Mycobacterium genus and tRNA arrays

The presence of tRNA array, a region with high tRNA gene number and density, has been demonstrated in Mycobacterium genus. However, a recent phylogenomic study revealed the existence of five distinct monophyletic groups (genera) within this genus. Considering this new scenario, and based on in-silico analyses, we have identified and characterised the abundance and diversity of tRNA array units within Mycobacterium, Mycolicibacterium gen. nov., Mycolicibacillus gen. nov., and Mycobacteroides gen. nov. The occurrence and prevalence of tRNA arrays among the genera belonging to Actinobacteria indicate their possible role in the organismal fitness.

Exploring tRNA gene cluster in archaea

BACKGROUND Shared traits between prokaryotes and eukaryotes are helpful in the understanding of the tree of life evolution. In bacteria and eukaryotes, it has been shown a particular organisation of tRNA genes as clusters, but this trait has not been explored in the archaea domain. OBJECTIVE Explore the occurrence of tRNA gene clusters in archaea. METHODS In-silico analyses of complete and draft archaeal genomes based on tRNA gene isotype and synteny, tRNA gene cluster content and mobilome elements. FINDINGS We demonstrated the prevalence of tRNA gene clusters in archaea. tRNA gene clusters, composed of archaeal-type tRNAs, were identified in two Archaea class, Halobacteria and Methanobacteria from Euryarchaeota supergroup. Genomic analyses also revealed evidence of the association between tRNA gene clusters to mobile genetic elements and intra-domain horizontal gene transfer. MAIN CONCLUSIONS tRNA gene cluster occurs in the three domains of life, suggesting a role of this type of tRNA gene organisation in the biology of the living organisms.

Distinct subcellular localization of tRNA-derived fragments in the infective metacyclic forms of Trypanosoma cruzi

Small non-coding RNAs derived from transfer RNAs have been identified as a broadly conserved prokaryotic and eukaryotic response to stress. Their presence coincides with changes in developmental state associated with gene expression regulation. In the epimastigote form of Trypanosoma cruzi, tRNA fragments localize to posterior cytoplasmic granules. In the infective metacyclic form of the parasite, we found tRNA-derived fragments to be abundant and evenly distributed within the cytoplasm. The fragments were not associated with polysomes, suggesting that the tRNA-derived fragments may not be directly involved in translation control in metacyclics.

Enfermedad mitocondrial y ototoxicidad". Enfermedad mitocondrial sindrómica, mutación G7444A / Mitochondrial disease and ototoxicity

Como parte del programa de “screening” genético para la pérdida de la audición; se realizó el estudio de mutaciones en los genes rRNA 12S y tRNAser (UCN), a partir del DNA mitocondrial, que están asociadas con la pérdida de audición inducida por antibióticos aminoglucósidos (ATB-AG) y de presentación no sindrómica. Se estudiaron 40 pacientes con sorderaneurosensorial, la cual podría haber sido causada posterior al tratamiento con ATB-AG. El sujeto afectado y el control, luego del examen físico completo y extracción de DNA a partir de sangre periférica, se amplificó y estudió segmentos en el gen 12S rRNA y en el gen tRNAser (UCN) por PCR-RFLP. En presencia de una mutación, se analizó el genoma mitocondrial completo en el probando y su familia por línea materna. Estos resultadosse han correlacionado con los valores de la relación dela citocromo oxidasa / citrato sintasa, el cual indica unapobre actividad de la citocromo oxidasa. La clínica en el“pedigree” por línea materna y los estudios moleculares,bioquímicos y morfológicos, podría indicar que se tratade una presentación sindrómica de la mutación 7444G>Aen Córdoba - Argentina.

As part of the "screening" for genetic hearing loss,was performed to study mutations in 12S rRNA and tRNAser(UCN) genes from mitochondrial DNA, which are associatedwith hearing loss induced by antibiotics aminoglycosides(ATB-AG) and non-syndromic presentation. We studied40 patients with sensorineural hearing loss, which could have been caused subsequent to treatment with ATB - AG. The affected individual and control, after a completephysical examination and extraction of DNA fromperipheral blood, was amplified and studied segments inthe 12S rRNA and tRNAser (UCN) genes by PCR-RFLP. In the presence of a mutation, we analyzed the complete mitochondrial genome in the proband and his family from maternal line. These results werecorrelated with the values of the ratio of the cytochromeoxidase / citrate synthase, which indicates a poor activity of cytochrome oxidase. The clinic in the "pedigree" frommaternal line and molecular, biochemical and morphological might indicate that it is a syndromic presentation of the mutation 7444G> A in Córdoba - Argentina.

