Increased expression of CTP:phosphocholine cytidylyltransferase in maturing type II cells.
We previously reported that phosphatidylcholine synthesis increased in fetal rat lung type II cells with advancing gestation. This increase was accompanied by an increase in CTP:phosphocholine cytidylyltransferase activity, which catalyses a rate regulatory step in de novo phosphatidylcholine synthesis by fetal type II cells. To determine whether this increase in cytidylyltransferase activity is due to an increase in cytidylyltransferase protein levels, the gene and protein expression of cytidylyltransferase was investigated in maturing type II cells. The cytidylyltransferase cDNA was cloned from fetal rat type II cells and showed 99% sequence homology with rat liver cDNA. The cDNA detected two mRNA transcripts (1.8 and 7.5 kb) in fetal rat lung. By reverse-transcriptase polymerase chain reaction (RT-PCR) analysis, cytidyltransferase mRNA content increased three-fold in fetal type II cells with advancing gestation, whereas cytidylyltransferase mRNA levels in fibroblasts remained constant. An antibody against rat liver cytidylyltransferase was used to assess cytidylyltransferase protein. Western blotting revealed that cytidylyltransferase protein content increased threefold in the microsomal fraction of type II cells with advancing gestation. The enzyme protein levels in the cytosolic fraction did not significantly change with development. Enzyme activity studies confirmed these latter observations. We conclude that the increase in surfactant phosphatidylcholine synthesis by type II cells at late fetal gestation is due in part to an increase in the amount of cytidylyltransferase protein.
Hogan
M
M
Department of Paediatrics, Hospital for Sick Children Research Institute, University of Toronto, Ontario, Canada.
Zimmermann
L J
LJ
Wang
J
J
Kuliszewski
M
M
Liu
J
J
Post
M
M
D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D019916:Choline-Phosphate Cytidylyltransferase; D003001:Cloning, Molecular; D005260:Female; D015870:Gene Expression; D008168:Lung / Q000166:cytology / Q000196:embryology*; D008297:Male; D015335:Molecular Probes / Q000235:genetics; D008969:Molecular Sequence Data; D009713:Nucleotidyltransferases / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar
D015335:Molecular Probes; D009713:Nucleotidyltransferases; D019916:Choline-Phosphate Cytidylyltransferase