor (ER) positive breast cancer. Methods: A total of 188 cases with breast cancer after receiving tamoxifen treatment in the Tianjin Medical University Cancer Institute and Hospital from June 2008 to July 2013 were enrolled in this study.Using immunohistochemical SP method to detect AIB1and AR expression in breast cancer tissue, analyzing the relationship of AIB1 and AR expression and the effect of tamoxifen.And verify the results of the experiment through the GEPIA database. Results: The response of tamoxifen was 80.3%. The response rate in AR positive group and AR negative group was 79.6% and 82.4%, with no significant difference (P=0.669). The response rate in AIB1 High expression group and AIB1 Low expression group was 68.4% and 93.3%, respectively, with a significant difference (P<0.001).The response rate in AR negative and AIB1 Low expression group, AR negative and AIB1 High expression group, AR positive and AIB1 Low expression group, AR positive and AIB1High expression group was 89.7%, 71.4%, 96.7%, 66.2%respectively, with a significant difference (P<0.001). Conclusions: The expression level of AIB1 is correlated with the therapeutic effect of tamoxifen in breast cancer. Its high expression can cause tamoxifen resistance, while AR positive and High expression of AIB1 are more likely to cause tamoxifen resistance, and AIB1 can be used as an independent influencing factor for breast cancer tamoxifentreatment.

NCoA3 upregulation in breast cancer­associated adipocytes elicits an inflammatory profile.

Nuclear receptor coactivator 3 (NCoA3) is a transcriptional coactivator of NF­κB and other factors, which is expressed at relatively low levels in normal cells and is amplified or overexpressed in several types of cancer, including breast tumors. NCoA3 levels have been shown to be decreased during adipogenesis; however, its role in tumor­surrounding adipose tissue (AT) remains unknown. Therefore, the present study assessed the modulation of NCoA3 in breast cancer­associated adipocytes and evaluated its association with the expression of inflammatory markers. 3T3­L1 adipocytes were stimulated with conditioned medium from human breast cancer cell lines and the expression levels of NCoA3 were evaluated by reverse transcription­quantitative (q)PCR. NF­κB activation was measured by immunofluorescence, and tumor necrosis factor and monocyte chemoattractant protein 1 levels were analyzed by qPCR and dot blot assays. The results obtained from the in vitro model were supported using mammary AT (MAT) from female mice, MAT adjacent to tumors from patients with breast cancer and bioinformatics analysis. The results revealed that adipocytes expressing high levels of NCoA3 were mainly associated with a pro­inflammatory profile. In 3T3­L1 adipocytes, NCoA3 downregulation or NF­κB inhibition reversed the expression of inflammatory molecules. In addition, MAT from patients with a worse prognosis exhibited high levels of this coactivator. Notably, adipocyte NCoA3 levels could be modulated by inflammatory signals from tumors. The modulation of NCoA3 levels in synergy with NF­κB activity in MAT in a tumor context could be factors required to establish breast cancer­associated inflammation. As adipocytes are involved in the development and progression of breast cancer, this signaling network deserves to be further investigated to improve future tumor treatments.

Nuclear receptor coactivator 3 transactivates proinflammatory cytokines in collagen-induced arthritis.

Rheumatoid arthritis (RA) is an autoimmune disorder in which the immune system mistakenly attacks joints. The molecular mechanisms underlying RA pathology are still under investigation. In this study, we discovered overexpression of nuclear receptor coactivator 3 (NCOA3) in the joint tissues of type II collagen-induced arthritis (CIA) mice, an important autoimmune model of human RA. Administration of two NCOA3 inhibitors, gossypol (GSP) and SI-2 hydrochloride (SHC), significantly alleviated inflammation and improved the outcomes of CIA mice. In vivo and in vitro experiments revealed that NCOA3 assembled a transcriptional complex with a histone acetyltransferase p300 and two subunits of nuclear factor kappa B (NF-κB). This complex specifically controlled the expression of proinflammatory cytokine genes by binding to their promoters. Knockdown of NCOA3 or in vitro treatments with GSP and SHC impaired the assembly of NCOA3-p300-NF-κB complex and decreased the expression of proinflammatory cytokine genes. Taken together, our results demonstrated that NCOA3 acts as a mediator of proinflammatory cytokine genes in CIA mice and that inhibition of the NCOA3-p300-NF-κB complex may represent a new avenue for improving RA outcomes.

