Regulatory connection between the expression level of classical protein kinase C and pruning of climbing fibers from cerebellar Purkinje cells

摘要

Cerebellar Purkinje cells ( PC s) express two members of the classical protein kinase C (c PKC ) subfamily, namely, PKC α and PKC γ. Previous studies on PKC γ knockout ( KO ) mice have revealed a critical role of PKC γ in the pruning of climbing fibers ( CF s) from PC s during development. The question remains as to why only PKC γ and not PKC α is involved in CF synapse elimination from PC s. To address this question, we assessed the expression levels of PKC γ and PKC α in wild‐type ( WT ) and PKC γ KO PC s using PC ‐specific quantitative real‐time reverse transcription‐polymerase chain reaction, western blotting, and immunohistochemical analysis. The results revealed that the vast majority of cPKC s in PC s were PKC γ, whereas PKC α accounted for the remaining minimal fraction. The amount of PKC α was not up‐regulated in PKC γ KO PC s. Lentiviral expression of PKC α in PKC γ KO PC s resulted in a 10‐times increase in the amount of PKC α mRNA in the PKC γ KO PC s, compared to that in WT PC s. Our quantification showed that the expression levels of cPKC mRNA in PKC γ KO PC s increased roughly from 1% to 22% of that in WT PC s solely through PKC α expression. The up‐regulation of PKC α in PKC γ KO PC s significantly rescued the impaired CF synapse elimination. Although both PKC α and PKC γ are capable of pruning supernumerary CF synapses from developing PC s, these results suggest that the expression levels of cPKC s in PKC γ KO PC s are too low for CF pruning.