Expression of glutamic acid decarboxylase during human neuronal differentiation: studies using the NTera-2 culture system.
Human NTera-2N neurons, but not the parental NTera-2 teratocarcinoma line, decarboxylate [2-(15)N]glutamine to form gamma-[15N]aminobutyric acid (GABA). The reverse transcriptase-polymerase chain reaction (RT-PCR) followed by Southern blotting showed that NTera-2N neurons transcribe the glutamic acid decarboxylase p67 (GAD67) gene, and also demonstrated that there is developmentally regulated alternative splicing of GAD67 mRNA in NTera-2N neurons. As in rat central nervous system (CNS), this mRNA processing generates two RNA transcripts, owing to the inclusion or exclusion of an approximately 80 bp coding region insert. In embryonic day 16 (E16) rat brain, the larger of the two GAD67 mRNAs, which encodes a truncated, inactive apoenzyme, reaches a concentration almost equal to that of the smaller transcript, which encodes functional GAD67. In developing NTera-2N neurons, however, the larger transcript is barely detectable by RT-PCR. RT-PCR also revealed that rat CNS of all ages examined contains GAD65 mRNA, and that GAD65 mRNA is below the detectable range in NTera-2N neurons.
Yoshioka
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A
Section of Neurology, Children's Hospital of Philadelphia, PA, USA.
Yudkoff
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M
Pleasure
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D
D000818:Animals; D015139:Blotting, Southern; D002454:Cell Differentiation / Q000502:physiology; D002460:Cell Line; D002490:Central Nervous System / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D005968:Glutamate Decarboxylase / Q000235:genetics*; D006801:Humans; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D016133:Polymerase Chain Reaction / Q000379:methods; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D014158:Transcription, Genetic
D012333:RNA, Messenger; D005968:Glutamate Decarboxylase