LSD1 destabilizes FBXW7 and abrogates FBXW7 functions independent of its demethylase activity.

FBXW7 acts as a typical tumor suppressor, with loss-of-function alterations in human cancers, by promoting ubiquitylation and degradation of many oncoproteins. Lysine-specific demethylase 1 (LSD1) is a well-characterized histone demethylase. Whether LSD1 has demethylase-independent activity remains elusive. Here we report that LSD1 directly binds to FBXW7 to destabilize FBXW7 independent of its demethylase activity. Specifically, LSD1 is a pseudosubstrate of FBXW7 and LSD1-FBXW7 binding does not trigger LSD1 ubiquitylation, but instead promotes FBXW7 self-ubiquitylation by preventing FBXW7 dimerization. The self-ubiquitylated FBXW7 is subjected to degradation by proteasome as well as lysosome in a manner dependent on autophagy protein p62/SQSTM1. Biologically, LSD1 destabilizes FBXW7 to abrogate its functions in growth suppression, nonhomologous end-joining repair, and radioprotection. Collectively, our study revealed a previously unknown activity of LSD1, which likely contributes to its oncogenic function. Targeting LSD1 protein, not only its demethylase activity, might be a unique approach for LSD1-based drug discovery for anticancer application.

MEKK3 coordinates with FBW7 to regulate WDR62 stability and neurogenesis.

Mutations of WD repeat domain 62 (WDR62) lead to autosomal recessive primary microcephaly (MCPH), and down-regulation of WDR62 expression causes the loss of neural progenitor cells (NPCs). However, how WDR62 is regulated and hence controls neurogenesis and brain size remains elusive. Here, we demonstrate that mitogen-activated protein kinase kinase kinase 3 (MEKK3) forms a complex with WDR62 to promote c-Jun N-terminal kinase (JNK) signaling synergistically in the control of neurogenesis. The deletion of Mekk3, Wdr62, or Jnk1 resulted in phenocopied defects, including premature NPC differentiation. We further showed that WDR62 protein is positively regulated by MEKK3 and JNK1 in the developing brain and that the defects of wdr62 deficiency can be rescued by the transgenic expression of JNK1. Meanwhile, WDR62 is also negatively regulated by T1053 phosphorylation, leading to the recruitment of F-box and WD repeat domain-containing protein 7 (FBW7) and proteasomal degradation. Our findings demonstrate that the coordinated reciprocal and bidirectional regulation among MEKK3, FBW7, WDR62, and JNK1, is required for fine-tuned JNK signaling for the control of balanced NPC self-renewal and differentiation during cortical development.

Tumor-suppressor Fbxw7 targets SIK2 for degradation to interfere with TORC2-AKT signaling in pancreatic cancer.

The tumor suppressor F-box/WD repeat-containing protein 7 (Fbxw7) is a substrate-recognition subunit of a ubiquitin ligase complex. We have previously proposed that Fbxw7 inhibited pancreatic cancer cell proliferation and invasion by targeting ß-catenin. To identify other targets of Fbxw7 involved in pancreatic carcinogenesis, we screened the human protein database for Fbxw7 target candidates using the conserved Fbxw7-recognizing sequences. Twenty-three candidates are identified, including five known Fbxw7 targets and two cancer-related genes (salt inducible kinase 2 [SIK2] and ZMIZ1). We identified SIK2 as an Fbxw7 target for degradation by binding to the "TPPPS" motif of SIK2 in pancreatic cancer cells. We also demonstrated that SIK2 promoted proliferation and mitotic progression of pancreatic cancer cells. Moreover, endogenous Fbxw7 downregulates SIK2 protein level for controlling cell cycle progression, possibly by interfering the SIK2/TORC2/AKT signaling pathway to modulate p21 expression. Collectively, these data demonstrate that Fbxw7 targets the cell cycle controller, SIK2, for degradation, thereby leading to the disruption of downstream TORC2/AKT signaling to inhibit pancreatic cancer cell proliferation and cell cycle progression.

Genetic Deletion of Fbw7 in the mouse intestinal epithelium aggravated dextran sodium sulfate-induced colitis by modulating the inflammatory response of NF-κB pathway.

Fbw7 is a type of E3 ubiquitin ligase that targets various proteins for degradation and has been found to have a high expression level in progenitor cells. Deletion of Fbw7 in the intestine results in the accumulation of progenitor cells. Moreover, Fbw7 loss increases the susceptibility of colorectal cancer. However, the involvement of Fbw7 in the progress and development of inflammatory bowel disease (IBD) is still controversial. To identify the function of Fbw7 on dextran sodium sulfate (DSS)-induced colonic inflammation, we generated Fbw7ΔG mice, lacking Fbw7 specifically in intestinal epithelium. Colitis was induced in male Fbw7 ΔG and wild-type (WT) mice (both age and body weight matched) by treating with 3% DSS in drinking water. We demonstrate that deletion of Fbw7 in the mouse intestinal epithelium aggravates DSS-induced colitis, showing inflammatory response and reduced survival rate. Furthermore, we found that Fbw7 loss caused activation of NF-κB signaling. Thus, FBW7 plays a protective role in acute intestinal inflammation by modulating the inflammatory response of NF-κB pathway.

Exosomal microRNA-19b targets FBXW7 to promote colorectal cancer stem cell stemness and induce resistance to radiotherapy.

