Inhibition of Proteasomal Degradation of Rpn4 Impairs Nonhomologous End-Joining Repair of DNA Double-Strand Breaks

BACKGROUND: The proteasome homeostasis in Saccharomyces cerevisiae is regulated by a negative feedback circuit in which the transcription factor Rpn4 induces the proteasome genes and is rapidly degraded by the assembled proteasome. The integrity of the Rpn4-proteasome feedback loop is critical for cell viability under stressed conditions. We have demonstrated that inhibition of Rpn4 degradation sensitizes cells to DNA damage, particularly in response to high doses of DNA damaging agents. The underlying mechanism, however, remains unclear. METHODOLOGY/PRINCIPAL FINDINGS: Using yeast genetics and biochemical approach we show that inhibition of Rpn4 degradation displays a synthetic growth defect with deletion of the MEC1 checkpoint gene and sensitizes several checkpoint mutants to DNA damage. In addition, inhibition of Rpn4 degradation leads to a defect in repair of double-strand breaks (DSBs) by nonhomologous end-joining (NHEJ). The expression levels of several key NHEJ genes are downregulated and the recruitment of Yku70 to a DSB is reduced by inhibition of Rpn4 degradation. We find that Rpn4 and the proteasome are recruited to a DSB, suggesting their direct participation in NHEJ. Inhibition of Rpn4 degradation may result in a concomitant delay of release of Rpn4 and the proteasome from a DSB. CONCLUSION/SIGNIFICANCE: This study provides the first evidence for the role of proteasomal degradation of Rpn4 in NHEJ

Treatment of Unruptured Vertebral Artery Aneurysm Involving Posterior Inferior Cerebellar Artery With Pipeline Embolization Device

Background: Treatment of unruptured vertebral artery aneurysm involving posterior inferior cerebellar artery (PICA) is challenging. The experience of pipeline embolization device (PED) therapy for these lesions is still limited.Objective: To evaluate the safety and efficacy of the PED for unruptured vertebral artery aneurysm involving PICA.Methods: Thirty-two patients with unruptured vertebral artery aneurysm involving PICA underwent treatment with PED were retrospectively identified. Procedure-related complications, PICA patency, clinical, and angiographic outcomes were analyzed.Results: Thirty-two aneurysms were successfully treated without any procedure-related complications. Images were available in 30 patients (93.8%) during a period of 3–26 months follow-up (average 8.4 months), which confirmed complete occlusion in 17 patients (56.5%), near-complete occlusion in 9 patients (30%), and incomplete occlusion in one patient (3.3%). Parent artery occlusion (PAO) was occurred in 3 patients (10%). Twenty-eight of 30 PICA remained patent. The two occlusions of PICA were secondary to PAO. At a mean of 20.7 months (range 7–50 months) clinical follow-up, all the patients achieved a favorable outcome without any new neurological deficit.Conclusion: PED seems to be a safe and effective alternative endovascular option for patients with unruptured vertebral artery aneurysm involving PICA

Effects of FBXW7α on Expression and Localization of Heat Shock Transcription Factor 1 in Colorectal Cancer Cells

Objective To investigate the effect of FBXW7 on the expression and localization of HSF1 in colorectal cancer cells. Methods The expression levels of HSF1 and pHSF1Ser326 protein in FBXW7 deletion (KO) and wild-type (WT) FBXW7-expressing counterpart colorectal cancer cells were detected by Western blot. The nucleoprotein expression and localization of pHSF1Ser326 in heat-shocked or recovery stage cells were observed by Western blot and immunofluorescence method. Results The HSF1 expression level in DLD1 cells transfected with FBXW7α was decreased significantly (P < 0.01). The expression levels of HSF1 and pHSF1Ser326 protein in FBXW7 KO cells were higher than those in WT cells (all P < 0.05). HSF1 and pHSF1Ser326 in FBXW7 KO cells were mainly expressed in nucleus and weakly expressed in cytoplasm. After warm stimulation, the expression of HSF1 and pHSF1Ser326 in WT cells recovered to the unstimulated level, while the expression of HSF1 and pHSF1Ser326 in FBXW7 KO cells were higher in the nucleus (all P < 0.01). Conclusion Loss of FBXW7 could affect the nuclear HSF1 recovery after warm stimulation. It may be associated with FBXW7 deletion inhibiting the degradation of nuclear HSF1

Abstract 303: Host-derived CCL2 plays a critical role in prostate cancer growth and skeletal metastasis

