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Proteolytic elimination of N-myristoyl modifications by the Shigella virulence factor IpaJ


Protein N-myristoylation is a 14-carbon fatty-acid modification that is conserved across eukaryotic species and occurs on nearly 1% of the cellular proteome1,2. The ability of the myristoyl group to facilitate dynamic protein–protein and protein–membrane interactions (known as the myristoyl switch) makes it an essential feature of many signal transduction systems3. Thus pathogenic strategies that facilitate protein demyristoylation would markedly alter the signalling landscape of infected host cells. Here we describe an irreversible mechanism of protein demyristoylation catalysed by invasion plasmid antigen J (IpaJ), a previously uncharacterized Shigella flexneri type III effector protein with cysteine protease activity. A yeast genetic screen for IpaJ substrates identified ADP-ribosylation factor (ARF)1p and ARF2p, small molecular mass GTPases that regulate cargo transport through the Golgi apparatus4. Mass spectrometry showed that IpaJ cleaved the peptide bond between N-myristoylated glycine-2 and asparagine-3 of human ARF1, thereby providing a new mechanism for host secretory inhibition by a bacterial pathogen5,6. We further demonstrate that IpaJ cleaves an array of N-myristoylated proteins involved in cellular growth, signal transduction, autophagasome maturation and organelle function. Taken together, these findings show a previously unrecognized pathogenic mechanism for the site-specific elimination of N-myristoyl protein modification.

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Figure 1: Shigella IpaJ and VirA disrupt Golgi morphology.
Figure 2: IpaJ belongs to the C39-like family of cysteine proteases and targets ARF-family GTPases.
Figure 3: IpaJ cleaves the N-myristoylated glycine of lipidated substrates.
Figure 4: Conformational-dependent cleavage of lipidated substrates by IpaJ.

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We thank our colleagues at University of Texas Southwestern Medical Center, specifically K. Orth, Y. M. Chook and J. Seeman, for discussions in preparing this manuscript. We are particularly indebted to L. Kinch, N. Grishin, D. Mitchel, X. Guo, D. Trudgian and S. Perelman for their contributions. We acknowledge the services of the University of Texas Southwestern Medical Center proteomics core, supported by a Cancer Prevention and Research Institute of Texas grant RP120613. A.S.S. was supported by the Howard Hughes Medical Institute International Student Research fellowship. B.A.W. was supported by a National Institutes of Health training grant (NIAID; 5T32AI007520) and S.S.W. by the Burroughs Wellcome Fund. This work was supported by grants from the National Institutes of Health (NIAID; RO1AI083359 and NIGMS; R01GM100486), the Welch Foundation (I-1704) and the Burroughs Wellcome Fund to N.M.A.

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Authors and Affiliations



N.B. and N.M.A. conceived the general ideas for this work. N.B. and N.M.A. planned, performed and interpreted experiments. T.G.F. and A.S.S. generated bacterial knockout strains and performed GTPase assays, respectively. J.M.E. and S.S.W. performed mouse infection studies. B.A.W. performed the bioinformatics studies on IpaJ. D.A.P. and S.M.P. performed top-down mass spectrometry. N.M.A. and N.B. wrote the manuscript and all authors provided editorial input.

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Correspondence to Neal M. Alto.

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The authors declare no competing financial interests.

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Burnaevskiy, N., Fox, T., Plymire, D. et al. Proteolytic elimination of N-myristoyl modifications by the Shigella virulence factor IpaJ. Nature 496, 106–109 (2013).

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