title: Aspirin acetylates multiple cellular proteins in HCT-116 colon cancer cells: Identification of novel targets.
abstract: Epidemiological and clinical observations provide consistent evidence that regular intake of aspirin may effectively inhibit the occurrence of epithelial tumors; however, the molecular mechanisms are not completely understood. In the present study, we determined the ability of aspirin to acetylate and post-translationally modify cellular proteins in HCT-116 human colon cancer cells to understand the potential mechanisms by which it may exerts anti-cancer effects. Using anti-acetyl lysine antibodies, here we demonstrate that aspirin causes the acetylation of multiple proteins whose molecular weight ranged from 20 to 200 kDa. The identity of these proteins was determined, using immuno-affinity purification, mass spectrometry and immuno-blotting. A total of 33 cellular proteins were potential targets of aspirin-mediated acetylation, while 16 were identified as common to both the control and aspirin-treated samples. These include enzymes of glycolytic pathway, cytoskeleton proteins, histones, ribosomal and mitochondrial proteins. The glycolytic enzymes which were identified include aldolase, glyceraldehyde-3-phosphate dehydrogenase, enolase, pyruvate kinase M2, and lactate dehydrogenase A and B chains. Immunoblotting experiment showed that aspirin also acetylated glucose-6-phosphate dehydrogenase and transketolase, both enzymes of pentose phosphate pathway involved in ribonucleotide biosynthesis. In vitro assays of these enzymes revealed that aspirin did not affect pyruvate kinase and lactate dehydrogenase activity; however, it decreased glucose 6 phosphate dehydrogenase activity. Similar results were also observed in HT-29 human colon cancer cells. Selective inhibition of glucose-6-phosphate dehydrogenase may represent an important mechanism by which aspirin may exert its anti-cancer effects through inhibition of ribonucleotide synthesis.
authors: Department of Biomedical Sciences, Texas Tech University Health Sciences Center, Amarillo, TX 79106, USA.
authors: Srinivasan
authors: S
authors: Marimuthu
authors: Raghavender S V
authors: RS
authors: Chivukula
authors: Lloyd F
authors: LF
authors: Alfonso
authors: Majid
authors: M
authors: Moridani
authors: Fred K
authors: FK
authors: Hagen
authors: G Jayarama
authors: GJ
authors: Bhat
mesh_terms: D000107:Acetylation / Q000187:drug effects; D001241:Aspirin / Q000494:pharmacology*; D003110:Colonic Neoplasms / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D005954:Glucosephosphate Dehydrogenase / Q000378:metabolism; D006031:Glycosylation / Q000187:drug effects; D045325:HCT116 Cells; D019073:HT29 Cells; D006801:Humans; D045022:Lactate Dehydrogenases / Q000378:metabolism; D013058:Mass Spectrometry; D053858:Metabolic Networks and Pathways / Q000187:drug effects; D010427:Pentose Phosphate Pathway / Q000187:drug effects; D010766:Phosphorylation / Q000187:drug effects; D011506:Proteins / Q000032:analysis / Q000378:metabolism*; D011770:Pyruvate Kinase / Q000378:metabolism
chemical_list: D011506:Proteins; D045022:Lactate Dehydrogenases; D005954:Glucosephosphate Dehydrogenase; D011770:Pyruvate Kinase; D001241:Aspirin