Editors' ChoicePosttranslational modification

Spreading AMPylation?

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Sci. Signal.  14 Apr 2009:
Vol. 2, Issue 66, pp. ec131
DOI: 10.1126/scisignal.266ec131

The bacterial pathogen Histophilus somni secretes IbpA (immunoglobulin-binding protein A), a surface antigen with a domain that resembles YopT (a cytotoxic protein produced by pathogenic Yersinia species). Interested in determining whether the IbpA YopT-like domain functions like YopT (a cysteine protease that targets Rho family GTPases, thereby disrupting the actin cytoskeleton), Worby et al. expressed a fluorescently labeled fusion protein containing the region of IbpA that includes the YopT-like domain (IbpA-COOH) in HeLa cells. Although IbpA-COOH disrupted stress fibers and elicited rounding, the phenotype was less severe than that produced by YopT. Moreover, mutational analysis revealed that the effects of IbpA-COOH depended not on the YopT-like domain but on two Fic (filamentation induced by cAMP) domains—a domain recently implicated in Vibrio parahaemolyticus pathogenesis, through addition of adenosine 5'-monophosphate (AMP) to a threonine residue of Rho GTPases. IbpA Fic 1 and Fic2 inhibited binding of activated RhoA and Rac to downstream effectors, whereas a noncytotoxic form of Fic2, in which a conserved histidine was substituted with an alanine, failed to do so. Mass spectrometric analysis revealed that, when GST-labeled Fic2 was incubated with ATP, Mg2+, and GST-labeled RhoA, Rac, or Cdc42, an AMP residue was added to a conserved tyrosine found in all three GTPases. Consistent with these observations, 32P was transferred by Fic2 from α32P-ATP to RhoA, Rac, and Cdc42 but not to forms in which the conserved tyrosine was substituted with phenylalanine (or to dominant-negative forms). BLAST analysis identified Fic domains in bacteria, prophages, and eukaryotes, including huntingtin yeast interacting protein E (HYPE) in humans. HYPE, the only human protein with a Fic domain, retained the ability to AMPylate Rho GTPases, although it did not elicit HeLa cell rounding and was less effective at inhibiting RhoA-effector interaction. Thus, the authors conclude that AMPylation is a Fic domain–mediated posttranslational modification that has been evolutionarily conserved.

C. A. Worby, S. Mattoo, R. P. Kruger, L. B. Corbeil, A. Koller, J. C. Mendez, B. Zekarias, C. Lazar, J. E. Dixon, The Fic domain: Regulation of cell signaling by adenylylation. Mol. Cell 34, 93–103 (2009). [Online Journal]

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