A heat-activated and thermoresistant telomerase activity in Leishmania major Friedlin

Galindo, María Mercedes; Rodriguez, Evelyn J.; Rojas, M.G.; Figarella, Katherine; Campelo, Riward; Ramı́rez, José Luis

doi:10.1016/j.actatropica.2009.02.002

Cited by 5 publications

(6 citation statements)

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“…In their work, they noticed a low processivity for this enzyme compared with their Trypanosoma brucei and Leishmania tarentolae counterparts. In a more recent work, we showed that a high activity in L. major telomerase can be obtained when the assay conditions are adjusted, including specific extension primers and heating of the nuclear extracts prior to the assay reaction (9). The LmTERT subunit is encoded by a single gene located at chromosome 36 of this parasite (32); the size of its ORF corresponds to 1,451 amino acids (aa), which is larger than for the Trypanosoma brucei or Trypanosoma cruzi counterparts (approximately 1,198 aa) or the human protein (1,132 aa).…”

Section: Discussionmentioning

confidence: 99%

“…Electroblotting was done overnight at 4°C, and membranes were then exposed to the following primary antibodies for 1 h at room temperature (RT): (i) a polyclonal rabbit antibody produced against the L. major telomerase N-terminal domain including the GQ motif (anti-TEL-1) (see Fig. S1 in the supplemental material) at a 1:2,000 dilution (this antibody was purified by immunoabsorption using a recombinant L. major TERT protein fragment produced in E. coli cells as previously described [9]; anti-TEL-1 was able to immunoprecipitate a protein band with the expected molecular mass of L. major TERT [LmTERT] and with telomerase activity [see Fig. S2 in the supplemental material]), (ii) a monoclonal mouse antibody produced against human laminin B (Santa Cruz Biotechnology) tested at a 1:500 dilution (see Fig.…”

Section: Methodsmentioning

confidence: 99%

“…There have been subsequent reports for Leishmania amazonensis (8), L. major (9), Trypanosoma cruzi (10,11), and Trypanosoma brucei (7,12). Trypanosomatid TERT DNA sequences show identities of 51% within the Trypanosoma genus, 87% in species included in the Leishmania genus, and 33% between these two genera.…”

mentioning

confidence: 99%

“…In a previous study, we characterized molecularly and biochemically the telomerase of L. major promastigotes (9). During those experiments, we noted that in nuclear preparations, despite showing apparently good integrity, half of the total telomerase activity remained in the supernatant.…”

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confidence: 99%

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Leishmania major Telomerase TERT Protein Has a Nuclear/Mitochondrial Eclipsed Distribution That Is Affected by Oxidative Stress

et al. 2015

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In its canonical role the reverse transcriptase telomerase recovers the telomeric repeats that are lost during DNA replication. Other locations and activities have been recently described for the telomerase protein subunit TERT in mammalian cells. In the present work, using biochemistry, molecular biology, and electron microscopy techniques, we found that in the human parasite Leishmania major, TERT (and telomerase activity) shared locations between the nuclear, mitochondrial, and cytoplasmic compartments. Also, some telomerase activity and TERT protein could be found in ϳ100-nm nanovesicles. In the mitochondrial compartment, TERT appears to be mainly associated with the kinetoplast DNA. When Leishmania cells were exposed to H 2 O 2 , TERT changed its relative abundance and activity between the nuclear and mitochondrial compartments, with the majority of activity residing in the mitochondrion. Finally, overexpression of TERT in Leishmania transfected cells not only increased the parasitic cell growth rate but also increased their resistance to oxidative stress. Mammalian telomeres are nucleoprotein complexes with double and single DNA strand (telomeric overhang) runs of TTAGGG repeats, a chromatin organization containing methylated H3 and H4 histones, and a heavily methylated DNA, which are features of a transcriptionally repressed zone. At the ends of linear chromosomes, since DNA polymerases do not initiate DNA synthesis de novo, the new DNA synthesized over the lagging strand lacks the primer for the completion of its synthesis, leaving a gap; how to seal this gap has been named the end replication problem (1). Most eukaryotes solve the problem using mechanisms in which the enzyme telomerase plays an important role (2). Telomerase has been described for most eukaryotic cells, and in fact, the first report of telomerase was made for the protozoan Tetrahymena (2). The telomerase core has two components: a reverse transcriptase (TERT) protein and an RNA subunit (TER) that provides the template for copying the hexameric repeats to the telomeric overhang (2). In addition, mammalian telomeres contain a six-protein complex called shelterin that regulates the telomerase activity and protects the chromosomal ends from nucleolytic degradation and chromosomal rearrangements (3).The TERT component has the typical motifs of reverse transcriptase enzymes (1, 2, A, B=, C, and E) and a telomerase-specific T-domain (4), whereas the TER subunit has been sequenced in a large number of organisms, and the length of the RNA varies from ϳ200 to 1,300 nucleotides (nt) (5).Growing evidence indicates that TERT plays multiple roles in addition to its telomeric function, including (i) acting as a transcriptional modulator of the ␤-catenin signaling pathway in mice; (ii) interacting with an RNA component of mitochondrial RNA processing endoribonuclease (RMRP), creating a new activity that synthetizes RNA in a RNA-dependent manner and generating a double-stranded RNA (dsRNA) that enters into the RNA interference (RNAi) cascade; (iii) regu...

