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Kinetoplastids

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https://www.readbyqxmd.com/read/28631386/toward-establishing-model-organisms-for-marine-protists-successful-transfection-protocols-for-parabodo-caudatus-kinetoplastida-excavata
#1
Fatma Gomaa, Paulo A Garcia, Jennifer Delaney, Peter R Girguis, Cullen R Buie, Virginia P Edgcomb
We developed protocols for, and demonstrated successful transfection of, the free-living kinetoplastid flagellate Parabodo caudatus with three plasmids carrying a fluorescence reporter gene (pEF-GFP with the EF1 alpha promoter, pUB-GFP with Ubiquitin C promoter, and pEYFP-Mitotrap with CMV promoter). We evaluated three electroporation approaches: 1) a square-wave electroporator designed for eukaryotes, 2) a novel microfluidic transfection system employing hydrodynamically-controlled electric field waveforms, and 3) a traditional exponential decay electroporator...
June 19, 2017: Environmental Microbiology
https://www.readbyqxmd.com/read/28623350/multiplexed-spliced-leader-sequencing-a-high-throughput-selective-method-for-rna-seq-in-trypanosomatids
#2
Bart Cuypers, Malgorzata A Domagalska, Pieter Meysman, Géraldine de Muylder, Manu Vanaerschot, Hideo Imamura, Franck Dumetz, Thomas Wolf Verdonckt, Peter J Myler, Gowthaman Ramasamy, Kris Laukens, Jean-Claude Dujardin
High throughput sequencing techniques are poorly adapted for in vivo studies of parasites, which require prior in vitro culturing and purification. Trypanosomatids, a group of kinetoplastid protozoans, possess a distinctive feature in their transcriptional mechanism whereby a specific Spliced Leader (SL) sequence is added to the 5'end of each mRNA by trans-splicing. This allows to discriminate Trypansomatid RNA from mammalian RNA and forms the basis of our new multiplexed protocol for high-throughput, selective RNA-sequencing called SL-seq...
June 16, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28573017/a-crispr-cas9-high-throughput-genome-editing-toolkit-for-kinetoplastids
#3
Tom Beneke, Ross Madden, Laura Makin, Jessica Valli, Jack Sunter, Eva Gluenz
Clustered regularly interspaced short palindromic repeats (CRISPR), CRISPR-associated gene 9 (Cas9) genome editing is set to revolutionize genetic manipulation of pathogens, including kinetoplastids. CRISPR technology provides the opportunity to develop scalable methods for high-throughput production of mutant phenotypes. Here, we report development of a CRISPR-Cas9 toolkit that allows rapid tagging and gene knockout in diverse kinetoplastid species without requiring the user to perform any DNA cloning. We developed a new protocol for single-guide RNA (sgRNA) delivery using PCR-generated DNA templates which are transcribed in vivo by T7 RNA polymerase and an online resource (LeishGEdit...
May 2017: Royal Society Open Science
https://www.readbyqxmd.com/read/28559295/polyvalent-proteins-a-pervasive-theme-in-the-inter-genomic-biological-conflicts-of-bacteriophages-and-conjugative-elements
#4
Lakshminarayan M Iyer, A Maxwell Burroughs, Swadha Anand, Robson F de Souza, L Aravind
Intense biological conflicts between prokaryotic genomes and their genomic parasites have resulted in an arms-race in terms of the molecular "weaponry" deployed on both sides. Using a recursive computational approach we uncovered a remarkable class of multidomain proteins with 2-15 domains in the same polypeptide deployed by viruses and plasmids in such conflicts. Domain architectures and genomic contexts indicate that they are part of a widespread conflict strategy involving proteins injected into the host cell along with parasite DNA during the earliest phase of infection...
May 30, 2017: Journal of Bacteriology
https://www.readbyqxmd.com/read/28545140/the-role-of-the-zinc-finger-protein-zc3h32-in-bloodstream-form-trypanosoma-brucei
#5
Cornelia Klein, Monica Terrao, Christine Clayton
Kinetoplastids rely heavily on post-transcriptional mechanisms for control of gene expression, with regulation of mRNA processing, translation and degradation by RNA-binding proteins. ZC3H32 is a cytosolic mRNA-binding protein with three non-canonical CCCH zinc finger domains. It is much more abundant in bloodstream-form Trypanosoma brucei than in procyclic forms. Tethering of ZC3H32 to a reporter mRNA suppressed translation and resulted in mRNA degradation, and deletion analysis suggested that this activity was present in both the N- and C-terminal domains, but not the central zinc finger-containing domain...