Phylogenetic analysis of Biomphalaria tenagophila (Orbigny, 1835) (Mollusca: Gastropoda)

Mitochondrial DNA of Biomphalaria tenagophila, a mollusc intermediate host of Schistosoma mansoni in Brazil, was sequenced and characterised. The genome size found for B. tenagophila was 13,722 bp and contained 13 messenger RNAs, 22 transfer RNAs (tRNA) and two ribosomal RNAs (rRNA). In addition to sequencing, the mitochondrial DNA (mtDNA) genome organization of B. tenagophila was analysed based on its content and localization of both coding and non-coding regions, regions of gene overlap and tRNA nucleotide sequences. Sequences of protein, rRNA 12S and rRNA 16S nucleotides as well as gene organization were compared between B. tenagophila and Biomphalaria glabrata, as the latter is the most important S. mansoni intermediate host in Brazil. Differences between such species were observed regarding rRNA composition. The complete sequence of the B. tenagophila mitochondrial genome was deposited in GenBank (accession EF433576). Furthermore, phylogenetic relationships were estimated among 28 mollusc species, which had their complete mitochondrial genome deposited in GenBank, using the neighbour-joining method, maximum parsimony and maximum likelihood bootstrap. B. tenagophila was positioned at a branch close to B. glabrata and Pulmonata molluscs, collectively comprising a paraphyletic group, contrary to Opistobranchia, which was positioned at a single branch and constituted a monophyletic group.

Effects of Trypanosoma brucei tryptophanyl-tRNA synthetases silencing by RNA interference

The kinetoplast genetic code deviates from the universal code in that 90 percent of mitochondrial tryptophans are specified by UGA instead of UGG codons. A single nucleus-encoded tRNA Trp(CCA) is used by both nuclear and mitochondria genes, since all kinetoplast tRNAs are imported into the mitochondria from the cytoplasm. To allow decoding of the mitochondrial UGA codons as tryptophan, the tRNA Trp(CCA) anticodon is changed to UCA by an editing event. Two tryptophanyl tRNA synthetases (TrpRSs) have been identified in Trypanosoma brucei: TbTrpRS1 and TbTrpRS2 which localize to the cytoplasm and mitochondria respectively. We used inducible RNA interference (RNAi) to assess the role of TbTrpRSs. Our data validates previous observations of TrpRS as potential drug design targets and investigates the RNAi effect on the mitochondria of the parasite.

Análisis de la estructura primaria y secundaria del ARN de transferencia mitocondrial para serina en siete especies de Lutzomyia / Analysis of the primary and secondary structure of the mitochondrial serine transfer RNA in seven species of Lutzomyia

Introducción. Los insectos del género Lutzomyia son los responsables de la transmisión del parásito Leishmania spp. en América. La taxonomía de estos vectores se fundamenta en los caracteres morfológicos que exhiben los adultos, principalmente, en las estructuras pareadas de la cabeza y los genitales. Aunque estos caracteres permiten distinguir la mayoría de los taxones, la similitud en algunos subgéneros y grupos de especies pone límites a la identificación por criterios morfológicos. Objetivo. Evaluar la utilidad del ARN de transferencia mitocondrial para serina ARNtSer en la determinación taxonómica de Lutzomyia. Materiales y métodos. Se analizaron siete especies flebotomíneas, L. trinidadensis, L. panamensis, L. cayennensis cayennensis, L. dubitans, L. gomezi, L. rangeliana y L. evansi. A partir de cada individuo, se extrajo, amplificó y obtuvo la secuencia del gen mitocondrial que codifica para el ARNtSer, delimitado por los genes citocromo b y NAD deshidrogenasa uno. La estructura secundaria del ARNtSer se infirió teniendo como base las estructuras homólogas descritas en otros insectos del orden Diptera. Resultados. La longitud del gen ARNtSer osciló entre 66 pb en L. gomezi y 69 pb en L. trinidadensis. En el alineamiento nucleotídico de 70 posiciones, se detectaron 14 sitios polimórficos, incluyendo cuatro eventos indel. La mayoría de las sustituciones correspondieron a las lupas dihidrouridina, ribotimidina-pseudouridina-citosina y variable, así como al extremo basal del brazo anticodón. Conclusión. Los cambios en la secuencia primaria de nucleótidos y los rearreglos en la estructura secundaria del ARNtSer son potencialmente útiles para la discriminación taxonómica de las especies flebotomíneas estudiadas.

