Protein Kinase D1 Mediates Anchorage-dependent and -independent Growth of Tumor Cells via the Zinc Finger Transcription Factor Snail1.
ABSTRACT We here identify protein kinase D1 (PKD1) as a major regulator of anchorage-dependent and -independent growth of cancer cells controlled via the transcription factor Snail1. Using FRET, we demonstrate that PKD1, but not PKD2, efficiently interacts with Snail1 in nuclei. PKD1 phosphorylates Snail1 at Ser-11. There was no change in the nucleocytoplasmic distribution of Snail1 using wild type Snail1 and Ser-11 phosphosite mutants in different tumor cells. Regardless of its phosphorylation status or following co-expression of constitutively active PKD, Snail1 was predominantly localized to cell nuclei. We also identify a novel mechanism of PKD1-mediated regulation of Snail1 transcriptional activity in tumor cells. The interaction of the co-repressors histone deacetylases 1 and 2 as well as lysyl oxidase-like protein 3 with Snail1 was impaired when Snail1 was not phosphorylated at Ser-11, which led to reduced Snail1-associated histone deacetylase activity. Additionally, lysyl oxidase-like protein 3 expression was up-regulated by ectopic PKD1 expression, implying a synergistic regulation of Snail1-driven transcription. Ectopic expression of PKD1 also up-regulated proliferation markers such as Cyclin D1 and Ajuba. Accordingly, Snail1 and its phosphorylation at Ser-11 were required and sufficient to control PKD1-mediated anchorage-independent growth and anchorage-dependent proliferation of different tumor cells. In conclusion, our data show that PKD1 is crucial to support growth of tumor cells via Snail1.
- SourceAvailable from: Christoph Wille
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- "HeLa and Panc1 cells were seeded on glass coverslips, transfected as indicated, and processed as described in Eiseler et al. (2012). Samples were analyzed by a confocal laser scanning microscope, LSM710 (Zeiss, Jena, Germany) or TCS SP5 (Leica, Wetzlar, Germany ), equipped with respective 63× Plan Apo oil or 40× water immersion objective. "
ABSTRACT: Pancreatic cancer cell invasion, metastasis and angiogenesis are major challenges for the development of novel therapeutic strategies. Protein Kinase D (PKD) isoforms are involved in controlling tumor cell motility, angiogenesis and metastasis. In particular PKD2 expression is up-regulated in pancreatic cancer, whereas PKD1 expression is lower. We here report that both kinases control pancreatic cancer cell invasive properties in an isoform-specific manner. PKD2 enhances invasion in 3D-ECM cultures by stimulating expression and secretion of matrix-metalloproteinase 7 and 9 (MMP7/9), whereby MMP7 is likely to act upstream of MMP9. Knockdown of MMP7/9 blocks PKD2-mediated invasion in 3D-ECM assays and in-vivo utilizing tumors growing on chorioallantois membranes (CAM). Furthermore, MMP9 enhances PKD2-mediated tumor angiogenesis by releasing extracellular matrix-bound VEGF-A, thereby increasing its bio-availability and angiogenesis. Interestingly, specific knockdown of PKD1 in PKD2-expressing pancreatic cancer cells further enhanced the invasive properties in 3D-ECM systems by generating a high-motility phenotype. Loss of PKD1 thus may be beneficial for tumor cells to enhance their matrix-invading abilities. In conclusion, we define for the first time PKD1 and -2 isoform-selective effects on pancreatic cancer cell invasion and angiogenesis, in-vitro and in-vivo, addressing PKD isoform-specificity as a major factor for future therapeutic strategies.Molecular biology of the cell 12/2013; 25(3). DOI:10.1091/mbc.E13-06-0334 · 5.98 Impact Factor
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ABSTRACT: The fact that advanced NSCLC patients with wild type (wt) EGFR can benefit from erlotinib therapy makes it critical to find out biomarkers for effective selection of patients and improving the therapy effects. In present study, 3 NSCLC cell lines (U1752, Calu-6 and NCI-H292) with wt EGFR and different sensitivities to erlotinib were used for microarray analysis. The differential basal gene expression between 2 NSCLC cell lines was analyzed, about 353 genes were expression-altered with higher than 2-fold changes between Calu-6 and U1752. And Ingenuity Pathway Analysis (IPA) showed that these genes were mainly enriched in regulation of epithelial-mesenchymal transition (EMT) pathway, Wnt-β catenin signaling, Tec kinase signaling and some types of cancer-related signaling. More interestingly, RAF1 (c-raf), MAP2K1 (MEK1), SNAI and downstream signaling molecules ERK and AKT were predicted to be activated in erlotinib-resistant cell line by IPA. Subsequent immunoblotting experiments showed that the phosphorylation of ERK and AKT were exactly increased stepwise from erlotinib sensitive cell line to erlotinib resistant cell lines. Collectively, activation of RAF1-MEK1-ERK/AKT axis may determine the resistance of NSCLC cell lines bearing wt EGFR to erlotinib. Our work provides potential biomarkers and therapeutic targets for NSCLC patients harboring wt EGFR.International journal of clinical and experimental pathology 01/2013; 6(8):1493-504. · 1.78 Impact Factor
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ABSTRACT: Metastasis is the leading cause of cancer-associated death in most tumor types. Metastatic dissemination of cancer cells from the primary tumor is believed to be initiated by the reactivation of an embryonic development program referred to as epithelial-mesenchymal transition (EMT), whereby epithelial cells lose apicobasal polarity and cell-cell contacts, and gain mesenchymal phenotypes with increased migratory and invasive capabilities. EMT has also been implicated in the regulation of cancer stem cell property, immune suppression and cancer regression. Several transcription factors have been identified as master regulators of EMT, including the Snail, Zeb and Twist families, and their expression is tightly regulated at different steps of transcription, translation and protein stability control by a variety of cell-intrinsic pathways as well as extracellular cues. Here, we review the recent literature on the signaling pathways and mechanisms that control the expression of these master transcription factors during EMT and cancer progression.Oncogene advance online publication, 22 April 2013; doi:10.1038/onc.2013.128.Oncogene 04/2013; 33(14). DOI:10.1038/onc.2013.128 · 8.56 Impact Factor