2017: PloS One
https://www.readbyqxmd.com/read/28535252/trypanosome-rna-editing-mediator-complex-proteins-have-distinct-functions-in-grna-utilization
#6
Rachel M Simpson, Andrew E Bruno, Runpu Chen, Kaylen Lott, Brianna L Tylec, Jonathan E Bard, Yijun Sun, Michael J Buck, Laurie K Read
Uridine insertion/deletion RNA editing is an essential process in kinetoplastid parasites whereby mitochondrial mRNAs are modified through the specific insertion and deletion of uridines to generate functional open reading frames, many of which encode components of the mitochondrial respiratory chain. The roles of numerous non-enzymatic editing factors have remained opaque given the limitations of conventional methods to interrogate the order and mechanism by which editing progresses and thus roles of individual proteins...
May 23, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28495405/trypanosoma-cruzi-i-towards-the-need-of-genetic-subdivision-part-ii
#7
REVIEW
Juan David Ramírez, Carolina Hernández
Chagas disease is a complex zoonosis caused by the kinetoplastid parasite Trypanosoma cruzi. This protozoan exhibits remarkable genetic diversity evinced in at least six Discrete Typing Units (DTUs) with the foreseen emergence of a genotype associated to bats (TcBat). T. cruzi I is the DTU with the broadest geographical distribution and associated to severe cardiomyopathies. In 2011, we published a review questioning the need for genetic subdivision within TcI. However, after six years of intensive research herein, we attempted to determine if TcI should be subdivided or not in the light of the current genetic, biological, clinical and ecological data...
May 8, 2017: Acta Tropica
https://www.readbyqxmd.com/read/28493888/the-mitochondrial-sir2-related-protein-2-sir2rp2-impacts-leishmania-donovani-growth-and-infectivity
#8
Nimisha Mittal, Rohini Muthuswami, Rentala Madhubala
BACKGROUND: Leishmania donovani, a protozoan parasite is the major causative agent of visceral leishmaniasis. Increased toxicity and resistance to the existing repertoire of drugs has been reported. Hence, an urgent need exists for identifying newer drugs and drug targets. Previous reports have shown sirtuins (Silent Information Regulator) from kinetoplastids as promising drug targets. Leishmania species code for three SIR2 (Silent Information Regulator) related proteins. Here, we for the first time report the functional characterization of SIR2 related protein 2 (SIR2RP2) of L...
May 2017: PLoS Neglected Tropical Diseases
https://www.readbyqxmd.com/read/28483460/voacamine-alters-leishmania-ultrastructure-and-kills-parasite-by-poisoning-unusual-bi-subunit-topoisomerase-ib
#9
Somenath Roy Chowdhury, Ashish Kumar, Joseane Lima Prado Godinho, Sara Teixeira De Macedo Silva, Aline Araujo Zuma, Sourav Saha, Neha Kumari, Juliany Cola Fernandez Rodrigues, Shyam Sundar, Jean-Claude Dujardin, Syamal Roy, Wanderley De Souza, Sibabrata Mukhopadhyay, Hemanta K Majumder
Indole alkaloids possess a large spectrum of biological activities including anti-protozoal action. Here we report for the first time that voacamine, isolated from the plant Tabernaemontana coronaria, is an antiprotozoal agent effective against a large array of trypanosomatid parasites including Indian strain of Leishmania donovani and Brazilian strains of Leishmania amazonensis and Trypanosoma cruzi. It inhibits the relaxation activity of topoisomerase IB of L. donovani (LdTop1B) and stabilizes the cleavable complex...
May 5, 2017: Biochemical Pharmacology
https://www.readbyqxmd.com/read/28462832/6-nitro-2-3-dihydroimidazo-2-1-b-1-3-thiazoles-facile-synthesis-and-comparative-appraisal-against-tuberculosis-and-neglected-tropical-diseases
#10
Andrew M Thompson, Adrian Blaser, Brian D Palmer, Robert F Anderson, Sujata S Shinde, Delphine Launay, Eric Chatelain, Louis Maes, Scott G Franzblau, Baojie Wan, Yuehong Wang, Zhenkun Ma, William A Denny
As part of a quest for backups to the antitubercular drug pretomanid (PA-824), we investigated the unexplored 6-nitro-2,3-dihydroimidazo[2,1-b][1,3]-thiazoles and related -oxazoles. The nitroimidazothiazoles were prepared in high yield from 2-bromo-4-nitroimidazole via heating with substituted thiiranes and diisopropylethylamine. Equivalent examples of these two structural classes provided broadly comparable MICs, with 2-methyl substitution and extended aryloxymethyl side chains preferred; albeit, S-oxidised thiazoles were ineffective for tuberculosis...
