G1 Cyclins Research Articles

TQ inhibits hepatocellular carcinoma growth in vitro and in vivo via repression of Notch signaling.

Thymoquinone (TQ) has been reported to possess anti-tumor activity in various types of cancer. However, its effects and molecular mechanism of action in hepatocellular carcinoma (HCC) are still not completely understood. We observed that TQ inhibited tumor cell growth in vitro, where treatment with TQ arrested the cell cycle in G1 by upregulating p21 and downregulating cyclinD1 and CDK2 expression; moreover, TQ induced apoptosis by decreasing expression of Bcl-2 and increasing expression of Bax. Simultaneously, TQ demonstrated a suppressive impact on the Notch pathway, where overexpression of NICD1 reversed the inhibitory effect of TQ on cell proliferation, thereby attenuating the repressive effects of TQ on the Notch pathway, cyclinD1, CDK2 and Bcl-2, and also diminishing upregulation of p21 and Bax. In a xenograft model, TQ inhibited HCC growth in nude mice; this inhibitory effect in vivo, as well as of HCC cell growth in vitro, was associated with a discernible decline in NICD1 and Bcl-2 levels and a dramatic rise in p21 expression. In conclusion, TQ inhibits HCC cell growth by inducing cell cycle arrest and apoptosis, achieving these effects by repression of the Notch signaling pathway, suggesting that TQ represents a potential preventive or therapeutic agent in HCC patients.

Dilution of the cell cycle inhibitor Whi5 controls budding-yeast cell size.

Cell size fundamentally affects all biosynthetic processes by determining the scale of organelles and influencing surface transport1,2. Although extensive studies have identified many mutations affecting cell size, the molecular mechanisms underlying size control have remained elusive3. In budding yeast, size control occurs in G1 phase prior to Start, the point of irreversible commitment to cell division4,5. It was previously thought that activity of the G1 cyclin Cln3 increased with cell size to trigger Start by initiating the inhibition of the transcriptional inhibitor Whi56-8. However, while Cln3 concentration does modulate the rate at which cells pass Start, we found that its synthesis increases in proportion to cell size so that its total concentration is nearly constant during pre-Start G1. Rather than increasing Cln3 activity, we identify decreasing Whi5 activity — due to the dilution of Whi5 by cell growth — as a molecular mechanism through which cell size controls proliferation. Whi5 is synthesized in S/G2/M phases of the cell cycle in a largely size-independent manner. This results in smaller daughter cells being born with higher Whi5 concentrations that extend their pre-Start G1 phase. Thus, at its most fundamental level, budding yeast size control results from the differential scaling of Cln3 and Whi5 synthesis rates with cell size. More generally, our work shows that differential size-dependency of protein synthesis can provide an elegant mechanism to coordinate cellular functions with growth.

TORC1 controls G1–S cell cycle transition in yeast via Mpk1 and the greatwall kinase pathway

The target of rapamycin complex 1 (TORC1) pathway couples nutrient, energy and hormonal signals with eukaryotic cell growth and division. In yeast, TORC1 coordinates growth with G1–S cell cycle progression, also coined as START, by favouring the expression of G1 cyclins that activate cyclin-dependent protein kinases (CDKs) and by destabilizing the CDK inhibitor Sic1. Following TORC1 downregulation by rapamycin treatment or nutrient limitation, clearance of G1 cyclins and C-terminal phosphorylation of Sic1 by unknown protein kinases are both required for Sic1 to escape ubiquitin-dependent proteolysis prompted by its flagging via the SCFCdc4 (Skp1/Cul1/F-box protein) ubiquitin ligase complex. Here we show that the stabilizing phosphorylation event within the C-terminus of Sic1 requires stimulation of the mitogen-activated protein kinase, Mpk1, and inhibition of the Cdc55 protein phosphatase 2A (PP2ACdc55) by greatwall kinase-activated endosulfines. Thus, Mpk1 and the greatwall kinase pathway serve TORC1 to coordinate the phosphorylation status of Sic1 and consequently START with nutrient availability.

