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Anthrax edema factor toxicity is strongly mediated by the N-end rule.

Leysath CE, Phillips DD, Crown D, Fattah RJ, Moayeri M, Leppla SH - PLoS ONE (2013)

Bottom Line: We show that EF adenylate cyclase toxin activity is strongly mediated by the N-end rule, and thus is dependent on the identity of the N-terminal amino acid.EF variants having unfavorable, destabilizing N-terminal residues showed much greater activity in cells when the E1 ubiquitin ligase was inactivated or when proteasome inhibitors were present.Taken together, these results show that EF is uniquely affected by ubiquitination and/or proteasomal degradation.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, United States of America.

ABSTRACT
Anthrax edema factor (EF) is a calmodulin-dependent adenylate cyclase that converts adenosine triphosphate (ATP) into 3'-5'-cyclic adenosine monophosphate (cAMP), contributing to the establishment of Bacillus anthracis infections and the resulting pathophysiology. We show that EF adenylate cyclase toxin activity is strongly mediated by the N-end rule, and thus is dependent on the identity of the N-terminal amino acid. EF variants having different N-terminal residues varied by more than 100-fold in potency in cultured cells and mice. EF variants having unfavorable, destabilizing N-terminal residues showed much greater activity in cells when the E1 ubiquitin ligase was inactivated or when proteasome inhibitors were present. Taken together, these results show that EF is uniquely affected by ubiquitination and/or proteasomal degradation.

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Production of cAMP by purified EF N-terminal variants in RAW264.7 cells.Dilutions of EF N-terminal variants were incubated with 500 ng/mL PA on RAW264.7 cells for 2 h at 37°C before lysis and cAMP measurement. The concentration of EF estimated to induce 50 nM of cAMP production is presented here.
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pone-0074474-g002: Production of cAMP by purified EF N-terminal variants in RAW264.7 cells.Dilutions of EF N-terminal variants were incubated with 500 ng/mL PA on RAW264.7 cells for 2 h at 37°C before lysis and cAMP measurement. The concentration of EF estimated to induce 50 nM of cAMP production is presented here.

Mentions: Characterization of these six purified EF variants on RAW264.7 cells produced results consistent with the previous supernatant experiments (Figure 2). EF variants with histidine, aspartic acid, or arginine at the N-terminus were lower in activity than variants harboring cysteine, asparagine, or alanine as the first residue.


Anthrax edema factor toxicity is strongly mediated by the N-end rule.

Leysath CE, Phillips DD, Crown D, Fattah RJ, Moayeri M, Leppla SH - PLoS ONE (2013)

Production of cAMP by purified EF N-terminal variants in RAW264.7 cells.Dilutions of EF N-terminal variants were incubated with 500 ng/mL PA on RAW264.7 cells for 2 h at 37°C before lysis and cAMP measurement. The concentration of EF estimated to induce 50 nM of cAMP production is presented here.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3755998&req=5

pone-0074474-g002: Production of cAMP by purified EF N-terminal variants in RAW264.7 cells.Dilutions of EF N-terminal variants were incubated with 500 ng/mL PA on RAW264.7 cells for 2 h at 37°C before lysis and cAMP measurement. The concentration of EF estimated to induce 50 nM of cAMP production is presented here.
Mentions: Characterization of these six purified EF variants on RAW264.7 cells produced results consistent with the previous supernatant experiments (Figure 2). EF variants with histidine, aspartic acid, or arginine at the N-terminus were lower in activity than variants harboring cysteine, asparagine, or alanine as the first residue.

Bottom Line: We show that EF adenylate cyclase toxin activity is strongly mediated by the N-end rule, and thus is dependent on the identity of the N-terminal amino acid.EF variants having unfavorable, destabilizing N-terminal residues showed much greater activity in cells when the E1 ubiquitin ligase was inactivated or when proteasome inhibitors were present.Taken together, these results show that EF is uniquely affected by ubiquitination and/or proteasomal degradation.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, United States of America.

ABSTRACT
Anthrax edema factor (EF) is a calmodulin-dependent adenylate cyclase that converts adenosine triphosphate (ATP) into 3'-5'-cyclic adenosine monophosphate (cAMP), contributing to the establishment of Bacillus anthracis infections and the resulting pathophysiology. We show that EF adenylate cyclase toxin activity is strongly mediated by the N-end rule, and thus is dependent on the identity of the N-terminal amino acid. EF variants having different N-terminal residues varied by more than 100-fold in potency in cultured cells and mice. EF variants having unfavorable, destabilizing N-terminal residues showed much greater activity in cells when the E1 ubiquitin ligase was inactivated or when proteasome inhibitors were present. Taken together, these results show that EF is uniquely affected by ubiquitination and/or proteasomal degradation.

Show MeSH
Related in: MedlinePlus