CTP: phosphocholine cytidylyltransferase is both a nuclear and cytoplasmic protein in primary hepatocytes.
CTP:phosphocholine cytidylyltransferase (CT) has recently been reported to be a predominantly intranuclear enzyme in several cell lines (Wang et al., J. Biol. Chem. 268, 5899-5904 (1993)). This contrasts with previous reports that CT was a cytosolic protein that translocated to the endoplasmic reticulum upon activation. The aim of the present study was to compare the localization of CT in CHO cells and in primary rat hepatocytes. Indirect immunofluorescence of CHO cells revealed a largely nuclear localization of the CT. On the other hand, immunogold electron microscopy and biochemical studies showed a similar density of distribution of CT between the nucleus and cytoplasm. In primary rat hepatocytes immunofluorescence studies indicated that CT was largely cytoplasmic. Studies by immunogold electron microscopy of rat hepatocytes demonstrated that the enzyme was homogeneously distributed throughout all cytoplasmic regions and the nucleoplasm. This result was confirmed by biochemical studies using digitonin and streptolysin O, which permeabilizes the plasma membrane of cells. Enucleation studies indicated that in CHO cells 76% of the CT activity was in the nuclear fraction, whereas in hepatocytes only 32% was recovered in this fraction. The data indicate that CT is found both in nuclear and cytoplasmic fractions of primary hepatocytes and is not predominantly a nuclear enzyme.
Houweling
M
M
Department of Biochemistry, University of Alberta, Edmonton, Canada.
Cui
Z
Z
Anfuso
C D
CD
Bussière
M
M
Chen
M H
MH
Vance
D E
DE
D000818:Animals; D016466:CHO Cells; D002463:Cell Membrane Permeability; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D019916:Choline-Phosphate Cytidylyltransferase; D006224:Cricetinae; D003593:Cytoplasm / Q000378:metabolism; D004072:Digitonin / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D007150:Immunohistochemistry; D008099:Liver / Q000166:cytology; D008297:Male; D009687:Nuclear Proteins / Q000378:metabolism*; D009713:Nucleotidyltransferases / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley
D009687:Nuclear Proteins; D009713:Nucleotidyltransferases; D019916:Choline-Phosphate Cytidylyltransferase; D004072:Digitonin