Cyclin G2: A novel independent prognostic marker in pancreatic cancer.

Unlike other cyclins that positively regulate the cell cycle, cyclin G2 (CCNG2) regulates cell proliferation as a tumor suppressor gene. A decreased CCNG2 expression serves as a marker for poor prognosis in several types of cancer. The aim of the present study was to clarify the correlation of CCNG2 expression with overall survival and histopathological factors in pancreatic cancer patients. This retrospective analysis included data from 36 consecutive patients who underwent complete surgical resection for pancreatic cancer and did not undergo any preoperative therapies. The association between prognoses and the expression of CCNG2 was assessed using immunohistochemical staining. Multivariate analysis identified that the expression of CCNG2 is an independent prognostic factor. In addition, the Kaplan-Meier curve for overall survival revealed that decreased expression of CCNG2 was a consistent indicator of poor prognosis in pancreatic cancer patients (P=0.0198). A decreased CCNG2 expression significantly correlated with venous invasion in tumor specimens and the tumor invasion depth. In conclusion, CCNG2 expression inversely reflected cancer progression and may be a novel, independent prognostic marker in pancreatic cancer.

Upregulation of microRNA-122 by farnesoid X receptor suppresses the growth of hepatocellular carcinoma cells.

BackgroundmicroRNA-122 (miR-122) is the most abundant and specific miRNA in the liver. It acts as an important tumor suppressor in hepatocellular carcinoma (HCC) through regulating its target genes, but details of its own regulation are largely unknown. Farnesoid X receptor (FXR), a transcription factor with multiple functions, plays an important role in protecting against liver carcinogenesis, but it is unclear whether the anti-HCC effect of FXR is involved in the regulation of miR-122.MethodsThe levels of miR-122 and FXR in HCC tissues and cell lines were examined by quantitative real-time PCR (qRT-PCR). qRT-PCR was also used to detect the expression of miR-122 target genes at mRNA level, while Western blotting was used to analyze that of their protein products. The effect of FXR on the transcriptional activity of miR-122 promoter was evaluated by a luciferase reporter assay. Electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) assay were performed to identify the FXR binding site within miR-122 promoter region. The cell proliferation was analyzed by a CCK-8 assay. The influence of FXR on tumor growth and miR-122 expression in vivo was monitored using HCC xenografts in nude mice.ResultsThe expression of FXR was positively correlated with that of miR-122 in HCC tissues and cell lines. Activation of FXR in HCC cells upregulated miR-122 expression and in turn downregulated the expression of miR-122 target genes including insulin-like growth factor-1 receptor and cyclin G1. FXR bound directly to the DR2 element (−338 to −325) in miR-122 promoter region, and enhanced the promoter’s transcriptional activity. Functional experiments showed that the FXR-mediated upregulation of miR-122 suppressed the proliferation of HCC cells in vitro and the growth of HCC xenografts in vivo.ConclusionsmiR-122 is a novel target gene of FXR, and the upregulation of miR-122 by FXR represses the growth of HCC cells, suggesting that FXR may serve as a key transcriptional regulator for manipulating miR-122 expression, and the FXR/miR-122 pathway may therefore be a novel target for the treatment of HCC.Electronic supplementary materialThe online version of this article (doi:10.1186/s12943-015-0427-9) contains supplementary material, which is available to authorized users.