Colorectal cancer (CRC) continues to be one of the most malignant cancers with a high mortality rate to date. Promoting the radio-responsiveness of CRC is of great importance for local control and prognosis. In this study, we examined the roles of exosomal microRNA-19b (miR-19b) in CRC radioresistance. The regulatory role of miR-19b in CRC stem cells and radiotherapy-resistant cells was determined using miRNA microarray analysis, and its prognostic value was probed using the TCGA database. It was found that miR-19b was overexpressed in CRC tissues, which indicated a poor prognosis. CRC-derived exosomes (EXOs) enhanced the radio-resistance and stemness properties of CRC cells via delivery of miR-19b in vitro and in vivo. FBXW7 was identified as a putative target of miR-19b. On the contrary, reintroduction of FBXW7 reversed the effects of miR-19b on radioresistance and stemness properties. Furthermore, the Wnt/ß-catenin pathway activity was elevated in CRC cells upon EXOs treatment, decreased after miR-19b downregulation, and increased again after FBXW7 downregulation. These results suggest that miR-19b inhibition could enhance the efficacy of radiotherapy while reducing the stemness properties, thus presenting a promising strategy for sensitizing CRC cells to radiotherapy.

Honokiol antagonizes doxorubicin resistance in human breast cancer via miR-188-5p/FBXW7/c-Myc pathway.

BACKGROUND: Honokiol, a natural phenolic compound derived from Magnolia plants, is a promising anti-tumor compound that exerts a wide range of anti-cancer effects. Herein, we investigated the effect of honokiol on doxorubicin resistance in breast cancer. METHODS: Doxorubicin-sensitive (MCF-7 and MDA-MB-231) and doxorubicin-resistant (MCF-7/ADR and MDA-MB-231/ADR) breast cancer cell lines were treated with doxorubicin in the absence or presence of honokiol; then, the following tests were performed: flow cytometry for cell apoptosis, WST-1 assay for cell viability, qPCR and western blot for the expression of miR-188-5p, FBXW7, and c-Myc. MiR-188-5p mimic, miR-188-5p inhibitor, siFBXW7, and c-Myc plasmids were transfected into cancer cells to evaluate whether miR-188-5p and FBXW7/c-Myc signaling are involved in the effect of honokiol on doxorubicin resistance in breast cancer. A dual luciferase reporter system was used to study the direct interaction between miR-188-5p and FBXW7. RESULTS: Honokiol sensitized doxorubicin-resistant breast cancer cells to doxorubicin-induced apoptosis. Mechanically, upregulation of miR-188-5p was associated with doxorubicin resistance, and honokiol enhanced doxorubicin sensitivity by downregulating miR-188-5p. FBXW7 was confirmed to be a direct target gene of miR-188-5p. FBXW7/c-Myc signaling was involved in the chemosensitization effect of honokiol. Honokiol induced apoptosis in MCF-7/ADR and MDA-MB-231/ADR cells. However, FBXW7 silencing or c-Myc transfection resulted in resistance to the honokiol-induced apoptotic effect. CONCLUSION: These findings suggest that downregulation of miR-188-5p by honokiol enhances doxorubicin sensitivity through FBXW7/c-Myc signaling in human breast cancer. Our study finds an important role of miR-188-5p in the development of doxorubicin resistance in breast cancer, and enriches our understanding of the mechanism of action of honokiol in cancer therapy.

FBXW7 Mutations Promote Cell Proliferation, Migration, and Invasion in Cervical Cancer.

Aim: Cervical cancer is the most common gynecological cancer. Recent studies have revealed that the F-box and WD repeat domain containing 7 (FBXW7) gene, which encodes a subunit of Skp1-Cul1-F-box protein (SCF) ubiquitin ligase, is frequently mutated in cervical squamous cell carcinomas. In this study, we investigated whether Chinese cervical cancer cells also harbor these mutations. Methods: Using PCR and sequencing assays, a total of 190 specimens from Han Chinese patients with cervical cancer were analyzed for FBXW7 mutations. Results: Two FBXW7 mutations (p.R479P and p.L443H), were identified from a study of 145 (1.4%) cervical squamous cell carcinomas. The p.L443H somatic mutation has not been previously reported. Functional assays showed that both of these FBXW7 mutations could promote cell proliferation, migration, and invasion. Conclusion: A low frequency (1.4%) of cervical squamous cell carcinomas were identified with FBXW7 mutations. We did, however, identify a novel FBXW7 mutation. Our results also demonstrated that the identified FBXW7 mutations could promote cell proliferation, migration, and invasion in cervical cancer cells.

Physiological functions of FBW7 in cancer and metabolism.

FBW7 is one of the most well characterized F-box proteins that serve as substrate recognition subunits of SCF (Skp1-Cullin 1-F-box proteins) E3 ubiquitin ligase complexes. SCFFBW7 plays key roles in regulating cell cycle progression, differentiation, and stem cell maintenance largely through targeting a broad range of oncogenic substrates for proteasome-dependent degradation. The identification of an increasing number of FBW7 substrates for ubiquitination, and intensive in vitro and in vivo studies have revealed a network of signaling components controlled by FBW7 that contributes to metabolic regulation as well as its tumor suppressor role. Here we mainly focus on recent findings that highlight a critical role for FBW7 in cancer and metabolism.