MicroRNA-antagonism regulates breast cancer stemness and metastasis via TET-family-dependent chromatin remodeling.

Tumor cells metastasize to distant organs through genetic and epigenetic alterations, including changes in microRNA (miR) expression. Here we find miR-22 triggers epithelial-mesenchymal transition (EMT), enhances invasiveness and promotes metastasis in mouse xenografts. In a conditional mammary gland-specific transgenic (TG) mouse model, we show that miR-22 enhances mammary gland side-branching, expands the stem cell compartment, and promotes tumor development. Critically, miR-22 promotes aggressive metastatic disease in MMTV-miR-22 TG mice, as well as compound MMTV-neu or -PyVT-miR-22 TG mice. We demonstrate that miR-22 exerts its metastatic potential by silencing antimetastatic miR-200 through direct targeting of the TET (Ten eleven translocation) family of methylcytosine dioxygenases, thereby inhibiting demethylation of the mir-200 promoter. Finally, we show that miR-22 overexpression correlates with poor clinical outcomes and silencing of the TET-miR-200 axis in patients. Taken together, our findings implicate miR-22 as a crucial epigenetic modifier and promoter of EMT and breast cancer stemness toward metastasis.

Nuclear PTEN regulates the APC-CDH1 tumor-suppressive complex in a phosphatase-independent manner.

PTEN is a frequently mutated tumor suppressor gene that opposes the PI3K/AKT pathway through dephosphorylation of phosphoinositide-3,4,5-triphosphate. Recently, nuclear compartmentalization of PTEN was found as a key component of its tumor-suppressive activity; however its nuclear function remains poorly defined. Here we show that nuclear PTEN interacts with APC/C, promotes APC/C association with CDH1, and thereby enhances the tumor-suppressive activity of the APC-CDH1 complex. We find that nuclear exclusion but not phosphatase inactivation of PTEN impairs APC-CDH1. This nuclear function of PTEN provides a straightforward mechanistic explanation for the fail-safe cellular senescence response elicited by acute PTEN loss and the tumor-suppressive activity of catalytically inactive PTEN. Importantly, we demonstrate that PTEN mutant and PTEN null states are not synonymous as they are differentially sensitive to pharmacological inhibition of APC-CDH1 targets such as PLK1 and Aurora kinases. This finding identifies a strategy for cancer patient stratification and, thus, optimization of targeted therapies. PAPERCLIP:

Regulatory Mechanism of MicroRNA Expression in Cancer.

Altered gene expression is the primary molecular mechanism responsible for the pathological processes of human diseases, including cancer. MicroRNAs (miRNAs) are virtually involved at the post-transcriptional level and bind to 3' UTR of their target messenger RNA (mRNA) to suppress expression. Dysfunction of miRNAs disturbs expression of oncogenic or tumor-suppressive target genes, which is implicated in cancer pathogenesis. As such, a large number of miRNAs have been found to be downregulated or upregulated in human cancers and to function as oncomiRs or oncosuppressor miRs. Notably, the molecular mechanism underlying the dysregulation of miRNA expression in cancer has been recently uncovered. The genetic deletion or amplification and epigenetic methylation of miRNA genomic loci and the transcription factor-mediated regulation of primary miRNA often alter the landscape of miRNA expression in cancer. Dysregulation of the multiple processing steps in mature miRNA biogenesis can also cause alterations in miRNA expression in cancer. Detailed knowledge of the regulatory mechanism of miRNAs in cancer is essential for understanding its physiological role and the implications of cancer-associated dysfunction and dysregulation. In this review, we elucidate how miRNA expression is deregulated in cancer, paying particular attention to the cancer-associated transcriptional and post-transcriptional factors that execute miRNA programs.

miR-218 and miR-129 regulate breast cancer progression by targeting Lamins.

