Language：English   Publishing type：Research paper (scientific journal)  

The transcriptional repressor, BCL11A, is involved in hematological malignancies, B-cell development, and fetal-to-adult hemoglobin switching. However, the molecular mechanism by which it promotes the development of myeloid leukemia remains largely unknown. We find that Bcl11a cooperates with the pseudokinase, Trib1, in the development of acute myeloid leukemia (AML). Bcl11a promotes the proliferation and engraftment of Trib1-expressing AML cells both in vitro and in vivo. ChIP-seq analysis showed that upon DNA-binding, Bcl11a is significantly associated with PU.1, an inducer of myeloid differentiation, and that Bcl11a represses several PU.1 target genes, such as Asb2, Clec5a, and Fcgr3. Asb2, as a Bcl11a target gene that modulates cytoskeleton and cell-cell interaction, plays a key role in Bcl11a-induced malignant progression. The repression of PU.1 target genes by Bcl11a is achieved by both sequence-specific DNA-binding activity and recruitment of corepressors by Bcl11a. Suppression of the corepressor components, HDAC and LSD1, reverses the repressive activity. Moreover, treatment of AML cells with the HDAC inhibitor, pracinostat, and LSD1 inhibitor, GSK2879552, resulted in growth inhibition both in vitro and in vivo. High BCL11A expression is associated with worse prognosis in human AML patients. Blocking of BCL11A expression upregulates the expression of PU.1 target genes, and inhibits the growth of HL-60 cells and their engraftment to the bone marrow, suggesting that BCL11A is involved in human myeloid malignancies via the suppression of PU.1 transcriptional activity.

DOI： 10.1182/bloodadvances.2021004558

PubMed

Authorship：Lead author   Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1038/s41375-021-01384-1

PubMed

Authorship：Lead author   Language：English   Publishing type：Research paper (scientific journal)  

The pseudokinase Trib1 functions as a myeloid oncogene that recruits the E3 ubiquitin ligase COP1 to C/EBPα and interacts with MEK1 to enhance extracellular signal-regulated kinase (ERK) phosphorylation. A close genetic effect of Trib1 on Hoxa9 has been observed in myeloid leukemogenesis, where Trib1 overexpression significantly accelerates Hoxa9-induced leukemia onset. However, the mechanism underlying how Trib1 functionally modulates Hoxa9 transcription activity is unclear. Herein, we provide evidence that Trib1 modulates Hoxa9-associated super-enhancers. Chromatin immunoprecipitation sequencing analysis identified increased histone H3K27Ac signals at super-enhancers of the Erg, Spns2, Rgl1, and Pik3cd loci, as well as increased messenger RNA expression of these genes. Modification of super-enhancer activity was mostly achieved via the degradation of C/EBPα p42 by Trib1, with a slight contribution from the MEK/ERK pathway. Silencing of Erg abrogated the growth advantage acquired by Trib1 overexpression, indicating that Erg is a critical downstream target of the Trib1/Hoxa9 axis. Moreover, treatment of acute myeloid leukemia (AML) cells with the BRD4 inhibitor JQ1 showed growth inhibition in a Trib1/Erg-dependent manner both in vitro and in vivo. Upregulation of ERG by TRIB1 was also observed in human AML cell lines, suggesting that Trib1 is a potential therapeutic target of Hoxa9-associated AML. Taken together, our study demonstrates a novel mechanism by which Trib1 modulates chromatin and Hoxa9-driven transcription in myeloid leukemogenesis.

DOI： 10.1182/blood.2019004586

PubMed

Language：Japanese   Publishing type：Research paper (scientific journal)  

Cancer cells adapt to various stress conditions by optimizing gene expression profiles via transcriptional and translational regulation. However, whether and how EXOSC9, a component of the RNA exosome complex, regulates adaptation to stress conditions and tumorigenicity in cancer cells remain unclear. Here, we examined the effects of EXOSC9 depletion on cancer cell growth under various stress conditions. EXOSC9 depletion attenuated growth and survival under various stress conditions in cancer cells. Interestingly, this also decreased the number of P-bodies, which are messenger ribonucleoprotein particles (mRNPs) required for stress adaptation. Meanwhile, EXOSC2/EXOSC4 depletion also attenuated P-body formation and stress resistance with decreased EXOSC9 protein. EXOSC9-mediated stress resistance and P-body formation were found to depend on the intact RNA-binding motif of this protein. Further, RNA-seq analyses identified 343 EXOSC9-target genes, among which, APOBEC3G contributed to defects in stress resistance and P-body formation in MDA-MB-231 cells. Finally, EXOSC9 also promoted xenografted tumor growth of MDA-MB-231 cells in an intact RNA-binding motif-dependent manner. Database analyses further showed that higher EXOSC9 activity, estimated based on the expression of 343 target genes, was correlated with poorer prognosis in some cancer patients. Thus, drugs targeting activity of the RNA exosome complex or EXOSC9 might be useful for cancer treatment.

