A transient forward-targeting element for microneme-regulated secretion in Toxoplasma gondii.
BACKGROUND
Accurate sorting of proteins to the three types of secretory granules in Toxoplasma gondii is crucial for successful cell invasion by this obligate intracellular parasite. As in other eukaryotic systems, propeptide sequences are a common yet poorly understood feature of proteins destined for regulated secretion, which for Toxoplasma occurs through two distinct invasion organelles, rhoptries and micronemes. Microneme discharge during parasite apical attachment plays a pivotal role in cell invasion by delivering adhesive proteins for host receptor engagement.


RESULTS
We show here that the small micronemal proprotein MIC5 (microneme protein-5) undergoes proteolytic maturation at a site beyond the Golgi, and only the processed form of MIC5 is secreted via the micronemes. Proper cleavage of the MIC5 propeptide relies on an arginine residue in the P1' position, although P1' mutants are still cleaved to a lesser extent at an alternative site downstream of the primary site. Nonetheless, this aberrantly cleaved species still correctly traffics to the micronemes, indicating that correct cleavage is not necessary for micronemal targeting. In contrast, a deletion mutant lacking the propeptide was retained within the secretory system, principally in the ER (endoplasmic reticulum). The MIC5 propeptide also supported correct trafficking when exchanged for the M2AP propeptide, which was recently shown to also be required for micronemal trafficking of the TgMIC2 (T. gondii MIC2)-M2AP complex [Harper, Huynh, Coppens, Parussini, Moreno and Carruthers (2006) Mol. Biol. Cell 17, 4551-4563].


CONCLUSIONS
Our results illuminate common and unique features of micronemal propeptides in their role as trafficking facilitators.
Brydges
Susannah D
SD
W. Harry Feinstone Department of Molecular Microbiology and Immunology, Johns Hopkins Bloomberg School of Public Health, Baltimore, MD 21205, USA.
Harper
Jill M
JM
Parussini
Fabiola
F
Coppens
Isabelle
I
Carruthers
Vern B
VB
D000595:Amino Acid Sequence; D000818:Animals; D030801:Animals, Genetically Modified; D002614:Chelating Agents / Q000494:pharmacology; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D006056:Golgi Apparatus / Q000378:metabolism; D006801:Humans; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D015388:Organelles / Q000378:metabolism*; D010957:Plasmids; D017510:Protein Folding; D011499:Protein Processing, Post-Translational; D021381:Protein Transport; D015800:Protozoan Proteins / Q000032:analysis / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D022142:Secretory Vesicles / Q000378:metabolism; D014122:Toxoplasma / Q000378:metabolism* / Q000648:ultrastructure
D002614:Chelating Agents; D015800:Protozoan Proteins; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid