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Jingqi Chen,
Yandan Yao,
Chang Gong,
Fengyan Yu,
Shicheng Su,
Jianing Chen,
Bodu Liu, Hui Deng,
Fengsong Wang,
Ling Lin,
Herui Yao,
Fengxi Su,
Karen S Anderson,
Qiang Liu,
Mark E Ewen,
Xuebiao Yao,
Erwei Song
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ABSTRACT: Tumor-associated macrophages (TAMs) can influence cancer progression and metastasis, but the mechanism remains unclear. Here, we show that breast TAMs abundantly produce CCL18, and its expression in blood or cancer stroma is associated with metastasis and reduced patient survival. CCL18 released by breast TAMs promotes the invasiveness of cancer cells by triggering integrin clustering and enhancing their adherence to extracellular matrix. Furthermore, we identify PITPNM3 as a functional receptor for CCL18 that mediates CCL18 effect and activates intracellular calcium signaling. CCL18 promotes the invasion and metastasis of breast cancer xenografts, whereas suppressing PITPNM3 abrogates these effects. These findings indicate that CCL18 derived from TAMs plays a critical role in promoting breast cancer metastasis via its receptor, PITPNM3.
Cancer cell 04/2011; 19(4):541-55. · 25.29 Impact Factor
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Yong Chen,
Dongmei Wang,
Zhen Guo,
Jun Zhao,
Bing Wu, Hui Deng,
Ti Zhou,
Hongjun Xiang,
Fei Gao,
Xue Yu, [......],
Peng Xia,
Chibuzo Emenari,
Xia Ding,
Winston Thompson,
Kelong Ma,
Jingde Zhu,
Felix Aikhionbare,
Kefen Dou,
Shi-Yuan Cheng,
Xuebiao Yao
[show abstract]
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ABSTRACT: During progression of hepatocellular carcinoma, multiple genetic and epigenetic alterations act to posttranslationally modulate the function of proteins that promote cancer invasion and metastasis. To define such abnormalities that contribute to liver cancer metastasis, we carried out a proteomic comparison of primary hepatocellular carcinoma and samples of intravascular thrombi from the same patient. Mass spectrometric analyses of the liver cancer samples revealed a series of acidic phospho-isotypes associated with the intravascular thrombi samples. In particular, we found that Thr567 hyperphosphorylation of the cytoskeletal protein ezrin was tightly correlated to an invasive phenotype of clinical hepatocellular carcinomas and to poor outcomes in tumor xenograft assays. Using phospho-mimicking mutants, we showed that ezrin phosphorylation at Thr567 promoted in vitro invasion by hepatocarcinoma cells. Phospho-mimicking mutant ezrinT567D, but not the nonphosphorylatable mutant ezrinT567A, stimulated formation of membrane ruffles, suggesting that Thr567 phosphorylation promotes cytoskeletal-membrane remodeling. Importantly, inhibition of Rho kinase, either by Y27632 or RNA interference, resulted in inhibition of Thr567 phosphorylation and a blockade to cell invasion, implicating Rho kinase-ezrin signaling in hepatocellular carcinoma cell invasion. Our findings suggest a strategy to reduce liver tumor metastasis by blocking Rho kinase-mediated phosphorylation of ezrin.
Cancer Research 03/2011; 71(5):1721-9. · 7.86 Impact Factor
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Xia Ding, Hui Deng,
Dongmei Wang,
Jiajia Zhou,
Yuejia Huang,
Xuannv Zhao,
Xue Yu,
Ming Wang,
Fengsong Wang,
Tarsha Ward,
Felix Aikhionbare,
Xuebiao Yao
[show abstract]
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ABSTRACT: The ezrin-radixin-moesin proteins provide a regulated linkage between membrane proteins and the cortical cytoskeleton and also participate in signal transduction pathways. Ezrin is localized to the apical membrane of parietal cells and couples the protein kinase A activation cascade to the regulated HCl secretion. Our recent proteomic study revealed a protein complex of ezrin-ACAP4-ARF6 essential for volatile membrane remodeling (Fang, Z., Miao, Y., Ding, X., Deng, H., Liu, S., Wang, F., Zhou, R., Watson, C., Fu, C., Hu, Q., Lillard, J. W., Jr., Powell, M., Chen, Y., Forte, J. G., and Yao, X. (2006) Mol. Cell Proteomics 5, 1437-1449). However, knowledge of whether ACAP4 physically interacts with ezrin and how their interaction is integrated into membrane-cytoskeletal remodeling has remained elusive. Here we provide the first evidence that ezrin interacts with ACAP4 in a protein kinase A-mediated phosphorylation-dependent manner through the N-terminal 400 amino acids of ACAP4. ACAP4 locates in the cytoplasmic membrane in resting parietal cells but translocates to the apical plasma membrane upon histamine stimulation. ACAP4 was precipitated with ezrin from secreting but not resting parietal cell lysates, suggesting a phospho-regulated interaction. Indeed, this interaction is abolished by phosphatase treatment and validated by an in vitro reconstitution assay using phospho-mimicking ezrin(S66D). Importantly, ezrin specifies the apical distribution of ACAP4 in secreting parietal cells because either suppression of ezrin or overexpression of non-phosphorylatable ezrin prevents the apical localization of ACAP4. In addition, overexpressing GTPase-activating protein-deficient ACAP4 results in an inhibition of apical membrane-cytoskeletal remodeling and gastric acid secretion. Taken together, these results define a novel molecular mechanism linking ACAP4-ezrin interaction to polarized epithelial secretion.
