Kinetic study of the plasma-membrane potential in procyclic and bloodstream forms of <i>Trypanosoma brucei brucei</i> using the fluorescent probe bisoxonol

Defrise-Quertain, Fabienne; Fraser-L’Hostis, Chantal; Coral, Danièle; Deshusses, J.

doi:10.1042/bj3140595

Cited by 19 publications

(15 citation statements)

References 33 publications

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“…1, pink curves). The effect of exogenous K ϩ on ⌿ p in T. brucei has been controversial (11,37). Our results indicate that in T. brucei, bloodstream as well as procyclic forms, the plasma membrane is not immediately penetrable for K ϩ .…”

Section: Phenotypic Characterization Of Kmentioning

confidence: 60%

A Trk/HKT-Type K ⁺ Transporter from Trypanosoma brucei

et al. 2010

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The molecular mechanisms of K ؉ homeostasis are only poorly understood for protozoan parasites. Trypanosoma brucei subsp. parasites, the causative agents of human sleeping sickness and nagana, are strictly extracellular and need to actively concentrate K ؉ from their hosts' body fluids. The T. brucei genome contains two putative K ؉ channel genes, yet the trypanosomes are insensitive to K ؉ antagonists and K ؉ channelblocking agents, and they do not spontaneously depolarize in response to high extracellular K ؉ concentrations. However, the trypanosomes are extremely sensitive to K ؉ ionophores such as valinomycin. Surprisingly, T. brucei possesses a member of the Trk/HKT superfamily of monovalent cation permeases which so far had only been known from bacteria, archaea, fungi, and plants. The protein was named TbHKT1 and functions as a Na ؉ -independent K ؉ transporter when expressed in Escherichia coli, Saccharomyces cerevisiae, or Xenopus laevis oocytes. In trypanosomes, TbHKT1 is expressed in both the mammalian bloodstream stage and the Tsetse fly midgut stage; however, RNA interference (RNAi)-mediated silencing of TbHKT1 expression did not produce a growth phenotype in either stage. The presence of HKT genes in trypanosomatids adds a further piece to the enigmatic phylogeny of the Trk/HKT superfamily of K ؉ transporters. Parsimonial analysis suggests that the transporters were present in the first eukaryotes but subsequently lost in several of the major eukaryotic lineages, in at least four independent events.

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Section: Phenotypic Characterization Of Kmentioning

confidence: 60%

A Trk/HKT-Type K ⁺ Transporter from Trypanosoma brucei

et al. 2010

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“…A multifunctional intracellular ATPase in trypanosomes, TbVCP, is sensitive to NEM inhibition (40). In bloodstream form T. brucei brucei, NEM caused depolarization of the plasma membrane potential, presumably through inhibition of H ϩ -ATPases (41). In addition to ATPases, phosphatases often contain redox-active cysteine residues that are sensitive to oxidation (42).…”

Section: Discussionmentioning

confidence: 99%

Trypanosome Lytic Factor-1 Initiates Oxidation-stimulated Osmotic Lysis of Trypanosoma brucei brucei

Greene

Hajduk

2016

Journal of Biological Chemistry

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Human innate immunity against the veterinary pathogenTrypanosoma brucei brucei is conferred by trypanosome lytic factors (TLFs), against which human-infective T. brucei gambiense and T. brucei rhodesiense have evolved resistance. TLF-1 is a subclass of high density lipoprotein particles defined by two primate-specific apolipoproteins: the ion channel-forming toxin ApoL1 (apolipoprotein L1) and the hemoglobin (Hb) scavenger Hpr (haptoglobin-related protein). The role of oxidative stress in the TLF-1 lytic mechanism has been controversial. Here we show that oxidative processes are involved in TLF-1 killing of T. brucei brucei. The lipophilic antioxidant N,N-diphenyl-p-phenylenediamine protected TLF-1-treated T. brucei brucei from lysis. Conversely, lysis of TLF-1-treated T. brucei brucei was increased by the addition of peroxides or thiol-conjugating agents. Previously, the Hpr-Hb complex was postulated to be a source of free radicals during TLF-1 lysis. However, we found that the iron-containing heme of the Hpr-Hb complex was not involved in TLF-1 lysis. Furthermore, neither high concentrations of transferrin nor knock-out of cytosolic lipid peroxidases prevented TLF-1 lysis. Instead, purified ApoL1 was sufficient to induce lysis, and ApoL1 lysis was inhibited by the antioxidant DPPD. Swelling of TLF-1-treated T. brucei brucei was reminiscent of swelling under hypotonic stress. Moreover, TLF-1-treated T. brucei brucei became rapidly susceptible to hypotonic lysis. T. brucei brucei cells exposed to peroxides or thiol-binding agents were also sensitized to hypotonic lysis in the absence of TLF-1. We postulate that ApoL1 initiates osmotic stress at the plasma membrane, which sensitizes T. brucei brucei to oxidation-stimulated osmotic lysis.

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“…These two properties make recording the membrane voltage with microelectrodes difficult. As an alternative strategy we monitored the membrane voltage of the host cells with the voltage‐sensitive fluorescent dye bisoxonol [14,15]. A cell suspension was incubated in a solution of 1 μM bisoxonol.…”

Section: Pbcv‐1 Infection May Induce Membrane Depolarization In Host mentioning

confidence: 99%

Possible function for virus encoded K⁺ channel Kcv in the replication of chlorella virus PBCV‐1

et al. 2003

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The K + channel Kcv is encoded by the chlorella virus PBCV-1. There is evidence that this channel plays an essential role in the replication of the virus, because both PBCV-1 plaque formation and Kcv channel activity in Xenopus oocytes have similar sensitivities to inhibitors. Here we report circumstantial evidence that the Kcv channel is important during virus infection. Recordings of membrane voltage in the host cells Chlorella NC64A reveal a membrane depolarization within the ¢rst few minutes of infection. This depolarization displays the same sensitivity to cations as Kcv conductance; depolarization also requires the intact membrane of the virion. Together these data are consistent with the idea that the virus carries functional K + channels in the virion and inserts them into the host cell plasma membrane during infection.

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Kinetic study of the plasma-membrane potential in procyclic and bloodstream forms of Trypanosoma brucei brucei using the fluorescent probe bisoxonol

Cited by 19 publications

References 33 publications

A Trk/HKT-Type K ⁺ Transporter from Trypanosoma brucei

A Trk/HKT-Type K ⁺ Transporter from Trypanosoma brucei

Trypanosome Lytic Factor-1 Initiates Oxidation-stimulated Osmotic Lysis of Trypanosoma brucei brucei

Possible function for virus encoded K⁺ channel Kcv in the replication of chlorella virus PBCV‐1

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Kinetic study of the plasma-membrane potential in procyclic and bloodstream forms of Trypanosoma brucei brucei using the fluorescent probe bisoxonol

Cited by 19 publications

References 33 publications

A Trk/HKT-Type K + Transporter from Trypanosoma brucei

A Trk/HKT-Type K + Transporter from Trypanosoma brucei

Trypanosome Lytic Factor-1 Initiates Oxidation-stimulated Osmotic Lysis of Trypanosoma brucei brucei

Possible function for virus encoded K+ channel Kcv in the replication of chlorella virus PBCV‐1

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A Trk/HKT-Type K ⁺ Transporter from Trypanosoma brucei

A Trk/HKT-Type K ⁺ Transporter from Trypanosoma brucei

Possible function for virus encoded K⁺ channel Kcv in the replication of chlorella virus PBCV‐1