Identification of a post-transcriptional regulatory element that responds to glucose in the African trypanosome

Qiu, Yefeng; Shankar, Vijay; Noorai, Rooksana E.; Yeung, Nelson; Sg, Mcalpine; Morris, James C.

doi:10.1101/327346

“…Its co-regulation with Hexokinase and THT1 hexose transporter in TfR-depleted cells suggests it might also have broader roles in general nutrient sensing. For example, it has been reported to be upregulated in glucose-starved cells (65), a finding that we have reproduced experimentally (C. Tiengwe, unpublished). Identifying its mRNA targets globally and molecular partners will be important for revealing its cellular dynamics.…”

Section: Iron Homeostasis In African Trypanosomes 18supporting

confidence: 78%

Novel aspects of iron homeostasis in pathogenic bloodstream form Trypanosoma brucei

Carbajo

¹

,

Cornell

²

,

Madbouly

³

et al. 2021

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Iron is an essential regulatory signal for virulence factors in many pathogens. Mammals and bloodstream form (BSF) Trypanosoma brucei obtain iron by receptor-mediated endocytosis of transferrin bound to receptors (TfR) but the mechanisms by which T. brucei subsequently handles iron remains enigmatic. Here, we analyse the transcriptome of T. brucei cultured in iron-rich and iron-poor conditions. We show that adaptation to iron-deprivation induces upregulation of TfR, a cohort of parasite-specific genes (ESAG3, PAGS), genes involved in glucose uptake and glycolysis (THT1 and hexokinase), endocytosis (Phosphatidic Acid Phosphatase, PAP2), and most notably a divergent RNA binding protein RBP5, indicative of a non-canonical mechanism for regulating intracellular iron levels. We show that cells depleted of TfR by RNA silencing import free iron as a compensatory survival strategy. The TfR and RBP5 iron response are reversible by genetic complementation, the response kinetics are similar, but the regulatory mechanisms are distinct. Increased TfR protein is due to increased mRNA. Increased RBP5 expression, however, occurs by a post-transcriptional feedback mechanism whereby RBP5 interacts with its own, and with PAP2 mRNAs. Further observations suggest that increased RBP5 expression in iron-deprived cells has a maximum threshold as ectopic overexpression above this threshold disrupts normal cell cycle progression resulting in an accumulation of anucleate cells and cells in G2/M phase. This phenotype is not observed with overexpression of RPB5 containing a point mutation (F61A) in its single RNA Recognition Motif. Our experiments shed new light on how T. brucei BSFs reorganise their transcriptome to deal with iron stress revealing the first iron responsive RNA binding protein that is co-regulated with TfR, is important for cell viability and iron homeostasis; two essential processes for successful proliferation.

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“…Its co-regulation with Hexokinase and THT1 hexose transporter in TfR-depleted cells suggests it might also have broader roles in general nutrient sensing. For example, it has been reported to be upregulated in glucose-starved cells [ 63 ]. Identifying its mRNA targets globally and molecular partners will be important for revealing its cellular dynamics.…”

Section: Discussionmentioning

confidence: 99%

Novel aspects of iron homeostasis in pathogenic bloodstream form Trypanosoma brucei

