TY - JOUR A1 - Yanku, Yifat A1 - Bitman-Lotan, Eliya A1 - Zohar, Yaniv A1 - Kurant, Estee A1 - Zilke, Norman A1 - Eilers, Martin A1 - Orian, Amir T1 - Drosophila HUWE1 ubiquitin ligase regulates endoreplication and antagonizes JNK signaling during salivary gland development JF - Cells N2 - The HECT-type ubiquitin ligase HECT, UBA and WWE Domain Containing 1, (HUWE1) regulates key cancer-related pathways, including the Myc oncogene. It affects cell proliferation, stress and immune signaling, mitochondria homeostasis, and cell death. HUWE1 is evolutionarily conserved from Caenorhabditis elegance to Drosophila melanogaster and Humans. Here, we report that the Drosophila ortholog, dHUWE1 (CG8184), is an essential gene whose loss results in embryonic lethality and whose tissue-specific disruption establishes its regulatory role in larval salivary gland development. dHUWE1 is essential for endoreplication of salivary gland cells and its knockdown results in the inability of these cells to replicate DNA. Remarkably, dHUWE1 is a survival factor that prevents premature activation of JNK signaling, thus preventing the disintegration of the salivary gland, which occurs physiologically during pupal stages. This function of dHUWE1 is general, as its inhibitory effect is observed also during eye development and at the organismal level. Epistatic studies revealed that the loss of dHUWE1 is compensated by dMyc proeitn expression or the loss of dmP53. dHUWE1 is therefore a conserved survival factor that regulates organ formation during Drosophila development. KW - HECT KW - HUWE1 KW - ubiquitin KW - salivary gland KW - endoreplication KW - JNK KW - dMyc KW - dmP53 Y1 - 2018 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-197630 SN - 2073-4409 VL - 7 IS - 10 ER - TY - JOUR A1 - Wiegering, Armin A1 - Pfann, Christina A1 - Uthe, Friedrich Wilhelm A1 - Otto, Christoph A1 - Rycak, Lukas A1 - Mäder, Uwe A1 - Gasser, Martin A1 - Waaga-Gasser, Anna-Maria A1 - Eilers, Martin A1 - Germer, Christoph-Thomas T1 - CIP2A Influences Survival in Colon Cancer and Is Critical for Maintaining Myc Expression JF - PLoS ONE N2 - The cancerous inhibitor of protein phosphatase 2A (CIP2A) is an oncogenic factor that stabilises the c-Myc protein. CIP2A is overexpressed in several tumours, and expression levels are an independent marker for long-term outcome. To determine whether CIP2A expression is elevated in colon cancer and whether it might serve as a prognostic marker for survival, we analysed CIP2A mRNA expression by real-time PCR in 104 colon cancer samples. CIP2A mRNA was overexpressed in colon cancer samples and CIP2A expression levels correlated significantly with tumour stage. We found that CIP2A serves as an independent prognostic marker for disease-free and overall survival. Further, we investigated CIP2A-dependent effects on levels of c-Myc, Akt and on cell proliferation in three colon cancer cell lines by silencing CIP2A using small interfering (si) and short hairpin (sh) RNAs. Depletion of CIP2A substantially inhibited growth of colon cell lines and reduced c-Myc levels without affecting expression or function of the upstream regulatory kinase, Akt. Expression of CIP2A was found to be dependent on MAPK activity, linking elevated c-Myc expression to deregulated signal transduction in colon cancer. KW - caco-2 cells KW - carcinomas KW - colon KW - colorectal cancer KW - MAPK signaling cascades KW - metastasis KW - protein expression KW - small interferring RNA Y1 - 2013 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-97252 ER - TY - JOUR A1 - Teutschbein, Janka A1 - Haydn, Johannes M. A1 - Samans, Birgit A1 - Krause, Michael A1 - Eilers, Martin A1 - Schartl, Manfred A1 - Meierjohann, Svenja T1 - Gene expression analysis after receptor tyrosine kinase activation reveals new potential melanoma proteins N2 - Background: Melanoma is an aggressive tumor with increasing incidence. To develop accurate prognostic markers and targeted therapies, changes leading to malignant transformation of melanocytes need to be understood. In the Xiphophorus melanoma model system, a mutated version of the EGF receptor Xmrk (Xiphophorus melanoma receptor kinase) triggers melanomagenesis. Cellular events downstream of Xmrk, such as the activation of Akt, Ras, B-Raf or Stat5, were also shown to play a role in human melanomagenesis. This makes the elucidation of Xmrk downstream targets a useful method