Immunohistochemical localization of protein- O-carboxylmethyltransferase in rat brain neurons

Abstract

The distribution of the enzyme protein- O-carboxylmethyltransferase (EC 2.1.1.24) has been investigated in the rat brain using both immunohistochemical and biochemical techniques. The enzyme, which carboxylmethylates free aspartic and glutamic acid residues of protein substrates, was localized in neurons, but not other cell types throughout the brain. The highest immunoreactivity was detected throughout the cortex, followed by the hippocampus, the corpus striatum, the thalamus and the amygdala. Immunoreactive cells were detected in other brain regions but were not as prominent as those regions listed above. The distribution of immunoreactivity in the hippocampus was most striking, with considerable labelling of the pyramidal and granule cells in all regions. Numerous pyramidal cells were labelled in the cerebral cortex, with some ascending processes exhibiting immunoreactivity. The corpus striatum was uniformly labelled, suggesting that the enzyme was not localized to any specific neurotransmitter system. The antisera employed in this study was generated against purified bovine brain protein- O-carboxylmethyltransferase and Western immunoblot analysis showed cross reactivity against both rat brain and human erythrocyte forms of the enzyme. Enzyme activity and methyl acceptor protein capacity were examined in 1.5 mm coronal sections of rat brain. The regions with highest enzyme activities were found in cross-sections containing cortex and corpus striatum or cortex and hippocampus. The lowest enzyme activities were noted in slices of brainstem and cerebellum, areas exhibiting low amounts of immunoreactive protein- O-carboxylmethyltransferase. Methyl acceptor protein capacity was highest in slices of cortex and corpus striatum, cortex and hippocampus and was lowest in slices of brainstem and cerebellum. These results demonstrate that protein- O-carboxylmethyltransferase has an unique neuronal pattern of distribution in the rodent central nervous system, and suggest that the carboxylmethylation of proteins may be of functional significance in these neurons.