Energy metabolism and protein phosphorylation during apoptosis: a phosphorylation study of tau and high-molecular-weight tau in differentiated PC12 cells

Apoptosis has been characterized as a regulated, energy-dependent process. Specific protein-phosphorylation events have been demonstrated previously to occur during apoptosis and may play an important role in the regulation of this death process. In this study, energy metabolism and protein phosphorylation during apoptosis of neuronal PC 12 cells induced by nerve growth factor and serum deprivation was examined using [P-32]P-i-labelling techniques. Although ATP levels were maintained at control levels during apoptosis, [32P]P-i incorporation into ATP was decreased significantly, coinciding with an almost identical decrease in Na+-dependent phosphate uptake. During neuronal PC 12-cell apoptosis, increased phosphorylation of tau and high-molecular-weight (HMW) tau was observed within the epitope of Tau-l, a phosphate-dependent tau antibody that only recognizes the unphosphorylated form of its epitope. In addition, based on two-dimensional phosphopeptide mapping, [P-32]P-i incorporation into a phosphopeptide of tau and HMW tau from apoptotic cells increased. Whereas [32P]P-i incorporation into total protein decreased to 23% of the control during apoptosis, [P-32]P-i incorporation into tau and HMW tau was significantly higher, indicating a preferential phosphorylation of specific proteins during the apoptotic process. This study provides novel information about phosphate uptake, incorporation of [32P]P-i into ATP, and protein phosphorylation events during apoptosis.