for identifying processes involved in melanoma formation. Methods: Here, we analyzed Xmrk-induced gene expression using a microarray approach. Several highly expressed genes were confirmed by realtime PCR, and pathways responsible for their induction were revealed using small molecule inhibitors. The expression of these genes was also monitored in human melanoma cell lines, and the target gene FOSL1 was knocked down by siRNA. Proliferation and migration of siRNA-treated melanoma cell lines were then investigated. Results: Genes with the strongest upregulation after receptor activation were FOS-like antigen 1 (Fosl1), early growth response 1 (Egr1), osteopontin (Opn), insulin-like growth factor binding protein 3 (Igfbp3), dual-specificity phosphatase 4 (Dusp4), and tumor-associated antigen L6 (Taal6). Interestingly, most genes were blocked in presence of a SRC kinase inhibitor. Importantly, we found that FOSL1, OPN, IGFBP3, DUSP4, and TAAL6 also exhibited increased expression levels in human melanoma cell lines compared to human melanocytes. Knockdown of FOSL1 in human melanoma cell lines reduced their proliferation and migration. Conclusion: Altogether, the data show that the receptor tyrosine kinase Xmrk is a useful tool in the identification of target genes that are commonly expressed in Xmrk-transgenic melanocytes and melanoma cell lines. The identified molecules constitute new possible molecular players in melanoma development. Specifically, a role of FOSL1 in melanomagenic processes is demonstrated. These data are the basis for future detailed analyses of the investigated target genes. KW - Melanoma Y1 - 2010 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-67900 ER - TY - JOUR A1 - Schülein-Völk, Christina A1 - Wolf, Elmar A1 - Zhu, Jing A1 - Xu, Wenshan A1 - Taranets, Lyudmyla A1 - Hellmann, Andreas A1 - Jänicke, Laura A. A1 - Diefenbacher, Markus E. A1 - Behrens, Axel A1 - Eilers, Martin A1 - Popov, Nikita T1 - Dual Regulation of Fbw7 Function and Oncogenic Transformation by Usp28 JF - CELL REPORTS N2 - Fbw7, the substrate recognition subunit of SCF(Fbw7) ubiquitin ligase, mediates the turnover of multiple proto-oncoproteins and promotes its own degradation. Fbw7-dependent substrate ubiquitination is antagonized by the Usp28 deubiquitinase. Here, we show that Usp28 preferentially antagonizes autocatalytic ubiquitination and stabilizes Fbw7, resulting in dose-dependent effects in Usp28 knockout mice. Monoallelic deletion of Usp28 maintains stable Fbw7 but drives Fbw7 substrate degradation. In contrast, complete knockout triggers Fbw7 degradation and leads to the accumulation of Fbw7 substrates in several tissues and embryonic fibroblasts. On the other hand, overexpression of Usp28 stabilizes both Fbw7 and its substrates. Consequently, both complete loss and ectopic expression of Usp28 promote Ras-driven oncogenic transformation. We propose that dual regulation of Fbw7 activity by Usp28 is a safeguard mechanism for maintaining physiological levels of proto-oncogenic Fbw7 substrates, which is equivalently disrupted by loss or overexpression of Usp28. KW - Fbw7 KW - oncogenic transformation Y1 - 2014 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-118219 SN - 2211-1247 N1 - The sequencing data have been submitted to the GEO repository under accession number GSE59354. VL - 9 IS - 3 ER - TY - JOUR A1 - Sanz-Moreno, Adrian A1 - Fuhrmann, David A1 - Wolf, Elmar A1 - von Eyss, Björn A1 - Eilers, Martin A1 - Elsässer, Hans-Peter T1 - Miz1 Deficiency in the Mammary Gland Causes a Lactation Defect by Attenuated Stat5 Expression and Phosphorylation JF - PLOS ONE N2 - Miz1 is a zinc finger transcription factor with an N-terminal POZ domain. Complexes with Myc, Bcl-6 or Gfi-1 repress expression of genes like Cdkn2b (p15(Ink4)) or Cd-kn1a (p21(Cip1)). The role of Miz1 in normal mammary gland development has not been addressed so far. Conditional knockout of the Miz1 POZ domain in luminal cells during pregnancy caused a lactation defect with a transient reduction of glandular tissue, reduced proliferation and attenuated differentiation. This was recapitulated in vitro using mouse mammary gland derived HC11 cells. Further analysis revealed decreased Stat5 activity in Miz1 Delta POZ mammary glands and an attenuated expression of Stat5 targets. Gene expression of the Prolactin receptor (PrlR) and ErbB4, both critical for Stat5 phosphorylation (pStat5) or