Breast cancer is the most frequently diagnosed life-threatening cancer in women. Triple-negative breast cancer (TNBC) has an aggressive clinical behavior, but the treatment of TNBC remains challenging. MicroRNAs (miRNAs) have emerged as a potential target for the diagnosis, therapy and prognosis of breast cancer. However, the precise role of miRNAs and their targets in breast cancer remain to be elucidated. Here we show that miR-218 is downregulated and miR-129 is upregulated in TNBC samples and their expressions confer prognosis to patients. Gain-of-function and loss-of-function analysis reveals that miR-218 has a tumor suppressive activity, while miR-129 acts as an oncomir in breast cancer. Notably, miR-218 and miR-129 directly target Lamin B1 and Lamin A, respectively, which are also found to be deregulated in human breast tumors. Finally, we demonstrate Lamins as the major factors in reliable miR-218 and miR-129 functions for breast cancer progression. Our findings uncover a new miRNA-mediated regulatory network for different Lamins and provide a potential therapeutic target for breast cancer.

MicroRNAs in the pathogenesis of myelodysplastic syndromes and myeloid leukaemia.

PURPOSE OF REVIEW: This review highlights recent insights into the roles of microRNAs (miRNAs) in pathogenesis of myeloid malignancies and tantalising prospects of miRNA therapy. RECENT FINDINGS: New roles for miRNAs in biological and disease processes are constantly being discovered. Although great effort has been put into identifying and cataloguing aberrantly expressed miRNAs in leukaemia, very little is known about the functional consequences of their deregulation in myeloid malignancies. This review will discuss the significance of powerful oncogenic miRNAs such as miR-22 in self-renewal and transformation of haematopoietic stem cells, as well as their ability to induce epigenetic alterations in the pathogenesis of the stem cell disorder myelodysplastic syndromes and myeloid leukaemia. SUMMARY: Improved understanding of biological roles of miRNAs in the pathogenesis of haematological malignancies will allow rational stratification of patients and provide new therapeutic entries for the treatment of myelodysplastic syndromes and leukaemia.

The correlation between the expression of ubiquitin-conjugating enzyme 2C and prostate cancer prognosis.

PURPOSE: Ubiquitin-conjugating enzyme E2 C (UBE2C) is known to show a causal relationship with cancer development and advancement. The role of UBE2C is to control the mitotic spindle checkpoint. Excess UBE2C has been identified in patients with advanced prostate cancer. The objective of the present study was to examine positive connections between the expression of UBE2C and prognostic factors for prostate cancer. METHODS: Prostate cancer patients' clinical data were analysed. Tissue microarrays (TMAs) were also performed for human prostate cancer tissues (n = 335) and adjacent non-neoplastic tissues (n = 22). TMA slides were incubated with antibodies against UBE2C. Cores were scored by a pathologist who was blind to cancer results. RESULTS: Of 335 prostate cancer patients, 200 could be assessed for biochemical recurrence, clinical recurrence, and overall survival. Human prostate cancer tissues showed higher expression of UBE2C than adjacent non-neoplastic tissues. High expression level of UBE2C showed a strong positive relationship with a high prostate-specific antigen (PSA), Gleason's score, and pathological stage of prostate cancer. Patients with a higher UBE2C grade demonstrated greater lymphatic engagement of prostate cancer than those with a lower UBE2C grade. CONCLUSION: The expression of UBE2C has positive correlations with several prognostic factors for prostate cancer. Thus, investigating the expression level of UBE2C staining is a promising tool for predicting prostate cancer prognosis.

The Correlations between the Intensity of Histopathological Ubiquitin-Specific Protease 11 Staining and Progression of Prostate Cancer.