Journal of Biological Chemistry 04/2010; 285(24):18769-80. · 4.77 Impact Factor
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Ya Liu,
Xia Ding,
Dongmei Wang, Hui Deng,
Mingye Feng,
Min Wang,
Xue Yu,
Kai Jiang,
Tarsha Ward,
Felix Aikhionbare,
Zhen Guo,
John G Forte,
Xuebiao Yao
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ABSTRACT: Syntaxin and Munc18 are essential for regulated exocytosis in all eukaryotes. It was shown that Munc18 inhibition of neuronal syntaxin 1 can be overcome by CDK5 phosphorylation, indicating that structural change disrupts the syntaxin-Munc18 interaction. Here, we show that this phosphorylation promotes the assembly of Munc18b-syntaxin 3-SNAP25 tripartite complex and membrane fusion machinery SNARE. Using siRNAs to screen for genes required for regulated epithelial secretion, we identified the requirements of CDK5 and Munc18b in cAMP-dependent gastric acid secretion. Biochemical characterization revealed that Munc18b bears a syntaxin 3-selective binding site located at its most C-terminal 53 amino acids. Significantly, the phosphorylation of Thr572 by CDK5 attenuates Munc18b-syntaxin 3 interaction and promotes formation of Munc18b-syntaxin 3-SNAP25 tripartite complex, leading to an assembly of functional Munc18b-syntaxin 3-SNAP25-VAMP2 membrane fusion machinery. Thus, our studies suggest a novel regulatory mechanism in which phosphorylation of Munc18b operates vesicle docking and fusion in regulated exocytosis.
FEBS Letters 10/2007; 581(22):4318-24. · 3.54 Impact Factor
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Zhiyou Fang,
Yong Miao,
Xia Ding, Hui Deng,
Siqi Liu,
Fengsong Wang,
Rihong Zhou,
Charles Watson,
Chuanhai Fu,
Qicong Hu,
James W Lillard,
Michael Powell,
Yong Chen,
John G Forte,
Xuebiao Yao
[show abstract]
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ABSTRACT: ARF6 GTPase is a conserved regulator of membrane trafficking and actin-based cytoskeleton dynamics at the leading edge of migrating cells. A key determinant of ARF6 function is the lifetime of the GTP-bound active state, which is orchestrated by GTPase-activating protein (GAP) and GTP-GDP exchanging factor. However, very little is known about the molecular mechanisms underlying ARF6-mediated cell migration. To systematically analyze proteins that regulate ARF6 activity during cell migration, we performed a proteomic analysis of proteins selectively bound to active ARF6 using mass spectrometry and identified a novel ARF6-specific GAP, ACAP4. ACAP4 encodes 903 amino acids and contains two coiled coils, one pleckstrin homology domain, one GAP motif, and two ankyrin repeats. Our biochemical characterization demonstrated that ACAP4 has a phosphatidylinositol 4,5-bisphosphate-dependent GAP activity specific for ARF6. The co-localization of ACAP4 with ARF6 occurred in ruffling membranes formed upon AIF(4) and epidermal growth factor stimulation. ACAP4 overexpression limited the recruitment of ARF6 to the membrane ruffles in the absence of epidermal growth factor stimulation. Expression of GTP hydrolysis-resistant ARF6(Q67L) resulted in accumulations of ACAP4 and ARF6 in the cytoplasmic membrane, suggesting that GTP hydrolysis is required for the ARF6-dependent membrane remodeling. Significantly the depletion of ACAP4 by small interfering RNA or inhibition of ARF6 GTP hydrolysis by overexpressing GAP-deficient ACAP4 suppressed ARF6-dependent cell migration in wound healing, demonstrating the importance of ACAP4 in cell migration. Thus, our study sheds new light on the biological function of ARF6-mediated cell migration.
Molecular & Cellular Proteomics 09/2006; 5(8):1437-49. · 7.40 Impact Factor
