Analysis of genetic elements regulating the methionine adenosyltransferase gene in Leishmania infantum

Garcı́a-Estrada, Carlos; Pérez-Pertejo, Yolanda; Ordóñez, David; Balaña‐Fouce, Rafael; Reguera, Rosa M.

doi:10.1016/j.gene.2006.11.003

Cited by 7 publications

(5 citation statements)

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“…This locus contains two genes, each with two copies, LinJ.30.3550/70 and METK ( LinJ.30.3560/80 ). Arrangement, transcriptional regulation and expression have been studied for this gene cluster in L. infantum by García-Estrada et al (2003, 2007, 2008), establishing among other findings that both genes are expressed throughout the parasite's life cycle, with possible stage-specific post transcriptional regulation of their level. The METK gene encodes S-adenosylmethionine synthetase, a key enzyme in the synthesis of S-adenosylmethionine (AdoMet), a molecule involved in critical cellular functions such as trans -methylation, trans -sulphuration and polyamine synthesis in L. infantum (Reguera et al, 2002, 2007).…”

Section: Discussionmentioning

confidence: 96%

Resistance of Leishmania infantum to allopurinol is associated with chromosome and gene copy number variations including decrease in the S-adenosylmethionine synthetase (METK) gene copy number

Yasur-Landau

Jaffe

David

et al. 2018

International Journal for Parasitology: Drugs and Drug Resistan

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Leishmania infantum is one of the causative agents of visceral leishmaniasis (VL), a widespread, life-threatening disease. This parasite is responsible for the majority of human VL cases in Brazil, the Middle East, China, Central Asia and the Mediterranean basin. Its main reservoir are domestic dogs which, similar to human patients, may develop severe visceral disease and die if not treated. The drug allopurinol is used for the long-term maintenance of dogs with canine leishmaniasis. Following our report of allopurinol resistance in treated relapsed dogs, we investigated the mechanisms and markers of resistance to this drug. Whole genome sequencing (WGS) of clinical resistant and susceptible strains, and laboratory induced resistant parasites, was carried out in order to detect genetic changes associated with resistance. Significant gene copy number variation (CNV) was found between resistant and susceptible isolates at several loci, including a locus on chromosome 30 containing the genes LinJ.30.3550 through LinJ.30.3580. A reduction in copy number for LinJ.30.3560, encoding the S-adenosylmethionine synthetase (METK) gene, was found in two resistant clinical isolates and four induced resistant clonal strains. Using quantitative real time PCR, this reduction in METK copy number was also found in three additional resistant clinical isolates. Furthermore, inhibition of S-adenosylmethionine synthetase encoded by the METK gene in allopurinol susceptible strains resulted in increased allopurinol resistance, confirming its role in resistance to allopurinol. In conclusion, this study identified genetic changes associated with L. infantum resistance to allopurinol and the reduction in METK copy number identified may serve as a marker for resistance in dogs, and reduced protein activity correlated with increased allopurinol resistance.

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Section: Discussionmentioning

confidence: 96%

Resistance of Leishmania infantum to allopurinol is associated with chromosome and gene copy number variations including decrease in the S-adenosylmethionine synthetase (METK) gene copy number

Yasur-Landau

Jaffe

David

et al. 2018

International Journal for Parasitology: Drugs and Drug Resistan

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“…Amongst the Leishmania, Leishmania infantum, a trypanosomatid protozoan parasite, has two MAT genes in a tandem array. Two AG dinucleotide trans-splicing sites are located in its 5!-UTR, and the +420/+905 region of the 3´-UTR contains cis-regulatory elements [144]. These genes are expressed as a single transcript [145]; the resultant protein forms a dimer.…”

Section: Mat In Parasitesmentioning

confidence: 99%

Methionine Adenosyltransferase (S‐Adenosylmethionine Synthetase)