BACKGROUND: Ubiquitin-specific protease 11 (USP11), one of the principal phosphatase and tensin homolog (PTEN) deubiquitinases, can reserve PTEN polyubiquitination to maintain PTEN protein integrity and inhibit PI3K/AKT pathway activation. The aim of the current study was to investigate the associations between immunohistochemical USP11 staining intensities and prognostic indicators in individuals with prostate cancer. METHODS: Tissue microarrays (TMAs) were performed for human prostate cancer and normal tissue (control) samples. Data on patient's age, Gleason score, plasma prostate-specific antigen (PSA) titer, disease stage, and presence of seminal vesicles, lymph nodes, and surgical margin involvement were collected. A pathologist who was blinded to the clinical outcome data scored the TMA for USP11 staining intensity as either positive or negative. RESULTS: Cancerous tissues exhibited lower USP11 staining intensity, whereas the neighboring benign peri-tumoral tissues showed higher USP11 staining intensity. The degree of USP11 staining intensity was lower in patients with a higher PSA titer, higher Gleason score, or more advanced disease stage. Patients who showed positive USP11 staining were more likely to have more optimal clinical and biochemical recurrence-free survival statistics. CONCLUSIONS: USP11 staining intensity in patients with prostate cancer is negatively associated with several prognostic factors such as an elevated PSA titer and a high Gleason score. It also reflects both biochemical and clinical recurrence-free survival in such patients. Thus, USP11 staining is a valuable prognostic factor in patients with prostate cancer.

The tumour suppressor RASSF1A promotes MDM2 self-ubiquitination by disrupting the MDM2-DAXX-HAUSP complex.

The tumour suppressor p53, which accumulates in response to DNA damage and induces cell-cycle arrest and apoptosis, has a key function in the maintenance of genome integrity. Under normal conditions, the antiproliferative effects of p53 are inhibited by MDM2, a ubiquitin ligase that promotes p53 ubiquitination and degradation. MDM2 is also self-ubiquitinated and degraded. Here, we show that the tumour suppressor RASSF1A regulates G(1)-S cell-cycle progression in a p53-dependent manner by promoting MDM2 self-ubiquitination and preventing p53 degradation. Importantly, RASSF1A associates with MDM2 and death-domain-associated protein (DAXX) in the nucleus, thereby disrupting the interactions between MDM2, DAXX, and the deubiquitinase, HAUSP, and enhancing the self-ubiquitin ligase activity of MDM2. Moreover, RASSF1A partially contributes to p53-dependent checkpoint activation at early time points in response to DNA damage. These findings reveal a new and important function for RASSF1A in regulating the p53-MDM2 pathway.

The equilibrium of tumor suppression: DUBs as active regulators of PTEN.

PTEN is among the most commonly lost or mutated tumor suppressor genes in human cancer. PTEN, a bona fide lipid phosphatase that antagonizes the highly oncogenic PI3K-AKT-mTOR pathway, is considered a major dose-dependent tumor suppressor. Although PTEN function can be compromised by genetic mutations in inherited syndromes and cancers, posttranslational modifications of PTEN may also play key roles in the dynamic regulation of its function. Notably, deregulated ubiquitination and deubiquitination lead to detrimental impacts on PTEN levels and subcellular partitioning, promoting tumorigenesis. While PTEN can be targeted by HECT-type E3 ubiquitin ligases for nuclear import and proteasomal degradation, studies have shown that several deubiquitinating enzymes, including HAUSP/USP7, USP10, USP11, USP13, OTUD3 and Ataxin-3, can remove ubiquitin from ubiquitinated PTEN in cancer-specific contexts and thus reverse ubiquitination-mediated PTEN regulation. Researchers continue to reveal the precise molecular mechanisms by which cancer-specific deubiquitinases of PTEN regulate its roles in the pathobiology of cancer, and new methods of pharmacologically for modulating PTEN deubiquitinases are critical areas of investigation for cancer treatment and prevention. Here, we assess the mechanisms and functions of deubiquitination as a recently appreciated mode of PTEN regulation and review the link between deubiquitinases and PTEN reactivation and its implications for therapeutic strategies.

The Positive Correlations between the Expression of Histopathological Ubiquitin-Conjugating Enzyme 2O Staining and Prostate Cancer Advancement.

