記述言語：英語  

FBXW7 (also known as Fbw7, Sel10, hCDC4, or hAgo) is a tumor suppressor and the most frequently mutated member of the F-box protein family in human cancers. FBXW7 functions as the substrate recognition component of an SCF-type E3 ubiquitin ligase. It specifically controls the proteasome-mediated degradation of many oncoproteins such as c-MYC, NOTCH, KLF5, cyclin E, c-JUN, and MCL1. In this review, we summarize the molecular and biological features of FBXW7 and its substrates as well as the impact of mutations of FBXW7 on cancer development. We also address the clinical potential of anticancer therapy targeting FBXW7.

DOI： 10.1016/j.semcancer.2020.02.017

記述言語：英語  

Inflammation plays an essential role in the development and progression of most cancers. Chemokine C-C motif chemokine 2 (CCL2) and its receptor C-C chemokine receptor type 2 (CCR2) constitute a key signaling axis in inflammation that has recently attracted much interest on the basis of evidence showing its association with cancer progression. Propagermanium (3-oxygermylpropionic acid polymer) is an organogermanium compound that is given for the treatment of hepatitis B in Japan and which inhibits the CCL2-CCR2 signaling pathway. Herein, we review the importance of the CCL2-CCR2 axis as a target in cancer treatment as shown by studies in mice and humans with pharmacological agents including propagermanium.

DOI： 10.1111/cas.14075

記述言語：英語  

The biological relation between ubiquitin ligases and their substrates has been largely unclear. We previously developed a method-differential proteomics-based identification of ubiquitylation substrates (DiPIUS)-for the comprehensive identification of substrates for a given ubiquitin ligase. We have now applied DiPIUS to the F-box protein Fbxw7 in three cell lines (mHepa, Neuro2A and C2C12) and thereby identified Krüppel-like factor 7 (KLF7) as a candidate substrate of the SCFFbxw7 ubiquitin ligase complex. KLF7 was shown to interact with Fbxw7 and to undergo Fbxw7-mediated polyubiquitylation. The stability of KLF7 was increased by depletion of Fbxw7, mutation of a putative Cdc4 phosphodegron (CPD) of KLF7 or exposure to inhibitors of glycogen synthase kinase-3 (GSK-3). Over-expression of Fbxw7 in Neuro2A cells down-regulated expression of the p21Cip1 gene, which is a transcriptional target of KLF7 in neuronal differentiation and maintenance. Despite the presence of an almost identical CPD sequence in KLF6, the closest paralog of KLF7, mutation of this sequence affected neither the interaction of KLF6 with Fbxw7 nor its half-life. Our results suggest that KLF7, but not KLF6, is a bona fide substrate of SCFFbxw7 , and that control of KLF7 abundance by SCFFbxw7 might contribute to the regulation of neuronal differentiation and maintenance.

DOI： 10.1111/gtc.12680

記述言語：英語  

The ubiquitin-proteasome system regulates the abundance of many cellular proteins by mediating their targeted degradation. We previously developed a method-differential proteomics-based identification of ubiquitylation substrates (DiPIUS)-for the comprehensive identification of substrates for a given F-box protein subunit of SCF-type ubiquitin ligases. We have now applied DiPIUS to the F-box protein Fbxw7 in three cell lines (mHepa, Neuro2A, and C2C12) and thereby identified myelin regulatory factor (MyRF), an endoplasmic reticulum-anchored transcription factor that is essential for myelination of nerves in the central nervous system, as a candidate substrate of Fbxw7 specifically in mHepa cells. Co-immunoprecipitation analysis confirmed that the NH2-terminal cytoplasmic domain of MyRF interacted with Fbxw7 in these cells. Furthermore, an in vitro ubiquitylation assay revealed that MyRF undergoes polyubiquitylation in the presence of purified recombinant SCFFbxw7 In addition, the stability of MyRF in mHepa cells was increased by mutation of a putative phosphodegron sequence or by exposure of the cells to an inhibitor of glycogen synthase kinase-3 (GSK-3). We found that MyRF mRNA is not restricted to the central nervous system but is instead distributed widely among mouse tissues. Furthermore, with the use of RNA sequencing in mHepa cells overexpressing or depleted of MyRF, we identified many novel potential target genes of MyRF. Our results thus suggest that Fbxw7 controls the transcription of MyRF target genes in various tissues through regulation of MyRF protein stability in a manner dependent on MyRF phosphorylation by GSK-3.

