Person:

Wei, Wenyi

Loss of the Fbw7 tumor suppressor is common in diverse human cancer types, including T-Cell Acute Lymphoblastic Leukemia (T-ALL), although the mechanistic basis of its anti-oncogenic activity remains largely unclear. We recently reported that SCF(^{Fbw7}) regulates cellular apoptosis by controlling the ubiquitination and destruction of the pro-survival protein, Mcl-1, in a GSK3 phosphorylation-dependent manner. We found that human T-ALL cell lines displayed a close relationship between Fbw7 loss and Mcl-1 overexpression. More interestingly, T-ALL cell lines that are deficient in Fbw7 are particularly sensitive to sorafenib, a multi-kinase inhibitor that has been demonstrated to reduce Mcl-1 expression through an unknown mechanism. On the other hand, Fbw7-deficient T-ALL cell lines are much more resistant to the Bcl-2 antagonist, ABT-737. Furthermore, reconstitution of Fbw7 or depletion of Mcl-1 in Fbw7-deficient cells restores ABT-737 sensitivity, suggesting that elevated Mcl-1 expression is important for Fbw7-deficient cells to evade apoptosis. Therefore, our work provides a novel molecular mechanism for the tumor suppression function of Fbw7. Furthermore, it provides the rationale for targeted usage of Mcl-1 antagonists to treat Fbw7-deficient T-ALL patients.

The Mdm2/p53 pathway is compromised in more than 50% of all human cancers, therefore it is an intensive area of research to understand the upstream regulatory pathways governing Mdm2/p53 activity. Mdm2 is frequently overexpressed in human cancers while the molecular mechanisms underlying the timely destruction of Mdm2 remain unclear. We recently reported that Casein Kinase I phosphorylates Mdm2 at multiple sites to trigger Mdm2 interaction with, and subsequent ubiquitination and destruction by the SCF(^{\beta-TRCP}) E3 ubiquitin ligase. We also demonstrated that the E3 ligase activity-deficient Mdm2 was still unstable in the G1 phase and could be efficiently degraded by SCF(^{\beta-TRCP}). Thus our finding expands the current knowledge on how Mdm2 is tightly regulated by both self- and SCF(^{\beta-TRCP})-dependent ubiquitination to control p53 activity in response to stress. It further indicates that loss of β-TRCP or Casein Kinase I function contributes to elevated Mdm2 expression that is frequently found in various types of tumors.

CYLD negatively regulates the NF-κB signaling pathway and osteoclast differentiation largely through antagonizing TNF receptor-associated factor (TRAF)-mediated K63-linkage polyubiquitination in osteoclast precursor cells. CYLD activity is controlled by IκB kinase (IKK), but the molecular mechanism(s) governing CYLD protein stability remains largely undefined. Here, we report that SCFβ-TRCP regulates the ubiquitination and degradation of CYLD, a process dependent on prior phosphorylation of CYLD at Ser432/Ser436 by IKK. Furthermore, depletion of β-TRCP induced CYLD accumulation and TRAF6 deubiquitination in osteoclast precursor cells, leading to suppression of RANKL-induced osteoclast differentiation. Therefore, these data pinpoint the IKK/β-TRCP/CYLD signaling pathway as an important modulator of osteoclastogenesis.

Anaphase-promoting complex/cyclosome/Cdh1 is a multi-subunit ubiquitin E3 ligase that drives M to G1 cell cycle progression through primarily earmarking various substrates for ubiquitination and subsequent degradation by the 26S proteasome. Notably, emerging evidence suggested that Cdh1 could also function in various cellular processes independent of anaphase-promoting complex/cyclosome. To this end, we recently identified an anaphase-promoting complex/cyclosome-independent function of Cdh1 in modulating osteoblast differentiation through activating Smurf1, one of the NEDD4 family of HECT domain-containing E3 ligases. However, it remains largely unknown whether Cdh1 could exert its tumor suppressor role through similarly modulating the E3 ligase activities of other NEDD4 family members, most of which have characterized important roles in tumorigenesis. Here we report that in various tumor cells, Cdh1, conversely, suppresses the E3 ligase activity of WWP2, another NEDD4 family protein, in an anaphase-promoting complex/cyclosome-independent manner. As such, loss of Cdh1 activates WWP2, leading to reduced abundance of WWP2 substrates including PTEN, which subsequently activates PI3K/Akt oncogenic signaling to facilitate tumorigenesis. This study expands the non-anaphase-promoting complex/cyclosome function of Cdh1 in regulating the NEDD4 family E3 ligases, and further suggested that enhancing Cdh1 to inhibit the E3 ligase activity of WWP2 could be a promising strategy for treating human cancers.

The Mdm2 oncoprotein promotes p53 ubiquitination and destruction. Yet, exact molecular mechanisms of Mdm2 destruction itself, under DNA damaging conditions, remain unclear. Recently, we identified SCFβ-TRCP as a novel E3 ligase that targets Mdm2 for ubiquitination and destruction in a Casein Kinase Iδ (CKIδ)-dependent manner. However, it remains elusive how the β-TRCP/CKIδ/Mdm2 signaling axis is regulated by DNA damage signals to govern p53 activity. Consistent with previous studies, we found that inactivation of the Ataxia Telangiectasia Mutated (ATM) kinase, in turn, impaired DNA damage-induced Mdm2 destruction. Although phosphorylation of Mdm2 at Ser395 (an ATM phosphorylation site) facilitated Mdm2 interaction with β-TRCP, Ser395A-Mdm2 was degraded non-distinguishably from WT-Mdm2 by SCFβ-TRCP upon DNA damaging treatments. This indicates that in addition to phosphorylating Mdm2 at Ser395, ATM may govern Mdm2 stability through other unknown mechanisms. We further demonstrated that DNA damage-induced activation of ATM directly phosphorylated CKIδ at two well-conserved S/TQ sites, which promotes CKIδ nuclear localization to increase CKIδ-mediated phosphorylation of Mdm2, thereby facilitating subsequent Mdm2 ubiquitination by SCFβ-TRCP. Our studies provide a molecular mechanism of how ATM could govern DNA damage-induced destruction of Mdm2 in part by phosphorylating both Mdm2 and CKIδ to modulate SCFβ-TRCP–mediated Mdm2 ubiquitination. Given the pivotal role of Mdm2 in the negative regulation of p53, this work will also provide a rationale for developing CKIδ or ATM agonists as anti-cancer agents.