Pajares

Markham

2011

Advances in Enzymology - And Related Areas of Molecular Biology

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10.3. Thermal Unfolding 10.4. Association of MAT III to form MAT I: Role of the Disulfide C35-C61 10.5. Complementary Folding Data Obtained in MAT Proteins of Different Origins 11. MAT Inhibitors 12. Role of MAT in Diseases 12.1. Cognitive and Neurodegenerative Diseases 12.2. Human Mutations in MAT1A Gene: Hypermethioninemia and Demyelination 12.3. MAT and Cancer 12.4. MAT and Ethanol 2. MAMMALIAN MATs Early studies on mammalian MATs showed the existence of this activity in all the tissues studied, the highest being observed in liver. However, complex kinetics were obtained which made the results difficult to interpret until the presence of several isoenzymes was realized. Separations carried out on phenyl Sepharose columns showed elution of three peaks with different hydrophobic character; these were named according to the order of elution as MAT I, II and III. All these isoenzymes share some characteristics such as the Mg 2+ dependence of their 9/14/2012 Page 6 of 80 activity (S 0.5 !1 mM), stimulation by K + ions, and their AdoMet-inducible tripolyphosphatase activity [4, 5]. 2.1. MAT I/III isoenzymes Since their discovery, these isoenzymes were known as liver-specific forms of MAT, until Lu et al. in 2003 [6] showed their presence also in pancreas. Several laboratories have reported in depth studies of MAT I and III which have been carried out using adult liver. The differences can be summarized as follows: i) MAT I is a tetramer whereas MAT III is a dimer, with respective Mr appearing as 200 and 110 kDa upon gel filtration chromatography, ii) their Km values for methionine are in the micromolar range for MAT I (60-120 µM) and in the millimolar range for MAT III (1 mM) [7, 8]; iii) MAT III can be activated by DMSO at physiological concentrations of methionine (60 µM) and is highly hydrophobic (eluting from phenyl Sepharose columns at 50% DMSO (v/v)); and, iv) AdoMet inhibits MAT I and activates MAT III. Some of these differences are normally used to distinguish between isoenzymes in activity assays. Both purified proteins show a common single band (designated "1) of approximately 48 kDa on SDS-PAGE, and antibodies raised against the dimer form recognized both, thus suggesting that a single type of subunit arranged in two assemblies could explain the presence of both oligomers [7]. Further experiments that supported this hypothesis included peptide mapping of the purified isoenzymes [7], dissociation of the tetramer to the dimer using LiBr [9] or N-ethylmaleimide (NEM) modification [10]. The final confirmation came upon cloning of a single cDNA [11, 12] followed by overexpresion in E. coli cells [13], which showed in vivo production of dimers and tetramers. This single subunit type is a product of the MAT1A gene that contains nine exons and eight introns spanning !20 kb [14], and has been localized by fluorescence in situ hybridization to the human chromosome 10q.22 [15]. Animal models, as well as the analysis of human samples, led to the identification of changes in the MAT I/III ratio in pathological conditi...

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“…MAT2 genes are arranged in tandem and alternate with another gene, named LORIEN , which encodes a protein of unknown function with a SP-RING/Miz zinc-finger motif. The two copies of the LORIEN gene are located 3250-bp upstream of each MAT2 gene copy (García-Estrada et al 2007). Genomic studies have shown that these two genes are transcribed into large polycistronic nascent RNAs, which are post-transcriptionally processed into monocistronic mRNAs by sequential addition of a 39-nt spliced leader at the 5′-end and a poly(A) tail at the 3′-end (García-Estrada et al .…”

Section: Introductionmentioning

confidence: 99%

Leishmania donovani: proteasome-mediated down-regulation of methionine adenosyltransferase

et al. 2011

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Methionine adenosyltransferase (MAT) is an important enzyme for metabolic processes, to the extent that its product, S-adenosylmethionine (AdoMet), plays a key role in trans-methylation, trans-sulphuration and polyamine synthesis. Previous studies have shown that a MAT-overexpressing strain of Leishmania donovani controls AdoMet production, keeping the intracellular AdoMet concentration at levels that are compatible with cell survival. This unexpected result, together with the fact that MAT activity and abundance changed with time in culture, suggests that different regulatory mechanisms acting beyond the post-transcriptional level are controlling this protein. In order to gain an insight into these mechanisms, several experiments were carried out to explain the MAT abundance during promastigote cell growth. Determination of MAT turnover in cycloheximide (CHX)-treated cultures resulted in a surprising 5-fold increase in MAT turnover compared to CHX-untreated cultures. This increase agrees with a stabilization of the MAT protein, whose integrity was maintained during culture. The presence of proteasome inhibitors, namely MG-132, MG-115, epoxomycin and lactacystin in the culture medium prevented MAT degradation in both MAT-overexpressing and 'mock-transfected' leishmanial strains. The role of the ubiquitin (Ub) pathway in MAT down-regulation was supported using immunoprecipitation experiments. Immunoprecipitated MAT cross-reacted with anti-Ub antibodies, which provides evidence of a proteasome-mediated down-regulation of the leishmanial MAT abundance.

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Analysis of genetic elements regulating the methionine adenosyltransferase gene in Leishmania infantum

Cited by 7 publications

References 46 publications

Resistance of Leishmania infantum to allopurinol is associated with chromosome and gene copy number variations including decrease in the S-adenosylmethionine synthetase (METK) gene copy number

Resistance of Leishmania infantum to allopurinol is associated with chromosome and gene copy number variations including decrease in the S-adenosylmethionine synthetase (METK) gene copy number

Methionine Adenosyltransferase (S‐Adenosylmethionine Synthetase)

Leishmania donovani: proteasome-mediated down-regulation of methionine adenosyltransferase

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