FXYD5 is a Na+-K+-ATPase regulator, expressed in a variety of normal epithelia. In parallel, it has been found to be associated with several types of cancer and effect lethal outcome by promoting metastasis. However, the molecular mechanism underlying FXYD5 mediated invasion has not yet been identified. In this study, using in vivo 4T1 murine breast cancer model, we found that FXYD5-specific shRNA significantly inhibited lung cancer metastasis, without having a substantial effect on primary tumor growth. Our study reveals that FXYD5 participates in multiple stages of metastatic development and exhibits more than one mode of E-cadherin regulation. We provide the first evidence that FXYD5-related morphological changes are mediated through its interaction with Na+-K+-ATPase. Experiments in cultured 4T1 cells have indicated that FXYD5 expression may downregulate the ß1 isoform of the pump. This behavior could have implications on both transcellular interactions and intracellular events. Further studies suggest that differential localization of the adaptor protein Annexin A2 in FXYD5-expressing cells may correlate with matrix metalloproteinase 9 secretion and adhesion changes in 4T1 wild-type cells.
Gene Expression Regulation, Neoplastic , Lung Neoplasms/genetics , Mammary Neoplasms, Experimental/genetics , Membrane Proteins/genetics , Neoplasms, Adipose Tissue/genetics , Sodium-Potassium-Exchanging ATPase/genetics , Animals , Annexin A2/genetics , Annexin A2/metabolism , Cadherins/genetics , Cadherins/metabolism , Cell Adhesion , Cell Line, Tumor , Cell Movement , Disease Models, Animal , Female , Ion Channels , Lung Neoplasms/metabolism , Lung Neoplasms/secondary , Mammary Glands, Animal/metabolism , Mammary Glands, Animal/pathology , Mammary Neoplasms, Experimental/metabolism , Mammary Neoplasms, Experimental/pathology , Matrix Metalloproteinase 9/genetics , Matrix Metalloproteinase 9/metabolism , Membrane Proteins/antagonists & inhibitors , Membrane Proteins/metabolism , Mice , Mice, Inbred BALB C , Microfilament Proteins , Neoplasms, Adipose Tissue/metabolism , Neoplasms, Adipose Tissue/pathology , Protein Subunits/genetics , Protein Subunits/metabolism , RNA, Small Interfering/genetics , RNA, Small Interfering/metabolism , Signal Transduction , Sodium-Potassium-Exchanging ATPase/metabolism
FXYD5 (Dysadherin, RIC) is a single span type I membrane protein that plays multiple roles in regulation of cellular functions. It is expressed in a variety of epithelial tissues and acts as an auxiliary subunit of the Na(+)/K(+)-ATPase. During the past decade, a correlation between enhanced expression of FXYD5 and tumor progression has been established for various tumor types. In this review, current knowledge on FXYD5 is discussed, including experimental data on the functional effects of FXYD5 on the Na(+)/K(+)-ATPase. FXYD5 modulates cellular junctions, influences chemokine production, and affects cell adhesion. The accumulated data may provide a basis for understanding the molecular mechanisms underlying FXYD5 mediated phenotypes.
The FXYD proteins are a family of small membrane proteins that share an invariant four amino acid signature motif F-X-Y-D and act as tissue-specific regulatory subunits of the Na,K-ATPase. FXYD5 (also termed dysadherin or RIC) is a structurally and functionally unique member of the FXYD family. As other FXYD proteins, FXYD5 specifically interacts with the Na,K-ATPase and alters its kinetics by increasing Vmax However, unlike other family members FXYD5 appears to have additional functions, which cannot be readily explained by modulation of transport kinetics. Knockdown of FXYD5 in MDA-MB-231 breast cancer cells largely decreases expression and secretion of the chemokine CCL2 (MCP-1). A related effect has also been observed in renal cell carcinoma cells. The current study aims to further characterize the relationship between the expression of FXYD5 and CCL2 secretion. We demonstrate that transfection of M1 epithelial cell line with FXYD5 largely increases lipopolysaccharide (LPS) stimulated CCL2 mRNA and secretion of the translated protein. We have completed a detailed analysis of the molecular events leading to the above response. Our key findings indicate that FXYD5 generates a late response by increasing the surface expression of the TNFα receptor, without affecting its total protein level, or mRNA transcription. LPS administration to mice demonstrates induced secretion of CCL2 and TNFα in FXYD5-expressing lung peripheral tissue, which suggests a possible role for FXYD5 in normal epithelia during inflammation.
Membrane Glycoproteins/metabolism , Membrane Proteins/metabolism , Neoplasm Proteins/metabolism , Animals , Cell Line , Cell Line, Tumor , Chemokine CCL2/genetics , Chemokine CCL2/metabolism , Epithelial Cells/metabolism , Gene Expression/drug effects , Gene Knockdown Techniques , Humans , Inflammation Mediators/metabolism , Ion Channels , Kinetics , Lipopolysaccharides/pharmacology , Membrane Glycoproteins/antagonists & inhibitors , Membrane Glycoproteins/genetics , Membrane Proteins/genetics , Mice , Microfilament Proteins , Neoplasm Proteins/antagonists & inhibitors , Neoplasm Proteins/genetics , RNA, Messenger/genetics , RNA, Messenger/metabolism , Receptors, Tumor Necrosis Factor/genetics , Receptors, Tumor Necrosis Factor/metabolism , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Signal Transduction/drug effects