Abstract We have previously reported that the presence of CC chemokine ligand 2 (CCL2) produced by prostate cancer (PCa) epithelial cells, stroma/osteoblasts, and bone marrow endothelial cells, is pro-tumorigenic for prostate cancer cells. CCL2 increases prostate cancer cell proliferation, migration, and survival and induces osteoclast formation. Moreover, the key role of CCL2 in the tumor growth and metastasis has been linked to its regulatory role in mediating monocyte/macrophage infiltration into the tumor microenvironment. Using a model of human PCa cells implanted in mice, Loberg R. et al. reported that anti-murine CCL2 neutralizing antibodies resulted in greater reduction of overall tumor burden compared to the use of anti-human antibody (anti-tumor cell-derived). This suggests that host-derived CCL2 may play a key role in prostate tumor growth and metastasis. Therefore, in this study, we have employed CCL2 knockouts which were interbred to SCID mice to generate colonies of homozygous SCID/CCL2−/− mice. PCa cells (PC3Luc) were injected into the left ventricle (Intracardiac-injection) of SCID/CCL2−/- and wild-type controls (n=20/group). The effects of deletion of CCL2 in the host on prostate cancer cell homing, growth in bone and bone destruction were evaluated for 4 weeks. In a parallel experiment, PC3Luc cells were subcutaneously implanted and tumor size was monitored weekly for 4 weeks. This parallel study addressed bone-specific vs. generalized influences of host-derived CCL2 on prostate cancer growth. We found that skeletal metastases occurred in 4/20 SCID/CCL2−/− mice vs 14/20 WT mice. In addition, X-ray photographs showed significantly fewer bone resorptive lesions in SCID/CCL2−/− mice vs. WT mice. Histology examinations confirmed x-ray findings. In the parallel experiment, PC3Luc tumor grew slower in SCID/CCL2−/- mice vs. WT mice. These results indicate that host-derived CCL2 production defines a permissive microenvironment for prostate cancer growth and skeletal metastasis. These findings may significantly impact the therapeutic field through targeting “the host” instead of “the tumor cells” in prostate cancer. Supported by U.S. Department of Defense PC061231 (J. Zhang), National Natural Sciences Foundation Key Project (81130046, J. Zhang) and National Natural Science Foundation Project (81171993, Y. Lu). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 303. doi:1538-7445.AM2012-303</jats:p

Abstract 729: The role of skp2 in paclitaxel-resistant prostate cancer cells

Abstract Prostate cancer is the most common cancer and the second most frequent cause of cancer-related death of men in the United States. It has been shown that the F-box protein Skp2(S-phase kinase associated protein 2) plays an important role in the development and progression of prostate cancer. The molecular mechanism responsible for paclitaxel-resistant in prostate cancer cells remain unclear. Interestingly, we found that the expression of Skp2 is higher in the paclitaxel-resistant prostate cancer cells DU145-TXR or PC-3-TXR compared with their parental cells DU145 or PC-3 respectively.This study was aimed to investigate the role of Skp2 in prostate cancer cell resistance to paclitaxel and the progress of epithelial-to-mesenchymal transition(EMT). Skp2 encoding plasmid was stably transfected into PC-3 and DU145 cells. Meanwhile, Skp2 was silenced by shRNA in DU145-TXR or PC-3-TXR cells. MTS assay was used to measure the cytotoxicity effect of Skp2 overexpression or knockdown of Skp2 in cells after treatment with paclitaxel. We found that when downregulating Skp2 in DU145-TXR and PC-3-TXR, the cells showed a more sensitive response to paclitaxel. Moreover, Overexpression of Skp2 enhances prostate cancer cells resistance to paclitaxel. EMT markers E-cadherin was decreased while vimentin was increased in PC-3-TXR or DU145-TXR cells. The further study showed that when downregulating Skp2 in PC-3 or DU145, the expression of E-cadherin was increased. However, E-cadherin has no significant change when knockdown of Skp2 in PC-3-TXR or DU145-TXR cells. CyclinD1, P21 and P27 were found to increase while CyclinB1 and CyclinE2 to decrease when downregulating Skp2 in PC-3-TXR or DU145-TXR cells. These data provide evidence that Skp2 is associated with prostate cancer cell resistance to paclitaxel. Skp2 could be a potential therapeutic target for prostate cancer. This study is supported by NSFC key project 81130046, NSFC projects 81201669, 81171993 and 81272415. Note: This abstract was not presented at the meeting. Citation Format: Yeguo Yang, Qiong Song, Yi Lu, Jian Zhang, Jiejun Fu. The role of skp2 in paclitaxel-resistant prostate cancer cells. [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 729. doi:10.1158/1538-7445.AM2015-729</jats:p

Role-based intelligent application state computing for OpenFlow distributed controllers in software-defined networking.

