TgPRELID, a Mitochondrial Protein Linked to Multidrug Resistance in the ParasiteToxoplasma gondii.
New drugs to control infection with the protozoan parasiteToxoplasma gondiiare needed as current treatments exert toxic side effects on patients. Approaches to develop novel compounds for drug development include screening of compound libraries and targeted inhibition of essential cellular pathways. We identified two distinct compounds that display inhibitory activity against the parasite's replicative stage: F3215-0002, which we previously identified during a compound library screen, and I-BET151, an inhibitor of bromodomains, the "reader" module of acetylated lysines. In independent studies, we sought to determine the targets of these two compounds using forward genetics, generating resistant mutants and identifying the determinants of resistance with comparative genome sequencing. Despite the dissimilarity of the two compounds, we recovered resistant mutants with nonsynonymous mutations in the same domain of the same gene,TGGT1_254250, which we found encodes a protein that localizes to the parasite mitochondrion (designated TgPRELID after the name of said domain). We found that mutants selected with one compound were cross resistant to the other compound, suggesting a common mechanism of resistance. To further support our hypothesis thatTgPRELIDmutations facilitate resistance to both I-BET151 and F3215-0002, CRISPR (clustered regularly interspaced short palindromic repeat)/CAS9-mediated mutation ofTgPRELIDdirectly led to increased F3215-0002 resistance. Finally, all resistance mutations clustered in the same subdomain of TgPRELID. These findings suggest thatTgPRELIDmay encode a multidrug resistance factor or that I-BET151 and F3215-0002 have the same target(s) despite their distinct chemical structures.IMPORTANCEWe report the discovery of TgPRELID, a previously uncharacterized mitochondrial protein linked to multidrug resistance in the parasiteToxoplasma gondii. Drug resistance remains a major problem in the battle against parasitic infection, and understanding how TgPRELID mutations augment resistance to multiple, distinct compounds will reveal needed insights into the development of new therapies for toxoplasmosis and other related parasitic diseases.
Department of Pharmacology and Toxicology, Indiana University School of Medicine, Indianapolis, Indiana, USA.
Victoria
0000-0003-1233-1088
V
Jeffers
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania, USA.
Edwin T
ET
Kamau
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania, USA.
Ananth R
AR
Srinivasan
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania, USA.
Jonathan
J
Harper
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania, USA.
Preethi
P
Sankaran
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania, USA.
Sarah E
SE
Post
Department of Pharmacology and Toxicology, Indiana University School of Medicine, Indianapolis, Indiana, USA.
Joseph M
JM
Varberg
Department of Pharmacology and Toxicology, Indiana University School of Medicine, Indianapolis, Indiana, USA; Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, Indiana, USA.
William J
WJ
Sullivan
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania, USA.
Jon P
JP
Boyle
PRELI domain; Toxoplasma gondii; mitochondrial protein import; multidrug resistance