Regulation of Nucleocytoplasmic Shuttling of the Transcriptional Regulator Human Mesoderm Induction Early Response 1 α in Breast Carcinoma Cells

Li, Shengnan (2018) Regulation of Nucleocytoplasmic Shuttling of the Transcriptional Regulator Human Mesoderm Induction Early Response 1 α in Breast Carcinoma Cells. Doctoral (PhD) thesis, Memorial University of Newfoundland.

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Temporal and spatial regulation of the subcellular distribution of transcriptional regulators is important to ensure their proper functioning in a cell. Mesoderm induction early response 1 α (MIER1α) has been implicated as a tumour suppressor in breast cancer. Analysis of MIER1α subcellular localization in breast samples revealed a stepwise translocation from the nucleus to the cytoplasm during progression to invasive carcinoma (McCarthy et al., 2008). Therefore, an investigation of MIER1α nucleocytoplasmic shuttling is critical to unraveling its role in breast cancer progression. Structurally, MIER1α has conserved domains found in a number of other transcriptional regulators, including N-terminal acidic stretches, ELM2 and SANT domains. However, none of these domains contain the predicted nuclear import or export signals. In this thesis, I show that MIER1α localizes in the nucleus in breast carcinoma MCF7 cells without an intrinsic nuclear localization signal (NLS). Although MIER1α has been shown to bind to ERα, active nuclear import of MIER1α is not through interaction with ERα; instead, it depends on interaction and co-transport with HDAC1/2 through a “piggyback” mechanism. Deletion analysis demonstrated that the entire ELM2 (aa164-283) is required and sufficient for nuclear targetting of MIER1α and that a simple mutation, 214W→A in the ELM2 domain abolishes both the interaction between MIER1α and HDAC1/2 and its nuclear localization. Further investigation revealed that MIER1α is exported out of the nucleus when cells are treated with insulin, IGF-1, EGF or FGF, but not with 17β-estradiol, and this export out of the nucleus is mediated by CRM1. HDAC1 & 2 nuclear localization were not affected by MIER1α export, suggesting they are only involved in MIER1α nuclear import. Both Mitogen-activated protein kinase (MAPK) and phosphoinositide 3-kinase B/Akt (PI3’K/AKT) pathways are activated upon treatment with growth factors, and it was further confirmed MIER1α nuclear export is triggered by the MAPK pathway, but not the PI3’K/AKT pathway. However, the mutation of predicted ERK1/2 consensus phosphorylation sites S10-P and/or S377-P motifs in the MIER1α sequence had no effect on its localization. MIER1α returns to the nucleus when activation of MAPK pathway diminishes, suggesting this process is transient and reversible. Deletion analysis narrowed the required sequence for export to the N-terminal region, aa1-163, containing acidic stretches. Overall, these results provide details of the mechanism responsible for MIER1α nucleocytoplasmic shuttling in a breast cancer carcinoma cell line; a similar mechanism may be operating during breast cancer progression.

Item Type: Thesis (Doctoral (PhD))
Item ID: 13406
Additional Information: Includes bibliographical references.
Keywords: MIER1; nucleocytoplasmic shuttling; HDAC; growth factor; ER
Department(s): Medicine, Faculty of > Biomedical Sciences
Date: May 2018
Date Type: Submission
Library of Congress Subject Heading: Active Transport, Cell Nucleus; Breast Neoplasms; Mesoderm -- Cytology.

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