Introduction. Lutzomyia sand flies are involved in the transmission of the parasite Leishmania spp. in America. The taxonomy of these vectors is traditionally based on morphological features of the adult stage, particularly the paired structures of the head and genitalia. Although these characters are useful to distinguish most species of Lutzomyia, morphological identification may be complicated by the similarities within subgenera and species group. Objective. To evaluate the utility of mitochondrial serine transfer RNA tRNASer for taxonomic identification of Lutzomyia. Materials and methods. Seven sand fly species, each representing one of the 27 taxonomic subdivisions in genus Lutzomyia, were analyzed including L. trinidadensis (Oswaldoi group), L. (Psychodopygus) panamensis, L.(Micropygomyia) cayennensis cayennensis, L. dubitans (Migonei group), L. (Lutzomyia) gomezi, L. rangeliana (ungrouped) and L. evansi (Verrucarum group). The mitochondrial tRNASer gene, flanked by the cytochrome b and NAD dehydrogenase subunit one genes, was extracted, amplified and sequenced from each specimen. Secondary structure of the tRNASer was predicted by comparisons with previously described homologous structures from other dipteran species. Results. The tRNASer gene ranged in size from 66 base pairs in L. gomezi to 69 base pairs in L. trinidadensis. Fourteen polymorphic sites, including four insertion-deletion events, were observed in the aligned 70 nucleotide positions. The majority of the substitutions were located in the dihydrouridine, ribothymidine-pseudouridine-cytosine and variable loops, as well as in the basal extreme of the anticodon arm. Conclusion. Changes of primary sequence of the tRNASer provided useful molecular characters for taxonomic identification of the sand fly species under consideration.

Mitochondrial tRNA gene translocations in highly eusocial bees

Mitochondrial gene rearrangement events, especially involving tRNA genes, have been described more frequently as more complete mitochondrial genome sequences are becoming available. In the present work, we analyzed mitochondrial tRNA gene rearrangements between two bee species belonging to the tribes Apini and Meliponini within the "corbiculate Apidae". Eleven tRNA genes are in different genome positions or strands. The molecular events responsible for each translocation are explained. Considering the high number of rearrangements observed, the data presented here contradict the general rule of high gene order conservation among closely related organisms, and also represent a powerful molecular tool to help solve questions about phylogeny and evolution in bees.

Molecular investigation of tRNA genes integrity and its relation to pathogenicity islands in Shiga toxin-producing Escherichia coli (STEC) strains

tRNA genes are known target sites for the integration of pathogenicity islands (PAI) and other genetic elements, such as bacteriophages, into bacterial genome. In most STEC (Shiga toxin-producing Escherichia coli), the PAI called LEE (locus of enterocyte effacement) is related to bacterial virulence and is mostly associated to the tRNA genes selC and pheU. In this work, we first investigated the relationship of LEE with tRNA genes selC and pheU in 43 STEC strains. We found that 28 strains (65 percent) had a disrupted selC and/or pheU. Three of these strains (637/1, 650/5 and 654/3) were chosen to be submitted to a RAPD-PCR technique modified by the introduction of specific primers (corresponding to the 5'end of genes selC and pheU) into the reaction, which we called "anchored RAPD-PCR". The PCR fragments obtained were transferred onto membranes, and those fragments which hybridized to selC and pheU probes were isolated. One of these fragments from strain 637/1 was partially sequenced. An 85-nucleotide sequence was found to be similar to the cfxA2 gene that encodes a beta-lactamase and is part of transposon Tn4555, a pathogenicity island originally integrated into the Bacteroides genome.

