ABSTRACT
The neurotransmitter dopamine (DA) is capable of inducing apoptosis in post-mitotic sympathetic neurons via its oxidative metabolites. The differential display method was applied to cultured sympathetic neurons in an effort to detect genes whose expression is transcriptionally regulated during the early stages of DA-triggered apoptosis. One of the up-regulated genes was identified as the chick homologue to T-complex polypeptide-1delta (TCP-1delta), a member of the molecular chaperone family of proteins. Each chaperone protein is a complex of seven to nine different subunits. A full-length clone of 1.9 kilobases was isolated containing an open reading frame of 536 amino acids with a predicted molecular weight of 57,736. Comparison with the mouse TCP-1delta revealed 78 and 91% homology on the DNA and protein levels, respectively. Northern blot analysis disclosed a steady and significant increase in mRNA levels of TCP-1delta after DA administration, reaching a peak between 4 and 9 h and declining thereafter. Induction of the TCP-1delta protein levels was also observed as a function of DA treatment. Overexpression of TCP-1delta in sympathetic neurons accelerated DA-induced apoptosis; inhibition of TCP-1delta expression in these neurons using antisense technology significantly reduced DA-induced neuronal death. These findings suggest a functional role for TCP-1delta as a positive mediator of DA-induced neuronal apoptosis.
Subject(s)
Apoptosis/drug effects , Chaperonins/genetics , Chaperonins/metabolism , Dopamine/pharmacology , Neurons/cytology , Neurons/drug effects , Sympathetic Nervous System/embryology , Amino Acid Sequence , Animals , Base Sequence , Cells, Cultured , Chaperonin Containing TCP-1 , Chaperonins/chemistry , Chick Embryo , Cloning, Molecular , Gene Expression Regulation, Developmental/drug effects , Molecular Sequence Data , Nerve Growth Factor/deficiency , Nerve Growth Factor/metabolism , Neurons/metabolism , Oligonucleotides, Antisense/genetics , Oligonucleotides, Antisense/pharmacology , Protein Subunits , RNA, Messenger/genetics , RNA, Messenger/metabolism , Sequence Alignment , Sequence Homology, Amino Acid , Sympathetic Nervous System/cytologyABSTRACT
It was suggested that neuronal degeneration in Parkinson's Disease (PD) is linked to dopamine (DA) toxicity. Dopamine has been shown to induce programmed cell death in both neuronal and non-neuronal cell types. We examined the molecular changes associated with dopamine-triggered apoptosis in sympathetic neurons using the differential display approach, and isolated 14 different DA responsive genes whose expression is altered during the early stages of the apoptotic process. Nine of these genes are upregulated and five are downregulated in response to DA exposure. Two of the upregulated genes were identified as cyclin B2 and a chicken homologue of chaperonin, a member of the heat shock protein family. Total increase in mRNA expression of both genes after 12 hours of exposure to DA was 40%. These two genes participate in cell cycle control and are specifically involved in determining entry of dividing cells into mitosis. Upregulation of mitosis-related genes in postmitotic sympathetic neurons undergoing apoptosis, may be indicative of an abortive attempt of these neurons to re-enter the cell cycle prior to their death. Possible implications to neuronal degeneration in PD are discussed.