Steroid receptor coactivator-3 inhibition generates breast cancer antitumor immune microenvironment.

BACKGROUND: The tumor immune microenvironment (TIME) generated by cancer-infiltrating immune cells has a crucial role in promoting or suppressing breast cancer progression. However, whether the steroid receptor coactivator-3 (SRC-3) modulates TIME to progress breast cancer is unclear. Therefore, the present study evaluates whether SRC-3 generates a tumor-promoting TIME in breast tumors using a syngeneic immune-intact mouse model of breast cancer. METHODS: We employed E0771 and 4T1 breast cancer in immune-intact syngeneic female C57BL/6 and BALB/c mice, respectively. SI-2, a specific small-molecule inhibitor of SRC-3, was administered daily (2.5 mg/kg) to E0771 and 4T1 breast tumor-bearing immune-intact mice. In addition, SRC-3 knockdown (KD)-E0771 and SRC-3 KD-4T1 cells and their parental breast cancer cells were injected into their syngeneic immune-intact female mice versus immune-deficiency mice to validate that the host immune system is required for breast tumor suppression by SRC-3 KD in immune-intact mice. Furthermore, tumor-infiltrating immune cells (such as CD4+, CD8+, CD56+, and Foxp3+ cells) in E0771 and 4T1 breast cancers treated with SI-2 and in SRC-3 KD E0771 and 4T1 breast cancers were determined by immunohistochemistry. Additionally, cytokine levels in SI-2-treated and SRC-3 KD E0771 breast tumors and their control cancers were defined with a Mouse Cytokine Array. RESULTS: SRC-3 inhibition by SI-2 significantly suppressed the progression of breast cancer cells (E0771 and 4T1) into breast cancers in immune-intact syngeneic female mice. SRC-3 KD-E0771 and -4T1 breast cancer cells did not produce well-developed tumors in immune-intact syngeneic female mice compared to their parental cells, but SRC-3 KD breast cancers were well developed in immune-defective host mice. SRC-3 inhibition by SI-2 and SRC-3 KD effectively increased the numbers of cytotoxic immune cells, such as CD4+ and CD8+ T cells and CD56+ NK cells, and Interferon γ (Ifng) in breast cancers compared to vehicle. However, SI-2 treatment reduced the number of tumor-infiltrating CD4+/Foxp3+ regulatory T (Treg) cells compared to vehicle treatment. In addition, SRC-3 inhibition by SI-2 and SRC-3 KD increased C-X-C motif chemokine ligand 9 (Cxcl9) expression in breast cancer to recruit C-X-C motif chemokine receptor 3 (Cxcr3)-expressing cytotoxic immune cells into breast tumors. CONCLUSIONS: SRC-3 is a critical immunomodulator in breast cancer, generating a protumor immune microenvironment. SRC-3 inhibition by SI-2 or SRC-3 KD activates the Cxcl9/Cxcr3 axis in breast tumors and enhances the antitumor immune microenvironment to suppress breast cancer progression.

SRC-3 deficiency prevents atherosclerosis development by decreasing endothelial ICAM-1 expression to attenuate macrophage recruitment.

Steroid receptor coactivator 3 (SRC-3) is a member of the p160 SRC family. This factor can interact with multiple nuclear hormone receptors and transcription factors to regulate the expression of their target genes. Although many physiological roles of SRC-3 have been revealed, its role in atherosclerosis is not clear. In this study, we found that SRC-3-/-ApoE-/- mice have reduced atherosclerotic lesions and necrotic areas in their aortas and aortic roots compared with SRC-3+/+ApoE-/- mice after Western diet (WD) feeding for 12 weeks. RNA-Seq and Western blot analyses of the aorta revealed that SRC-3 was required for maintaining the expression of ICAM-1, which was required for macrophage recruitment and atherosclerosis development. siRNA-mediated knockdown of SRC-3 in endothelial cells significantly reduced WD-induced atherosclerotic plaque formation. Additionally, treatment of ApoE-/- mice with SRC-3 inhibitor bufalin prevented atherosclerotic plaque development. SRC-3 deficiency reduced aortic macrophage recruitment. Accordingly, ICAM-1 expression was markedly decreased in the aortas of SRC-3-/-ApoE-/- mice and ApoE-/- mice with endothelial SRC-3 knockdown mediated by AAV9-shSRC-3 virus. Mechanistically, SRC-3 coactivated NF-κB p65 to increase ICAM-1 transcription in endothelial cells. Collectively, these findings demonstrate that inhibiting SRC-3 ameliorates atherosclerosis development, at least in part through suppressing endothelial activation by decreasing endothelial ICAM-1 expression via reducing NF-κB signaling.

