bromochloroacetic-acid and sphingosine-phosphorylcholine

Sphingosylphosphorylcholine induces keratin phosphorylation and reorganization, and increases viscoelasticity of metastatic cancer cells such as PANC-1 cells. However, the mechanism involved in sphingosylphosphorylcholine-induced keratin phosphorylation and reorganization is largely unknown. Sphingosylphosphorylcholine dose- and time-dependently induces the expression of RhebL1. The involvement of RhebL1 in sphingosylphosphorylcholine-induced events including keratin 8 (K8) phosphorylation, reorganization, migration and invasion was examined. Gene silencing of RhebL1 suppressed the sphingosylphosphorylcholine-induced events and overexpression of RhebL1 enhanced those events even without sphingosylphosphorylcholine treatment. We examined whether the G protein function of RhebL1 induces K8 phosphorylation using constitutively active RhebL1Q64L and dominant negative RhebL1D60K. G protein activity of RhebL1 is involved in sphingosylphosphorylcholine-induced K8 phosphorylation. We found that RhebL1 binds and activates AKT1. G protein activity of RhebL1 is involved in the binding and activation of AKT1. MK2206 (AKT inhibitor) and gene silencing of AKT1 inhibited the sphingosylphosphorylcholine-induced events, whereas overexpression of activated-AKT1 induced K8 phosphorylation, reorganization, migration and invasion even without sphingosylphosphorylcholine treatment.The collective results indicate that RhebL1 is involved in sphingosylphosphorylcholine-induced events in A549 lung cancer cells via binding to AKT1 leading to activation of it. These results suggest that suppression of RhebL1 or inhibition of RhebL1's binding to AKT1 might be a novel way that prevents changes in the physical properties of metastatic cancer cells.

Topics: Apoptosis; Biomarkers, Tumor; Cell Proliferation; Gene Expression Regulation, Neoplastic; GTP-Binding Proteins; Humans; Keratins; Lung Neoplasms; Neoplasm Staging; Phosphorylation; Phosphorylcholine; Prognosis; Proto-Oncogene Proteins c-akt; ras Proteins; Sphingosine; Survival Rate; Tumor Cells, Cultured

Sphingosylphosphorylcholine (SPC) is a naturally occurring bioactive lipid that is present in high density lipoproteins (HDL) particles and found at increased levels in blood and malignant ascites of patients with ovarian cancer. Here, we show that incubation of human epithelial tumour cells with SPC induces a perinuclear reorganization of intact keratin 8-18 filaments. This effect is specific for SPC, largely independent of F-actin and microtubules, and is accompanied by keratin phosphorylation. In vivo visco-elastic probing of single cancer cells demonstrates that SPC increases cellular elasticity. Accordingly, SPC stimulates migration of cells through size-limited pores in a more potent manner than lysophosphatidic acid (LPA). LPA induces actin stress fibre formation, but does not reorganize keratins in cancer cells and hence increases cellular stiffness. We propose that reorganization of keratin by SPC may facilitate biological phenomena that require a high degree of elasticity, such as squeezing of cells through membranous pores during metastasis.

Topics: Actin Cytoskeleton; Carcinoma; Cell Movement; Cell Size; Cytoskeleton; Elasticity; Fluorescent Antibody Technique; Humans; Keratins; Microscopy, Electron; Neoplasm Metastasis; Pancreatic Neoplasms; Phosphorylcholine; Sphingosine; Stress, Mechanical; Tumor Cells, Cultured