Allosteric regulation of an essential trypanosome polyamine biosynthetic enzyme by a catalytically dead homolog

Willert, Erin K.; Fitzpatrick, Richard; Phillips, Margaret A.

doi:10.1073/pnas.0701111104

Cited by 81 publications

(121 citation statements)

References 45 publications

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“…Instead, we recently discovered a unique mechanism for the regulation of AdoMetDC in trypanosomatids. T. brucei AdoMetDC is activated 1,200-fold by forming a heterodimer with an inactive homolog termed prozyme, which is found only in the trypanosomatids (39). Gene silencing by RNAi or the chemical inhibition of AdoMetDC led to 25-fold and 7-fold increases in the protein levels of prozyme and ODC, respectively, with the data suggesting that the expression of these enzymes is translationally regulated (40).…”

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confidence: 79%

RNA Interference-Mediated Silencing of Ornithine Decarboxylase and Spermidine Synthase Genes in Trypanosoma brucei Provides Insight into Regulation of Polyamine Biosynthesis

2009

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Polyamine biosynthesis is a drug target for the treatment of African sleeping sickness; however, mechanisms regulating the pathway in Trypanosoma brucei are not well understood. Recently, we showed that RNA interference (RNAi)-mediated gene silencing or the inhibition of S-adenosylmethionine decarboxylase (AdoMetDC) led to the upregulation of the AdoMetDC activator, prozyme, and ornithine decarboxylase (ODC) proteins. To determine if this regulatory response is specific to AdoMetDC, we studied the effects of the RNAi-induced silencing of the spermidine synthase (SpdSyn) and ODC genes in bloodstream form T. brucei. The knockdown of either gene product led to the depletion of the polyamine and trypanothione pools and to cell death. Decarboxylated AdoMet levels were elevated, while AdoMet was not affected. There was no significant effect on the protein levels of other polyamine pathway enzymes. The treatment of parasites with the ODC inhibitor ␣-difluoromethylornithine gave similar results to those observed for ODC knockdown. Thus, the cellular response to the loss of AdoMetDC activity is distinctive, suggesting that AdoMetDC activity controls the expression levels of the other spermidine biosynthetic enzymes. RNAi-mediated cell death occurred more rapidly for ODC than for SpdSyn. Further, the ODC RNAi cells were rescued by putrescine, but not spermidine, suggesting that the depletion of both putrescine and spermidine is more detrimental than the depletion of spermidine alone. This finding may contribute to the effectiveness of ODC as a target for the treatment of African sleeping sickness, thus providing important insight into the mechanism of action of a key antitrypanosomal agent.

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confidence: 79%

RNA Interference-Mediated Silencing of Ornithine Decarboxylase and Spermidine Synthase Genes in Trypanosoma brucei Provides Insight into Regulation of Polyamine Biosynthesis

2009

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“…Furthermore, the kinetoplastids and Entamoeba encode two DHS genes, and it was shown for T. brucei that the two gene products (one catalytically active but impaired and the other catalytically inactive) associate to form a heterotetrameric enzyme, which is the functionally active form of the enzyme (26). This novel mechanism of enzyme activation by a catalytically inactive paralog was also observed for T. brucei S-adenosylmethionine decarboxylase, which is required for formation of the DHS substrate spermidine (28,29). The hydroxylase reaction is not found in all eukaryotes; however, the kinetoplastids do contain a homolog of DOHH, and it has recently been shown to be a functional enzyme in Leishmania (30).…”

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confidence: 86%

Deoxyhypusine Modification of Eukaryotic Translation Initiation Factor 5A (eIF5A) Is Essential for Trypanosoma brucei Growth and for Expression of Polyprolyl-containing Proteins

Nguyen

Leija

Kinch

et al. 2015

Journal of Biological Chemistry

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Background: eIF5A facilitates translation of mRNAs encoding proteins with polyprolyl repeats. Results: The protozoan parasite Trypanosoma brucei contains polyprolyl tracts in 15% of its proteome, and both eIF5A and its post-translational modification with deoxyhypusine are essential. Conclusion: Loss of eIF5A leads to abnormal cell morphology, abnormal cell division, and decreased flagellar attachment. Significance: Inhibitors of hypusine biosynthesis would disrupt multiple essential cellular processes.