DOI： 10.1038/s41598-020-66455-2

Web of Science

Scopus

PubMed

Language：Japanese   Publishing type：Research paper (scientific journal)  

Unlike most cells, cancer cells activate hypoxia inducible factor-1 (HIF-1) to use glycolysis even at normal oxygen levels, or normoxia. Therefore, HIF-1 is an attractive target in cancer therapy. However, the regulation of HIF-1 during normoxia is not well characterised, although Mint3 was recently found to activate HIF-1 in cancer cells and macrophages by suppressing the HIF-1 inhibitor, factor inhibiting HIF-1 (FIH-1). In this study, we analysed Mint3-binding proteins to investigate the mechanism by which Mint3 regulates HIF-1. Yeast two-hybrid screening using Mint3 as bait identified N-terminal EF-hand calcium binding protein 3 (NECAB3) as a novel factor regulating HIF-1 activity via Mint3. NECAB3 bound to the phosphotyrosine-binding domain of Mint3, formed a ternary complex with Mint3 and FIH-1, and co-localised with Mint3 at the Golgi apparatus. Depletion of NECAB3 decreased the expression of HIF-1 target genes and reduced glycolysis in normoxic cancer cells. NECAB3 mutants that binds Mint3 but lacks an intact monooxygenase domain also inhibited HIF-1 activation. Inhibition of NECAB3 in cancer cells by either expressing shRNAs or generating a dominant negative mutant reduced tumourigenicity. Taken together, the data indicate that NECAB3 is a promising new target for cancer therapy.

DOI： 10.1038/srep22784

Web of Science

Scopus

PubMed

Authorship：Lead author   Language：English   Publishing type：Research paper (scientific journal)  

Loss of anchorage to the extracellular matrix leads to apoptosis (anoikis) in normal cells, but cancerous cells are usually resistant to such stress. Here we report the pivotal role of an E3 ubiquitin ligase, ring-finger protein 126 (RNF126), in the resistance of cancer cells to the stress associated with non-adherent conditions. Non-adherent cancer cells exhibited increased flux through the tricarboxylic acid cycle via increased conversion of pyruvate to acetyl-CoA. RNF126 was found to act as a ubiquitin ligase for pyruvate dehydrogenase kinases (PDKs), resulting in their proteasomal degradation. This decrease in PDK levels allowed pyruvate dehydrogenases to catalyze the conversion of pyruvate to acetyl-CoA. Moreover, depletion of RNF126 or increased expression of PDK1 in cancer cells suppressed colony formation in soft agar as well as tumorigenicity in mice. RNF126 expression in cancer cells was found to be under the control of the extracellular signal-regulated kinase signaling pathway, which is essential for anoikis resistance. Thus, RNF126 is an attractive molecule for treating cancer by selectively targeting anchorage-independent growth.

DOI： 10.1038/celldisc.2016.19

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

Hypoxia-inducible factor 1 (HIF-1) plays a key role in the cellular adaptation to hypoxia. Although HIF-1 is usually strongly suppressed by posttranslational mechanisms during normoxia, HIF-1 is active and enhances tumorigenicity in malignant tumor cells that express the membrane protease MT1-MMP. The cytoplasmic tail of MT1-MMP, which can bind a HIF-1 suppressor protein called factor inhibiting HIF-1 (FIH-1), promotes inhibition of FIH-1 by Mint3 during normoxia. To explore possible links between HIF-1 activation by MT1-MMP/Mint3 and tumor growth signals, we surveyed a panel of 252 signaling inhibitors. The mTOR inhibitor rapamycin was identified as a possible modulator, and it inhibited the mTOR-dependent phosphorylation of Mint3 that is required for FIH-1 inhibition. A mutant Mint3 protein that cannot be phosphorylated exhibited a reduced ability to inhibit FIH-1 and promoted tumor formation in mice. These data suggest a novel molecular link between the important hub proteins MT1-MMP and mTOR that contributes to tumor malignancy.

DOI： 10.1128/MCB.01169-13

PubMed

Authorship：Lead author   Language：English   Publishing type：Research paper (scientific journal)  

Oxygen is a vital requirement for multi-cellular organisms to generate energy and cells have developed multiple compensatory mechanisms to adapt to stressful hypoxic conditions. Such adaptive mechanisms are intricately interconnected with other signaling pathways that regulate cellular functions such as cell growth. However, our understanding of the overall system governing the cellular response to the availability of oxygen remains limited. To identify new genes involved in the response to hypoxic stress, we have performed a genome-wide gene knockdown analysis in human lung carcinoma PC8 cells using an shRNA library carried by a lentiviral vector. The knockdown analysis was performed under both normoxic and hypoxic conditions to identify shRNA sequences enriched or lost in the resulting selected cell populations. Consequently, we identified 56 candidate genes that might contribute to the cellular response to hypoxia. Subsequent individual knockdown of each gene demonstrated that 13 of these have a significant effect upon oxygen-sensitive cell growth. The identification of BCL2L1, which encodes a Bcl-2 family protein that plays a role in cell survival by preventing apoptosis, validates the successful design of our screen. The other selected genes have not previously been directly implicated in the cellular response to hypoxia. Interestingly, hypoxia did not directly enhance the expression of any of the identified genes, suggesting that we have identified a new class of genes that have been missed by conventional gene expression analyses to identify hypoxia response genes. Thus, our genetic screening method using a genome-wide shRNA library and the newly-identified genes represent useful tools to analyze the cellular systems that respond to hypoxic stress.

DOI： 10.1371/journal.pone.0035590

PubMed

Authorship：Lead author  

Language：Japanese   Publisher：(有)科学評論社  

Authorship：Lead author   Publisher：(有)科学評論社  

Authorship：Lead author