DOI： 10.1074/jbc.RA117.000741

記述言語：英語   掲載種別：研究論文（学術雑誌）  

How stem cells maintain their stemness or initiate exit from the stem cell state for differentiation remains largely unknown. Aldehyde dehydrogenase (ALDH) activity is a hallmark of stem cells-including embryonic, adult tissue, and cancer stem cells-and is essential for their maintenance. The mechanisms by which ALDH activity is regulated in stem cells have remained poorly understood, however. We now show that the ubiquitin-dependent degradation of ALDH3 mediated by FBXL12 (F box and leucine-rich repeat protein 12) is essential for execution of the differentiation program of trophoblast stem cells (TSCs). FBXL12 is present only in eutherian mammals, and its expression is largely restricted to the placenta during mouse embryogenesis. FBXL12 was found to interact specifically with members of the ALDH3 family and to mediate their polyubiquitylation. Most mice deficient in FBXL12 died during the embryonic or perinatal period probably as a result of abnormal development of the placenta, characterized by impaired formation of the junctional zone. ALDH3 accumulated in the FBXL12-deficient placenta, and forced expression of ALDH3 in wild-type TSCs phenocopied the differentiation defect of FBXL12-deficient TSCs. Conversely, inhibition of ALDH3 activity by gossypol rescued the phenotype of FBXL12 deficiency. Our results suggest that FBXL12 plays a key role in the downregulation of ALDH3 activity in TSCs and thereby initiates trophoblast differentiation during placental development.

DOI： 10.1002/stem.2088

記述言語：英語  

Although identification of substrates for ubiquitin ligase (E3) is important for understanding its biological functions, detection of the interaction between an E3 and its substrates has remained challenging. We recently developed a new approach, termed differential proteomics-based identification of ubiquitylation substrates (DiPIUS), for the discovery of substrates of a given E3 ligase. We have now applied this approach to an uncharacterized human F-box protein, FBXO21, which serves as the substrate-recognition subunit of a SKP1-CUL1-F-box protein (SCF)-type E3, thereby identifying EID1 (EP300-interacting inhibitor of differentiation 1) as a candidate substrate. The central and COOH-terminal portion of FBXO21 was found to interact with the COOH-terminal region of EID1 in transfected cells. Over-expression of FBXO21 resulted in the down-regulation of EID1, whereas disruption of the FBXO21 gene with the CRISPR/Cas9 system stabilized EID1 and led to its accumulation in both the cytoplasm and nucleus. An in vitro ubiquitylation assay showed that EID1 is a direct substrate of SCF(FBXO)(21). Collectively, our results suggest that EID1 is a bona fide substrate of FBXO21 and that the control of EID1 abundance by SCF(FBXO)(21) might affect the transcriptional repression activity of EID1.

DOI： 10.1111/gtc.12260

記述言語：英語  

Fbxw7 has been identified as an oncosuppressor protein in many types of cancer. We have recently shown that loss of Fbxw7 in bone marrow-derived stromal cells (BMSCs) promotes cancer metastasis by increasing production of the chemokine CCL2, which attracts monocytic myeloid-derived suppressor cells (Mo-MDSCs) and macrophages to the metastatic niche.

