Date of Completion

6-8-2011

Embargo Period

7-7-2011

Advisors

Richard Bruno; Ji-Young Lee

Field of Study

Nutritional Science

Degree

Master of Science

Open Access

Open Access

Abstract

LIV-1, the estrogen-regulated gene encoding a member of the ZIP family of zinc transporters (SLC39A6), has been implicated in both growth and metastasis of estrogen-receptor (ER) positive (+) and negative (-) breast cancer cells. In ER+ breast cancer cells, LIV-1 expression is induced by both estrogen and insulin. In embryonic cells, LIV-1 has also been negatively associated with the expression of the cell adhesion protein, E-cadherin (CDH1), thought to play a role in metastasis. In the present study, we investigated the regulation of LIV-1 and its relationship to CDH1 in MDA-MB-231 ER- breast cancer cells. Cells were treated with insulin and EGF for 24 hours and the mRNA concentrations of LIV-1 and CDH1 were analyzed by RT-PCR. EGF treatment induced LIV-1 mRNA by about 2-fold whereas insulin had minor effects. The mRNA expression of CDH1 paralleled that of LIV-1 with these treatments. MDA-MB-231 cells were also transfected with an shRNA construct designed to knock down LIV-1 expression. This procedure reduced LIV-1 mRNA concentrations by approximately 40% and expression of CDH1 was reduced to a similar extent. This positive association between LIV-1 and CDH1 expression in MDA-MB-231 cells suggests that LIV-1 may be a regulator of CDH1 in ER- cells.

To further understand the linkage between LIV-1 and CDH1, expression of SNAI1, thought to be a transcriptional repressor of CDH1, was measured. However, neither EGF nor LIV-1 knockdown altered the expression of SNAI1 suggesting that this factor does not mediate influences of LIV-1 on CDH1 in these cells. We next investigated the expression of another Snail member, SNAI2, to explain the observed effects. EGF tended to decrease SNAI2 expression and LIV-1 knockdown increased its expression. Thus SNAI2 may be a modulator of a more aggressive and hormonally independent type of breast cancer cells. We also used 65Zn to characterize the role of LIV-1 in zinc uptake. Knockdown of LIV-1 reduced 65Zn uptake in parallel to the reduction in LIV-1 expression, demonstrating the important role of this transporter for zinc homeostasis in MDA-MB-231 cells. Overall, our current findings suggest that LIV-1 is regulated by the EGFR signaling pathway in ER- breast cancer cells. SNAI2 is inversely regulated by LIV-1 to repress CDH1, increasing the potential for metastasis.

Major Advisor

Hedley C. Freake

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