The tyrosine transporter of Toxoplasma gondii is a member of the newly defined apicomplexan amino acid transporter (ApiAT) family

Abstract

Apicomplexan parasites are auxotrophic for a range of amino acids which must be salvaged from their host cells, either through direct uptake or degradation of host proteins. Here, we describe a family of plasma membrane-localized amino acid transporters, termed the Apicomplexan Amino acid Transporters (ApiATs), that are ubiquitous in apicomplexan parasites. Functional characterization of the ApiATs of Toxoplasma gondii indicate that several of these transporters are important for intracellular growth of the tachyzoite stage of the parasite, which is responsible for acute infections. We demonstrate that the ApiAT protein Tg ApiAT5-3 is an exchanger for aromatic and large neutral amino acids, with particular importance for L-tyrosine scavenging and amino acid homeostasis, and that Tg ApiAT5-3 is critical for parasite virulence. Our data indicate that T . gondii expresses additional proteins involved in the uptake of aromatic amino acids, and we present a model for the uptake and homeostasis of these amino acids. Our findings identify a family of amino acid transporters in apicomplexans, and highlight the importance of amino acid scavenging for the biology of this important phylum of intracellular parasites.

Substrate-mediated regulation of the arginine transporter ofToxoplasma gondii

“…We examined the effects of the concentration of a range of other nutrients, including L-tyrosine 141 (Tyr), L-glutamine (Gln) and D-glucose, on the nanoLUC:fLUC luminescence ratio in the dual 142 reporter strain and on TgApiAT1-HA3 protein abundance. At the lowest concentration of each 143 nutrient tested, we observed decreased TgApiAT1-HA3 protein abundance and a significantly 144 concentrations tested for Tyr and Gln were close to the minimal amount of those nutrients 146 required for optimal parasite growth [14,17]. This is consistent with the hypothesis that 147…”

“…To explore further the relationship 185 between parasite [Arg] and TgApAT1 regulation, we introduced a 'knockout' frameshift 186 mutation in the TgApiAT1 locus of the dual luciferase reporter strain, generating a strain we 187 termed apiAT1 ∆54-534 . As demonstrated previously for parasites lacking TgApiAT1 [14,15], 188 apiAT1 ∆54-534 parasites exhibited reduced proliferation over an 8-day growth assay in 189…”

“…TgApiAT1 if the concentration of Lys, a competitive inhibitor of Arg uptake via the transporter, 155 is low [15]. Our unpublished research indicates that this transporter is TgApiAT6-1 ( Figure 156 4A; Rajendran, Fairweather, et al, in preparation), another member of the TgApiAT family 157 that localises to the parasite plasma membrane [14]. Using an HA-tagged TgApiAT6-1 strain 158 [14], we asked whether TgApiAT6-1-HA3 abundance is regulated during growth in media 159 containing a range of amino acid concentrations.…”

“…Our unpublished research indicates that this transporter is TgApiAT6-1 ( Figure 156 4A; Rajendran, Fairweather, et al, in preparation), another member of the TgApiAT family 157 that localises to the parasite plasma membrane [14]. Using an HA-tagged TgApiAT6-1 strain 158 [14], we asked whether TgApiAT6-1-HA3 abundance is regulated during growth in media 159 containing a range of amino acid concentrations. We found that the abundance of TgApiAT6-160 1-HA3 did not differ in any of the tested Arg or Lys concentrations (Figure 4B-C To distinguish between the two possibilities, we generated a regulatable TgApiAT6-1 172 (rTgApiAT6-1) parasite strain, in which TgApiAT6-1 expression can be knocked down 173 through the addition of anhydrotetracycline (ATc; Figure S4).…”

“…Amino acid Transporter (ApiAT) family [14]. We have demonstrated that one member of this 59 family, TgApiAT1, is an Arg transporter that is essential for T. gondii virulence [15].…”