BACKGROUND: The mTOR signaling pathway is inactivated by AMPK's tumor-suppressing function. It is recognized that ubiquitin conjugating enzyme 2O (UBE2O), which directly targets AMPK for ubiquitination and degradation, is intensified in human cancers. METHODS: This study investigated the clinical data about prostate cancer. Examination was also carried out into tissue microarrays (TMA) of human prostate cancer (n = 382) and adjacent non-neoplastic tissues around prostate cancer (n = 61). The TMA slides were incubated with antibodies against UBE2O, and the cores were scored by the pathologist blind to cancer results. RESULTS: Very strong positive correlations were identified between the expression of UBE2O staining and high PSA and pathological stage of prostate cancer. Cox's proportional hazard analysis established correlations between the following: (1) positive surgical margin and biochemical recurrence free survival, (2) PSA grade and clinical recurrence free survival, (3) regional lymph node positive and clinical recurrence free survival, (4) adjuvant treatment and overall survival, and (5) pathological T stage and overall survival. CONCLUSION: There is a positive correlation between the expression of UBE2O staining and prognosis for prostate cancer. Thus, a prostate cancer prognosis can be assessed with the expression of UBE2O staining.

NEAT1 is essential for metabolic changes that promote breast cancer growth and metastasis.

Accelerated glycolysis is the main metabolic change observed in cancer, but the underlying molecular mechanisms and their role in cancer progression remain poorly understood. Here, we show that the deletion of the long noncoding RNA (lncRNA) Neat1 in MMTV-PyVT mice profoundly impairs tumor initiation, growth, and metastasis, specifically switching off the penultimate step of glycolysis. Mechanistically, NEAT1 directly binds and forms a scaffold bridge for the assembly of PGK1/PGAM1/ENO1 complexes and thereby promotes substrate channeling for high and efficient glycolysis. Notably, NEAT1 is upregulated in cancer patients and correlates with high levels of these complexes, and genetic and pharmacological blockade of penultimate glycolysis ablates NEAT1-dependent tumorigenesis. Finally, we demonstrate that Pinin mediates glucose-stimulated nuclear export of NEAT1, through which it exerts isoform-specific and paraspeckle-independent functions. These findings establish a direct role for NEAT1 in regulating tumor metabolism, provide new insights into the Warburg effect, and identify potential targets for therapy.

Interplay between c-Src and the APC/C co-activator Cdh1 regulates mammary tumorigenesis.

The Anaphase Promoting Complex (APC) coactivator Cdh1 drives proper cell cycle progression and is implicated in the suppression of tumorigenesis. However, it remains elusive how Cdh1 restrains cancer progression and how tumor cells escape the inhibition of Cdh1. Here we report that Cdh1 suppresses the kinase activity of c-Src in an APC-independent manner. Depleting Cdh1 accelerates breast cancer cell proliferation and cooperates with PTEN loss to promote breast tumor progression in mice. Hyperactive c-Src, on the other hand, reciprocally inhibits the ubiquitin E3 ligase activity of APCCdh1 through direct phosphorylation of Cdh1 at its N-terminus, which disrupts the interaction between Cdh1 and the APC core complex. Furthermore, pharmacological inhibition of c-Src restores APCCdh1 tumor suppressor function to repress a panel of APCCdh1 oncogenic substrates. Our findings reveal a reciprocal feedback circuit of Cdh1 and c-Src in the crosstalk between the cell cycle machinery and the c-Src signaling pathway.

A muscle-specific UBE2O/AMPKα2 axis promotes insulin resistance and metabolic syndrome in obesity.

Ubiquitin-conjugating enzyme E2O (UBE2O) is expressed preferentially in metabolic tissues, but its role in regulating energy homeostasis has yet to be defined. Here we find that UBE2O is markedly upregulated in obese subjects with type 2 diabetes and show that whole-body disruption of Ube2o in mouse models in vivo results in improved metabolic profiles and resistance to high-fat diet-induced (HFD-induced) obesity and metabolic syndrome. With no difference in nutrient intake, Ube2o-/- mice were leaner and expended more energy than WT mice. In addition, hyperinsulinemic-euglycemic clamp studies revealed that Ube2o-/- mice were profoundly insulin sensitive. Through phenotype analysis of HFD mice with muscle-, fat-, or liver-specific knockout of Ube2o, we further identified UBE2O as an essential regulator of glucose and lipid metabolism programs in skeletal muscle, but not in adipose or liver tissue. Mechanistically, UBE2O acted as a ubiquitin ligase and targeted AMPKα2 for ubiquitin-dependent degradation in skeletal muscle; further, muscle-specific heterozygous knockout of Prkaa2 ablated UBE2O-controlled metabolic processes. These results identify the UBE2O/AMPKα2 axis as both a potent regulator of metabolic homeostasis in skeletal muscle and a therapeutic target in the treatment of diabetes and metabolic disorders.

