p62/IMP2 is an oncofetal protein that is overexpressed in several types of cancer, and is a member of the family of insulin-like growth factor 2 mRNA binding proteins. breast cancer cell lines: MDA-MB-231 and LM2-4. Using assays we found that overexpression of p62/IMP2 can increase cell migration, and reduce cell adhesion to extracellular matrix (ECM) proteins. A Human Extracellular Matrix and Adhesion Molecules qPCR array was performed with our generated variants, and it identified a group of mRNAs whose expression was SKI-606 altered with p62/IMP2 overexpression, including SKI-606 connective tissue growth factor (CTGF) mRNA C which we show to be a p62/IMP2 binding partner. Overall, our results provide new insights into the molecular mechanism by which p62/IMP2 can contribute to breast cancer progression. assays to evaluate relative cell adhesion, cell migration, and cell proliferation, compared to controls. The results from the wound healing assay suggested an increased migration of p62/IMP2 positive cells compared to controls (Figure ?(Figure5A).5A). Thus, overexpression of p62/IMP2 in breast cancer cells increased wound closure by 50% to 70% (Figure ?(Figure5B).5B). We also tested cell adhesion to collagen, and to fibronectin. We found that p62/IMP2 positive cells were less adherent compared to control cells (Figure ?(Figure5C).5C). Thus overexpression of p62/IMP2 reduced cell adhesion by 30%-50% (Figure ?(Figure5D).5D). Relative cell growth was also examined using a proliferation assay, but no impact of p62/IMP2 expression on breast cancer cell proliferation was observed over a 6C7 day monitoring period (Figure ?(Figure5E).5E). Taken together, these data indicate that p62/IMP2 increases cell migration and reduces cell adhesion in breast cancer cells < 0.01). c-myc mRNA, which is a well-studied target of p62/IMP2, was used as a positive control (Figure ?(Figure7B7B and Figure ?Figure7C).7C). The half-life of c-myc mRNA was 0.3 0.19 hours in clone 5, and the half-life of c-myc mRNA in clone 6 was 0.8 0.23 hours (= 0.044). In summary, our data indicated that p62/IMP2 can bind to and stabilize the mRNA of CTGF. Figure 7 CTGF SKI-606 mRNA is a novel target of p62/IMP2 DISCUSSION The results of this study indicate that p62/IMP2 overexpression is observed in breast cancer tissues. Furthermore, by ELISA we observed a high frequency of anti-p62/IMP2 autoantibody in sera from breast cancer patients. A previous study demonstrated that p62/IMP2 is developmentally regulated and is expressed in fetal liver tissues but not in adult livers, and yet it is also overexpressed in liver tissues from patients with HCC [13]. Detectable autoantibody to p62/IMP2 was found to be present in 21.1% of HCC patients from China but not in patients with precursor conditions such as chronic hepatitis and liver cirrhosis [9]. Liu et al. found the overexpression of p62/IMP2 in colon cancer tissues by IHC [10]. In a Rabbit Polyclonal to TF2H1 study of 82 patients with digestive canal tumors, 38.6% of patients were positive for autoantibody to p62/IMP2, and 33.7% of those patients had metastatic disease, which suggests that the anti-p62/IMP2 autoantibody might be a predictive marker for cancer metastasis [20]. Overall these data indicate a role of p62/IMP2 in various human tumor types. Anti-TAA autoantibodies are stable and can readily be detected by using ELISA, which can make these autoantibodies as potential markers of diseases. And number of anti-TAA autoantibodies have been reported to be present in sera from cancer patients [21]. Furthermore, the elevation of anti-TAA autoantibody levels also correlates with the occurrence of some cancers, so the detection of the anti-TAA autoantibody can be used for the immunodiagnosis of some types of cancer [9, 11]. Furthermore, using p62/IMP2 transfected cells, we observed that p62/IMP2 can increase cell migration and reduce cell adhesion to the extracellular matrix. Since increased cell migration and reduced cell adhesion are important in the process of tumor cell invasion, and tumor spread [22, 23], our results suggest some mechanism by which p62/IMP2 overexpression may contribute to breast cancer progression. To uncover the mechanisms by which p62/IMP2 regulates cell migration and cell adhesion, a Human Extracellular Matrix & Adhesion Molecules RT2 Profiler PCR Array was carried out to observe mRNA changes regulated by p62/IMP2. We found that most of genes with altered expression.