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Regulation of prostaglandin synthesis and cell adhesion by a tryptophan catabolizing enzyme.

Marshall B, Keskin DB, Mellor AL - BMC Biochem. (2001)

Bottom Line: Therefore, we investigated the circumstances under which IDO is expressed in vitro together with the effects of overexpression of IDO on the growth and morphology of cells.In the case of RAW cells, IDO effects on cell growth could be reversed by adding back prostaglandins.These results suggest that catabolism of the rarest essential amino acid may regulate processes such as cell adhesion and prostaglandin synthesis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Program in Molecular Immunology, Institute for Molecular Medicine and Genetics, Medical College of Georgia, CB 2803, 1120 15th Street, Augusta, GA 30912-3175, USA. marshall@immagene.mcg.edu

ABSTRACT

Background: The tryptophan catabolizing enzyme, indoleamine 2,3, dioxygenase (IDO) is one of two mammalian enzymes, which can catabolize the rarest essential amino acid, tryptophan. IDO is inducible by cytokines such as interferon-gamma and plays a role in inflammation and maternal tolerance of fetal allografts, although its exact mode of action is unclear. Therefore, we investigated the circumstances under which IDO is expressed in vitro together with the effects of overexpression of IDO on the growth and morphology of cells.

Results: Overexpression of IDO in the murine macrophage cell line RAW 264.7 and the murine fibrosarcoma cell line MC57, resulted in the growth of macroscopic cell foci, with altered cell adhesion properties. The expression of IDO was also detected during adhesion of wild type, nontransfected cells in tissue culture to standard cell growth substrates. Inhibition of this expression, likewise resulted in alterations in cell adhesion. Overexpression of IDO or inhibition of endogenous IDO expression was accompanied by changes in metalloproteinase expression and also in the expression and activity of the cyclooxygenase enzymes. In the case of RAW cells, IDO effects on cell growth could be reversed by adding back prostaglandins.

Conclusions: These results suggest that catabolism of the rarest essential amino acid may regulate processes such as cell adhesion and prostaglandin synthesis.

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Effect of IDO on cyclooxygenase expression clones. (A) Expression of COX-1 and COX-2 protein by vector-only transfected (Vo) and IDO expressing clones. (B) Effect of LPS treatment on expression of COX-1 and COX-2 in IDO-expressing clones. Vector-only and IDO-expressing RAW cells were treated with 1 ng/ml LPS for 12 hours. (C) COX-2 expression in vector-only or IDO-expressing MC57 cells.
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Figure 8: Effect of IDO on cyclooxygenase expression clones. (A) Expression of COX-1 and COX-2 protein by vector-only transfected (Vo) and IDO expressing clones. (B) Effect of LPS treatment on expression of COX-1 and COX-2 in IDO-expressing clones. Vector-only and IDO-expressing RAW cells were treated with 1 ng/ml LPS for 12 hours. (C) COX-2 expression in vector-only or IDO-expressing MC57 cells.

Mentions: In IDO-expressing RAW cells, COX-1 protein levels were unchanged compared to the vector-only control (Fig 8A). In contrast, COX-2 was not expressed by vector only controls or RAW clones 6, 8 and 22 but COX-2 mRNA and protein was induced in the RAW clone expressing the greatest amount of IDO (clone 11) (Fig 8A). Although COX-2 is not usually expressed in RAW cells, it can be strongly induced with lipopolysaccharide (LPS) [32, 33]. Therefore, we treated IDO transfected RAW cells and controls with LPS and measured COX-2 expression 24 hours later. COX-2 mRNA was most strongly induced in vector only or low IDO-expressing clones (Fig 8B). Curiously, clones expressing higher levels of IDO (clones 22 and 11) showed lower levels of COX-2 mRNA induction. In contrast, COX-2 protein levels were higher in clones expressing lower amounts of IDO mRNA and lower in vector only controls, whereas COX-1 protein levels were unchanged by LPS treatment. MC57 cells expressed COX-2 constitutively, consistent with the domination of the PG profile by PG E2. However, IDO overexpressing clone 26 showed a reduced amount of COX-2 protein compared to the vector only control (Fig 8C).