PTEN self-regulates through USP11 via the PI3K-FOXO pathway to stabilize tumor suppression.

PTEN is a lipid phosphatase that antagonizes the PI3K/AKT pathway and is recognized as a major dose-dependent tumor suppressor. The cellular mechanisms that control PTEN levels therefore offer potential routes to therapy, but these are as yet poorly defined. Here we demonstrate that PTEN plays an unexpected role in regulating its own stability through the transcriptional upregulation of the deubiquitinase USP11 by the PI3K/FOXO pathway, and further show that this feedforward mechanism is implicated in its tumor-suppressive role, as mice lacking Usp11 display increased susceptibility to PTEN-dependent tumor initiation, growth and metastasis. Notably, USP11 is downregulated in cancer patients, and correlates with PTEN expression and FOXO nuclear localization. Our findings therefore demonstrate that PTEN-PI3K-FOXO-USP11 constitute the regulatory feedforward loop that improves the stability and tumor suppressive activity of PTEN.

Role of the tumor suppressor RASSF1A in Mst1-mediated apoptosis.

Mammalian sterile 20-like kinase 1 (Mst1) is activated by both caspase-mediated cleavage and phosphorylation in response to apoptotic stimuli, including Fas ligation. Here, we examined the possible role of the tumor suppressor RASSF1A in Mst1 activation and Mst1-mediated apoptosis induced by death receptor signaling. Immunoprecipitation and immunofluorescence analyses revealed that Mst1 was associated with RASSF1A in cultured mammalian cells, with both proteins colocalizing to microtubules throughout the cell cycle. Whereas purified recombinant RASSF1A inhibited the kinase activity of purified recombinant Mst1 in vitro, overexpression of RASSF1A increased the kinase activity of Mst1 in intact cells, suggesting that regulation of Mst1 by RASSF1A in vivo involves more than the simple association of the two proteins. Both the activation of Mst1 and the incidence of apoptosis induced by Fas ligation were markedly reduced in cells depleted of RASSF1A by RNA interference and were increased by restoration of RASSF1A expression in RASSF1A-deficient cells. Moreover, the stimulatory effect of RASSF1A overexpression on Fas-induced apoptosis was inhibited by depletion of Mst1. These findings indicate that RASSF1A facilitates Mst1 activation and thereby promotes apoptosis induced by death receptor signaling.

New Insights into the Role of E2s in the Pathogenesis of Diseases: Lessons Learned from UBE2O.

Intracellular communication via ubiquitin (Ub) signaling impacts all aspects of cell biology and regulates pathways critical to human development and viability; therefore aberrations or defects in Ub signaling can contribute to the pathogenesis of human diseases. Ubiquitination consists of the addition of Ub to a substrate protein via coordinated action of E1-activating, E2-conjugating and E3-ligating enzymes. Approximately 40 E2s have been identified in humans, and most are thought to be involved in Ub transfer; although little information is available regarding the majority of them, emerging evidence has highlighted their importance to human health and disease. In this review, we focus on recent insights into the pathogenetic roles of E2s (particularly the ubiquitin-conjugating enzyme E2O [UBE2O]) in debilitating diseases and cancer, and discuss the tantalizing prospect that E2s may someday serve as potential therapeutic targets for human diseases.

A new duet in cancer biology: AMPK the typical and UBE2O the atypical.