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Leishmania major Telomerase TERT Protein Has a Nuclear/Mitochondrial Eclipsed Distribution That Is Affected by Oxidative Stress

et al. 2015

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“…The telomerase component of Leishmania is mainly composed of telomerase RNA (TER) and telomerase reverse transcriptase (TERT) [24][25][26][27][28][29]. Leishmania TERT conserves all the canonical structural and functional domains found in most TERT but presents amino acid substitutions specific to the genus [24,26,[30][31][32][33].…”

Section: Introductionmentioning

confidence: 99%

Ablation of telomerase reverse transcriptase inLeishmania majorresults in a senescent-like phenotype and loss of infectivity

Shiburah,

de Oliveira,

Bisetegn

et al. 2023

Preprint

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The lack of efficient human vaccines and effective nontoxic drugs for leishmaniasis necessitates a search for new therapeutic targets. The telomere environment could provide potential targets against leishmaniasis. TERT, the telomerase reverse transcriptase component, has been on the radar for new therapeutic options against several diseases for more than two decades. In this study, we constructed a full deletion (LmTERT-/-) and an ORF disruption (LmN420) of the gene encoding the TERT component ofLeishmania major.LmTERT-/- andLmN420 parasites showed replicative and proliferative defects, growth impairment, cell cycle alterations, increased DNA damage, and progressive telomere shortening. Blockage of parasite altruism and the presence of autophagosomes characteristic of a senescent-like phenotype were also detected.LmTERT-/- andLmN420 parasites caused either micro lesion development or no visible lesions in mouse footpads and reduced infectivity in macrophages. While our checks to see if telomere erosion had reached theSCGgenes involved in lipophosphoglycan modification showed no changes, our proteomic assessment revealed a downregulation of a metacyclic-associated protein. Complementation of the knockout lineages using the WTLmTERT restored some of the lost phenotypes. Therefore, we speculate that the pleiotropic effects of the loss ofLmTERT advance the case for using it as a drug target against the parasite.

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Drug Targets, Drug Effectors, and Drug Targeting and Delivery

Loiseau

Barratt

2012

Drug Resistance in Leishmania Parasites

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A heat-activated and thermoresistant telomerase activity in Leishmania major Friedlin

Cited by 5 publications

References 10 publications

Leishmania major Telomerase TERT Protein Has a Nuclear/Mitochondrial Eclipsed Distribution That Is Affected by Oxidative Stress

Leishmania major Telomerase TERT Protein Has a Nuclear/Mitochondrial Eclipsed Distribution That Is Affected by Oxidative Stress

Ablation of telomerase reverse transcriptase inLeishmania majorresults in a senescent-like phenotype and loss of infectivity

Drug Targets, Drug Effectors, and Drug Targeting and Delivery

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