Carbajo

¹

,

Cornell

²

,

Madbouly

³

et al. 2021

View full text Add to dashboard Cite

Iron is an essential regulatory signal for virulence factors in many pathogens. Mammals and bloodstream form (BSF) Trypanosoma brucei obtain iron by receptor-mediated endocytosis of transferrin bound to receptors (TfR) but the mechanisms by which T. brucei subsequently handles iron remains enigmatic. Here, we analyse the transcriptome of T. brucei cultured in iron-rich and iron-poor conditions. We show that adaptation to iron-deprivation induces upregulation of TfR, a cohort of parasite-specific genes (ESAG3, PAGS), genes involved in glucose uptake and glycolysis (THT1 and hexokinase), endocytosis (Phosphatidic Acid Phosphatase, PAP2), and most notably a divergent RNA binding protein RBP5, indicative of a non-canonical mechanism for regulating intracellular iron levels. We show that cells depleted of TfR by RNA silencing import free iron as a compensatory survival strategy. The TfR and RBP5 iron response are reversible by genetic complementation, the response kinetics are similar, but the regulatory mechanisms are distinct. Increased TfR protein is due to increased mRNA. Increased RBP5 expression, however, occurs by a post-transcriptional feedback mechanism whereby RBP5 interacts with its own, and with PAP2 mRNAs. Further observations suggest that increased RBP5 expression in iron-deprived cells has a maximum threshold as ectopic overexpression above this threshold disrupts normal cell cycle progression resulting in an accumulation of anucleate cells and cells in G2/M phase. This phenotype is not observed with overexpression of RPB5 containing a point mutation (F61A) in its single RNA Recognition Motif. Our experiments shed new light on how T. brucei BSFs reorganise their transcriptome to deal with iron stress revealing the first iron responsive RNA binding protein that is co-regulated with TfR, is important for cell viability and iron homeostasis; two essential processes for successful proliferation.

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“…mRNA translation is then regulated by modifying their stability, localization and translationality. For that reason, RNA Binding Proteins (RBPs) and RNA regulons have shown to have essential roles in different cellular processes in these organisms, such as differentiation and infectivity [ 62 , 63 ], and responses to nutrient and oxygen availability [ 64 , 65 ]. One of our hypotheses regarding new roles for the AMPK complexes in trypanosomatids involves its interaction with the RBPs and influencing regulon activation.…”

Section: Discussionmentioning

confidence: 99%

An AMP-activated protein kinase complex with two distinctive alpha subunits is involved in nutritional stress responses in Trypanosoma cruzi

Sternlieb¹,

Schoijet

²

,

Genta³

et al. 2021

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Trypanosoma cruzi, the etiological agent of Chagas disease, has a digenetic life cycle. In its passage from the insect vector to the mammalian host, and vice versa, it must be prepared to cope with abrupt changes in environmental conditions, such as carbon source, pH, temperature and osmolarity, in order to survive. Sensing and signaling pathways that allow the parasite to adapt, have unique characteristics with respect to their hosts and other free-living organisms. Many of the canonical proteins involved in these transduction pathways have not yet been found in the genomes of these parasites because they present divergences either at the functional, structural and/or protein sequence level. All of this makes these pathways promising targets for therapeutic drugs. The AMP-activated protein kinase (AMPK) is a serine/threonine kinase activated by environmental stresses such as osmotic stress, hypoxia, ischaemia and exercise that results in reduction of ATP and increase of AMP levels. Thus, AMPK is regarded as a fuel gauge, functioning both as a nutrient and an energy sensor, to maintain energy homeostasis and, eventually, to protect cells from death by nutrient starvation. In the present study we report the characterization of AMPK complexes for the first time in T. cruzi and propose the function of TcAMPK as a novel regulator of nutritional stress in epimastigote forms. We show that there is phosphotransferase activity specific for SAMS peptide in epimastigotes extracts, which is inhibited by Compound C and is modulated by carbon source availability. In addition, TcAMPKα2 subunit has an unprecedented functional substitution (Ser x Thr) at the activation loop and its overexpression in epimastigotes led to higher autophagic activity during prolonged nutritional stress. Moreover, the over-expression of the catalytic subunits resulted in antagonistic phenotypes associated with proliferation. Together, these results point to a role of TcAMPK in autophagy and nutrient sensing, key processes for the survival of trypanosomatids and for its life cycle progression.

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Identification of a post-transcriptional regulatory element that responds to glucose in the African trypanosome

Cited by 4 publications

References 71 publications

Novel aspects of iron homeostasis in pathogenic bloodstream form Trypanosoma brucei

Novel aspects of iron homeostasis in pathogenic bloodstream form Trypanosoma brucei

Novel aspects of iron homeostasis in pathogenic bloodstream form Trypanosoma brucei

An AMP-activated protein kinase complex with two distinctive alpha subunits is involved in nutritional stress responses in Trypanosoma cruzi

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