This paper describes some properties of
glutamine oxidation and
glutaminase isoform expression in cell lines derived from human colorectal
adenomas and
carcinomas. The slow-growing
adenoma-derived cell line AA/C1, and the rapidly proliferating
carcinoma cell line HT29, both required
glutamine for growth. The rate of (14)CO(2) production from [U-(14)C]
glutamine was faster in AA/C1 cells than in HT29 cells. Conversely HT29 cells showed faster rates of
glucose oxidation and
lactate production. The activity of
glutaminase was 3 times higher in AA/C1
cell extracts than in extracts of HT29 cells.
Glutaminase activity in the two cell lines had similar K(m) values for
glutamine, but the activity in AA/C1 cells had a higher K(0.5) for activation by
phosphate.
Glutaminase activity in extracts of both cells was inhibited by
glutamate. Western blotting showed the presence, in both cell lines, of
isoform(s) of
glutaminase with an molecular mass of 63 kDa, intermediate between that of kidney
glutaminase and liver
glutaminase. PCR-based analysis showed that an
mRNA species identical to the kidney-type
isoform glutaminase C was present in both cell types as was an additional
mRNA species identical to the liver-type
glutaminase isoform from human breast tumour cells. Northern blotting using
isoform-specific
cDNA probes demonstrated that
mRNA for both
glutaminase isoforms was expressed at significant levels in both cell types. Similar results to those in AA/C1 cells and HT29 cells were obtained in two further
adenoma and
carcinoma cell lines respectively. These results contrast with those reported previously in hepatocyte/
hepatoma model systems with respect to fuel selection,
glutaminase activity and
isoform expression. They also constitute the first demonstration of simultaneous expression of two
glutaminase isoforms in a single cell type.