Abstract 4787: Cyclin G2, a novel target of the SNF5/BAF47 tumor suppressor gene

Abstract Malignant Rhabdoid Tumor (MRT) is a rare and aggressive pediatric cancer that most frequently appears in the kidney (RTK) and brain (AT/RT). They are generally characterized by the lack of SNF5 (SMARCB1/INI1) expression, a subunit of the SWI/SNF chromatin remodeling complex. Previous studies have shown that reintroducing SNF5 into MRT cell lines reconstitutes the SWI/SNF complex and results in an inhibition of growth through induction of p21WAF1/CIP1, p16INK4A and p57KIP2 expression. However, the mechanisms behind this growth arrest remain incompletely characterized. In this current report, we used a RT-PCR Cell Cycle SuperArray to identify six candidate genes that showed increased expression after SNF5 reexpression but not after expression of a constitutively active RB gene. One of these genes, Cyclin G2 (CCNG2), a member of the non-canonical Cyclin G family, is thought to act as a negative regulator of the cell cycle. Using a panel of RTK and AT/RT cell lines, we confirmed that SNF5 reexpression leads to induction of CCGN2 in additional MRT cell lines. To determine if CCNG2 is a direct target of SNF5, we used chromatin immunoprecipitation analyses (ChIP) to verify that SNF5 binds to the CCNG2 promoter with peak binding at the transcription start site (TSS) after its reexpression in a MRT cell line. Importantly, primary MRT samples display reduced CCNG2 expression when compared to normal brain tissue or other types of pediatric brain cancers. Therefore, CCNG2 represents a new SNF5 target gene whose downregulation may play a role during MRT development. Citation Format: Darmood Wei, Aubri Charboneau, Ho-yoon Chung, Donastas Sakellariou-Thompson, Brian Davies, Dennis A. Simpson, Yasumichi Kuwahara, William K. Kaufmann, Charles W. M. Roberts, Bernard E. Weissman. Cyclin G2, a novel target of the SNF5/BAF47 tumor suppressor gene. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4787. doi:10.1158/1538-7445.AM2015-4787

Abstract 2601: A natural small molecule, catechol, induces c-Myc degradation by directly targeting ERK2 in lung cancer in vitro and in vivo

Abstract For decades, lung cancer has been a leading cause of death in the U.S., mainly due to oncogenic signaling pathways activated by carcinogens such as smoking. Various carcinogens induce EGFR/RAS/MAPK signaling that is critical in the development of lung cancer. In particular, constitutive activation of extracellular signal-regulated kinase 2 (ERK2) is observed in many lung cancer patients, and therefore developing compounds capable of targeting ERK2 against lung carcinogenesis will be beneficial to these patients. Herein, we examined the therapeutic effects of catechol (also known as pyrocatechol or 1,2-dihydroxybenzene), a naturally occurring chemical in many foods such as apple, banana, apricot, grape, plum, avocado and mushroom. Co-crystallography results suggested that ERK2 is a direct target of catechol. In agreement with the co-crystallography result, catechol was confirmed to inhibit ERK2 kinase activity in vitro. Catechol inhibited anchorage-independent KP2 and H460 lung cancer cell growth in a dose-dependent manner. Phosphorylation of c-Myc, a substrate of ERK2, was reduced in catechol-treated lung cancer cells compared to untreated controls. Decreased c-Myc phosphorylation resulted in its reduced protein stability and subsequent down-regulation of total c-Myc levels. Treatment with catechol induced G1 cell cycle arrest and resulted in a decrease of G1 phase related proteins, CDK2 and cyclin D1. In vivo study results also revealed that catechol inhibited growth of KP2 and H460 xenograft tumors. Phosphorylation levels of c-Myc were also strongly decreased by catechol treatment in vivo. Taken together, we have shown that catechol exerts inhibitory effects on the ERK2/c-Myc signaling axis in lung cancer cells to reduce tumor growth in vitro and in vivo. These findings suggest that catechol, a natural small molecule, possesses potential as a novel therapeutic agent against lung carcinogenesis. Citation Format: Do Young Lim, Seung Ho Shin, Margarita Malakhova, Mee-Hyun Lee, Ann M. Bode, Zigang Dong. A natural small molecule, catechol, induces c-Myc degradation by directly targeting ERK2 in lung cancer in vitro and in vivo. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2601. doi:10.1158/1538-7445.AM2015-2601

Neisseria meningitidis causes cell cycle arrest of human brain microvascular endothelial cells at S phase via p21 and cyclin G2.