Hypoxia induces chemoresistance to proteasome inhibitors through orchestrating deSUMOylation and ubiquitination of SRC-3 in multiple myeloma.

The bone marrow microenvironment in multiple myeloma (MM) is hypoxic and provides multi-advantages for the initiation of chemoresistance, but the underlying mechanisms and key regulators are still indistinct. In the current study, we found that hypoxia stimulus easily induced chemoresistance to proteasome inhibitors (PIs), and the steroid receptor coactivator 3 (SRC-3) expression was remarkably augmented at posttranslational level. Protein interactome analysis identified SENP1 as a key modifier of SRC-3 stability, as SENP1-mediated deSUMOylation attenuated the K11-linked polyubiquitination of SRC-3. SENP1 depletion in the SENP1fl/flCD19Cre/+ B cells showed impaired SRC3 stability, and knockdown of SENP1 in MM cells by CRISPR/cas9 sgRNA accelerated the degradation of SRC-3 and remarkably overcame the resistance to PIs. In the Vk*Myc and 5TGM1 mouse models as well as patient-derived xenograft (PDX) of myeloma, SENP1 inhibitor Momordin Ιc (Mc) increased the sensitivity to PIs in MM cells. Importantly, SENP1 level was positively correlated with SRC-3 level in the tissues from refractory/relapsed MM, as well as in xenograft tissues from mice treated with bortezomib and Mc. Taken together, our findings suggest that hypoxia-induced SENP1 is a crucial regulator of chemoresistance to PIs, and shed light on developing therapeutic strategies to overcome chemoresistance by using small molecules targeting SENP1 or SRC-3.

Bone marrow mesenchymal stem cells-derived exosomes mediate nuclear receptor coactivator-3 expression in osteoblasts by delivering miR-532-5p to influence osteonecrosis of the femoral head development.

Exosomes (Exo) originated from bone marrow mesenchymal stem cells (BMSCs) have therapeutic impacts on osteonecrosis of the femoral head (ONFH), and microRNA (miR)-532-5p has been confirmed to participate in ONFH progression. In the research, it was figured out whether BMSCs-Exo could relieve ONFH by delivering miR-532-5p. MG-63 cells were treated with DEX to construct an ONFH cell model in vitro. The effects of Exo and miR-532-5p on the cell viability, lactate dehydrogenase (LDH) content, and apoptosis of BMSCs were detected. The ONFH rat model was established, and the effect of BMSCs-Exo delivering miR-532-5p on the pathological damage of ONFH rats was evaluated. Changes in nuclear receptor coactivator-3 (NCOA3) and apoptotic proteins were assessed by western blot. The relationship between miR-532-5p and NCOA3 was verified by dual luciferase reporter experiments. miR-532-5p was elevated in vivo and in vitro ONFH-models, while NCOA3 expression was reduced. Overexpression of miR-532-5p aggravated DEX toxicity in osteoblasts, decreased cell viability, and promoted apoptosis. Knockdown of miR-532-5p made Exo further attenuate the toxic effect of DEX on osteoblasts and inhibited apoptosis. The protective effect of miR-532-5p-delivering Exo on osteoblasts was reversed by NCOA3 silencing. In addition, in vivo experiments also confirmed that knockdown of miR-532-5p enhanced the therapeutic effect of Exo on ONFH rats. This study demonstrates that miR-532-5p-delivering BMSCs-Exo inhibits osteoblast viability and promote apoptosis by targeting NCOA3, thereby aggravating ONFH development.

SRC3 deficiency exacerbates lipopolysaccharide-induced acute respiratory distress syndrome in mice.

