Cyclin G2 in macrophages triggers CTL-mediated antitumor immunity and antiangiogenesis via interferon-gamma.

BACKGROUND: IFN-γ is a key mediator of tumor immunity that can induce macrophage polarization to suppress tumor growth. Cyclin G2 functions as a tumor suppressor in various cancer cells; however, its role in macrophages remains unclear. This study aimed to investigate the role and underlying mechanisms of cyclin G2 in macrophages in vitro and in vivo. METHODS: Mouse tumor models were used to determine the effect of cyclin G2 in macrophages on tumor growth in vivo following IFN-γ treatment. Immunohistochemistry staining, immunofluorescence staining and flow cytometry were used to evaluate the number of cytotoxic T lymphocytes (CTLs) and blood vessels in the mouse tumors. Moreover, the biological roles of cyclin G2 in macrophages with regard to CTL chemotaxis, cytotoxic function, and vascular endothelial cell tube formation were assessed using in vitro functional experiments. Immunoprecipitation (IP), real-time PCR, and enzyme-linked immunosorbent assays (ELISAs) were conducted to investigate the underlying mechanisms by which cyclin G2 regulates CTLs and vascular endothelial cells. RESULTS: We found that cyclin G2 expression was upregulated in macrophages after IFN-γ treatment. Upregulated cyclin G2 inhibited lung and colon cancer growth by increasing the secretion of its downstream effector CXCL9, which promoted CTL chemotaxis and suppressed vascular endothelial cell tube formation. Moreover, cyclin G2 increased CXCL9 mRNA levels by promoting STAT1 nuclear translocation. In addition, cyclin G2 promoted the activation of the STAT1 signaling pathway, which was dependent on PP2Ac. CONCLUSIONS: Cyclin G2 is upregulated by IFN-γ in macrophages, promotes the secretion of CXCL9 to increase CTL chemotaxis and inhibit angiogenesis to suppress tumor growth. Our findings suggest that targeting cyclin G2 could benefit future immunotherapy.

Cyclin G2 reverses immunosuppressive tumor microenvironment and potentiates PD-1 blockade in glioma.

BACKGROUND: Expression of aberrant cyclin G2 is a key factor contributing to cancer biological processes, including glioma. However, the potential underlying mechanisms of cyclin G2 in the glioma tumor immune microenvironment remain unclear. METHODS: Co-immunoprecipitation (co-IP), in situ proximity ligation assay (PLA), and in vitro kinase assay were conducted to reveal the underlying mechanism by which cyclin G2 regulates Y10 phosphorylation of LDHA. Further, the biological roles of cyclin G2 in cell proliferation, migration, invasion capacity, apoptosis, glycolysis, and immunomodulation were assessed through in vitro and in vivo functional experiments. Expressions of cyclin G2 and Foxp3 in glioma specimens was determined by immunohistochemistry. RESULTS: In this study, we found that cyclin G2 impeded the interaction between LDHA and FGFR1, thereby decreasing Y10 phosphorylation of LDHA through FGFR1 catalysis. Cyclin G2 inhibited proliferation, migration, invasion capacity, and glycolysis and promoted apoptosis glioma cells via suppressing Y10 phosphorylation of LDHA. Moreover, we further verified that cyclin G2 reversed the immunosuppressive to antitumor immune microenvironment through inhibiting lactate production by glioma cells. Besides, cyclin G2 potentiated PD-1 blockade and exerted strong antitumor immunity in the glioma-bearing mice model. CONCLUSIONS: Cyclin G2 acts as a potent tumor suppressor in glioma and enhances responses to immunotherapy. Our findings may be helpful in selecting glioma patients for immunotherapy trials in the future.

Cyclin G2 Inhibits Oral Squamous Cell Carcinoma Growth and Metastasis by Binding to IGFBP3 and Regulating the FAK-SRC-STAT Signaling Pathway.