Microbial pathogens have developed several mechanisms to modulate and interfere with host cell cycle progression. In this study, we analysed the effect of the human pathogen Neisseria meningitidis on cell cycle in a brain endothelial cell line as well as in primary brain endothelial cells. We found that N. Meningitidis causes an accumulation of cells in the S phase early at 3 and at 24 h post-infection that was paralleled by a decrease of cells in G2/M phase. Importantly, the outer membrane proteins of the colony opacity-associated (Opa) protein family as well as the Opc protein proved to trigger the accumulation of cells in the S phase. A focused cell cycle reverse transcription quantitative polymerase chain reaction-based array and integrated network analysis revealed changes in the abundance of several cell cycle regulatory mRNAs, including the cell cycle inhibitors p21(WAF1/CIP1) and cyclin G2. These alterations were reflected in changes in protein expression levels and/or relocalization in N. meningitidis-infected cells. Moreover, an increase in p21(WAF1/CIP1) expression was found to be p53 independent. Genetic ablation of p21(WAF1/CIP1) and cyclin G2 abrogated N. meningitidis-induced S phase accumulation. Finally, by measuring the levels of the biomarker 8-hydroxydeoxyguanosine and phosphorylation of the histone variant H2AX, we provide evidence that N. meningitidis induces oxidative DNA damage in infected cells.

Tannic acid inhibits the Jak2/STAT3 pathway and induces G1/S arrest and mitochondrial apoptosis in YD-38 gingival cancer cells.

Tannic acid (TA), is a potent anti-oxidant, showing anti-proliferative effects on numerous cancers. The ability of TA to induce proliferation inhibition on the rare tumor, gingival squamous cell carcinoma (GSCC), comprising <10% of all head and neck squamous cell carcinomas was studied in the YD-38 cell line. The main goal was to modulate the Jak2/STAT3 pathway using TA and to induce cell cycle arrest and apoptosis in GSCC. TA treatment induced G1 arrest and apoptosis in YD-38 cells. Molecular analysis revealed that TA inhibits Jak2/STAT3 pathway by preventing their expression as well as phosphorylation. This inhibition of STAT3 phosphorylation prevented the nuclear translocation and DNA binding capability of STAT3. Together with the inhibition of transcriptional regulatory function of STAT3, TA inhibited the expression of G1 phase modulators CDK-4, CDK-6, cyclinD1 and cyclinE. It is also evidenced that TA exerted an intense activation of p21Waf1/Cip1, p27Kip1 and p53 genes confirming its role in G1 phase inhibition. Additionally, upon treatment with TA, the expression of mitochondrial pore factors Bax, Bcl-2 and Bcl-XL were changed. We observed inhibition of Bcl-2 and an increase in mitochondrial localization of Bax leading to the loss of mitochondrial membrane potential, resulting in the release of cytochrome c to the cytosol. In addition, we perceived the activation of caspases upon TA treatment. Specific inhibition of caspase protected the cells from TA induced apoptosis. Taken together, this study reveals that TA significantly inhibits the Jak2/STAT3 signaling pathway and induces G1 arrest and mitochondrial apoptosis in YD-38 cells.

A cross-talk between Hepatitis B virus and host mRNAs confers viral adaptation to liver.

