Translation modulation of acid beta-glucosidase in HepG2 cells: participation of the PKC pathway.
Acid beta-glucosidase (GCase) is the enzyme deficient in Gaucher disease, a prototypical inherited metabolic error for enzyme and gene therapy. An 80 kDa mammalian cytoplasmic translational control protein (TCP80) modulates GCase translation in vitro and ex vivo by interacting with the 5' coding region of GCase RNA. Ten predicted PKC phosphorylation sites (Ser- or Thr-) are in the TCP80 protein. Phosphorylation of TCP80 in vitro by PKC greatly enhanced its translational inhibitory function using in vitro translation assays; binding of GCase mRNA to TCP80 was unaltered. Conversely, de-phosphorylation of TCP80 reduced its translational inhibitory function. Phosphorylation-related modulation of GCase mRNA translation also was studied in HepG2 cells. GCase expression (protein and activity levels) in HepG2 cells increased (>2-fold) in cells treated with bisindolylmaleimide (BIM), a highly selective PKC specific inhibitor. This correlated with a 90% reduction in TCP80 phosphorylation in the presence of BIM. The amount of TCP80 protein in cytoplasm and its RNA-binding activity were unchanged. These experiments indicate that GCase mRNA translation is modulated by PKC signaling pathways that are mediated through TCP80. These findings indicate potential broader impacts of the TCP/PKC system on expression of this and other genes of therapeutic interest.
Xu
You-Hai
YH
Division of Human Genetics, The Children's Hospital Research Foundation, Cincinnati, OH 45229-3039, USA.
Grabowski
Gregory A
GA
D001483:Base Sequence; D002460:Cell Line; D017931:DNA Primers; D047468:Immunoprecipitation; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015398:Signal Transduction; D001617:beta-Glucosidase / Q000235:genetics*
D017931:DNA Primers; D011493:Protein Kinase C; D001617:beta-Glucosidase