Regulation of prostaglandin synthesis and cell adhesion by a tryptophan catabolizing enzyme.

Marshall B, Keskin DB, Mellor AL - BMC Biochem. (2001)

Effect of IDO on cyclooxygenase expression clones. (A) Expression of COX-1 and COX-2 protein by vector-only transfected (Vo) and IDO expressing clones. (B) Effect of LPS treatment on expression of COX-1 and COX-2 in IDO-expressing clones. Vector-only and IDO-expressing RAW cells were treated with 1 ng/ml LPS for 12 hours. (C) COX-2 expression in vector-only or IDO-expressing MC57 cells.
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Related In: Results  -  Collection

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Figure 8: Effect of IDO on cyclooxygenase expression clones. (A) Expression of COX-1 and COX-2 protein by vector-only transfected (Vo) and IDO expressing clones. (B) Effect of LPS treatment on expression of COX-1 and COX-2 in IDO-expressing clones. Vector-only and IDO-expressing RAW cells were treated with 1 ng/ml LPS for 12 hours. (C) COX-2 expression in vector-only or IDO-expressing MC57 cells.
Mentions: In IDO-expressing RAW cells, COX-1 protein levels were unchanged compared to the vector-only control (Fig 8A). In contrast, COX-2 was not expressed by vector only controls or RAW clones 6, 8 and 22 but COX-2 mRNA and protein was induced in the RAW clone expressing the greatest amount of IDO (clone 11) (Fig 8A). Although COX-2 is not usually expressed in RAW cells, it can be strongly induced with lipopolysaccharide (LPS) [32, 33]. Therefore, we treated IDO transfected RAW cells and controls with LPS and measured COX-2 expression 24 hours later. COX-2 mRNA was most strongly induced in vector only or low IDO-expressing clones (Fig 8B). Curiously, clones expressing higher levels of IDO (clones 22 and 11) showed lower levels of COX-2 mRNA induction. In contrast, COX-2 protein levels were higher in clones expressing lower amounts of IDO mRNA and lower in vector only controls, whereas COX-1 protein levels were unchanged by LPS treatment. MC57 cells expressed COX-2 constitutively, consistent with the domination of the PG profile by PG E2. However, IDO overexpressing clone 26 showed a reduced amount of COX-2 protein compared to the vector only control (Fig 8C).

Bottom Line: Therefore, we investigated the circumstances under which IDO is expressed in vitro together with the effects of overexpression of IDO on the growth and morphology of cells.In the case of RAW cells, IDO effects on cell growth could be reversed by adding back prostaglandins.These results suggest that catabolism of the rarest essential amino acid may regulate processes such as cell adhesion and prostaglandin synthesis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Program in Molecular Immunology, Institute for Molecular Medicine and Genetics, Medical College of Georgia, CB 2803, 1120 15th Street, Augusta, GA 30912-3175, USA. marshall@immagene.mcg.edu

ABSTRACT

Background: The tryptophan catabolizing enzyme, indoleamine 2,3, dioxygenase (IDO) is one of two mammalian enzymes, which can catabolize the rarest essential amino acid, tryptophan. IDO is inducible by cytokines such as interferon-gamma and plays a role in inflammation and maternal tolerance of fetal allografts, although its exact mode of action is unclear. Therefore, we investigated the circumstances under which IDO is expressed in vitro together with the effects of overexpression of IDO on the growth and morphology of cells.

Results: Overexpression of IDO in the murine macrophage cell line RAW 264.7 and the murine fibrosarcoma cell line MC57, resulted in the growth of macroscopic cell foci, with altered cell adhesion properties. The expression of IDO was also detected during adhesion of wild type, nontransfected cells in tissue culture to standard cell growth substrates. Inhibition of this expression, likewise resulted in alterations in cell adhesion. Overexpression of IDO or inhibition of endogenous IDO expression was accompanied by changes in metalloproteinase expression and also in the expression and activity of the cyclooxygenase enzymes. In the case of RAW cells, IDO effects on cell growth could be reversed by adding back prostaglandins.

Conclusions: These results suggest that catabolism of the rarest essential amino acid may regulate processes such as cell adhesion and prostaglandin synthesis.

Show MeSH
Related in: MedlinePlus