Hepatitis B virus (HBV) chronically infects approximately 350 million people worldwide. The replication of HBV which genome is only 3.2 kb long relies heavily on host factors. Previous studies demonstrated that a highly expressed liver-specific microRNA (miRNA) miR-122 suppresses HBV expression and replication in multiple ways. In this study, we found that the miR-122 response elements in viral genome facilitate HBV expression and replication in miR-122 highly-expressed hepatocytes. Moreover, mutations in miR-122 response elements are correlated with viral loads and disease progression in HBV-infected patients. We next found that HBV mRNA with miR-122 response elements alone could lead to altered expression of multiple host genes by whole genome expression analysis. HBV mRNA-mediated miR-122 down-regulation plays a major role in HBV mRNA-induced differential gene expression. HBV mRNA could enhance viral replication via miR-122 degradation and the up-regulation of its target cyclin G1. Our study thereby reveals that under the unique condition of high abundance of miR-122 and viral mRNAs and much lower level of miR-122 target in HBV infection, HBV may have evolved to employ the miRNA-mediated virus and host mRNAs network for optimal fitness within hepatocytes.

Arsenic-induced S phase cell cycle lengthening is associated with ROS generation, p53 signaling and CDC25A expression

Cellular response to arsenic is strongly dependent on p53 functional status. Primarily arresting the cell cycle in G1 or G2/M phases, arsenic treatment also induces an increase in the S-phase time in wild-type p53 cells. In contrast, cells with a non-functional p53 display only a subtle increase in the S phase, indicating arsenic differentially affects the cell cycle depending on p53 status. Importantly, it has been reported that arsenic induces reactive oxygen species (ROS), a process counteracted by p53. To evaluate the participation of p53 in the lengthening of the S phase and the connection between the transient cell cycle arrest and oxidative stress, we evaluated the cell response to arsenic in MCF-7 and H1299 cells, and analyzed p53’s role as a transcription factor in regulating genes involved in ROS reduction and S phase transition. Herein, we discovered that arsenic induced an increase in the population of S phase cells that was dependent on the presence and transcriptional activity of p53. Furthermore, for the first time, we demonstrate that arsenic activates p53-dependent transcription of ROS detoxification genes, such as SESN1, and by an indirect mechanism involving ATF3, genes that could be responsible for the S phase cell cycle arrest, such as CDC25A.

Induction of cell death in prostate cancer cells by escopoletin, a promising treatment strategy

&lt;p&gt;Escopoletin, a phenolic compound belonging to anthocyanin family shows promising antioxidant activities. In the present study, anti-cancer effects of escopoletin treatment in DU145 cells were investigated. The sulphorhodamine-B staining and annexin V and propidium iodide were respectively used for the analysis of cell viability and death. The results revealed a significantly higher cytotoxicity by escopoletin that caused cell death in DU145 cells. Escopoletin treatment in DU145 cells markedly inhibited cell growth through non-apoptotic cell death and induced significant reactive oxygen species (ROS) production. It also induced G1 cell cycle arrest and cyclin D1 accumulation through the enhanced expression of p21. However, the effect of escopoletin on DU145 cells was reversed by pretreatment with glutathione antioxidant. This suggests that escopoletin induced generation of ROS is responsible for the increased cytotoxicity in DU145 cells. Thus, escopoletin exhibits potential therapeutic efficacy for the treatment of prostate cancer.&lt;/p&gt;&lt;p&gt; &lt;/p&gt;

Vitamin K2 and cotylenin A synergistically induce monocytic differentiation and growth arrest along with the suppression of c-MYC expression and induction of cyclin G2 expression in human leukemia HL-60 cells.

Although all-trans retinoic acid (ATRA) is a standard and effective drug used for differentiation therapy in acute promyelocytic leukemia, ATRA-resistant leukemia cells ultimately emerge during this treatment. Therefore, the development of new drugs or effective combination therapy is urgently needed. We demonstrate that the combined treatment of vitamin K2 and cotylenin A synergistically induced monocytic differentiation in HL-60 cells. This combined treatment also synergistically induced NBT-reducing activity and non-specific esterase-positive cells as well as morphological changes to monocyte/macrophage-like cells. Vitamin K2 and cotylenin A cooperatively inhibited the proliferation of HL-60 cells in short-term and long-term cultures. This treatment also induced growth arrest at the G1 phase. Although 5 μg/ml cotylenin A or 5 μM vitamin K2 alone reduced c-MYC gene expression in HL-60 cells to approximately 45% or 80% that of control cells, respectively, the combined treatment almost completely suppressed c-MYC gene expression. We also demonstrated that the combined treatment of vitamin K2 and cotylenin A synergistically induced the expression of cyclin G2, which had a positive effect on the promotion and maintenance of cell cycle arrest. These results suggest that the combination of vitamin K2 and cotylenin A has therapeutic value in the treatment of acute myeloid leukemia.

