Activation of choline kinase by extracellular Ca2+ is Ca(2+)-sensing receptor, Galpha12 and Rho-dependent in breast cancer cells.
Breast cancer cell metastases to bone result in osteolysis and release of large quantities of Ca2+ into the bone microenviroment. Extracellular Ca2+ (Ca(o)2+) acting through the Ca(2+)-sensing receptor (CaR), a member of G protein-coupled receptor superfamily, plays an important role in the regulation of multiple signaling pathways. Here, we find that expression of the CaR and Galpha(12) is significantly up-regulated in breast cancer cells (MDA-MB-231 and MCF-7) compared with nonmalignant breast cells (Hs 578Bst and MCF-10A). Ca(o)2+ induces a significant increase in extracellular [(3)H]phosphocholine (P-cho) production in breast cancer cells. Using an anti-CaR antibody to block Ca(o)2+ binding to the CaR and small interfering RNA (siRNA) to silence CaR gene expression, our data demonstrate that [(3)H]P-cho production in response to Ca(o)(2+)-stimulation is CaR-dependent. By analyzing cellular lipid profiles and using siRNA to silence choline kinase (ChoK) expression, we determine that the production of [3H]P-cho is primarily related to CaR-induced ChoK activation, and not degradation of choline phospholipids. Finally, by pretreatment of the cells with either pertussis toxin or C3 exoenzyme, co-immunoprecipiation of Galpha(i), Galpha(q) or Galpha12 with the CaR, and RhoA translocation, we found that the enhancement of ChoK activation and P-cho production in breast cancer cells occurs via a CaR-Galpha12-Rho signaling pathway.
Huang
Chunfa
C
Louis Stokes Cleveland Veteran Affairs Medical Center, Cleveland, Ohio 44106, United States. chunfa.huang@case.edu
Hydo
Lindsey M
LM
Liu
Shiguo
S
Miller
R Tyler
RT
D001943:Breast Neoplasms / Q000201:enzymology* / Q000235:genetics; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D045744:Cell Line, Tumor; D002797:Choline Kinase / Q000378:metabolism*; D004789:Enzyme Activation; D005260:Female; D044365:GTP-Binding Protein alpha Subunits, G12-G13 / Q000235:genetics / Q000378:metabolism*; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D010767:Phosphorylcholine / Q000378:metabolism; D044169:Receptors, Calcium-Sensing / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation; D020741:rho GTP-Binding Proteins / Q000378:metabolism
D044169:Receptors, Calcium-Sensing; D010767:Phosphorylcholine; D002797:Choline Kinase; D044365:GTP-Binding Protein alpha Subunits, G12-G13; D020741:rho GTP-Binding Proteins; D002118:Calcium