Acute respiratory distress syndrome (ARDS) is a severe disease. Inflammation is the key element implicated in ARDS. Steroid receptor coactivator 3 (SRC3), a coactivator protein for transcription, is involved in regulation of inflammatory response. Here we explored the potential roles of SRC3 in ARDS. We utilized the SRC3 deficient (SRC3-/-) mice and established the lipopolysaccharides (LPS)-induced ARDS model. The mortality, lung injury, leucocytes infiltration and inflammatory cytokine production were compared between wild type (WT) and SRC3-/- mice. The NF-κB activation in lung of WT and SRC3-/- mice was measured. After LPS treatment, SRC3-/- mice had higher mortality and more severe lung damage than WT mice. LPS-treated SRC3-/- mice had more leucocytes infiltration and upregulated inflammatory cytokine production. LPS-treated SRC3-/- mice had elevated NF-κB activation. SRC3-/- mice had exacerbated ARDS in LPS-treated mice.

Aib1 deficiency exacerbates inflammatory responses in acute myocardial infarction mice.

Acute myocardial infarction (AMI) is one of the major causes of death throughout the world, while inflammation has been known as a major contributor to the pathophysiology of AMI. Inhibition of inflammation is shown to protect from AMI. Amplified in breast 1 (Aib1) is a transcriptional coactivator protein which can suppress inflammation. The anti-inflammatory activities of Aib1 imply its potential effects against AMI; however, to date the role of Aib1 in AMI has not been described yet. Here we explored the potential functions of Aib1 in AMI. We induced AMI in both wild-type (WT) and Aib1-/- mice. The expression levels of Aib1 and inflammatory cytokines in the AMI WT mice were measured by RT-PCR and Western blot. The heart infarction area and cardiac functions were compared between the AMI WT and Aib1-/- mice. The expression levels of inflammatory cytokines including IL-6, IL-1ß, TNF-α, and MCP-1 in heart tissues were compared between the AMI WT and Aib1-/- mice by ELISA and RT-PCR. AMI induced the production of inflammatory cytokines whereas suppressed the expression of Aib1 in WT mice. AMI Aib1-/- mice displayed increased infarct area and aggravated heart dysfunction, as well as upregulated levels of Il-6, Il-1ß, Tnf-α, and Mcp-1 in heart tissues. Aib1 deficiency exacerbates inflammation in AMI mice. KEY MESSAGES: AMI induced inflammation in the heart tissue of mice. Aib1 knockout exacerbated infarction in AMI mice. Aib1 knockout exacerbated cardiac dysfunction in AMI mice. Aib1 knockout exacerbated inflammation in AMI mice.

ZFP64::NCOA3 gene fusion defines a novel subset of spindle cell rhabdomyosarcoma.

Spindle cell rhabdomyosarcoma represents a rare neoplasm characterized by monomorphic spindle cells with a fascicular architecture and variable skeletal muscle differentiation. Following incidental identification of a ZFP64::NCOA3 gene fusion in an unclassified spindle cell sarcoma resembling adult-type fibrosarcoma, we performed a retrospective archival review and identified four additional cases with a similar histology and identical gene fusion. All tumors arose in adult males (28-71 years). The neoplasms were found in the deep soft tissues, two were gluteal, and one each arose in the thigh, abdominal wall, and chest wall. Morphologically, the tumors were characterized by spindle cells with a distinctive herringbone pattern and variable collagenous to myxoid stroma. The nuclei were relatively monomorphic with variable mitotic activity. Three tumors had immunoreactivity for MyoD1, and four contained variable expression of desmin and smooth muscle actin. All cases tested for myogenin, CD34, S100, pankeratin, and epithelial membrane antigen were negative. Targeted RNA sequencing revealed a ZFP64::NCOA3 fusion product in all five tumors. Three patients developed distant metastases, and two ultimately succumbed to their disease within 2 years of initial diagnosis. This study suggests ZFP64::NCOA3 fusions define a novel subtype of rhabdomyosarcoma with a spindle cell morphology and aggressive clinical behavior. The potential for morphologic and immunohistochemical overlap with several other sarcoma types underscores the value of molecular testing as a diagnostic adjunct to ensure accurate classification and management of these neoplasms.

Spatial transcriptomics using combinatorial fluorescence spectral and lifetime encoding, imaging and analysis.

