SCF泛素连接酶靶蛋白识别组分FBW7

FBW7（F-box and WD repeat domain-containing7）是F-box蛋白家族成员,为SCF（SKP1-CUL1-F-box）型泛素连接酶的靶蛋白识别组分。FBW7通过靶降解周期蛋白E、Myc、Jun等多种癌蛋白,对细胞周期进程、细胞生长、分化起重要调控作用。在多种人类肿瘤中已发现FBW7突变,FBW7功能缺失会引起染色体不稳定及肿瘤发生,表明FBW7是一种肿瘤抑制因子。在FBW7缺失所致的肿瘤发生过程中,周期蛋白E、Myc等靶蛋白活性升高、p53功能缺失有重要作用。     

SCF targets cyclin E2 for ubiquitin-dependent proteolysis

E-type cyclins (E1 and E2) regulate the S phase program in the mammalian cell division cycle. Expression of cyclin E1 and E2 is frequently deregulated in a variety of cancer types and a wealth of experimental evidence supports an oncogenic role of these proteins in human tumorigenesis. Although the molecular mechanisms responsible for cyclin E1 deregulation in cancer are well defined, little is known regarding cyclin E2. Here we report that cyclin E2 is targeted for ubiquitin-dependent proteolysis by the ubiquitin ligase SCF^Fbxw7/hCdc4. Ubiquitylation is triggered by phosphorylation of cyclin E2 on residues Thr392 and Ser396, and to a lesser extent Thr74, contained in two consensus Cdc4-phosphodegrons. Furthermore, we found that ectopic expression of cyclin E1 enhances the ubiquitin-dependent proteolysis of cyclin E2 in vivo, suggesting a potential cross-talk in the regulation of E-type cyclin activity. Since SCF^Fbxw7/hCdc4 is functionally inactivated in several human cancer types, alteration of this molecular pathway could contribute to the deregulation of cyclin E2 in tumorigenesis.     

FBW7泛素化修饰Snail促进上皮间质转化的实验研究

目的:探讨FBW7(F-box/WD repeat-containing protein 7)是否参与转录抑制因子Snail的泛素化修饰,通过调节上皮间质转化(EMT)进而导致非小细胞肺癌的侵袭和转移,为治疗非小细胞肺癌(NSCLC)患者晚期转移提供新的思路。方法:首先,通过Western Blot方法检测多种人肺细胞系中Snail的表达水平。上调(药物处理)及下调(设计并合成特异性sh RNA转染)H460细胞中FBW7的表达后,检测Snail的表达水平。采用平板克隆及Transwell方法检测下调FBW7的H460细胞形态改变和侵袭转移能力变化。结果:人非小细胞肺癌H460细胞中Snail表达水平较高。上调FBW7表达可使Snail表达下降,经蛋白酶抑制剂MG132处理后Snail表达升高;而下调FBW7表达后Snail表达增加。下调FBW7表达细胞后,细胞形态改变,侵袭转移能力增强。结论:FBW7参与了Snail泛素化修饰,进而经蛋白酶体途径将其降解引起EMT,导致肿瘤转移和进展的发生。     

泛素连接酶FBW7的肿瘤抑制作用和机制

泛素-蛋白酶体系统是一个主要的蛋白质降解调节通路,在细胞分裂过程中发挥着重要作用,其成员在肿瘤中频繁存在着表达异常现象.FBW7又名AGO、hCDC4、FBXW7和SEL-10,是一种拥有7串联WD40重复结构域的F-box蛋白,可作为SCF型泛素连接酶（E3）复合物的底物识别亚基发挥作用.FBW7是一种肿瘤抑制蛋白,其基因在多种肿瘤包括直肠癌、胃癌、卵巢癌和白血病中存在着基因突变或缺失.FBW7可直接结合和靶向作用多种转录激活因子或原癌基因,如周期蛋白E、c-Myc、c-Jun、Notch、MCL1、KLF5 和mTOR等并对其进行泛素化修饰和随后的26S蛋白酶体降解.肿瘤抑制蛋白FBW7的研究对肿瘤发生机制的理解具有重要意义,同时也为肿瘤的诊断和治疗提供了新的靶点.本文综述了FBW7的特征、肿瘤抑制作用及机制.     