DOI： 10.1080/2162402X.2015.1022308

記述言語：英語   掲載種別：研究論文（学術雑誌）  

A GATA family transcription factor, GATA-binding protein 2 (GATA2), participates in cell growth and differentiation of various cells, such as hematopoietic stem cells. Although its expression level is controlled by transcriptional induction and proteolytic degradation, the responsible E3 ligase has not been identified. Here, we demonstrate that F-box/WD repeat-containing protein 7 (Fbw7/Fbxw7), a component of Skp1, Cullin 1, F-box-containing complex (SCF)-type E3 ligase, is an E3 ligase for GATA2. GATA2 contains a cell division control protein 4 (Cdc4) phosphodegron (CPD), a consensus motif for ubiquitylation by Fbw7, which includes Thr(176). Ectopic expression of Fbw7 destabilized GATA2 and promoted its proteasomal degradation. Substitution of threonine 176 to alanine in GATA2 inhibited binding with Fbw7, and the ubiquitylation and degradation of GATA2 by Fbw7 was suppressed. The CPD kinase, which mediates the phosphorylation of Thr(176), was cyclin B-cyclin-dependent kinase 1 (CDK1). Moreover, depletion of endogenous Fbw7 stabilized endogenous GATA2 in K562 cells. Conditional Fbw7 depletion in mice increased GATA2 levels in hematopoietic stem cells and myeloid progenitors at the early stage. Increased GATA2 levels in Fbw7-conditional knock-out mice were correlated with a decrease in a c-Kit high expressing population of myeloid progenitor cells. Our results suggest that Fbw7 is a bona fide E3 ubiquitin ligase for GATA2 in vivo.

DOI： 10.1074/jbc.M114.613018

記述言語：英語  

The gene encoding F-box protein FBXW7 is frequently mutated in many human cancers. Although most previous studies have focused on the tumor-suppressive capacity of FBXW7 in tumor cells themselves, we determined that FBXW7 in the host microenvironment also suppresses cancer metastasis. Deletion of Fbxw7 in murine BM-derived stromal cells induced accumulation of NOTCH and consequent transcriptional activation of Ccl2. FBXW7-deficient mice exhibited increased serum levels of the chemokine CCL2, which resulted in the recruitment of both monocytic myeloid-derived suppressor cells and macrophages, thereby promoting metastatic tumor growth. Administration of a CCL2 receptor antagonist blocked the enhancement of metastasis in FBXW7-deficient mice. Furthermore, in human breast cancer patients, FBXW7 expression in peripheral blood was associated with serum CCL2 concentration and disease prognosis. Together, these results suggest that FBXW7 antagonizes cancer development in not only a cell-autonomous manner, but also a non-cell-autonomous manner, and that modulation of the FBXW7/NOTCH/CCL2 axis may provide a potential approach to suppression of cancer metastasis.

DOI： 10.1172/JCI78782

記述言語：英語   掲載種別：研究論文（学術雑誌）  

MDM2 mediates the ubiquitylation and thereby triggers the proteasomal degradation of the tumor suppressor protein p53. However, genetic evidence suggests that MDM2 contributes to multiple regulatory networks independently of p53 degradation. We have now identified the DEAD-box RNA helicase DDX24 as a nucleolar protein that interacts with MDM2. DDX24 was found to bind to the central region of MDM2, resulting in the polyubiquitylation of DDX24 both in vitro and in vivo. Unexpectedly, however, the polyubiquitylation of DDX24 did not elicit its proteasomal degradation but rather promoted its association with preribosomal ribonucleoprotein (pre-rRNP) processing complexes that are required for the early steps of pre-rRNA processing. Consistently with these findings, depletion of DDX24 in cells impaired pre-rRNA processing and resulted both in abrogation of MDM2 function and in consequent p53 stabilization. Our results thus suggest an unexpected role of MDM2 in the nonproteolytic ubiquitylation of DDX24, which may contribute to the regulation of pre-rRNA processing.

DOI： 10.1128/MCB.00320-14

記述言語：英語  

The Skp1-Cul1-F-box protein (SCF) complex is one of the most well characterized types of ubiquitin ligase (E3), with the E3 activity of the complex being regulated in part at the level of complex formation. Fbxl3 is an F-box protein that is responsible for the ubiquitylation and consequent degradation of cryptochromes (Crys) and thus regulates oscillation of the circadian clock. Here we show that formation of the SCF(Fbxl3) complex is regulated by substrate binding in vivo. Fbxl3 did not associate with Skp1 and Cul1 to a substantial extent in transfected mammalian cells. Unexpectedly, however, formation of the SCF(Fbxl3) complex was markedly promoted by forced expression of its substrate Cry1 in these cells. A mutant form of Fbxl3 that does not bind to Cry1 was unable to form an SCF complex, suggesting that interaction of Cry1 with Fbxl3 is essential for formation of SCF(Fbxl3). In contrast, recombinant Fbxl3 associated with recombinant Skp1 and Cul1 in vitro even in the absence of recombinant Cry1. Domain-swap analysis revealed that the COOH-terminal leucine-rich repeat domain of Fbxl3 attenuates the interaction of Skp1, suggesting that a yet unknown protein associated with the COOH-terminal domain of Fbxl3 and inhibited SCF complex formation. Our results thus provide important insight into the regulation of both SCF ubiquitin ligase activity and circadian rhythmicity.

