Antisense glutaminase inhibition modifies the O-GlcNAc pattern and flux through the hexosamine pathway in breast cancer cells

J Cell Biochem. 2008 Feb 15;103(3):800-11. doi: 10.1002/jcb.21449.


Glutamine behaves as a key nutrient for tumors and rapidly dividing cells. Glutaminase is the main glutamine-utilizing enzyme in these cells, and its activity correlates with glutamine consumption and growth rate. We have carried out the antisense L-type glutaminase inhibition in human MCF7 breast cancer cells, in order to study its effect on the hexosamine pathway and the pattern of protein O-glycosylation. The antisense mRNA glutaminase expressing cells, named ORF19, presented a 50% lower proliferation rate than parental cells, showing a more differentiated phenotype. ORF19 cells had an 80% reduction in glutamine:fructose-6-P amidotransferase activity, which is the rate-limiting step of the hexosamine pathway. Although the overall cellular protein O-glycosylation did not change, the O-glycosylation status of several key proteins was altered. O-glycosylation of O-GlcNAc transferase (OGT), the enzyme that links N-acetylglucosamine to proteins, was fivefold lower in ORF19 than in wild type cells. Inhibition of glutaminase also provoked a 10-fold increase in Sp1 expression, and a significant decrease in the ratio of O-glycosylated to total protein for both Sp1 and the Rpt2 proteasome component. These changes were accompanied by a higher Sp1 transcriptional activity. Proteome analysis of O-glycosylated proteins permitted the detection of two new OGT target proteins: the chaperonin TCP-1 theta and the oncogene Ets-related protein isoform 7. Taken together, our results support the hexosamine pathway and the O-glycosylation of proteins being a sensor mechanism of the nutritional and energetic states of the cell.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Breast Neoplasms / enzymology*
  • Breast Neoplasms / genetics*
  • Breast Neoplasms / pathology
  • Gene Expression / genetics
  • Glutaminase / antagonists & inhibitors
  • Glutaminase / genetics
  • Glutaminase / metabolism*
  • Glutamine / metabolism
  • Hexosamines / biosynthesis
  • Hexosamines / metabolism*
  • Humans
  • Mice
  • N-Acetylglucosaminyltransferases / metabolism
  • Protein Processing, Post-Translational / genetics
  • Proteomics
  • RNA Interference
  • Sp1 Transcription Factor / biosynthesis*
  • Sp1 Transcription Factor / metabolism
  • Tumor Cells, Cultured
  • Uridine Diphosphate N-Acetylglucosamine / metabolism


  • Hexosamines
  • Sp1 Transcription Factor
  • Glutamine
  • Uridine Diphosphate N-Acetylglucosamine
  • N-Acetylglucosaminyltransferases
  • O-GlcNAc transferase
  • Glutaminase