Emerging roles of the FBW7 tumour suppressor in stem cell differentiation

FBW7 is a ubiquitin E3 ligase substrate adaptor that targets many important oncoproteins-such as Notch, c-Myc, cyclin E and c-Jun-for ubiquitin-dependent proteolysis. By doing so, it plays crucial roles in many cellular processes, including cell cycle progression, cell growth, cellular metabolism, differentiation and apoptosis. Loss of FBW7 has been observed in many types of human cancer, and its role as a tumour suppressor was confirmed by genetic ablation of FBW7 in mice, which leads to the induction of tumorigenesis. How FBW7 exerts its tumour suppression function, and whether loss of FBW7 leads to de-differentiation or acquisition of stemness-a process frequently seen in human carcinomas-remains unclear. Emerging evidence shows that FBW7 controls stem cell self-renewal, differentiation, survival and multipotency in various stem cells, including those of the haematopoietic and nervous systems, liver and intestine. Here, we focus on the function of FBW7 in stem cell differentiation, and its potential relevance to human disease and therapeutics.     

The two faces of FBW7 in cancer drug resistance

Chemotherapy is an important therapeutic approach for cancer treatment. However, drug resistance is an obstacle that often impairs the successful use of chemotherapies. Therefore, overcoming drug resistance would lead to better therapeutic outcomes for cancer patients. Recently, studies by our own and other groups have demonstrated that there is an intimate correlation between the loss of the F-box and WD repeat domain-containing 7 (FBW7) tumor suppressor and the incurring drug resistance. While loss of FBW7 sensitizes cancer cells to certain drugs, FBW7-/- cells are more resistant to other types of chemotherapies. FBW7 exerts its tumor suppressor function by promoting the degradation of various oncoproteins that regulate many cellular processes, including cell cycle progression, cellular metabolism, differentiation, and apoptosis. Since loss of the FBW7 tumor suppressor is linked to drug resistance, FBW7 may represent a novel therapeutic target to increase drug sensitivity of cancer cells to conventional chemotherapeutics. This paper thus focuses on the new functional aspects of FBW7 in drug resistance.     

KLF6 degradation after apoptotic DNA damage

Krüppel-like factor 6 (KLF6) is a cancer gene (www.sanger.ac.uk/genetics/CGP/Census/). Here, we demonstrate that KLF6 protein is rapidly degraded when apoptosis is induced via the intrinsic pathway by cisplatin, adriamycin, or UVB irradiation in multiple cell lines (HCT116, SW40, HepG2, PC3-M, Skov3, NIH-3T3, 293T, GM09706, and MEF, IMR-90). KLF6 degradation occurred in the presence or absence of p53, was associated with ubiquitination, mediated by the proteasome (half-life 16 min, unstimulated), and independent of caspases and calpain. KLF6 was unchanged by apoptosis via the extrinsic/death-receptor pathway. Deregulation of KLF6 stability may alter its tumor suppressor function and/or the response of tumors to chemotherapeutics.     

ERK kinase phosphorylates and destabilizes the tumor suppressor FBW7 in pancreatic cancer

F-box and WD repeat domain-containing 7 (FBW7) is the substrate recognition component of the Skp1-Cul1-F-box (SCF) ubiquitin ligase complex and functions as a major tumor suppressor by targeting various oncoproteins for degradation. Genomic deletion or mutation of FBW7 has frequently been identified in many human cancers but not in pancreatic ductal adenocarcinoma. Thus it is important to know how the tumor suppressive function of FBW7 is impaired in pancreatic cancer. In this study, we first observed that low FBW7 expression correlated significantly with ERK activation in pancreatic cancer clinical samples, primarily due to KRAS mutations in pancreatic cancer. We further showed that ERK directly interacted with FBW7 and phosphorylated FBW7 at Thr205, which sequentially promoted FBW7 ubiquitination and proteasomal degradation. Furthermore, the phospho-deficient T205A FBW7 mutant is resistant to ERK activation and could significantly suppress pancreatic cancer cell proliferation and tumorigenesis. These results collectively demonstrate how the oncogenic KRAS mutation inhibits the tumor suppressor FBW7, thus revealing an important function of KRAS mutations in promoting pancreatic cancer progression.     