Metastasis suppressor 1 (MTSS1) is an important tumor suppressor protein, and loss of MTSS1 expression has been observed in several types of human cancers. Importantly, decreased MTSS1 expression is associated with more aggressive forms of breast and prostate cancers, and with poor survival rate. Currently, it remains unclear how MTSS1 is regulated in cancer cells, and whether reduced MTSS1 expression contributes to elevated cancer cell proliferation and migration. Here we report that the SCFβ-TRCP regulates MTSS1 protein stability by targeting it for ubiquitination and subsequent destruction via the 26S proteasome. Notably, depletion of either Cullin 1 or β-TRCP1 led to increased levels of MTSS1. We further demonstrated a crucial role for Ser322 in the DSGXXS degron of MTSS1 in governing SCFβ-TRCP-mediated MTSS1 degradation. Mechanistically, we defined that Casein Kinase Iδ (CKIδ) phosphorylates Ser322 to trigger MTSS1's interaction with β-TRCP for subsequent ubiquitination and degradation. Importantly, introducing wild-type MTSS1 or a non-degradable MTSS1 (S322A) into breast or prostate cancer cells with low MTSS1 expression significantly inhibited cellular proliferation and migration. Moreover, S322A-MTSS1 exhibited stronger effects in inhibiting cell proliferation and migration when compared to ectopic expression of wild-type MTSS1. Therefore, our study provides a novel molecular mechanism for the negative regulation of MTSS1 by β-TRCP in cancer cells. It further suggests that preventing MTSS1 degradation could be a possible novel strategy for clinical treatment of more aggressive breast and prostate cancers.

The incidence of human papillary thyroid cancer (PTC) is increasing and an aggressive subtype of this disease is resistant to treatment with vascular endothelial growth factor receptor 2 (VEGFR2) inhibitor. VEGFR2 promotes angiogenesis by triggering endothelial cell proliferation and migration. However, the molecular mechanisms governing VEGFR2 stability in vivo remain unknown. Additionally, whether VEGFR2 influences PTC cell migration is not clear. We show that the ubiquitin E3 ligase SCF(^{β-TRCP}) promotes ubiquitination and destruction of VEGFR2 in a casein kinase I (CKI)–dependent manner. β-TRCP knockdown or CKI inhibition causes accumulation of VEGFR2, resulting in increased activity of signaling pathways downstream of VEGFR2. β-TRCP–depleted endothelial cells exhibit enhanced migration and angiogenesis in vitro. Furthermore, β-TRCP knockdown increased angiogenesis and vessel branching in zebrafish. Importantly, we found an inverse correlation between β-TRCP protein levels and angiogenesis in PTC. We also show that β-TRCP inhibits cell migration and decreases sensitivity to the VEGFR2 inhibitor sorafenib in poorly differentiated PTC cells. These results provide a new biomarker that may aid a rational use of tyrosine kinase inhibitors to treat refractory PTC.

The Skp2 (S-phase kinase associated protein 2) oncoprotein is often highly expressed in various types of human cancers. However, the mechanistic basis of its oncogenic function, as well as the upstream regulatory pathway(s) that control Skp2 activities remains not fully understood. Recently, we reported that p300 acetylates Skp2 at two conserved lysine residues K68 and K71 within its NLS (Nuclear localization signal). This modification leads to increased Skp2 stability and cytoplasmic translocation, thus contributing to elevated Skp2 oncogenic potential. Moreover, we found that the SIRT3 tumor suppressor serves as the physiological deacetylase that antagonizes p300-mediated Skp2 acetylation. Furthermore, we showed that Skp2 governs E-cadherin ubiquitination and degradation in the cytosol. Consistent with this, we observed an inverse correlation between Skp2 and E-cadherin expression in clinical breast tumor samples. Therefore, our work elucidates a novel acetylation-dependent regulatory mechanism for Skp2 oncogenic functions.

The mechanistic target of rapamycin (mTOR) functions as a critical regulator of cellular growth and metabolism by forming multi-component, yet functionally distinct complexes mTORC1 and mTORC2. Although mTORC2 has been implicated in mTORC1 activation, little is known about how mTORC2 is regulated. Here we report that phosphorylation of Sin1 at T86 and T398 suppresses mTORC2 kinase activity by dissociating Sin1 from mTORC2. Importantly, Sin1 phosphorylation, triggered by S6K or Akt, in a cellular context-dependent manner, inhibits not only insulin/IGF-1-mediated, but also PDGF or EGF-induced Akt phosphorylation by mTORC2, demonstrating a negative regulation of mTORC2 independent of IRS-1 and Grb10. Lastly, a cancer patient-derived Sin1-R81T mutation impairs Sin1 phosphorylation, leading to hyper-mTORC2 activation via bypassing this negative regulation. Together, our work reveals a Sin1 phosphorylation-dependent mTORC2 regulation, providing a potential molecular mechanism by which mutations in the mTORC1/S6K/Sin1 signaling axis might cause aberrant hyper-activation of mTORC2/Akt that facilitates tumorigenesis.