March 27, 2017: Bioorganic & Medicinal Chemistry Letters
https://www.readbyqxmd.com/read/28459575/7-substituted-2-nitro-5-6-dihydroimidazo-2-1-b-1-3-oxazines-novel-antitubercular-agents-lead-to-a-new-preclinical-candidate-for-visceral-leishmaniasis
#11
Andrew M Thompson, Patrick D O'Connor, Andrew J Marshall, Vanessa Yardley, Louis Maes, Suman Gupta, Delphine Launay, Stephanie Braillard, Eric Chatelain, Scott G Franzblau, Baojie Wan, Yuehong Wang, Zhenkun Ma, Christopher B Cooper, William A Denny
Within a backup program for the clinical investigational agent pretomanid (PA-824), scaffold hopping from delamanid inspired the discovery of a novel class of potent antitubercular agents that unexpectedly possessed notable utility against the kinetoplastid disease visceral leishmaniasis (VL). Following the identification of delamanid analogue DNDI-VL-2098 as a VL preclinical candidate, this structurally related 7-substituted 2-nitro-5,6-dihydroimidazo[2,1-b][1,3]oxazine class was further explored, seeking efficacious backup compounds with improved solubility and safety...
May 25, 2017: Journal of Medicinal Chemistry
https://www.readbyqxmd.com/read/28429321/euglena-transcript-processing
#12
David C McWatters, Anthony G Russell
RNA transcript processing is an important stage in the gene expression pathway of all organisms and is subject to various mechanisms of control that influence the final levels of gene products. RNA processing involves events such as nuclease-mediated cleavage, removal of intervening sequences referred to as introns and modifications to RNA structure (nucleoside modification and editing). In Euglena, RNA transcript processing was initially examined in chloroplasts because of historical interest in the secondary endosymbiotic origin of this organelle in this organism...
2017: Advances in Experimental Medicine and Biology
https://www.readbyqxmd.com/read/28397756/anti-trypanosomatid-elemanolide-sesquiterpene-lactones-from-vernonia-lasiopus-o-hoffm
#13
Njogu M Kimani, Josphat C Matasyoh, Marcel Kaiser, Reto Brun, Thomas J Schmidt
Sleeping sickness or human African trypanosomiasis (HAT) is a neglected tropical disease (NTD) threatening millions of peoples' lives with thousands infected. The disease is endemic in poorly developed regions of sub-Saharan Africa and is caused by the kinetoplastid "protozoan" parasite Trypanosoma brucei. The parasites are transmitted to humans through bites of infected tsetse flies of the genus Glossina. The few available drugs for treatment of this disease are highly toxic, difficult to administer, costly and unavailable to poor rural communities bearing the major burden of this infection...
April 8, 2017: Molecules: a Journal of Synthetic Chemistry and Natural Product Chemistry
https://www.readbyqxmd.com/read/28381627/characterization-of-rbp9-and-rbp10-two-developmentally-regulated-rna-binding-proteins-in-trypanosoma-brucei
#14
Luis Miguel De Pablos, Steve Kelly, Janaina de Freitas Nascimento, Jack Sunter, Mark Carrington
The fate of an mRNA is determined by its interaction with proteins and small RNAs within dynamic complexes called ribonucleoprotein complexes (mRNPs). In Trypanosoma brucei and related kinetoplastids, responses to internal and external signals are mainly mediated by post-transcriptional processes. Here, we used proximity-dependent biotin identification (BioID) combined with RNA-seq to investigate the changes resulting from ectopic expression of RBP10 and RBP9, two developmentally regulated RNA-binding proteins (RBPs)...
April 2017: Open Biology
https://www.readbyqxmd.com/read/28369657/origin-and-evolutionary-alteration-of-the-mitochondrial-import-system-in-eukaryotic-lineages
#15
Yoshinori Fukasawa, Toshiyuki Oda, Kentaro Tomii, Kenichiro Imai
Protein transport systems are fundamentally important for maintaining mitochondrial function. Nevertheless, mitochondrial protein translocases such as the kinetoplastid ATOM complex have recently been shown to vary in eukaryotic lineages. Various evolutionary hypotheses have been formulated to explain this diversity. To resolve any contradiction, estimating the primitive state and clarifying changes from that state are necessary. Here, we present more likely primitive models of mitochondrial translocases, specifically the translocase of the outer membrane (TOM) and translocase of the inner membrane (TIM) complexes, using scrutinized phylogenetic profiles...