The cell cycle protein cyclin G2 is considered a tumor suppressor. However, its regulatory effects and potential mechanisms in oral cancers are not well understood. This study aimed to investigate the effect of cyclin G2 on oral squamous cell carcinoma (OSCC). The data from 80 patients with OSCC were utilized to predict the abnormal expression of cyclin G2. The proliferation and metastasis were determined by a cell counting Kit-8 assay, flow cytometry, a wound-healing assay, and a cell invasion assay. The expression of key proteins and genes associated with the cyclin G2 signaling pathways was determined by western blotting and real-time PCR, respectively. The orthotopic nude mice model was established by a mouth injection of SCC9 cells overexpressing cyclin G2. We showed that the low level of cyclin G2 in OSCC, which is negatively correlated with clinical staging, was a negative prognostic factor for the disease. We also found that cyclin G2 inhibited the proliferation, metastasis, and blocked the cell cycle at G1/S of OSCC cells, suggesting that cyclin G2 has an inhibitory effect in OSCC. Mechanistically, cyclin G2 inhibited the growth and metastasis of OSCC by binding to insulin-like growth factor binding protein 3 (IGFBP3) and regulating the focal adhesion kinase (FAK) -SRC-STAT signal transduction pathway. Cyclin G2 competed with integrin to bind to IGFBP3; the binding between integrin and IGFBP3 was reduced after cyclin G2 overexpression, thereby inhibiting the phosphorylation of FAK and SRC. These results showed that cyclin G2 inhibited the progression of OSCC by interacting with IGFBP3 and that it may be a new target for OSCC treatment.

Cyclin G2 regulates canonical Wnt signalling via interaction with Dapper1 to attenuate tubulointerstitial fibrosis in diabetic nephropathy.

Cyclin G2 (CCNG2) is an atypical cyclin that inhibits cell cycle progression and is often dysregulated in human cancers. Cyclin G2 in the occurrence and development of diabetic nephropathy (DN), one of the most severe diabetic complications, has not been fully identified. In this study, we investigated the function and regulatory mechanism of cyclin G2 in DN. In vivo studies revealed that a deficiency of cyclin G2 significantly increased albuminuria and promoted tubulointerstitial fibrosis in established DN. Cyclin G2 regulated the expression of fibrosis-related proteins via the canonical Wnt signalling pathway in renal tubular epithelial cells. Moreover, the binding of cyclin G2 to Dapper1 (Dpr1/DACT1), a protein involved in Wnt signalling, decreased the phosphorylation of Dpr1 at Ser762 by casein kinase 1 (CK1) and suppressed the Wnt signalling pathway. These findings reveal that cyclin G2 can protect against renal injury and fibrosis associated with DN and, thus, is a new target for the prevention and treatment of diabetic complications.

To control or to be controlled? Dual roles of CDK2 in DNA damage and DNA damage response.

CDK2 (cyclin-dependent kinase 2), a member of the CDK family, has been shown to play a role in many cellular activities including cell cycle progression, apoptosis and senescence. Recently, accumulating evidence indicates that CDK2 is involved in DNA damage and DNA repair response (DDR). When DNA is damaged by internal or external genotoxic stresses, CDK2 activity is required for proper DNA repair in vivo and in vitro, whereas inactivation of CDK2 by siRNA techniques or by inhibitors could result in DNA damage and stimulate DDR. Hence, CDK2 seems to play dual roles in DNA damage and DDR. On one aspect, it is activated and stimulates DDR to repair DNA damage when DNA damage occurs; on the other hand, its inactivation directly leads to DNA damage and evokes DDR. Here, we describe the roles of CDK2 in DNA damage and DDR, and discuss the potential application of CDK2 inhibitors as anti-cancer agents.

Cyclin G2 Inhibits the Warburg Effect and Tumour Progression by Suppressing LDHA Phosphorylation in Glioma.

Cyclin G2 has been identified as a tumour suppressor in several cancers. However, its regulatory roles and underlying mechanisms in tumours are still unknown. In this study, we demonstrated that cyclin G2 was expressed at low levels in glioma, which was as a poor prognostic factor for this disease. We also found that, cyclin G2 could suppress cell proliferation, initiate cell apoptosis and reduce aerobic glycolysis, suggesting that cyclin G2 plays a tumour suppressive role in glioma. Mechanistically, cyclin G2 could negatively regulate tyrosine-10 phosphorylation of a critical glycolytic enzyme, lactate dehydrogenase A, through direct interaction. Taken together, these results indicate that cyclin G2 acts as a tumour suppressor in glioma by repressing glycolysis and tumour progression through its interaction with LDHA.