The role of Cyclin G1 in cellular proliferation and apoptosis of human epithelial ovarian cancer.

Cyclin G1 plays an essential role in the development of human carcinoma. Here, we characterized the clinical significance of Cyclin G1 and investigated its role in cellular proliferation and apoptosis of epithelial ovarian cancer (EOC). Western blot was used to evaluate the expression of Cyclin G1 in nine fresh EOC tissues and three fresh normal ovarian tissues. Immunohistochemistry analysis was performed on formalin-fixed paraffin-embedded section of 119 cases of EOCs. Using cell counting kit (CCK)-8 and colony formation assays, we analyzed the effect of Cyclin G1 in cellular proliferation of EOC. Besides, the immunofluorescence and flow cytometry analysis was performed to study the role of Cyclin G1 in cellular apoptosis of EOC. We found Cyclin G1 was up-regulated in EOC tissues compared with the normal ovary tissues. Cyclin G1 expression in EOC was closely correlated with differentiation grade (P = 0.009) and malignant tumor cells in ascites (P = 0.009). The Kaplan-Meier curve showed that higher expression of Cyclin G1 was associated with significantly shorter survival in EOC patients. Multivariate analysis suggested Cyclin G1 expression was an independent prognostic factor for overall survival. CCK-8 and colony formation assays revealed that depletion of Cyclin G1 inhibited the proliferation and clone formation. Combined immunofluorescence and flow cytometry analysis showed that silencing of Cyclin G1 with shRNA could promote apoptosis of ovarian cancer cells. Additionally, the result of immunoprecipitation test showed Cyclin G1 interacted with CDK2 in EOC cells. In summary, our findings suggest that Cyclin G1 may be involved in the prognosis of EOC patients and be a useful therapeutic target for EOC.

A eudesmane-type sesquiterpene isolated from Pluchea odorata (L.) Cass. combats three hallmarks of cancer cells: Unrestricted proliferation, escape from apoptosis and early metastatic outgrowth in vitro

Pluchea odorata is ethno pharmaceutically used to treat inflammation-associated disorders. The dichloromethane extract (DME) was tested in the carrageenan-induced rat paw oedema assay investigating its effect on inflammation that was inhibited by 37%. Also an in vitro anti-neoplastic potential was reported. However, rather limited information about the bio-activity of purified compounds and their cellular mechanisms are available. Therefore, two of the most abundant eudesmanes in P. odorata were isolated and their anti-neoplastic and anti-intravasative activities were studied. HL-60 cells were treated with P. odorata compounds and metabolic activity, cell number reduction, cell cycle progression and apoptosis induction were correlated with relevant protein expression. Tumour cell intravasation through lymph endothelial monolayers was measured and potential causal mechanisms were analyzed by Western blotting. Compound PO-1 decreased the metabolic activity of HL-60 cells (IC50=8.9μM after 72h) and 10μM PO-1 induced apoptosis, while PO-2 showed just weak anti-neoplastic activities at concentrations beyond 100μM. PO-1 arrested the cell cycle in G1 and this correlated with induction of JunB expression. Independent of this mechanism 25μM PO-1 decreased MCF-7 spheroid intravasation through the lymph endothelial barrier. Hence, PO-1 inhibits an early step of metastasis, impairs unrestricted proliferation and induces apoptosis at low micromolar concentrations. These results warrant further testing in vivo to challenge the potential of PO-1 as novel lead compound.