July 1, 2017: Molecular Biology and Evolution
https://www.readbyqxmd.com/read/28336470/bioassay-guided-isolation-of-active-principles-from-nigerian-medicinal-plants-identifies-new-trypanocides-with-low-toxicity-and-no-cross-resistance-to-diamidines-and-arsenicals
#16
Godwin Unekwuojo Ebiloma, John Ogbaji Igoli, Evangelos Katsoulis, Anne-Marie Donachie, Anthonius Eze, Alexander Ian Gray, Harry P de Koning
ETHNOPHARMACOLOGICAL RELEVANCE: Leaves from the plant species studied herein are traditionally used in northern Nigeria against various protozoan infections. However, none of these herbal preparations have been standardized, nor have their toxicity to mammalian cells been investigated. In search of improved and non-toxic active antiprotozoal principles that are not cross-resistant with current anti-parasitics, we here report the results of the in vitro screening of extracts from seven selected medicinal plant species (Centrosema pubescens, Moringa oleifera, Tridax procumbens, Polyalthia longifolia, Newbouldia laevis, Eucalyptus maculate, Jathropha tanjorensis), used traditionally to treat kinetoplastid infections in Nigeria, and the isolation of their bioactive principles...
March 20, 2017: Journal of Ethnopharmacology
https://www.readbyqxmd.com/read/28334821/in-vivo-cleavage-specificity-of-trypanosoma-brucei-editosome-endonucleases
#17
Jason Carnes, Suzanne McDermott, Atashi Anupama, Brian G Oliver, D Noah Sather, Kenneth Stuart
RNA editing is an essential post-transcriptional process that creates functional mitochondrial mRNAs in Kinetoplastids. Multiprotein editosomes catalyze pre-mRNA cleavage, uridine (U) insertion or deletion, and ligation as specified by guide RNAs. Three functionally and compositionally distinct editosomes differ by the mutually exclusive presence of the KREN1, KREN2 or KREN3 endonuclease and their associated partner proteins. Because endonuclease cleavage is a likely point of regulation for RNA editing, we elucidated endonuclease specificity in vivo...
May 5, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28320630/early-polymerase-chain-reaction-detection-of-chagas-disease-reactivation-in-heart-transplant-patients
#18
Priscilla Almeida da Costa, Marcela Segatto, Danielle Fernandes Durso, Wagson José de Carvalho Moreira, Lucas Lodi Junqueira, Fábio Morato de Castilho, Silvio Amadeu de Andrade, Cláudio Léo Gelape, Egler Chiari, Andréa Teixeira-Carvalho, Sergio Danilo Junho Pena, Carlos Renato Machado, Gloria Regina Franco, Geraldo Brasileiro Filho, Maria da Consolação Vieira Moreira, Andréa Mara Macedo
BACKGROUND: Heart transplantation is a valuable therapeutic option for Chagas disease patients with severe cardiomyopathy. During patient follow-up, the differential diagnosis between cardiac transplant rejection and Chagas disease infection reactivation remains a challenging task, which hinders rapid implementation of the appropriate treatment. Herein we investigate whether polymerase chain reaction (PCR) strategies could facilitate early detection of Trypanosoma cruzi (T cruzi) in transplanted endomyocardial biopsies (EMBs)...
February 24, 2017: Journal of Heart and Lung Transplantation
https://www.readbyqxmd.com/read/28285595/evaluation-of-four-molecular-methods-to-detect-leishmania-infection-in-dogs
#19
Andreia Albuquerque, Lenea Campino, Luís Cardoso, Sofia Cortes
BACKGROUND: Canine leishmaniasis, a zoonotic disease caused by Leishmania infantum vectored by phlebotomine sand flies, is considered a relevant veterinary and public health problem in various countries, namely in the Mediterranean basin and Brazil, where dogs are considered the main reservoir hosts. Not only diseased dogs but also those subclinically infected play a relevant role in the transmission of L. infantum to vectors; therefore, early diagnosis is essential, under both a clinical and an epidemiological perspective...
March 13, 2017: Parasites & Vectors
https://www.readbyqxmd.com/read/28246372/a-novel-stage-specific-glycosomal-nucleoside-diphosphate-kinase-from-trypanosoma-cruzi
#20
Maria de Los Milagros Camara, Leon Bouvier, Chantal Reigada, Fabio A Digirolamo, Melisa Saye, Claudio A Pereira
Nucleoside diphosphate kinases (NDPK) are key enzymes involved in the intracellular nucleotide maintenance in all living organisms, especially in trypanosomatids which are unable to synthesise purines de novo. Four putative NDPK isoforms were identified in the Trypanosoma cruzi Chagas, 1909 genome but only two of them were characterised so far. In this work, we studied a novel isoform from T. cruzi called TcNDPK3. This enzyme presents an atypical N-terminal extension similar to the DM10 domains. In T. cruzi, DM10 sequences targeted other NDPK isoform (TcNDPK2) to the cytoskeleton, but TcNDPK3 was localised in glycosomes despite lacking a typical peroxisomal targeting signal...
February 17, 2017: Folia Parasitologica
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