Cyclin G2 suppresses Wnt/ß-catenin signaling and inhibits gastric cancer cell growth and migration through Dapper1.

BACKGROUND: Gastric cancer is one of the most common malignant tumors. Cyclin G2 has been shown to be associated with the development of multiple types of tumors, but its underlying mechanisms in gastric tumors is not well-understood. The aim of this study is to investigate the role and the underlying mechanisms of cyclin G2 on Wnt/ß-catenin signaling in gastric cancer. METHODS: Real-time PCR, immunohistochemistry and in silico assay were used to determine the expression of cyclin G2 in gastric cancer. TCGA datasets were used to evaluate the association between cyclin G2 expression and the prognostic landscape of gastric cancers. The effects of ectopic and endogenous cyclin G2 on the proliferation and migration of gastric cancer cells were assessed using the MTS assay, colony formation assay, cell cycle assay, wound healing assay and transwell assay. Moreover, a xenograft model and a metastasis model of nude mice was used to determine the influence of cyclin G2 on gastric tumor growth and migration in vivo. The effects of cyclin G2 expression on Wnt/ß-catenin signaling were explored using a TOPFlash luciferase reporter assay, and the molecular mechanisms involved were investigated using immunoblots assay, yeast two-hybrid screening, immunoprecipitation and Duolink in situ PLA. Ccng2-/- mice were generated to further confirm the inhibitory effect of cyclin G2 on Wnt/ß-catenin signaling in vivo. Furthermore, GSK-3ß inhibitors were utilized to explore the role of Wnt/ß-catenin signaling in the suppression effect of cyclin G2 on gastric cancer cell proliferation and migration. RESULTS: We found that cyclin G2 levels were decreased in gastric cancer tissues and were associated with tumor size, migration and poor differentiation status. Moreover, overexpression of cyclin G2 attenuated tumor growth and metastasis both in vitro and in vivo. Dpr1 was identified as a cyclin G2-interacting protein which was required for the cyclin G2-mediated inhibition of ß-catenin expression. Mechanically, cyclin G2 impacted the activity of CKI to phosphorylate Dpr1, which has been proved to be a protein that acts as a suppressor of Wnt/ß-catenin signaling when unphosphorylated. Furthermore, GSK-3ß inhibitors abolished the cyclin G2-induced suppression of cell proliferation and migration. CONCLUSIONS: This study demonstrates that cyclin G2 suppresses Wnt/ß-catenin signaling and inhibits gastric cancer cell growth and migration through Dapper1.

Cyclin G2 Suppresses Glomerulosclerosis by Regulating Canonical Wnt Signalling.

Recent data has shown that cyclin G2 (CCNG2) is an atypical cyclin that inhibits cell cycle progression and is often dysregulated in human cancers. The involvement of cyclin G2 in the occurrence and development of diabetic nephropathy (DN) has not been determined. In the present study, we conducted cyclin G2 knockout studies to determine whether this protein regulates glomerulosclerosis in DN mice. We found that cyclin G2 regulated the expression of renal glomerulosclerosis-related proteins via the canonical Wnt signalling pathway in glomerular mesangial cells. A cyclin G2 deficiency resulted in more severe renal injury in DN mice. These findings provided new insight into the pathogenesis of DN, revealing that cyclin G2 has a protective role in glomerulosclerosis and is a potential new target for the prevention and treatment of DN.

Cyclin G2 suppresses estrogen-mediated osteogenesis through inhibition of Wnt/ß-catenin signaling.