Multiplexed mRNA profiling in the spatial context provides new information enabling basic research and clinical applications. Unfortunately, existing spatial transcriptomics methods are limited due to either low multiplexing or complexity. Here, we introduce a spatialomics technology, termed Multi Omic Single-scan Assay with Integrated Combinatorial Analysis (MOSAICA), that integrates in situ labeling of mRNA and protein markers in cells or tissues with combinatorial fluorescence spectral and lifetime encoded probes, spectral and time-resolved fluorescence imaging, and machine learning-based decoding. We demonstrate MOSAICA's multiplexing scalability in detecting 10-plex targets in fixed colorectal cancer cells using combinatorial labeling of five fluorophores with facile error-detection and removal of autofluorescence. MOSAICA's analysis is strongly correlated with sequencing data (Pearson's r = 0.96) and was further benchmarked using RNAscopeTM and LGC StellarisTM. We further apply MOSAICA for multiplexed analysis of clinical melanoma Formalin-Fixed Paraffin-Embedded (FFPE) tissues. We finally demonstrate simultaneous co-detection of protein and mRNA in cancer cells.

NCOA3 is a critical oncogene in thyroid cancer via the modulation of major signaling pathways.

PURPOSE: The Nuclear Receptor Coactivator (NCOA3, also known as SRC-3, AIB1, p/CIP, RAC3, ACTR, and TRAM1), acts as an oncogene in multiple tumors, but its biological function in thyroid cancer remains unclear. This study was designed to explore the role of NCOA3 in thyroid cancer. METHODS: The study assessed NCOA3 expression in thyroid cancer and their matched non-cancerous thyroid tissues at mRNA and protein levels. Then we evaluated the effect of NCOA3 on malignant activities of thyroid cancer cells. To better understand the oncogenic role of NCOA3 in thyroid tumorigenesis, we tested the effect of NCOA3 on major proteins related to thyroid cancer. RESULTS: Our data demonstrated that protein expression of NCOA3 was significantly upregulated in thyroid cancer tissues. NCOA3 knockdown inhibited cell proliferation and invasion, and induced cell cycle arrest and apoptosis in thyroid cancer. Conversely, ectopic expression of NCOA3 promoted cell proliferation and invasiveness in thyroid cancer. Mechanistically, NCOA3 could improve the survival and invasiveness of thyroid cancer cells through the modulation of the ErbB, AKT, ERK, and ß-catenin pathways. CONCLUSION: Collectively, these findings suggest that NCOA3 is critical in the initiation and development of thyroid cancer, and might be a possible marker for prognosis and therapy.

Circ_0001667 knockdown blocks cancer progression and attenuates adriamycin resistance by depleting NCOA3 via releasing miR-4458 in breast cancer.

Accumulating evidence suggests that developmental chemoresistance in cancers is closely associated with the dysregulation of circular RNA transcriptions. The objective of this study is to disclose the role of circ_0001667 and provide a potential functional mechanism in breast cancer. Quantitative real-time PCR was applied for the analysis of circ_0001667, microRNA-4458 (miR-4458) and nuclear receptor coactivator 3 (NCOA3) expression. In adriamycin (ADM)-resistant cell lines, we investigated cell proliferation using 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay and colony formation assay. Cell migration and cell invasion were determined by transwell assay. The protein levels of multi-drug resistance-1, matrix metalloproteinases-9, cleaved-caspase3, cleaved-caspase9 and NCOA3 were detected by western blot. ADM resistance was ascertained by IC50 value using MTT assay. Cell apoptosis was checked by flow cytometry assay. The putative relationship between miR-4458 and circ_0001667 and NCOA3 was validated by pull-down assay, dual-luciferase reporter assay or RNA Immunoprecipitation assay. Circ_0001667 knockdown inhibited MCF-7/ADM and MDA-MB-231/ADM cell proliferation, migration, invasion and ADM resistance. MiR-4458 was a target of circ_0001667, and its expression was decreased in ADM-resistant tumor tissues and cells. MiR-4458 inhibition reversed the effects of circ_0001667 knockdown. In depth, NCOA3 was a target of circ_0001667, and circ_0001667 knockdown weakened NCOA3 expression by releasing miR-4458. MCF-7/ADM and MDA-MB-231/ADM cell proliferation, migration, invasion, and ADM resistance inhibited by miR-4458 restoration were recovered by NCOA3 overexpression. Circ_0001667 knockdown also repressed tumor growth and ADM resistance in vivo. Circ_0001667 knockdown blocks cancer progression and attenuates ADM resistance by depleting NCOA3 via releasing miR-4458 in breast cancer.

A-Kinase Anchoring Protein 2 Promotes Protection against Myocardial Infarction.