Methylation of miR-19b-3p promoter exacerbates inflammatory responses in sepsis-induced ALI via targeting KLF7

Sepsis-induced acute lung injury is associated with dysregulated inflammatory reactions. MiR-19b-3p level was reported to be downregulated in patients with sepsis. To evaluate the role of miR-19b-3p in sepsis, cecum ligation and puncture-induced mouse sepsis model and lpopolysaccharide (LPS)-treated pulmonary microvascular endothelial cells (PMVECs) were used. For in vivo study, lung tissue was harvested for hematoxylin and eosin (H&E) staining, tumor necrosis factor-α, interleukin-6 (IL-6), IL-1β, and p-p65, p-IκB measuring. Cell apoptosis was assessed by TUNEL assay. For in vitro study, cell proliferation and apoptosis were detected by CCK-8 and flow cytometry, respectively. Methylation of miR-19b-3p promoter was measured by methylation-specific PCR (MSP) assay. The target of miR-19b-3p was determined by dual-luciferase reporter gene assay. The level of miR-19b-3p was determined to be downregulated in vitro and in vivo. In addition, miR-19b-3p protected mice from inflammation injury through inhibiting NF-κB signaling pathway. Overexpression of miR-19b-3p increased cell viability, decreased apoptosis, and proinflammatory cytokines secretion in LPS-treated PMVECs. Besides these, Krüppel-like factor 7 (KLF7) was confirmed as the target of miR-19b-3p. And methylation of miR-19b-3p was the reason of decreased miR-19b-3p level. In conclusion, miR-19b-3p protected cells from sepsis-induced inflammation injury via inhibiting NF-κB signaling pathway, and KLF7 was a potential target.     

Mcl‐1 inhibition overcomes BET inhibitor resistance induced by low FBW7 expression in breast cancer

While the promise of bromodomains and extraterminal (BET) protein inhibitors (BETis) is emerging in breast cancer (BC) therapy, resistance in these cells to BETis conspicuously curbs their therapeutic potential. FBW7 is an important tumour suppressor. However, the role of FBW7 in BC is not clear. In the current study, our data indicated that the low expression of FBW7 contributes to the drug resistance of BC cells upon JQ1 treatment. shRNA‐mediated FBW7 silencing in FBW7 WT BC cells suppressed JQ1‐induced apoptosis. Mechanistically, it was revealed that this diminished FBW7 level leads to Mcl‐1 stabilization, while Mcl‐1 upregulation abrogates the killing effect of JQ1. Mcl‐1 knockdown or inhibition resensitized the BC cells to JQ1‐induced apoptosis. Moreover, FBW7 knockdown in MCF7 xenografted tumours demonstrated resistance to JQ1 treatment. The combination of JQ1 with a Mcl‐1 inhibitor ({"type":"entrez-nucleotide","attrs":{"text":"S63845","term_id":"400540","term_text":"S63845"}}S63845) resensitized the FBW7 knockdown tumours to JQ1 treatment in vivo. Our study paves the way for a novel therapeutic potential of BETis with Mcl‐1 inhibitors for BC patients with a low FBW7 expression.     

Ubiquitin-like proteins and Rpn10 play cooperative roles in ubiquitin-dependent proteolysis

Rpn10, a subunit of the 26S proteasome, has been proposed to act as a receptor for multiubiquitin chains in ubiquitin-dependent proteolysis. However, studies on RPN10-deleted mutants in yeasts have suggested the presence of other multiubiquitin chain-binding factors functioning in ubiquitin-dependent proteolysis. Here, we report that a mutant with a triple deletion of RAD23, DSK2, and RPN10 genes accumulates large amounts of polyubiquitinated proteins, as is the case with a mutant with RAD23 and DSK2 deletions under restrictive conditions. Dsk2, Rad23, and Rpn10 have different capacities to bind multiubiquitin chains. Another ubiquitin-like protein, Ddi1, has similar activity to those of Rad23 and Dsk2. Taken together, the results suggest that ubiquitin-like proteins, Rad23, Dsk2, possibly Ddi1, and Rpn10 play cooperative roles in ubiquitin-dependent proteolysis, serving as multiubiquitin chain-binding proteins.     

The CUL7 E3 ubiquitin ligase targets insulin receptor substrate 1 for ubiquitin-dependent degradation

Recent genetic studies have documented a pivotal growth-regulatory role played by the Cullin 7 (CUL7) E3 ubiquitin ligase complex containing the Fbw8-substrate-targeting subunit, Skp1, and the ROC1 RING finger protein. In this report, we identified insulin receptor substrate 1 (IRS-1), a critical mediator of the insulin/insulin-like growth factor 1 signaling, as a proteolytic target of the CUL7 E3 ligase in a manner that depends on mammalian target of rapamycin and the p70 S6 kinase activities. Interestingly, while embryonic fibroblasts of Cul7-/- mice were found to accumulate IRS-1 and exhibit increased activation of IRS-1's downstream Akt and MEK/ERK pathways, these null cells grew poorly and displayed phenotypes reminiscent of those associated with oncogene-induced senescence. Taken together, our findings demonstrate a key role for the CUL7 E3 in targeting IRS-1 for degradation, a process that may contribute to the regulation of cellular senescence.