Estrogen plays an important role in the maintenance of bone formation, and deficiency in the production of estrogen is directly linked to postmenopausal osteoporosis. To date, the underlying mechanisms of estrogen-mediated osteogenic differentiation are not well understood. In this study, a pluripotent mesenchymal precursor cell line C2C12 was used to induce osteogenic differentiation and subjected to detection of gene expressions or to manipulation of cyclin G2 expressions. C57BL/6 mice were used to generate bilateral ovariectomized and sham-operated mice for analysis of bone mineral density and protein expression. We identified cyclin G2, an unconventional member of cyclin, is involved in osteoblast differentiation regulated by estrogen in vivo and in vitro. In addition, the data showed that ectopic expression of cyclin G2 suppressed expression of osteoblast transcription factor Runx2 and osteogenic differentiation marker genes, as well as ALP activity and in vitro extracellular matrix mineralization. Mechanistically, Wnt/ß-catenin signaling pathway is essential for cyclin G2 to inhibit osteogenic differentiation. To the best of our knowledge, the current study presents the first evidence that cyclin G2 serves as a negative regulator of both osteogenesis and Wnt/ß-catenin signaling. Most importantly, the basal and 17ß-estradiol-induced osteogenic differentiation was restored by overexpression of cyclin G2. These results taken together suggest that cyclin G2 may function as an endogenous suppressor of estrogen-induced osteogenic differentiation through inhibition of Wnt/ß-catenin signaling.

Overexpression of DOC-1R inhibits cell cycle G1/S transition by repressing CDK2 expression and activation.

DOC-1R (deleted in oral cancer-1 related) is a novel putative tumor suppressor. This study investigated DOC-1R antitumor activity and the underlying molecular mechanisms. Cell phenotypes were assessed using flow cytometry, BrdU incorporation and CDK2 kinase assays in DOC-1R overexpressing HeLa cells. In addition, RT-PCR and Western blot assays were used to detect underlying molecular changes in these cells. The interaction between DOC-1R and CDK2 proteins was assayed by GST pull-down and immunoprecipitation-Western blot assays. The data showed that DOC-1R overexpression inhibited G1/S phase transition, DNA replication and suppressed CDK2 activity. Molecularly, DOC-1R inhibited CDK2 expression at the mRNA and protein levels, and there were decreased levels of G1-phase cyclins (cyclin D1 and E) and elevated levels of p21, p27, and p53 proteins. Meanwhile, DOC-1R associated with CDK2 and inhibited CDK2 activation by obstructing its association with cyclin E and A. In conclusion, the antitumor effects of DOC-1R may be mediated by negatively regulating G1 phase progression and G1/S transition through inhibiting CDK2 expression and activation.

When MAGE meets RING: insights into biological functions of MAGE proteins.

The melanoma antigen (MAGE) family proteins are well known as tumor-specific antigens and comprise more than 60 genes, which share a conserved MAGE homology domain (MHD). Type I MAGEs are highly expressed cancer antigens, and they play an important role in tumorigenesis and cancer cell survival. Recently, several MAGE proteins were identified to interact with RING domain proteins, including a sub-family of E3 ubiquitin ligases. The binding mode between MAGEs and RING proteins was investigated and one important structure of these MAGE-RING complexes was solved: the MAGE-G1-NSE1 complex. Structural and biochemical studies indicated that MAGE proteins could adjust the E3 ubiquitin ligase activity of its cognate RING partner both in vitro and in vivo. However, the underlying mechanism was not fully understood. Here, we review these exciting advances in the studies on MAGE family, suggest potential mechanisms by which MAGEs activate the E3 activity of their binding RING proteins and highlight the anticancer potential of this family proteins.

MAGE-RING protein complexes comprise a family of E3 ubiquitin ligases.

The melanoma antigen (MAGE) family consists of more than 60 genes, many of which are cancer-testis antigens that are highly expressed in cancer and play a critical role in tumorigenesis. However, the biochemical and cellular functions of this enigmatic family of proteins have remained elusive. Here, we identify really interesting new gene (RING) domain proteins as binding partners for MAGE family proteins. Multiple MAGE family proteins bind E3 RING ubiquitin ligases with specificity. The crystal structure of one of these MAGE-RING complexes, MAGE-G1-NSE1, reveals structural insights into MAGE family proteins and their interaction with E3 RING ubiquitin ligases. Biochemical and cellular assays demonstrate that MAGE proteins enhance the ubiquitin ligase activity of RING domain proteins. For example, MAGE-C2-TRIM28 is shown to target p53 for degradation in a proteasome-dependent manner, consistent with its tumorigenic functions. These findings define a biochemical and cellular function for the MAGE protein family.