DOI： 10.1074/jbc.M113.511303

記述言語：英語  

Although identification of substrates for an enzyme is a key step in elucidation of its biological functions, detection of the interaction between enzymes and substrates remains challenging. We recently developed a new approach, termed differential proteomics-based identification of ubiquitylation substrates (DiPIUS), for the discovery of substrates of ubiquitin ligases. We have now applied this approach to Fbxw7, the F-box protein component of an Skp1-Cul1-F-box protein-type ubiquitin ligase and, thereby, identified two similar transcription factors, old astrocyte specifically induced substance (OASIS) and BBF2 human homolog on chromosome 7 (BBF2H7), as candidate substrates. Coimmunoprecipitation analysis confirmed that the α and γ isoforms of Fbxw7 interact with OASIS and BBF2H7 in vivo. Sustained overexpression of Fbxw7 resulted in marked down-regulation of OASIS and BBF2H7, whereas RNAi-mediated Fbxw7 depletion stabilized both proteins. Mutation of a putative Cdc4 phosphodegron in OASIS and BBF2H7 attenuated their association with Fbxw7 and resulted in their stabilization. Depletion of Fbxw7 promoted the differentiation of mouse C2C12 mesenchymal cells into osteoblasts in association with the accumulation of OASIS. Conversely, overexpression of Fbxw7 in C2C12 cells resulted in down-regulation of Col1A1 mRNA, a target of OASIS. Conditional ablation of Fbxw7 in primary mouse mesenchymal cells promoted chondrogenesis in association with up-regulation of BBF2H7, whereas overexpression of Fbxw7 inhibited chondrogenesis in ATDC5 cells. Collectively, our results suggest that OASIS and BBF2H7 are bona fide substrates of Fbxw7 and that Fbxw7 controls osteogenesis and chondrogenesis by targeting OASIS and BBF2H7, respectively, for degradation.

DOI： 10.1074/jbc.M113.465179

記述言語：英語   掲載種別：研究論文（学術雑誌）  

In the mammalian circadian clockwork, CRY1 and CRY2 repressor proteins are regulated by posttranslational modifications for temporally coordinated transcription of clock genes. Previous studies revealed that FBXL3, an F-box-type E3 ligase, ubiquitinates CRYs and mediates their degradation. Here, we found that FBXL21 also ubiquitinates CRYs but counteracts FBXL3. Fbxl21(-/-) mice exhibited normal periodicity of wheel-running rhythms with compromised organization of daily activities, while an extremely long-period phenotype of Fbxl3(-/-) mice was attenuated in Fbxl3/Fbxl21 double-knockout mice. The double knockout destabilized the behavioral rhythms progressively and sometimes elicited arrhythmicity. Surprisingly, FBXL21 stabilized CRYs and antagonized the destabilizing action by FBXL3. Predominantly cytosolic distribution of FBXL21 contrasts with nuclear localization of FBXL3. These results emphasize the physiological importance of antagonizing actions between FBXL21 and FBXL3 on CRYs, and their combined actions at different subcellular locations stabilize oscillation of the circadian clock.

