Two separate, conserved acidic amino acid domains within the Toxoplasma gondii MIC2 cytoplasmic tail are required for parasite survival.
Apicomplexan parasites rely on actin-based motility to drive host cell invasion. Motility and invasion also require thrombospondin-related anonymous protein (TRAP) adhesins, which are secreted apically and translocated to the posterior end of the parasite before they are shed by the activity of a rhomboid protease. TRAP orthologs, including Toxoplasma gondii MIC2 (microneme protein 2), possess a short cytoplasmic tail, which is essential for motility. Previous studies have shown that aldolase forms a critical bridge between actin filaments and the cytoplasmic domains of MIC2 and TRAP. The cytoplasmic tails of TRAP family members harbor a conserved penultimate tryptophan, which is essential for aldolase binding, and clustered acidic residues. Herein, we determined the role of the conserved acidic residues by using alanine point mutants to investigate aldolase binding in vitro and to test functionality in the parasite. Our studies revealed two separate acidic residue clusters in the cytoplasmic domain of MIC2 that are essential for parasite survival. One region, located at the extreme C terminus, is required for the direct interaction with aldolase, whereas the second upstream acidic region is not necessary for aldolase binding but is nonetheless essential to parasite survival. Both acidic domains are conserved throughout TRAP orthologs, implicating a central role for these motifs in apicomplexan motility.
Starnes
G Lucas
GL
Department of Molecular Microbiology, Washington University School of Medicine, St. Louis, Missouri 63130-1093, USA.
Jewett
Travis J
TJ
Carruthers
Vern B
VB
Sibley
L David
LD
D000409:Alanine / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D003593:Cytoplasm / Q000378:metabolism*; D004252:DNA Mutational Analysis; D005634:Fructose-Bisphosphate Aldolase / Q000737:chemistry; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D008957:Models, Genetic; D008969:Molecular Sequence Data; D017354:Point Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D015800:Protozoan Proteins / Q000737:chemistry* / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014122:Toxoplasma / Q000378:metabolism*; D014364:Tryptophan / Q000737:chemistry
C408878:MIC2 protein, Toxoplasma gondii; D008565:Membrane Proteins; D015800:Protozoan Proteins; D014364:Tryptophan; D005634:Fructose-Bisphosphate Aldolase; D000409:Alanine