pStat5 nuclear translocation, was decreased in Miz1 Delta POZ females. Microarray, ChIP-Seq and gene set enrichment analysis revealed a down-regulation of Miz1 target genes being involved in vesicular transport processes. Our data suggest that deranged intracellular transport and localization of PrlR and ErbB4 disrupt the Stat5 signalling pathway in mutant glands and cause the observed lactation phenotype. KW - C-MYC KW - transcription factor MIZ-1 KW - breast-cancer cells KW - gene expression KW - epithelial cells KW - prolactin KW - transgenic mice KW - growth KW - differentiation KW - proliferation Y1 - 2014 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-117286 VL - 9 IS - 2 ER - TY - JOUR A1 - Sander, Bodo A1 - Xu, Wenshan A1 - Eilers, Martin A1 - Popov, Nikita A1 - Lorenz, Sonja T1 - A conformational switch regulates the ubiquitin ligase HUWE1 JF - eLife N2 - The human ubiquitin ligase HUWE1 has key roles in tumorigenesis, yet it is unkown how its activity is regulated. We present the crystal structure of a C-terminal part of HUWE1, including the catalytic domain, and reveal an asymmetric auto-inhibited dimer. We show that HUWE1 dimerizes in solution and self-associates in cells, and that both occurs through the crystallographic dimer interface. We demonstrate that HUWE1 is inhibited in cells and that it can be activated by disruption of the dimer interface. We identify a conserved segment in HUWE1 that counteracts dimer formation by associating with the dimerization region intramolecularly. Our studies reveal, intriguingly, that the tumor suppressor p14ARF binds to this segment and may thus shift the conformational equilibrium of HUWE1 toward the inactive state. We propose a model, in which the activity of HUWE1 underlies conformational control in response to physiological cues—a mechanism that may be exploited for cancer therapy. KW - Medicine KW - Structural Biology KW - Molecular Biophysics KW - HUWE1 KW - HECT Ligase KW - Ubiquitin KW - P14ARF KW - X-Ray Chrystallography KW - Enzyme Regulation Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-171862 VL - 6 ER - TY - JOUR A1 - Prieto‐Garcia, Cristian A1 - Hartmann, Oliver A1 - Reissland, Michaela A1 - Braun, Fabian A1 - Fischer, Thomas A1 - Walz, Susanne A1 - Schülein‐Völk, Christina A1 - Eilers, Ursula A1 - Ade, Carsten P. A1 - Calzado, Marco A. A1 - Orian, Amir A1 - Maric, Hans M. A1 - Münch, Christian A1 - Rosenfeldt, Mathias A1 - Eilers, Martin A1 - Diefenbacher, Markus E. T1 - Maintaining protein stability of ∆Np63 via USP28 is required by squamous cancer cells JF - EMBO Molecular Medicine N2 - The transcription factor ∆Np63 is a master regulator of epithelial cell identity and essential for the survival of squamous cell carcinoma (SCC) of lung, head and neck, oesophagus, cervix and skin. Here, we report that the deubiquitylase USP28 stabilizes ∆Np63 and maintains elevated ∆NP63 levels in SCC by counteracting its proteasome‐mediated degradation. Impaired USP28 activity, either genetically or pharmacologically, abrogates the transcriptional identity and suppresses growth and survival of human SCC cells. CRISPR/Cas9‐engineered in vivo mouse models establish that endogenous USP28 is strictly required for both induction and maintenance of lung SCC. Our data strongly suggest that targeting ∆Np63 abundance via inhibition of USP28 is a promising strategy for the treatment of SCC tumours. KW - ∆Np63 KW - NOTCH KW - squamous cell carcinoma KW - 28 Y1 - 2020 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-218303 VL - 12 IS - 4 ER - TY - JOUR A1 - Peter, Stefanie A1 - Bultinck, Jennyfer A1 - Myant, Kevin A1 - Jaenicke, Laura A. A1 - Walz, Susanne A1 - Müller, Judith A1 - Gmachl, Michael A1 - Treu, Matthias A1 - Boehmelt, Guido A1 - Ade, Casten P. A1 - Schmitz, Werner A1 - Wiegering, Armin A1 - Otto, Christoph A1 - Popov, Nikita A1 - Sansom, Owen A1 - Kraut, Norbert A1 - Eilers, Martin T1 - H Tumor cell-specific inhibition of MYC function using small molecule inhibitors of the HUWE1 ubiquitin ligase JF - EMBO Molecular Medicine N2 - Deregulated expression of MYC is a driver of colorectal carcinogenesis, necessitating novel strategies to inhibit MYC function. The ubiquitin ligase HUWE1 (HECTH9, ARF-BP1, MULE) associates with both MYC and the MYC-associated protein MIZ1. We show here that HUWE1 is required for growth of colorectal cancer cells in culture and in orthotopic xenograft models. Using