Arginine and Lysine Transporters Are Essential for Trypanosoma brucei

Abstract

For Trypanosoma brucei arginine and lysine are essential amino acids and therefore have to be imported from the host. Heterologous expression in Saccharomyces cerevisiae mutants identified cationic amino acid transporters among members of the T. brucei AAAP (amino acid/auxin permease) family. TbAAT5-3 showed high affinity arginine uptake (Km 3.6 ± 0.4 μM) and high selectivity for L-arginine. L-arginine transport was reduced by a 10-times excess of L-arginine, homo-arginine, canavanine or arginine-β-naphthylamide, while lysine was inhibitory only at 100-times excess, and histidine or ornithine did not reduce arginine uptake rates significantly. TbAAT16-1 is a high affinity (Km 4.3 ± 0.5 μM) and highly selective L-lysine transporter and of the compounds tested, only L-lysine and thialysine were competing for L-lysine uptake. TbAAT5-3 and TbAAT16-1 are expressed in both procyclic and bloodstream form T. brucei and cMyc-tagged proteins indicate localization at the plasma membrane. RNAi-mediated down-regulation of TbAAT5 and TbAAT16 in bloodstream form trypanosomes resulted in growth arrest, demonstrating that TbAAT5-mediated arginine and TbAAT16-mediated lysine transport are essential for T. brucei. Growth of induced RNAi lines could partially be rescued by supplementing a surplus of arginine or lysine, respectively, while addition of both amino acids was less efficient. Single and double RNAi lines indicate that additional low affinity uptake systems for arginine and lysine are present in T. brucei.

The Uptake and Metabolism of Amino Acids, and Their Unique Role in the Biology of Pathogenic Trypanosomatids

“…TbAAT5-3 and TbAAT16-1 have been heterologously expressed, showing high affinity for Arg and Lys (in both cases K M in the low µM range) and high selectivity (including stereoselectivity, the D-isomer does not affect the transport). Peculiarly, down-regulation of these transporters affected BSF growth, demonstrating they are essential for the parasites [ 45 ]. In addition, it has been shown that T. brucei has two high-affinity Orn transporters, TbAAT10-1 and TbAAT2-4.…”

Trypanosomatid Infections: How Do Parasites and Their Excreted–Secreted Factors Modulate the Inducible Metabolism of l-Arginine in Macrophages?

“…Interestingly, arginine kinase and arginine N -methyl transferase genes were overexpressed in Tbg isolated from tsetse flies ( 127 ), as if targeting l -arginine were a metabolic key in the developmental life cycle of African trypanosomes. l -Arginine transporters were also defined as essential for trypanosomes ( 128 ). Differences in arginase subcellular locations between Tbb and in T. cruzi have been reported ( 129 , 130 ), but their biological significance remains to be determined.…”

Crystal violet structural analogues identified by in silico drug repositioning present anti-Trypanosoma cruzi activity through inhibition of proline transporter TcAAAP069

“…The loss of TbAAT6 is sufficient to develop resistance to eflornithine, a drug used to treat human African trypanosomiasis (HAT) (19). Moreover, the importance of amino acid transporters in T. brucei has been demonstrated by RNAi-mediated knock-down of arginine and lysine transporters TbAAT5-3 and TbAAT16-1 from the AAAP family which resulted in growth arrest in bloodstream forms (20). In Leishmania donovani , the transporter LdAAP24 mediates proline and alanine uptake and regulates the response to osmotic stress and amino acid homeostasis (21).…”