title: Discovery of an ergosterol-signaling factor that regulates Trypanosoma brucei growth.
abstract: Ergosterol biosynthesis and homeostasis in the parasitic protozoan Trypanosoma brucei was analyzed by RNAi silencing and inhibition of sterol C24β-methyltransferase (TbSMT) and sterol 14α-demethylase [TbSDM (TbCYP51)] to explore the functions of sterols in T. brucei growth. Inhibition of the amount or activity of these enzymes depletes ergosterol from cells at <6 fg/cell for procyclic form (PCF) cells or <0.01 fg/cell for bloodstream form (BSF) cells and reduces infectivity in a mouse model of infection. Silencing of TbSMT expression by RNAi in PCF or BSF in combination with 25-azalanosterol (AZA) inhibited parasite growth and this inhibition was restored completely by adding synergistic cholesterol (7.8 μM from lipid-depleted media) with small amounts of ergosterol (1.2 μM) to the medium. These observations are consistent with the proposed requirement for ergosterol as a signaling factor to spark cell proliferation while imported cholesterol or the endogenously formed cholesta-5,7,24-trienol act as bulk membrane components. To test the potential chemotherapeutic importance of disrupting ergosterol biosynthesis using pairs of mechanism-based inhibitors that block two enzymes in the post-squalene segment, parasites were treated with AZA and itraconazole at 1 μM each (ED50 values) resulting in parasite death. Taken together, our results demonstrate that the ergosterol pathway is a prime drug target for intervention in T. brucei infection.
authors.affiliation: Center for Chemical Biology and Department of Chemistry and Biochemistry Texas Tech University, Lubbock, TX 79409.
authors.forename: Brad A
authors.initials: BA
authors.lastname: Haubrich
authors.affiliation: Department of Microbiology and Immunology, Meharry Medical College, Nashville, TN 37208.
authors.forename: Ujjal K
authors.initials: UK
authors.lastname: Singha
authors.affiliation: Center for Chemical Biology and Department of Chemistry and Biochemistry Texas Tech University, Lubbock, TX 79409.
authors.forename: Matthew B
authors.initials: MB
authors.lastname: Miller
authors.affiliation: Center for Chemical Biology and Department of Chemistry and Biochemistry Texas Tech University, Lubbock, TX 79409.
authors.forename: Craigen R
authors.initials: CR
authors.lastname: Nes
authors.affiliation: Center for Chemical Biology and Department of Chemistry and Biochemistry Texas Tech University, Lubbock, TX 79409.
authors.forename: Hosanna
authors.initials: H
authors.lastname: Anyatonwu
authors.affiliation: Laboratoire de Parasitologie Moléculaire, IBMM, Université Libre de Bruxelles, B6041 Gosselies, Belgium.
authors.forename: Laurence
authors.initials: L
authors.lastname: Lecordier
authors.affiliation: Center for Chemical Biology and Department of Chemistry and Biochemistry Texas Tech University, Lubbock, TX 79409.
authors.forename: Presheet
authors.initials: P
authors.lastname: Patkar
authors.affiliation: Center for Chemical Biology and Department of Chemistry and Biochemistry Texas Tech University, Lubbock, TX 79409 Institute of Chemistry and Biomedical Sciences, Nanjing University, Nanjing 210023, People's Republic of China.
authors.forename: David J
authors.initials: DJ
authors.lastname: Leaver
authors.affiliation: Department of Microbiology and Immunology, Meharry Medical College, Nashville, TN 37208.
authors.forename: Fernando
authors.initials: F
authors.lastname: Villalta
authors.affiliation: Laboratoire de Parasitologie Moléculaire, IBMM, Université Libre de Bruxelles, B6041 Gosselies, Belgium.
authors.forename: Benoit
authors.initials: B
authors.lastname: Vanhollebeke
authors.affiliation: Department of Microbiology and Immunology, Meharry Medical College, Nashville, TN 37208.
authors.forename: Minu
authors.initials: M
authors.lastname: Chaudhuri
authors.affiliation: Center for Chemical Biology and Department of Chemistry and Biochemistry Texas Tech University, Lubbock, TX 79409.
authors.forename: W David
authors.initials: WD
authors.lastname: Nes
mesh_terms: D000818:Animals; D002784:Cholesterol / Q000378:metabolism; D004875:Ergosterol / Q000378:metabolism*; D017964:Itraconazole / Q000494:pharmacology; D008297:Male; D008780:Methyltransferases / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism; D012313:RNA / Q000494:pharmacology; D014346:Trypanosoma brucei brucei / Q000187:drug effects / Q000378:metabolism*
chemical_list: D015800:Protozoan Proteins; D017964:Itraconazole; D012313:RNA; D002784:Cholesterol; D008780:Methyltransferases; D004875:Ergosterol
keywords: anti-parasite drugs; cholesterol; ergosterol biosynthesis; inhibitor; knockdown; ribonucleic acid interference; sparking function