Differential Role of Human Choline Kinase α and β Enzymes in Lipid Metabolism: Implications in Cancer Onset and Treatment
Figure 2
Differential activation of choline kinase α1 and β1 isoforms by Ras and Rho GTPases.
Choline kinase isoforms were expressed alone or in combination with the indicated Ras and Rho GTPases and the in vitro ChoK activity determined. A) Analysis of ectopic expression by Western Blot in HEK293T transfected cell extracts of ChoKα1 (52 KDa), ChoKβ1(45 KDa), RhoA-QL(22 KDa), Cdc42-QL(25 KDa) and H-rasV12 (23 KDa). Empty vectors were used as controls for the endogenous levels, and GAPDH as loading control. B) and C) In vitro choline kinase activity of ChoKα1 or ChoKβ1 in the presence of enhanced expression of constitutive active forms of RhoA, Cdc42 or H-Ras. D) and E) In vitro ethanolamine kinase activity of ChoKα or ChoKβ in the presence of each indicated constitutive active form of GTPase. The results are represented as fold induction of conversion to the corresponding phosphorylated metabolite determined as total cpm/µg of whole cellular extract, and normalized to the empty vector transfected cells as control. Data shown represent the mean values±SEM of 3 independent experiments, each one performed with duplicate samples. Statistical significance (p≤0.05) is marked by an asterisk comparing to the activity achieved when ChoKα1 or ChoKβ1, where appropriate, are transfected alone.