Apicomplexan rhomboids have a potential role in microneme protein cleavage during host cell invasion

Area of Science:

• Parasitology
• Molecular Biology
• Biochemistry

Background:

• Apicomplexan parasites, like *Toxoplasma gondii* and *Plasmodium* species, utilize transmembrane adhesive proteins for host cell invasion.
• These essential proteins undergo cleavage within their transmembrane domains by an unknown protease, microneme protein protease 1 (MPP1).
• The conserved cleavage site (IA↓GG) in *T. gondii* microneme proteins TgMIC2 and TgMIC6 suggests a conserved mechanism across apicomplexans.

Purpose of the Study:

• To identify and characterize the unknown protease responsible for cleaving apicomplexan transmembrane adhesive proteins.
• To investigate the role of the rhomboid family of intramembrane-cleaving serine proteases in this process within *T. gondii*.

Main Methods:

• Bioinformatic analysis to identify rhomboid-like proteins in *T. gondii*.
• Expression and functional assays of candidate rhomboid proteases using *Drosophila* Spitz and chimeric proteins containing apicomplexan transmembrane domains.

Main Results:

• Six *T. gondii* rhomboid-like (TgROM) genes were identified, with four localized to the secretory pathway.
• TgROM1, TgROM2, and TgROM5 demonstrated protease activity by cleaving the transmembrane domain of *Drosophila* Spitz.
• TgROM2 specifically cleaved chimeric proteins incorporating the transmembrane domains of *T. gondii* microneme proteins TgMIC2 and TgMIC12.

Conclusions:

• *Toxoplasma gondii* possesses active rhomboid proteases (TgROMs) capable of cleaving transmembrane domains.
• TgROM2 is a strong candidate for MPP1, the protease that processes apicomplexan microneme proteins for host cell invasion.
• These findings provide crucial insights into the molecular mechanisms of apicomplexan parasite invasion.