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The Effect of EHD2on Breast Cancer Cell Proliferation and Movement

Author: WuMin
Tutor: NiuRuiFang
School: Tianjin Medical University
Course: Biochemistry and Molecular Biology
Keywords: EHD2 small interference RNA breast cancer proliferation chemotaxsis
CLC: R737.9
Type: Master's thesis
Year: 2009
Downloads: 9
Quote: 0
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Abstract


OBJECTIVEMembrane trafficking including receptor trafficking takes important role in life processing. Receptors employ divergent endocytic pathways of transportation, which involve shared or divergent adaptors and regulators, depending on the physicochemical characteristics of the cargos per se.EHD is a recently identified subfamily of genes (including EHD1,2,3,4) believed to serve as regulators of endocytic trafficking including endocytosis and recycling. Our study reveals that EHD2is widely expressed in various cell lines except in more than50%breast cancer cell lines, implying that EHD2might be involved in breast cancer development. The current project is aimed at understanding the EHD2function at regulating cell proliferation and migration, through RNAi knock-down of EHD2endogenous expression in breast cancer cell lines.METHOD1. Western Blot was used to detect the EHD2protein expression level in cell lines.2. Construction and screening of RNAi plasmid vectors targeting EHD2expression.3. Establish stable knock-down cell clones with decreased EHD2protein expression.4. MTT assay to evaluate the cell proliferation property.5. Wound-healing and Boyden transwell chemotaxis assay to evaluate cell migration capacity. RESULTS1. EHD2was expressed normally in several cell lines including MDA-MB-231, T47D, Bcap37and Hela.2. The plasmid construct pSilencer-CMV-46was identified effective at targeting EHD2expression.3. Established EHD2knock-down clones of T47D and Bcap37cells.4. EHD2knock-down decreased overall EGFR protein level.5. Cell proliferation assay (MTT) shown increased growth of EHD2knock-down cells.6. Wound-healing and chemotaxis assays revealed impaired migration capacity of the EHD2knock-down cells.CONCLUSIONDown-regulation of EHD2protein expression accelerated breast cancer cell line proliferation, while cell migration was significantly inhibited. EHD2might affect breast cancer development through hypothetical regulation of growth factor or chemokine receptor such as EGFR trafficking and signaling, thus worth further investigation as a potential tumor suppressor which might reveal novel diagnostic and/or treatment strategies for breast cancer.

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CLC: > Medicine, health > Oncology > Genitourinary tumors > Breast tumor
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