DOI： 10.1016/j.cell.2013.01.054

記述言語：英語  

Induced pluripotent stem cells (iPSCs) share many biological properties with embryonic stem cells (ESCs), and are generated from somatic cells by expression of some transcription factors such as Oct3/4, Sox2, Klf4 and c-Myc. Among these factors, the abundance of c-Myc is strictly regulated by Fbxw7, a subunit of Skp1-Cul1-F-box protein-type ubiquitin ligase. We have now shown that the expression of Fbxw7 was increased as ESCs differentiated. To investigate the role of Fbxw7 in the ESCs/iPSCs, we examined the impact of Fbxw7 ablation in the efficiency in iPSC generation. The frequency of iPSC generation from mouse embryonic fibroblasts (MEFs) lacking Fbxw7 was markedly greater than that from control MEFs. Depletion of Fbxw7 also resulted in promotion of iPSC generation. Morphology of iPSC clonies from Fbxw7-depleted MEFs appeared more undifferentiated than that from MEFs overexpressing c-Myc. Additional depletion of c-Myc did not abrogate the effect of Fbxw7 depletion, suggesting that c-Myc accumulation is not necessarily required for the increased efficiency in iPSC generation by Fbxw7 ablation. Substrates of Fbxw7 other than c-Myc might therefore play a key role in iPSC generation. These results suggest that transient inhibition of Fbxw7 would be a more promising approach to efficient generation of iPSCs than c-Myc over-expression.

DOI： 10.1111/j.1365-2443.2012.01626.x

記述言語：英語  

Although elucidation of enzyme-substrate relations is fundamental to the advancement of biology, universal approaches to the identification of substrates for a given enzyme have not been established. It is especially difficult to identify substrates for ubiquitin ligases, given that most such substrates are immediately ubiquitylated and degraded as a result of their association with the enzyme. We here describe the development of a new approach, DiPIUS (differential proteomics-based identification of ubiquitylation substrates), to the discovery of substrates for ubiquitin ligases. We applied DiPIUS to Fbxw7α, Skp2, and Fbxl5, three of the most well-characterized F-box proteins, and identified candidate substrates including previously known targets. DiPIUS is thus a powerful tool for unbiased and comprehensive screening for substrates of ubiquitin ligases.

DOI： 10.1021/pr201216u

記述言語：英語   掲載種別：研究論文（学術雑誌）  

The budding yeast UbL-UBA protein Dsk2 has a UbL domain at its N-terminus and a UBA domain at its C-terminus, and thus functions as a shuttle protein in the ubiquitin-proteasome pathway. In this report we describe two isoforms of Xenopus Dsk2-related protein, XDRP1L and XDRP1S. Difference of the two proteins in sequence was that the UbL domain of XDRP1S lacks 15 residues in the middle part of that of XDRP1L. Both XDRP1L and XDRP1S were expressed in Xenopus eggs. XDRP1L and XDRP1S bound to polyubiquitinated proteins via their UBA domains. XDRP1L also bound to the proteasome via its UbL domain, whereas the XDRP1S UbL domain was less likely to bind to the proteasome. Instead, XDRP1S not XDRP1L bound to monomeric cyclin A and prevented its degradation. The existence of such Dsk2-isoforms in Xenopus eggs suggests that the shuttling function via the UbLUBA protein Dsk2 is evolutionally conserved across species. (c) 2006 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.bbrc.2006.09.123

記述言語：英語   掲載種別：研究論文（学術雑誌）  

The yeast UbL-UBA protein Dsk2 is thought to act as a shuttle protein that delivers polyubiquitinated proteins to the proteasome. Previously, we identified Xenopus Dsk2-related protein, XDRP1, as a cyclin A-interacting protein. Using Xenopus egg extracts, we further characterized its two isoforms, XDRP1L and XDRP1S, with respect to cyclin binding and its degradation. Polyubiquitinated cyclins bound to the UBA domain of XDRP1L and XDRP1S, whereas monomeric cyclins A and B bound to the UbL domain of XDRP1S but not to XDRP1L. Binding of XDRP1S with monomeric cyclins was affected by a Cdc2-mediated phosphorylation of either the XDRP1S UbL domain or cyclins. Degradation of cyclin B was also prevented by XDRP1S in a Cdc2-sensitive manner. Loss of the XDRP1S-cyclin interaction allowed cyclins to be degraded in calcium-treated CSF extracts. These results suggest that the shuttling pathway via the UbL-UBA protein XDRP1 participates in degradation of mitotic cyclins in Xenopus eggs. (c) 2006 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.bbrc.2006.09.122

記述言語：日本語   掲載種別：記事・総説・解説・論説等（学術雑誌）  

既存薬でがん転移抑制

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