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A novel role for Wnt/Ca2+ signaling in actin cytoskeleton remodeling and cell motility in prostate cancer.

Wang Q, Symes AJ, Kane CA, Freeman A, Nariculam J, Munson P, Thrasivoulou C, Masters JR, Ahmed A - PLoS ONE (2010)

Bottom Line: Wnt signaling is a critical regulatory pathway in development and disease.Live cell imaging showed that a functional consequence of CaMKII inhibition was 80% decrease in wound healing capacity and reduced cell motility in cancer cells.We propose that non-canonical Wnt/Ca(2+) signaling via CaMKII acts as a novel regulator of structural plasticity and cell motility in prostate cancer.

View Article: PubMed Central - PubMed

Affiliation: Prostate Cancer Research Centre and Division of Surgery, University College London, London, United Kingdom.

ABSTRACT
Wnt signaling is a critical regulatory pathway in development and disease. Very little is known about the mechanisms of Wnt signaling in prostate cancer, a leading cause of death in men. A quantitative analysis of the expression of Wnt5A protein in human tissue arrays, containing 600 prostate tissue cores, showed >50% increase in malignant compared to benign cores (p<0.0001). In a matched pair of prostate cancer and normal cell line, expression of Wnt5A protein was also increased. Calcium waves were induced in prostate cells in response to Wnt5A with a 3 fold increase in Flou-4 intensity. The activity of Ca(2+)/calmodulin dependent protein kinase (CaMKII), a transducer of the non-canonical Wnt/Ca(2+) signaling, increased by 8 fold in cancer cells; no change was observed in beta-catenin expression, known to activate the canonical Wnt/beta-catenin pathway. Mining of publicly available human prostate cancer oligoarray datasets revealed that the expression of numerous genes (e.g., CCND1, CD44) under the control of beta-catenin transcription is down-regulated. Confocal and quantitative electron microscopy showed that specific inhibition of CaMKII in cancer cells causes remodeling of the actin cytoskeleton, irregular wound edges and loose intercellular architecture and a 6 and 8 fold increase in the frequency and length of filopodia, respectively. Conversely, untreated normal prostate cells showed an irregular wound edge and loose intercellular architecture; incubation of normal prostate cells with recombinant Wnt5A protein induced actin remodeling with a regular wound edge and increased wound healing capacity. Live cell imaging showed that a functional consequence of CaMKII inhibition was 80% decrease in wound healing capacity and reduced cell motility in cancer cells. We propose that non-canonical Wnt/Ca(2+) signaling via CaMKII acts as a novel regulator of structural plasticity and cell motility in prostate cancer.

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Wnt5A induces calcium release in prostate cancer cells.A representative graph of calcium release in prostate cancer cell line (PC3) as a function of Fluo-4 intensity change over time using confocal live cell imaging. The green line represents the change in the Fluo-4 intensity (green line) in (A) control (after addition of vehicle PBS) or (B) recombinant Wnt5A peptide (100 ng/ml). There was a 3.1±0.1 fold increase in Fluo-4 intensity after addition of Wnt5A (n = 12). In some experiments Fura Red (red line) was loaded with Fluo-4. The ratio of change in Fluo-4 and Fura-red is plotted in (C).
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pone-0010456-g003: Wnt5A induces calcium release in prostate cancer cells.A representative graph of calcium release in prostate cancer cell line (PC3) as a function of Fluo-4 intensity change over time using confocal live cell imaging. The green line represents the change in the Fluo-4 intensity (green line) in (A) control (after addition of vehicle PBS) or (B) recombinant Wnt5A peptide (100 ng/ml). There was a 3.1±0.1 fold increase in Fluo-4 intensity after addition of Wnt5A (n = 12). In some experiments Fura Red (red line) was loaded with Fluo-4. The ratio of change in Fluo-4 and Fura-red is plotted in (C).

Mentions: Analysis of gene expression of CTNNB1 and TCF/LEF transcription targets, using Oncomine [27] and GeneSpring software and publicly available microarray datasets for normal (non-neoplastic or benign) vs cancer prostate tissue, showed that the expression of almost all Wnt/β-catenin/TCF targets analyzed, except c-myc, was decreased in prostate cancer (Fig. S3). These results are similar to those for prostate cell lines and demonstrate that β-catenin mediated increase in TCF transcription was not likely to be the mechanism of Wnt signaling in prostate cancer. We therefore tested the hypothesis that in the prostate cancer, Wnt signaling is transduced via Wnt/Ca2+ pathway. We performed experiments to establish if Wnt5A directly induced calcium release in prostate cells. Addition of Wnt5A peptide induced calcium waves, lasting upto 100s, in prostate cancer cell line with a 3.1±0.1 (n = 12) fold increase in the intensity of Flou-4 from the base line (Fig 3 and Movie S1).


