Profiling of myristoylation in Toxoplasma gondii reveals an N-myristoylated protein important for host cell penetration.

Malgorzata Broncel,Alex Hunt,Bethan A Wallbank,Moritz Treeck,Luis Vigetti,Joanna C Young,Stephanie D Nofal,Edward J Bartlett,Isabelle Tardieux,Bastien Touquet,Stephen Matthews,Edward W Tate,Caia Dominicus,Stefania Federico

doi:10.7554/elife.57861

Abstract

N-myristoylation is a ubiquitous class of protein lipidation across eukaryotes and N-myristoyl transferase (NMT) has been proposed as an attractive drug target in several pathogens. Myristoylation often primes for subsequent palmitoylation and stable membrane attachment, however, growing evidence suggests additional regulatory roles for myristoylation on proteins. Here we describe the myristoylated proteome of Toxoplasma gondii using chemoproteomic methods and show that a small-molecule NMT inhibitor developed against related Plasmodium spp. is also functional in Toxoplasma. We identify myristoylation on a transmembrane protein, the microneme protein 7 (MIC7), which enters the secretory pathway in an unconventional fashion with the myristoylated N-terminus facing the lumen of the micronemes. MIC7 and its myristoylation play a crucial role in the initial steps of invasion, likely during the interaction with and penetration of the host cell. Myristoylation of secreted eukaryotic proteins represents a substantial expansion of the functional repertoire of this co-translational modification.

Highlights

Toxoplasmosis currently affects approximately one third of the world’s population (RobertGangneux and Darde, 2012)
Utilizing conditional substrate depletion and complementation with wild-type and myristoylation mutant versions, we demonstrate that myristoylation of microneme protein 7 (MIC7) is functionally important in host cell invasion
The absence of palmitoylation cannot exclude the presence of other secondary signals, such as polybasic regions and protein-protein interactions (PPIs) sites, which could still aid in plasma membrane (PM) attachment, we predict that about half of the substrates we identified are likely only myristoylated at the N-terminus

Summary

Introduction

Toxoplasmosis currently affects approximately one third of the world’s population (RobertGangneux and Darde, 2012). Key steps in the successful propagation of Toxoplasma infection in the acute phase are orchestrated cycles of invasion and egress of tachyzoites from host cells (Black and Boothroyd, 2000). These crucial processes are regulated by several post-translational modifications (PTMs), such as phosphorylation (Gaji et al, 2015; Jacot and Soldati-Favre, 2012; Lourido et al, 2010; Treeck et al, 2014), ubiquitination (Silmon de Monerri et al, 2015), and protein lipidation, such as palmitoylation and myristoylation (Alonso et al, 2012; Frenal et al, 2014).

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