high-throughput screening, we identify small molecule inhibitors of HUWE1, which inhibit MYC-dependent transactivation in colorectal cancer cells, but not in stem and normal colon epithelial cells. Inhibition of HUWE1 stabilizes MIZ1. MIZ1 globally accumulates on MYC target genes and contributes to repression of MYC-activated target genes upon HUWE1 inhibition. Our data show that transcriptional activation by MYC in colon cancer cells requires the continuous degradation of MIZ1 and identify a novel principle that allows for inhibition of MYC function in tumor cells. KW - colorectal cancer KW - HUWE1 KW - MIZ1 KW - MYC KW - ubiquitination KW - cancer KW - digestive system KW - pharmacology KW - drug discovery Y1 - 2014 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-118132 SN - 1757-4684 VL - 6 IS - 12 ER - TY - JOUR A1 - Otto, Christoph A1 - Kastner, Carolin A1 - Schmidt, Stefanie A1 - Uttinger, Konstantin A1 - Baluapuri, Apoorva A1 - Denk, Sarah A1 - Rosenfeldt, Mathias T. A1 - Rosenwald, Andreas A1 - Roehrig, Florian A1 - Ade, Carsten P. A1 - Schuelein-Voelk, Christina A1 - Diefenbacher, Markus E. A1 - Germer, Christoph-Thomas A1 - Wolf, Elmar A1 - Eilers, Martin A1 - Wiegering, Armin T1 - RNA polymerase I inhibition induces terminal differentiation, growth arrest, and vulnerability to senolytics in colorectal cancer cells JF - Molecular Oncology N2 - Ribosomal biogenesis and protein synthesis are deregulated in most cancers, suggesting that interfering with translation machinery may hold significant therapeutic potential. Here, we show that loss of the tumor suppressor adenomatous polyposis coli (APC), which constitutes the initiating event in the adenoma carcinoma sequence for colorectal cancer (CRC), induces the expression of RNA polymerase I (RNAPOL1) transcription machinery, and subsequently upregulates ribosomal DNA (rDNA) transcription. Targeting RNAPOL1 with a specific inhibitor, CX5461, disrupts nucleolar integrity, and induces a disbalance of ribosomal proteins. Surprisingly, CX5461-induced growth arrest is irreversible and exhibits features of senescence and terminal differentiation. Mechanistically, CX5461 promotes differentiation in an MYC-interacting zinc-finger protein 1 (MIZ1)- and retinoblastoma protein (Rb)-dependent manner. In addition, the inhibition of RNAPOL1 renders CRC cells vulnerable towards senolytic agents. We validated this therapeutic effect of CX5461 in murine- and patient-derived organoids, and in a xenograft mouse model. These results show that targeting ribosomal biogenesis together with targeting the consecutive, senescent phenotype using approved drugs is a new therapeutic approach, which can rapidly be transferred from bench to bedside. KW - CRC KW - CX5461 KW - MIZ1 KW - MYC KW - ribosome KW - RNAPOL1 Y1 - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-312806 VL - 16 IS - 15 ER - TY - JOUR A1 - Muthalagu, Nathiya A1 - Junttila, Melissa R. A1 - Wiese, Kathrin E. A1 - Wolf, Elmar A1 - Morton, Jennifer A1 - Bauer, Barbara A1 - Evan, Gerard I. A1 - Eilers, Martin A1 - Murphy, Daniel J. T1 - BIM Is the Primary Mediator of MYC-Induced Apoptosis in Multiple Solid Tissues JF - Cell Reports N2 - MYC is one of the most frequently overexpressed oncogenes in human cancer, and even modestly deregulated MYC can initiate ectopic proliferation in many postmitotic cell types in vivo. Sensitization of cells to apoptosis limits MYC's oncogenic potential. However, the mechanism through which MYC induces apoptosis is controversial. Some studies implicate p19ARF-mediated stabilization of p53, followed by induction of proapoptotic BH3 proteins NOXA and PUMA, whereas others argue for direct regulation of BH3 proteins, especially BIM. Here, we use a single experimental system to systematically evaluate the roles of p19ARF and BIM during MYC-induced apoptosis, in vitro, in vivo, and in combination with a widely used chemotherapeutic, doxorubicin. We find a common specific requirement for BIM during MYC-induced apoptosis in multiple settings, which does not extend to the p53-responsive BH3 family member PUMA, and find no evidence of a role for p19ARF during MYC-induced apoptosis in the tissues examined. KW - ARF tumor-suppressor induced lymphomagenes KW - BCL-X-L P53 KW - C-MYC PUMA KW - in-vivo expression KW - cell-cycle arrest cancer therapy Y1 - 2014 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-115370 VL - 8 IS - 5 ER -