A novel role for Wnt/Ca2+ signaling in actin cytoskeleton remodeling and cell motility in prostate cancer.

Wang Q, Symes AJ, Kane CA, Freeman A, Nariculam J, Munson P, Thrasivoulou C, Masters JR, Ahmed A - PLoS ONE (2010)

Wnt5A induces calcium release in prostate cancer cells.A representative graph of calcium release in prostate cancer cell line (PC3) as a function of Fluo-4 intensity change over time using confocal live cell imaging. The green line represents the change in the Fluo-4 intensity (green line) in (A) control (after addition of vehicle PBS) or (B) recombinant Wnt5A peptide (100 ng/ml). There was a 3.1±0.1 fold increase in Fluo-4 intensity after addition of Wnt5A (n = 12). In some experiments Fura Red (red line) was loaded with Fluo-4. The ratio of change in Fluo-4 and Fura-red is plotted in (C).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2864254&req=5

pone-0010456-g003: Wnt5A induces calcium release in prostate cancer cells.A representative graph of calcium release in prostate cancer cell line (PC3) as a function of Fluo-4 intensity change over time using confocal live cell imaging. The green line represents the change in the Fluo-4 intensity (green line) in (A) control (after addition of vehicle PBS) or (B) recombinant Wnt5A peptide (100 ng/ml). There was a 3.1±0.1 fold increase in Fluo-4 intensity after addition of Wnt5A (n = 12). In some experiments Fura Red (red line) was loaded with Fluo-4. The ratio of change in Fluo-4 and Fura-red is plotted in (C).
Mentions: Analysis of gene expression of CTNNB1 and TCF/LEF transcription targets, using Oncomine [27] and GeneSpring software and publicly available microarray datasets for normal (non-neoplastic or benign) vs cancer prostate tissue, showed that the expression of almost all Wnt/β-catenin/TCF targets analyzed, except c-myc, was decreased in prostate cancer (Fig. S3). These results are similar to those for prostate cell lines and demonstrate that β-catenin mediated increase in TCF transcription was not likely to be the mechanism of Wnt signaling in prostate cancer. We therefore tested the hypothesis that in the prostate cancer, Wnt signaling is transduced via Wnt/Ca2+ pathway. We performed experiments to establish if Wnt5A directly induced calcium release in prostate cells. Addition of Wnt5A peptide induced calcium waves, lasting upto 100s, in prostate cancer cell line with a 3.1±0.1 (n = 12) fold increase in the intensity of Flou-4 from the base line (Fig 3 and Movie S1).

Bottom Line: Wnt signaling is a critical regulatory pathway in development and disease.Live cell imaging showed that a functional consequence of CaMKII inhibition was 80% decrease in wound healing capacity and reduced cell motility in cancer cells.We propose that non-canonical Wnt/Ca(2+) signaling via CaMKII acts as a novel regulator of structural plasticity and cell motility in prostate cancer.

View Article: PubMed Central - PubMed

Affiliation: Prostate Cancer Research Centre and Division of Surgery, University College London, London, United Kingdom.

ABSTRACT
Wnt signaling is a critical regulatory pathway in development and disease. Very little is known about the mechanisms of Wnt signaling in prostate cancer, a leading cause of death in men. A quantitative analysis of the expression of Wnt5A protein in human tissue arrays, containing 600 prostate tissue cores, showed >50% increase in malignant compared to benign cores (p<0.0001). In a matched pair of prostate cancer and normal cell line, expression of Wnt5A protein was also increased. Calcium waves were induced in prostate cells in response to Wnt5A with a 3 fold increase in Flou-4 intensity. The activity of Ca(2+)/calmodulin dependent protein kinase (CaMKII), a transducer of the non-canonical Wnt/Ca(2+) signaling, increased by 8 fold in cancer cells; no change was observed in beta-catenin expression, known to activate the canonical Wnt/beta-catenin pathway. Mining of publicly available human prostate cancer oligoarray datasets revealed that the expression of numerous genes (e.g., CCND1, CD44) under the control of beta-catenin transcription is down-regulated. Confocal and quantitative electron microscopy showed that specific inhibition of CaMKII in cancer cells causes remodeling of the actin cytoskeleton, irregular wound edges and loose intercellular architecture and a 6 and 8 fold increase in the frequency and length of filopodia, respectively. Conversely, untreated normal prostate cells showed an irregular wound edge and loose intercellular architecture; incubation of normal prostate cells with recombinant Wnt5A protein induced actin remodeling with a regular wound edge and increased wound healing capacity. Live cell imaging showed that a functional consequence of CaMKII inhibition was 80% decrease in wound healing capacity and reduced cell motility in cancer cells. We propose that non-canonical Wnt/Ca(2+) signaling via CaMKII acts as a novel regulator of structural plasticity and cell motility in prostate cancer.

Show MeSH
Related in: MedlinePlus