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“…In eukaryotes, the triamine spermidine is synthesized from the precursor diamine putrescine by the addition of an aminopropyl group donated by decarboxylated S-adenosylmethionine. The enzymes involved in eukaryotic spermidine biosynthesis, S-adenosylmethionine decarboxylase and spermidine synthase, are also present in many bacteria and archaea and have been extensively characterized (13)(14)(15)(16)(17)(18)(19). The decarboxylated S-adenosylmethionine pathway is also involved in norspermidine and norspermine biosynthesis in thermophilic Crenarchaeota and Clostridia (20,21) and probably in other bacterial species.…”

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Evolution and Multifarious Horizontal Transfer of an Alternative Biosynthetic Pathway for the Alternative Polyamine sym-Homospermidine

Shaw

Elliott

Kinch

et al. 2010

Journal of Biological Chemistry

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Polyamines are small flexible organic polycations found in almost all cells. They likely existed in the last universal common ancestor of all extant life, and yet relatively little is understood about their biological function, especially in bacteria and archaea. Unlike eukaryotes, where the predominant polyamine is spermidine, bacteria may contain instead an alternative polyamine, sym-homospermidine. We demonstrate that homospermidine synthase (HSS) has evolved vertically, primarily in the ␣-Proteobacteria, but enzymatically active, diverse HSS orthologues have spread by horizontal gene transfer to other bacteria, bacteriophage, archaea, eukaryotes, and viruses. By expressing diverse HSS orthologues in Escherichia coli, we demonstrate in vivo the production of co-products diaminopropane and N 1 -aminobutylcadaverine, in addition to sym-homospermidine. We show that sym-homospermidine is required for normal growth of the ␣-proteobacterium Rhizobium leguminosarum. However, sym-homospermidine can be replaced, for growth restoration, by the structural analogues spermidine and sym-norspermidine, suggesting that the symmetrical or unsymmetrical form and carbon backbone length are not critical for polyamine function in growth. We found that the HSS enzyme evolved from the alternative spermidine biosynthetic pathway enzyme carboxyspermidine dehydrogenase. The structure of HSS is related to lysine metabolic enzymes, and HSS and carboxyspermidine dehydrogenase evolved from the aspartate family of pathways. Finally, we show that other bacterial phyla such as Cyanobacteria and some ␣-Proteobacteria synthesize sym-homospermidine by an HSS-independent pathway, very probably based on deoxyhypusine synthase orthologues, similar to the alternative homospermidine synthase found in some plants. Thus, bacteria can contain alternative biosynthetic pathways for both spermidine and sym-norspermidine and distinct alternative pathways for sym-homospermidine.Polyamines are primordial, small flexible organic polycations found in almost all cells of bacteria, archaea, and eukaryotes (1). In bacteria and archaea, the key polyamines (see Fig. 1A) are the triamines spermidine, sym-norspermidine, and sym-homospermidine (referred to herein as norspermidine and homospermidine), and occasionally more than one triamine can be found in the same cell. In eukaryotes, which contain spermidine (and in some plants, yeasts, and animals, the tetraamine spermine), polyamines are required for growth, cell proliferation, and normal cellular physiology. Polyamine biosynthesis is essential in the fungi Saccharomyces cerevisiae (2), Schizosaccharomyces pombe (3), Aspergillus nidulans (4), and Ustilago maydis (5), the kinetoplastid parasites Trypanosoma brucei (6) and Leishmania donovani (7), and the diplomonad parasite Giardia lamblia (8). In mouse, polyamines are essential for early embryo development (9, 10), and they are also essential for seed development in the flowering plant Arabidopsis thaliana (11).The universal distribution of polyamines suggests that...

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Allosteric regulation of an essential trypanosome polyamine biosynthetic enzyme by a catalytically dead homolog

Cited by 81 publications

References 45 publications

RNA Interference-Mediated Silencing of Ornithine Decarboxylase and Spermidine Synthase Genes in Trypanosoma brucei Provides Insight into Regulation of Polyamine Biosynthesis

RNA Interference-Mediated Silencing of Ornithine Decarboxylase and Spermidine Synthase Genes in Trypanosoma brucei Provides Insight into Regulation of Polyamine Biosynthesis

Deoxyhypusine Modification of Eukaryotic Translation Initiation Factor 5A (eIF5A) Is Essential for Trypanosoma brucei Growth and for Expression of Polyprolyl-containing Proteins

Evolution and Multifarious Horizontal Transfer of an Alternative Biosynthetic Pathway for the Alternative Polyamine sym-Homospermidine

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