ABSTRACT
Although mounting evidence indicates the involvement of galectin-3 in cancer progression and metastasis, the underlying molecular mechanisms remain largely unknown. In this study, we investigated the effect and possible mechanism of galectin-3 on the migration and invasion of B16F10, a metastatic melanoma cell line, in which galectin-3 and matrix metalloproteinase-1 (MMP-1) were both found to be highly expressed. Knockdown of galectin-3 with specific siRNA reduced migration and invasion, which was associated with reduced expression of MMP-1. To further investigate the underlying mechanism, we examined the effect of galectin-3 knockdown on the activity of AP-1, a transcriptional factor regulating MMP-1 expression. We found that galectin-3 directly interacted with AP-1 and facilitated the binding of this complex to the MMP-1 promoter that drives MMP-1 transcription. Moreover, silencing of galectin-3 inhibited binding of fra-1 and c-Jun to promoter sites of MMP-1 gene. Consistent with these in vitro findings, our in vivo study demonstrated that galectin-3 shRNA treatment significantly reduced the total number of mouse lung metastatic nodules. Taken together, galectin-3 facilitates cell migration and invasion in melanoma in vitro and can induce metastasis in vivo, in part through, regulating the transcription activity of AP-1 and thereby up-regulating MMP-1 expression.
Subject(s)
Animals , Mice , Binding Sites/genetics , Cell Line, Tumor , Cell Movement , Cell Proliferation , Galectin 3/genetics , JNK Mitogen-Activated Protein Kinases/metabolism , Lung Neoplasms/drug therapy , Matrix Metalloproteinase 1/genetics , Melanoma, Experimental/metabolism , Mice, Inbred C57BL , NIH 3T3 Cells , Neoplasm Metastasis , Promoter Regions, Genetic , Proto-Oncogene Proteins c-fos/metabolism , RNA Interference , RNA, Small Interfering , Transcription Factor AP-1/genetics , Transcription, Genetic , Transcriptional ActivationABSTRACT
<p><b>OBJECTIVE</b>To investigate apoptosis-inducing effect and its mechanisms of HY-1, a carbazole alkaloid, on human erythroleukemia K562 cells.</p><p><b>METHODS</b>Cell proliferation was detected by sulforhodamine B (SRB) assay after treated with HY-1 at indicated doses. Cell cycle analysis was performed by flow cytometry, mitochondria membrane voltage change was assessed by rhodamine 123 staining, annexin V-PI apoptosis detecting kit and DNA agarose gel electrophoresis were used to identify apoptosis-inducing effect of HY-1. The alterations of apoptosis-relating proteins were detected by Western blot.</p><p><b>RESULTS</b>The IC(50) of HY-1 in K562 cells was (29.05 +/- 0.90) micromol/L by SRB assay. HY-1 had significant apoptotic inducing effect on K562 cells in a dose- and time-dependent manner as verified by appearance of Sub-G(1) peak on histogram of flow cytometry analysis, reduction of mitochondria membrane voltage, appearance of double positive cell group in Annexin V-PI apoptosis detecting test, and remarkable DNA ladder. The expression of cytosolic cytochrome c was apparently increased. Pro-caspase-9, pro-caspase-3 and PARP were all cleaved to active segments. There was no change in the expression of caspase-8.</p><p><b>CONCLUSION</b>HY-1 exerts its anticancer activity through triggering apoptosis of K562 cells by mitochondria-activating pathways.</p>