Hog1 targets Whi5 and Msa1 transcription factors to downregulate cyclin expression upon stress.

Yeast cells have developed complex mechanisms to cope with extracellular insults. An increase in external osmolarity leads to activation of the stress-activated protein kinase Hog1, which is the main regulator of adaptive responses, such as gene expression and cell cycle progression, that are essential for cellular survival. Upon osmostress, the G1-to-S transition is regulated by Hog1 through stabilization of the cyclin-dependent kinase inhibitor Sic1 and the downregulation of G1 cyclin expression by an unclear mechanism. Here, we show that Hog1 interacts with and phosphorylates components of the core cell cycle transcriptional machinery such as Whi5 and the coregulator Msa1. Phosphorylation of these two transcriptional regulators by Hog1 is essential for inhibition of G1 cyclin expression, for control of cell morphogenesis, and for maximal cell survival upon stress. The control of both Whi5 and Msa1 by Hog1 also revealed the necessity for proper coordination of budding and DNA replication. Thus, Hog1 regulates G1 cyclin transcription upon osmostress to ensure coherent passage through Start.

ELAS1-mediated inhibition of the cyclin G1-B'γ interaction promotes cancer cell apoptosis via stabilization and activation of p53.

Radiation therapy (RT) is useful for selectively killing cancer cells. However, because high levels of ionizing radiation (IR) are toxic to normal cells, RT cannot be applied repeatedly to cancer patients. Therefore, novel chemicals that enhance the efficacy of chemoradiotherapy (CRT) would be valuable. Here, we report that ELAS1, a peptide corresponding to the protein phosphatase 2A (PP2A) association domain of cyclin G1 (CycG1), can enhance the efficacy of CRT. ELAS1 interacts with the PP2A B'γ-subunit and competitively inhibits association with CycG1, thereby preventing the PP2A holoenzyme from dephosphorylating target proteins, Mdm2 (pT218) and p53 (pS46), following DNA double-strand break (DSB) insults. Doxycycline (Dox)-induced overexpression of Myc-ELAS1 caused γ-irradiation to induce apoptosis in human osteosarcoma (U2OS) cells, at 1/10th the effective dosage of γ-irradiation required for apoptosis in Myc-vector-expressing cells; ELAS1 peptide incorporation into U2OS cells also showed similar apoptotic effects. Moreover, administration of DSB-inducing chemicals, camptothecin (CPT) or irinotecan, to Myc-ELAS1-expressing U2OS cells also induced efficient apoptosis with only 1/100th (CPT) or 1/5th (irinotecan) of the amounts of drugs required for this effect in Myc-vector-expressing cells. Taken together, ELAS1 may be important for the design of ELAS1-mimetic compounds to improve CRT efficacy.

Bud-Localization of CLB2 mRNA Can Constitute a Growth Rate Dependent Daughter Sizer

Maintenance of cellular size is a fundamental systems level process that requires balancing of cell growth with proliferation. This is achieved via the cell division cycle, which is driven by the sequential accumulation and destruction of cyclins. The regulatory network around these cyclins, particularly in G1, has been interpreted as a size control network in budding yeast, and cell size as being decisive for the START transition. However, it is not clear why disruptions in the G1 network may lead to altered size rather than loss of size control, or why the S-G2-M duration also depends on nutrients. With a mathematical population model comprised of individually growing cells, we show that cyclin translation would suffice to explain the observed growth rate dependence of cell volume at START. Moreover, we assess the impact of the observed bud-localisation of the G2 cyclin CLB2 mRNA, and find that localised cyclin translation could provide an efficient mechanism for measuring the biosynthetic capacity in specific compartments: The mother in G1, and the growing bud in G2. Hence, iteration of the same principle can ensure that the mother cell is strong enough to grow a bud, and that the bud is strong enough for independent life. Cell sizes emerge in the model, which predicts that a single CDK-cyclin pair per growth phase suffices for size control in budding yeast, despite the necessity of the cell cycle network around the cyclins to integrate other cues. Size control seems to be exerted twice, where the G2/M control affects bud size through bud-localized translation of CLB2 mRNA, explaining the dependence of the S-G2-M duration on nutrients. Taken together, our findings suggest that cell size is an emergent rather than a regulatory property of the network linking growth and proliferation.

