POST-TRANSLATIONAL REGULATION OF ZEB1 CONTRIBUTES TO TGFβ-MEDIATED EMT

Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related death in the United States due in part to the affinity of tumors to metastasize. Understanding the process which contributes to metastasis provides promise for the discovery of novel therapeutic targets. Epithelial-to-mesenchymal transition (EMT) is a proposed model for the initiation of metastasis. During EMT epithelial cells lose their cell adhesion properties and acquire a mesenchymal-like phenotype, allowing tumor cells to migrate from their epithelial cell community and invade remote locations. EMT is mediated by several signaling pathways, with transforming growth factor-beta (TGF-β) receiving attention for its up-regulation in the metastatic tumor microenvironment. TGF-β facilitates EMT through a variety of mechanisms but a prominent feature of TGF-β induced EMT is the activation of the transcription factor zinc finger E-box-binding homeobox 1 (Zeb1). Although Zeb1 has an established role in EMT the mechanism by which Zeb1 is regulated has not been fully elucidated. Here we demonstrate that Zeb1 undergoes post-translational modification and that this modification contributes to protein stability. We also expose novel Zeb1 interactions using the BioID method for proximity-dependent labeling of proteins. These results lead us to hypothesize that by targeting factors which mediate Zeb1 post-translational modification we may provide a therapeutic approach for metastasis suppression in NSCLC

K811 acetylation regulates ZEB1 dimerization, protein stability, and NuRD complex interactions to promote lung adenocarcinoma progression and metastasis

View full abstracthttps://openworks.mdanderson.org/leading-edge/1010/thumbnail.jp

Uncovering the ZEB1 Interactome to Identify Novel Regulators of Metastatic Non-small Cell Lung Cancer

Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related death in the United States, due in part to the robust affinity of lung cancer cells to metastasize. Understanding the processes that contribute to metastasis provides promise for the discovery of novel therapeutic targets. Epithelial-tomesenchymal transition (EMT) is a proposed model for the initiation of metastasis. During EMT cell adhesion and polarity is reduced, allowing epithelial cancer cells to dissociate from the primary tumor and invade distant organs. The transcription factor zinc finger E-box-binding homeobox 1 (ZEB1) has been reported to uniquely correlate with NSCLC disease progression and to confer therapy resistance in multiple tumor types. Additionally, depletion of ZEB1 has been found to reverse therapy resistance, hence uncovering regulators of ZEB1 provides promise for innovative therapeutic strategies that may improve lung cancer patient outcome. Recent publications demonstrate that ZEB1 undergoes post-translational modification, suggesting a method for regulating ZEB1 function; however, the extent to which ZEB1 is modified as well as the purpose of ZEB1 modification has not been fully elucidated. Here, we apply two independent screens- BioID and an Epigenome shRNA dropout screen- to define ZEB1 interactors that regulate post-translational modification and are critical to metastatic NSCLC. These screens revealed an interaction amongst ZEB1 and the histone deacetylase (HDAC)-containing nucleosome and deacetylase remodeling complex (NuRD). Through treatment with class I HDAC inhibitors Trichostatin A, we identified that ZEB1 homodimerizes and determine that acetylation at lysine residue 811 regulates this association. Furthermore, we identify the NuRD complex as a novel ZEB1 co-repressor and the Rab22 GTPase activating protein TBC1D2b as a ZEB1/NuRD complex target. We find that TBC1D2b suppresses E-cadherin internalization, thus hindering cancer cell invasion and metastasis. Ultimately, this project provides a novel regulatory node for ZEB1 function and insight to the role of EMT in endocytosis

Expression and effects of inhibition of type I insulin-like growth factor receptor tyrosine kinase in mantle cell lymphoma

BACKGROUND: Type I insulin-like growth factor receptor (IGF-IR) tyrosine kinase induces significant oncogenic effects. Strategies to block IGF-IR signaling are being tested in clinical trials that include patients with aggressive solid malignancies. Mantle cell lymphoma is a B-cell neoplasm with poor prognosis and a tendency to develop resistance. The expression and potential significance of IGF-IR in mantle cell lymphoma are not known. DESIGN AND METHODS: We used reverse transcriptase polymerase chain reaction, quantitative real-time polymerase chain reaction, immunoprecipitation, western blotting, flow cytometry, and immunohistochemistry to analyze the expression of IGF-IR mRNA, and IGF-IR and pIGF-IR proteins in mantle cell lymphoma cell lines and patients’ specimens. Selective and specific blockade of IGF-IR was achieved using picropodophyllin and short-interfering RNA, respectively. Cell viability, apoptosis, cell cycle, cellular morphology, cell proliferation, and target proteins were then analyzed. RESULTS: We detected the expression of IGF-IR and pIGF-IR in mantle cell lymphoma cell lines. Notably, IGF-IR molecules/cell were markedly increased in mantle cell lymphoma cell lines compared with human B-lymphocytes. IGF-IR and pIGF-IR were also detected in 78% and 74%, respectively, of 23 primary mantle cell lymphoma specimens. Treatment of serum-deprived mantle cell lymphoma cell lines with IGF-I salvaged these cells from apoptosis. Selective inhibition of IGF-IR by picropodophyllin decreased the viability and proliferation of mantle cell lymphoma cell lines, and induced apoptosis and cell cycle arrest. Selective inhibition of IGF-IR was associated with caspase-3, caspase-8, caspase-9, and PARP cleavage, cytochrome c release, up-regulation of cyclin B1, and down-regulation of cyclin D1, pCdc2, pIRS-1, pAkt, and pJnk. Similar results were obtained by using IGF-IR short-interfering RNA. In addition, picropodophyllin decreased the viability and proliferation of primary mantle cell lymphoma cells that expressed IGF-IR. CONCLUSIONS: IGF-IR is up-regulated and frequently activated in mantle cell lymphoma. Our data suggest that IGF-IR could be a molecular target for the treatment of mantle cell lymphoma