Ubiquitin-conjugating enzyme E2O (UBE2O) is upregulated in human cancers. We have demonstrated that genetic deletion or pharmacological blockade of UBE2O reduces tumorigenesis through inhibiting the mammalian target of rapamycin complex 1-hypoxia-inducible factor 1-α pathway. Critically, UBE2O targets adenosine monophosphate (AMP)-activated protein kinase-α 2 (AMPKα2) for ubiquitination and degradation. We thus suggest the UBE2O-AMPKα2 axis as a potential therapeutic target for cancer.

MicroRNA, an Antisense RNA, in Sensing Myeloid Malignancies.

Myeloid malignancies, including myelodysplastic syndromes and acute myeloid leukemia, are clonal diseases arising in hematopoietic stem or progenitor cells. In recent years, microRNA (miRNA) expression profiling studies have revealed close associations of miRNAs with cytogenetic and molecular subtypes of myeloid malignancies, as well as outcome and prognosis of patients. However, the roles of miRNA deregulation in the pathogenesis of myeloid malignancies and how they cooperate with protein-coding gene variants in pathological mechanisms leading to the diseases have not yet been fully understood. In this review, we focus on recent insights into the role of miRNAs in the development and progression of myeloid malignant diseases and discuss the prospect that miRNAs may serve as a potential therapeutic target for leukemia.

A UBE2O-AMPKα2 Axis that Promotes Tumor Initiation and Progression Offers Opportunities for Therapy.

UBE2O is localized in the 17q25 locus, which is known to be amplified in human cancers, but its role in tumorigenesis remains undefined. Here we show that Ube2o deletion in MMTV-PyVT or TRAMP mice profoundly impairs tumor initiation, growth, and metastasis, while switching off the metabolic reprogramming of tumor cells. Mechanistically, UBE2O specifically targets AMPKα2 for ubiquitination and degradation, and thereby promotes activation of the mTOR-HIF1α pathway. Notably, inactivation of AMPKα2, but not AMPKα1, abrogates the tumor attenuation caused by UBE2O loss, while treatment with rapamycin or inhibition of HIF1α ablates UBE2O-dependent tumor biology. Finally, pharmacological blockade of UBE2O inhibits tumorigenesis through the restoration of AMPKα2, suggesting the UBE2O-AMPKα2 axis as a potential cancer therapeutic target.

The APC/C E3 Ligase Complex Activator FZR1 Restricts BRAF Oncogenic Function.

BRAF drives tumorigenesis by coordinating the activation of the RAS/RAF/MEK/ERK oncogenic signaling cascade. However, upstream pathways governing BRAF kinase activity and protein stability remain undefined. Here, we report that in primary cells with active APCFZR1, APCFZR1 earmarks BRAF for ubiquitination-mediated proteolysis, whereas in cancer cells with APC-free FZR1, FZR1 suppresses BRAF through disrupting BRAF dimerization. Moreover, we identified FZR1 as a direct target of ERK and CYCLIN D1/CDK4 kinases. Phosphorylation of FZR1 inhibits APCFZR1, leading to elevation of a cohort of oncogenic APCFZR1 substrates to facilitate melanomagenesis. Importantly, CDK4 and/or BRAF/MEK inhibitors restore APCFZR1 E3 ligase activity, which might be critical for their clinical effects. Furthermore, FZR1 depletion cooperates with AKT hyperactivation to transform primary melanocytes, whereas genetic ablation of Fzr1 synergizes with Pten loss, leading to aberrant coactivation of BRAF/ERK and AKT signaling in mice. Our findings therefore reveal a reciprocal suppression mechanism between FZR1 and BRAF in controlling tumorigenesis.Significance: FZR1 inhibits BRAF oncogenic functions via both APC-dependent proteolysis and APC-independent disruption of BRAF dimers, whereas hyperactivated ERK and CDK4 reciprocally suppress APCFZR1 E3 ligase activity. Aberrancies in this newly defined signaling network might account for BRAF hyperactivation in human cancers, suggesting that targeting CYCLIN D1/CDK4, alone or in combination with BRAF/MEK inhibition, can be an effective anti-melanoma therapy. Cancer Discov; 7(4); 424-41. ©2017 AACR.See related commentary by Zhang and Bollag, p. 356This article is highlighted in the In This Issue feature, p. 339.