Cyclin G2 Contributes to the Cell Cycle Arrest Response of Breast Cancer Cells to Estrogen Signaling‐Antagonists and the AMPK Agonist, Metformin

Resistance to estrogen (E2) receptor (ER) antagonists limits long‐term abatement of ER+ breast cancer (BC). Defining cell cycle control proteins that modify tumor cell resistance to ER‐antagonists tamoxifen and fulvestrant will improve therapeutic approaches. Cyclin G2 (CycG2) is encoded by the ER repressed gene, CCNG2, and upregulated during cell cycle arrest responses to cellular stresses. Here we show that blockade of ER‐signaling enhances the expression and nuclear localization of CycG2 in E2‐dependent BC cell lines. Knockdown of CycG2 in E2‐deprived and fulvestrant‐treated MCF7 cells dampened their cell cycle arrest response. Moreover, loss of CycG2 increases phospho‐activation of MEK1 and inhibition of RB. Our work also indicates that CycG2 forms complexes with the CDK linked to inhibition of tamoxifen resistance and modulation of RAF/MEK/MAPK signaling, CDK10. We reported that signaling through insulin and IGF‐1R receptors strongly represses CycG2 expression. Recent studies suggest the insulin‐sensitizer metformin (MTFN) inhibits BC cell growth by promoting AMPK–mediated suppression of mTOR. Patients taking MTFN exhibit a dose‐dependent reduction in cancer risk. We found that MTFN treatment stimulates CycG2 expression and potentiates fulvestrant‐mediated upregulation of CycG2 and growth arrest of MCF7 cells. Moreover knockdown of CycG2 blunts MTFN‐enhancement of fulvestrant effects. Importantly, analysis of BC tumor microarrays indicates that CCNG2 transcripts are reduced in aggressive, poor‐prognosis BC and that high CCNG2 expression correlates with longer relapse free survival.

Molecular cloning and tissue expression of the CCNG1 gene in sheep

The CCNG1 gene encodes cyclin G1, which is an important cell cycle regulator and has been reported to be involved in reproductive biological processes, such as oocyte maturation and granule cell proliferation in mammals. But the study of CCNG1 in sheep has been rarely reported. To examine the effects of CCNG1 on estrous control and seasonal breeding in sheep, we first cloned and characterized the expression level of the sheep CCNG1 gene. Then by Real-time PCR, we detected and analyzed the expressions of CCNG1 gene at mRNA levels in the hypothalamus-pituitary-ovary (HPO) axis in different stages of an estrous cycle in Duo Lang sheep (non-seasonal breeding) and Merino sheep (seasonal breeding). The results showed that the open reading frame of the sheep CCNG1 gene is 885 bp in length and encodes 294 amino acids. Bioinformatic analysis indicated that the secondary structure of the sheep CCNG1 protein contained multiple phosphorylation sites and some Protein Kinase C phosphorylation sites. CCNG1 mRNA was identified in all tissues tested, with the levels in ovary and kidney higher than others. The expression profiles of CCNG1 in the HPO axis in different stages of an estrous cycle were similar in different sheep breeds: the expression levels of CCNG1 in the ovary, uterus, pineal gland and pituitary gland all peaked in the estrus phase. But there were significant differences for expression change extent of CCNG1 in ovaries in the oestrus and metestrus phase between different sheep breeds. The results suggested that CCNG1 probably participated in the regulation of estrous behavior and seasonal reproduction through controling the growth and development of follicles in sheep.