The Trypanosoma brucei TbHrg protein is a heme transporter involved in the regulation of stage-specific morphological transitions

Abstract

The human parasite does not synthesize heme and instead relies entirely on heme supplied by its vertebrate host or its insect vector, the tsetse fly. In the host bloodstream scavenges heme via haptoglobin-hemoglobin (HpHb) receptor-mediated endocytosis occurring in the flagellar pocket. However, in the procyclic developmental stage, in which is confined to the tsetse fly midgut, this receptor is apparently not expressed, suggesting that takes up heme by a different, unknown route. To define this alternative route, we functionally characterized heme transporterHrg in the procyclic stage. RNAi-induced down-regulation of Hrg in heme-limited culture conditions resulted in slower proliferation, decreased cellular heme, and marked changes in cellular morphology so that the cells resemble mesocyclic trypomastigotes. Nevertheless, theHrg KO developed normally in the tsetse flies at rates comparable with wild-type cells. cells overexpressingHrg displayed up-regulation of the early procyclin GPEET and down-regulation of the late procyclin EP1, two proteins coating the surface in the procyclic stage. Light microscopy of immunostainedHrg indicated localization to the flagellar membrane, and scanning electron microscopy revealed more intense Hrg accumulation toward the flagellar pocket. Based on these findings, we postulate that senses heme levels via the flagellar Hrg protein. Heme deprivation in the tsetse fly anterior midgut might represent an environmental stimulus involved in the transformation of this important human parasite, possibly through metabolic remodeling.

Trypanosoma cruzisenses intracellular heme levels and regulates heme homeostasis by modulatingTcHTE protein expression

“…Summarizing these results, it is reasonable to postulate that TcHTE is localized in the flagellar pocket region where it facilitates heme transport in T. cruzi. Interestingly, we did not detect rTcHTE.His-GFP in internal vesicles as it was reported by the recombinant versions of Leishmania HRG protein (LaHR1 or LmHR1) (12,16) and T. brucei, TbHRG (16) or in the flagellar membrane of procyclic stage of T. brucei (17).…”

“…During the last years, several proteins involved in heme transport have been described in trypanosomatids, such as LHR1 in Leishmania spp. (12)(13)(14), TbHRG in Trypanosoma brucei (16,17), and TcHTE in Trypanosoma cruzi (15). And most recently a protein identified in L. major, named LmFLVCRb and member of a Major Facilitator Superfamily, was described as a heme importer in this organism (23).…”

“…This observation corroborates that TcHTE mRNA respond to heme as well as LHR1 (12), confirming TcHTE as a member of the Heme Response Gene (HRG) family (11). It is important to note that Leishmania (12) and T. cruzi HRGs respond to heme but not TbHRG (17), suggesting that those gene products might be under different regulations and/or play different roles in heme transport.…”

“…We described a homologous protein in T. cruzi, named TcHTE (T. cruzi Heme Transport Enhancer), that plays a critical role in heme transport and is found mainly in the flagellar pocket of T. cruzi (15). More recently, TbHRG (T. brucei Heme Responsive Gene) was described in T. brucei and postulated to be involved in heme transport as free-heme or in the salvage of heme from hemoglobin degradation (16,17). LHR1, TcHTE and TbHRG present similar growth performance when expressed in Saccharomyces cerevisiae hem1D, allowing this knock out cells to growth in a medium supplemented with low heme.…”

Modulation of mitochondrial metabolism as a biochemical trait in blood feeding organisms: the redox vampire hypothesis redux

“…While iron transport in bloodstream forms is mediated by endocytosis of transferrin/lactoferrin receptors (Steverding, ; Tanaka et al, ), transferrin‐independent reductive iron transport take place in procyclic forms (Mach et al, ). Heme transport in T. brucei is mediated by either endocytosis of haptoglobin‐hemoglobin receptor (HpHbR) in bloodstream forms (Vanhollebeke et al, ), or through the alternative transporter TbHRG in procyclic forms (Horáková et al, ), which do not express HpHbR. Regardless the transport mechanism, heme is eventually incorporated in parasite hemeproteins, mostly the cytochromes of mitochondrial ETS.…”