Software-defined networking, in which a network is programmable and controlled with soft computing techniques, is widely used in various network testbeds. To satisfy the demands of large networks, control planes are constructed with distributed controllers, which are a routine aspect of OpenFlow research. Distributed controllers are scalable and fault tolerant; thus, they can replace centralized controllers in large-scale networks. In the foreseeable future, there will emerge more applications based on soft computing techniques. Because there is scant research on application management, a distributed controller with a role-based mechanism for properly managing applications and their states based on their properties has yet to be developed. Thus, in this study, we propose a systematic approach for classifying applications according to their roles to dynamically deploy applications and their states. Both applications and their states are managed based on their properties, including CPU, memory and network bandwidth. This intelligent mechanism, which computes the overhead of applications, provides a compromise between storage and bandwidth usage in OpenFlow distributed controllers. We propose a hierarchical system to differentiate applications and design a controller module for dynamically determining the status of an application

Genistein Protects Genioglossus Myoblast Against Hypoxia-induced Injury through PI3K-Akt and ERK MAPK Pathways

AbstractObstructive sleep apnea and hypopnea syndrome (OSAHS) is a clinical syndrome characterized by recurrent episodes of obstruction of the upper airway during sleep that leads to a hypoxic condition. Genioglossus, an important pharyngeal muscle, plays an important role in maintaining an open upper airway for effective breathing. Our previous study found that genistein (a kind of phytoestrogen) protects genioglossus muscle from hypoxia-induced oxidative injury. However, the underlying mechanism is still unknown. In the present study, we examined the effects of hypoxia on genioglossus myoblast proliferation, viability and apoptosis, and the protective effect of genistein and its relationship with the PI3K/Akt and ERK MAPK pathways. Cell viability and Bcl-2 were reduced under hypoxic condition, while ROS generation, caspase-3, MDA, and DNA damage were increased following a hypoxia exposure. However, the effects of hypoxia were partially reversed by genistein in an Akt- and ERK- (but not estrogen receptor) dependent manner. In conclusion, genistein protects genioglossus myoblasts against hypoxia-induced oxidative injury and apoptosis independent of estrogen receptor. The PI3K-Akt and ERK1/2 MAPK signaling pathways are involved in the antioxidant and anti-apoptosis effect of genistein on genioglossus myoblasts.</jats:p

Abstract 2008: CUL7 promotes non-small cell lung cancer cells migration and invasion

Abstract Cullin7 (CUL7) is a scaffold protein of the CUL7 E3 ubiquitin ligase containing Fbxw8, Skp1 and ROC1. Germ line mutations of the CUL7 are linked to 3-M syndrome and Yakuts short stature syndrome, both of which are characterized by growth retardation and few bone abnormalities. We have previously shown that CUL7 promotes trophoblast cells migration and invasion. However, the role of CUL7 on cancer invasion and metastasis remains unknown. It has been reported that CUL7 mRNA is overexpressed in non-small cell lung cancer (NSCLC). In this study, we investigated the effects of CUL7 on NSCLC cells migration and invasion. Firstly, we evaluated CUL7 expression on tissue microarray slides by immunohistochemical staining. We found that CUL7 significantly overexpressed in NSCLC and its expression was associated with advanced TNM stage, nodal or bone metastasis and poor overall survival. Next, we determined CUL7, CUL1 and Fbxw8 expression in different NSCLC cell lines by real-time PCR and western blot. Higher level of CUL7 was observed in H1299, H358, H1437, and H1792 cells which derived from metastatic sites compared to H460 and human bronchial epithelial cells (BEAS2B). Knockdown CUL7 expression in NSCLC cells decreased cell motility and invasive ability using wound healing and matrigel invasion assay. Real-time PCR revealed that knockdown of CUL7 significantly decreased expression of CCL2, IL-6 and CXCL16. However, the expression of uPA, MMP-2 and MMP-9 were not changed. Furthermore, Knockdown of CUL7 in NSCLC cells decreased secretion of CCL2, CXCL16 and IL-6. This study indicates that CUL7 may be a potential biomarker as well as a novel therapeutic target in NSCLC progression. Supported by National Natural Sciences Foundation (NSFC) projects (81201669, 81171993 and 81272415); NSFC key project (81130046). Note: This abstract was not presented at the meeting. Citation Format: Qiong Song, Lihui Wang, Yi Lu, Jian Zhang, Jiejun Fu. CUL7 promotes non-small cell lung cancer cells migration and invasion. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2008. doi:10.1158/1538-7445.AM2014-2008</jats:p