Glutaminase isoform expression in cell lines derived from human colorectal adenomas and carcinomas.
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.
Turner
Abigail
A
Department of Biochemistry, School of Medical Sciences, University Walk, Bristol BS8 1TD, UK.
McGivan
John D
JD
D002455:Cell Division / Q000502:physiology; D015179:Colorectal Neoplasms / Q000201:enzymology* / Q000235:genetics; D005972:Glutaminase / Q000096:biosynthesis / Q000235:genetics*; D006801:Humans; D007527:Isoenzymes / Q000096:biosynthesis / Q000235:genetics; D014407:Tumor Cells, Cultured
D007527:Isoenzymes; D005972:Glutaminase