Myocardial infarction (MI) is a leading cause of maladaptive cardiac remodeling and heart failure. In the damaged heart, loss of function is mainly due to cardiomyocyte death and remodeling of the cardiac tissue. The current study shows that A-kinase anchoring protein 2 (AKAP2) orchestrates cellular processes favoring cardioprotection in infarcted hearts. Induction of AKAP2 knockout (KO) in cardiomyocytes of adult mice increases infarct size and exacerbates cardiac dysfunction after MI, as visualized by increased left ventricular dilation and reduced fractional shortening and ejection fraction. In cardiomyocytes, AKAP2 forms a signaling complex with PKA and the steroid receptor co-activator 3 (Src3). Upon activation of cAMP signaling, the AKAP2/PKA/Src3 complex favors PKA-mediated phosphorylation and activation of estrogen receptor α (ERα). This results in the upregulation of ER-dependent genes involved in protection against apoptosis and angiogenesis, including Bcl2 and the vascular endothelial growth factor a (VEGFa). In line with these findings, cardiomyocyte-specific AKAP2 KO reduces Bcl2 and VEGFa expression, increases myocardial apoptosis and impairs the formation of new blood vessels in infarcted hearts. Collectively, our findings suggest that AKAP2 organizes a transcriptional complex that mediates pro-angiogenic and anti-apoptotic responses that protect infarcted hearts.

An acetylation-enhanced interaction between transcription factor Sox2 and the steroid receptor coactivators facilitates Sox2 transcriptional activity and function.

SRY-box 2 (Sox2) is a transcription factor with critical roles in maintaining embryonic stem (ES) cell and adult stem cell functions and in tumorigenesis. However, how Sox2 exerts its transcriptional function remains unclear. Here, we used an in vitro protein-protein interaction assay to discover transcriptional regulators for ES cell core transcription factors (Oct4, Sox2, Klf4, and c-Myc) and identified members of the steroid receptor coactivators (SRCs) as Sox2-specific interacting proteins. The SRC family coactivators have broad roles in transcriptional regulation, but it is unknown whether they also serve as Sox2 coactivators. We demonstrated that these proteins facilitate Sox2 transcriptional activity and act synergistically with p300. Furthermore, we uncovered an acetylation-enhanced interaction between Sox2 and SRC-2/3, but not SRC-1, demonstrating it is Sox2 acetylation that promotes the interaction. We identified putative Sox2 acetylation sites required for acetylation-enhanced interaction between Sox2 and SRC-3 and demonstrated that acetylation on these sites contributes to Sox2 transcriptional activity and recruitment of SRC-3. We showed that activation domains 1 and 2 of SRC-3 both display a preferential binding to acetylated Sox2. Finally, functional analyses in mouse ES cells demonstrated that knockdown of SRC-2/3 but not SRC-1 in mouse ES cells significantly downregulates the transcriptional activities of various Sox2 target genes and impairs ES cell stemness. Taken together, we identify specific SRC family proteins as novel Sox2 coactivators and uncover the role of Sox2 acetylation in promoting coactivator recruitment and Sox2 transcriptional function.

Conformational Changes of RORγ During Response Element Recognition and Coregulator Engagement.

The retinoic acid receptor-related orphan receptor γ (RORγ) is a ligand-dependent transcription factor of the nuclear receptor super family that underpins metabolic activity, immune function, and cancer progression. Despite being a valuable drug target in health and disease, our understanding of the ligand-dependent activities of RORγ is far from complete. Like most nuclear receptors, RORγ must recruit coregulatory protein to enact the RORγ target gene program. To date, a majority of structural studies have been focused exclusively on the RORγ ligand-binding domain and the ligand-dependent recruitment of small peptide segments of coregulators. Herein, we examine the ligand-dependent assembly of full length RORγ:coregulator complexes on cognate DNA response elements using structural proteomics and small angle x-ray scattering. The results from our studies suggest that RORγ becomes elongated upon DNA recognition, preventing long range interdomain crosstalk. We also determined that the DNA binding domain adopts a sequence-specific conformation, and that coregulatory protein may be able to 'sense' the ligand- and DNA-bound status of RORγ. We propose a model where ligand-dependent coregulator recruitment may be influenced by the sequence of the DNA to which RORγ is bound. Overall, the efforts described herein will illuminate important aspects of full length RORγ and monomeric orphan nuclear receptor target gene regulation through DNA-dependent conformational changes.