Our long-term goal is to understand the molecular mechanisms regulating disabled-2 (Dab2) protein expression levels. Dab2 is a cytoplasmic adaptor molecule that is associated with clathrin mediated endocytosis. It was originally identified as a protein down-regulated in ovarian carcinoma and it has since been established that its expression levels have significant impact on cellular proliferation, differentiation, and cancer development. It has been designated as a putative tumor suppressor due to its decreased expression levels in ovarian, breast, prostate and pancreatic carcinoma cells. We have obtained preliminary data suggesting that, in addition to transcriptional regulation, cellular Dab2 expression levels are also maintained through post-transcriptional mechanisms involving transcript-specific translational control and ubiquitin-mediated proteasomal degradation. In two cellular differentiation systems, a TGFbeta-mediated epithelial to mesenchymal transdifferentiation model in mouse mammary epithelial (NMuMG) cells and a retinoic acid (RA)-induced visceral endoderm differentiation model in F9 teratocarcinoma cells, we present data suggesting that transcriptional induction of Dab2 mRNA alone can not account for induced Dab2 protein expression levels. We demonstrate that under basal .conditions, Dab2 expression levels are translationally silenced through a mechanism driven by sequence elements contained in the Dab2 3'-untranslated region (3'-UTR). Cytosolic extracts from TGFbeta-treated NMuMG or RA-treated F9 cells can overcome this 3'- UTR-mediated silencing, resulting in increased Dab2 protein expression. We further show that once synthesized, Dab2 levels are regulated through a GSK-3beta-dependent phosphorylation and ubiquitin-mediated degradation pathway. These results demonstrate that Dab2 expression is highly regulated and support the hypothesis that its expression levels underlie its proliferate and differentiative functions. In this proposal, we wish to further explore the molecular mechanisms through which Dab2 expression levels are regulated and determine whether dysregulation of these mechanisms might underlie its aberrant expression in cancer.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA055536-19
Application #
7756616
Study Section
Tumor Cell Biology Study Section (TCB)
Program Officer
Salnikow, Konstantin
Project Start
1992-02-01
Project End
2012-01-31
Budget Start
2010-02-01
Budget End
2011-01-31
Support Year
19
Fiscal Year
2010
Total Cost
$308,158
Indirect Cost
Name
Cleveland Clinic Lerner
Department
Other Basic Sciences
Type
Schools of Medicine
DUNS #
135781701
City
Cleveland
State
OH
Country
United States
Zip Code
44195
Howley, Breege V; Howe, Philip H (2018) TGF-beta signaling in cancer: post-transcriptional regulation of EMT via hnRNP E1. Cytokine :
Lv, Zongyang; Rickman, Kimberly A; Yuan, Lingmin et al. (2017) S. pombe Uba1-Ubc15 Structure Reveals a Novel Regulatory Mechanism of Ubiquitin E2 Activity. Mol Cell 65:699-714.e6
Grelet, Simon; Link, Laura A; Howley, Breege et al. (2017) A regulated PNUTS mRNA to lncRNA splice switch mediates EMT and tumour progression. Nat Cell Biol 19:1105-1115
Howley, B V; Hussey, G S; Link, L A et al. (2016) Translational regulation of inhibin ?A by TGF? via the RNA-binding protein hnRNP E1 enhances the invasiveness of epithelial-to-mesenchymal transitioned cells. Oncogene 35:1725-35
Link, Laura A; Howley, Breege V; Hussey, George S et al. (2016) PCBP1/HNRNP E1 Protects Chromosomal Integrity by Translational Regulation of CDC27. Mol Cancer Res 14:634-46
Brown, Andrew S; Mohanty, Bidyut K; Howe, Philip H (2016) Identification and characterization of an hnRNP E1 translational silencing motif. Nucleic Acids Res 44:5892-907
Jiang, Yong; Woosley, Alec N; Howe, Philip H (2016) Disabled-2; an autophagic and apoptotic switch. Cell Cycle 15:3319-3320
Jiang, Yong; Woosley, Alec N; Sivalingam, Nageswaran et al. (2016) Cathepsin-B-mediated cleavage of Disabled-2 regulates TGF-?-induced autophagy. Nat Cell Biol 18:851-63
Brown, Andrew S; Mohanty, Bidyut K; Howe, Philip H (2015) Computational Identification of Post Translational Modification Regulated RNA Binding Protein Motifs. PLoS One 10:e0137696
Hussey, George S; Link, Laura A; Brown, Andrew S et al. (2012) Establishment of a TGF?-induced post-transcriptional EMT gene signature. PLoS One 7:e52624

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