Localization of CCTbeta in rat brain and overexpression in insect cells.
Abstract
AIM: To study the localization of CTP: phosphocholine cytidylyltransferase beta
isoform (CCTbeta) in rat brain, its expression in insect cells and enzymatic
properties.
METHODS: Using digoxigenin-labeled CCTbeta probes, in situ hybridization was
carried out in rat brain wax sections. CCTbeta was overexpressed in Trichoplusia
Ni (Tn) cells using baculovirus expression system. CTP:phosphocholine
cytidylyltransferase assay (CT assay) and [3H] metabolic labeling experiment were
used to study its activity, properties, and the effect on phosphatidylcholine
(PC) synthesis.
RESULTS: (1) CCbeta was abundant in CA1, CA2, CA4, and dentate gyrus (DG) region
of hippocampus. (2) The content of CCTbeta in transfected Tn cells was over 1 104
times of that in rat brain, and CCTbeta increased the PC synthesis of Tn cells.
(3) Hexadecylphosphocholine as well as some ions like Zn2+ and PO3-4 could
inhibit the activity of CCTbeta, dCTP was another adaptive substrate of CCTbeta
besides CTP.
CONCLUSION: CCTbeta showed a similar localization in rat brain with the memory
enhancing peptide argipressin (4-8).
Keywords:
isoform (CCTbeta) in rat brain, its expression in insect cells and enzymatic
properties.
METHODS: Using digoxigenin-labeled CCTbeta probes, in situ hybridization was
carried out in rat brain wax sections. CCTbeta was overexpressed in Trichoplusia
Ni (Tn) cells using baculovirus expression system. CTP:phosphocholine
cytidylyltransferase assay (CT assay) and [3H] metabolic labeling experiment were
used to study its activity, properties, and the effect on phosphatidylcholine
(PC) synthesis.
RESULTS: (1) CCbeta was abundant in CA1, CA2, CA4, and dentate gyrus (DG) region
of hippocampus. (2) The content of CCTbeta in transfected Tn cells was over 1 104
times of that in rat brain, and CCTbeta increased the PC synthesis of Tn cells.
(3) Hexadecylphosphocholine as well as some ions like Zn2+ and PO3-4 could
inhibit the activity of CCTbeta, dCTP was another adaptive substrate of CCTbeta
besides CTP.
CONCLUSION: CCTbeta showed a similar localization in rat brain with the memory
enhancing peptide argipressin (4-8).