Rat choline acetyltransferase of the peripheral type differs from that of the common type in intracellular translocation.
Choline acetyltransferase (ChAT), the synthesizing enzyme for acetylcholine, has been implicated to involve multiple isoforms of ChAT mRNA in several animals. Since these isoforms are mostly non-coding splice variants, only a homologous ChAT protein of about 68 kDa has been shown to be produced in vivo. Recent evidence indicates the existence of a protein coding splice variant of ChAT mRNA, which lacks exons 6-9 of the rat ChAT gene. The encoded protein was designated ChAT of a peripheral type (pChAT), because of its preferential expression in the peripheral nervous system as confirmed by Western blot and immunohistochemistry. However, functional significance of pChAT is unknown. To obtain a clue to this question, we examined a possible difference in intracellular trafficking between pChAT and the well-known ChAT of the common type (cChAT) using green fluorescent protein (GFP) in living human embryonic kidney cells. Confocal laser scanning microscopy revealed that pChAT-GFP was detectable in the cytoplasm but not in the nucleus, whereas cChAT-GFP was found in both cytoplasm and nucleus. Following treatment with leptomycin B, a nuclear export pathway inhibitor, pChAT-GFP became detectable in both cytoplasm and nucleus, indicating that pChAT can be translocated to the nucleus. In contrast, the leptomycin B treatment did not seem to affect the content of intranuclear cChAT-GFP. After incubation with protein kinase C inhibitors, enhanced accumulation of pChAT-GFP but not cChAT-GFP occurred in the nucleus. These results clearly indicate that pChAT varies from cChAT in intracellular transportation, probably reflecting the difference in physiological roles between pChAT and cChAT.
Matsuo
Akinori
A
Molecular Neuroscience Research Center, Shiga University of Medical Science, Seta Tsukinowa-cho, Otsu, Shiga 520-2192, Japan.
Bellier
Jean-Pierre
JP
Hisano
Tadashi
T
Aimi
Yoshinari
Y
Yasuhara
Osamu
O
Tooyama
Ikuo
I
Saito
Naoaki
N
Kimura
Hiroshi
H
D000109:Acetylcholine / Q000096:biosynthesis*; D021581:Active Transport, Cell Nucleus / Q000187:drug effects / Q000502:physiology; D017398:Alternative Splicing / Q000502:physiology*; D000818:Animals; D002467:Cell Nucleus / Q000201:enzymology; D002478:Cells, Cultured; D002490:Central Nervous System / Q000201:enzymology*; D002795:Choline O-Acetyltransferase / Q000235:genetics / Q000378:metabolism*; D003342:Corpus Striatum / Q000201:enzymology; D003593:Cytoplasm / Q000201:enzymology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D005726:Ganglia, Parasympathetic / Q000201:enzymology; D049452:Green Fluorescent Proteins; D006801:Humans; D008297:Male; D017933:Peripheral Nervous System / Q000201:enzymology*; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects / Q000502:physiology; D051381:Rats
D004791:Enzyme Inhibitors; D005231:Fatty Acids, Unsaturated; D020033:Protein Isoforms; D049452:Green Fluorescent Proteins; D002795:Choline O-Acetyltransferase; D011493:Protein Kinase C; D000109:Acetylcholine; C038753:leptomycin B