Gene expression in Chromobacterium violaceum

The repertoire of 4,431 open reading frames (ORFs), eight rRNA operons and 98 tRNA genes of Chromobacterium violaceum must be expressed in a regulated manner for successful adaptation to a wide variety of environmental conditions. To accomplish this feat, the organism relies on protein machineries involved in transcription, RNA processing and translation. Analysis of the C. violaceum genome showed that transcription initiation, elongation and termination are performed by the five well-known RNA polymerase subunits, five categories of sigma 70 factors, one sigma 54 factor, as well as six auxiliary elongation and termination factors. RNA processing is performed by a variety of endonucleases and exonucleases, such as ribonuclease H, ribonuclease E, ribonuclease P, and ribonuclease III, in addition to poly(A) polymerase and specific methyltransferases and pseudouridine synthases. ORFs for all ribosomal proteins, except S22, were found. Only 19 aminoacyl-tRNA synthetases were found, in addition to three aminoacyl-tRNA synthetase-related proteins. Asparaginyl-tRNA (Asn) is probably obtained by enzymatic modification of a mischarged aminoacyl-tRNA. The translation factors IF-1, IF-2, IF-3, EF-Ts, EF-Tu, EF-G, RF-1, RF-2 and RF-3 are all present in the C. violaceum genome, although the absence of selB suggests that C. violaceum does not synthesize selenoproteins. The components of trans-translation, tmRNA and associated proteins, are present in the C. violaceum genome. Finally, a large number of ORFs related to regulation of gene expression were also found, which was expected, considering the apparent adaptability of this bacterium

The complete sequence of the mitochondrial genome of the Chinook salmon, Oncorhynchus tshawytscha

The complete sequence of the mitochondrial genome of Chinook salmon, Oncorhynchus tshawytscha, has been determined. The circular genome consisting of 16,644 base pairs encodes thirteen proteins, the 12S and 16S ribosomal RNAs, and 22 transfer RNAs. These genes are ordered in the same way as most other vertebrates. The nucleotide and amino acid sequences of the ribosomal RNAs and the thirteen protein-coding genes were compared with those of other salmonids such as Oncorhynchus mykiss, Salmo salar, Salvelinus fontinalis, Salvelinus alpinus and Coregonus lavaretus. The sequence features of the control region (D-loop), the origin of L-strand replication and a putative peptide codified by the 16S mitochondrial RNA are described and discussed

Efectos biológicos en células HEp-2, en la multiplicación del adenovirus tipo 6 y en la concentración de interferón en plasma humano en presencia de un RNA de transferencia fúngico / Biological effects in HEp-2 cells, adenovirus type 6 and interferon concentration in human plasma in the presence of a fungal transfer RNA

Introducción: Trabajos previos mostraron la capacidad antiviral in vitro, así como la utilidad como inductor de interferón natural (IFN-n) in vivo de un RNA de transferencia (tRNA) de origen fúngico, por este motivo se sigue estudiando esta molécula como una alternativa para el tratamiento antiviral. Objetivos: Estudiar el efecto del tRNA fúngico en la viabilidad, síntesis de DNA y en la multiplicación del adenovirus tipo 6 (AV-6) en células HEp-2, comparado su efecto con el producido por el polyl:polyC o por IFN-Ó. Valorar su capacidad como unductor a largo plazo de la síntesis de IFN-n in vivo. Material y métodos: Células HEp-2 se incubaron con diferentes concentraciones de las moléculas mencionadas durante 24 horas, y se determinó la viabilidad y la síntesis de DNA celular; adicionalmente cultivos tratados en las mismas condiciones se infectaron con 200 unidades formadoras de placas (ufp) del AV-6, se incubaron cinco días adicionales, y se valoró el grado de protección. In vivo a siete voluntarios clínicamente sanos se les administró intramuscularmente una dosis única de 100 mg del tRNA, y se determinó mediante la técnica de inhibición del efecto citopático (ECP) el nivel sérico de IFN-n, cinco días después de la inoculación. Resultados. El tRNA protegió a las células HEp-2 contra la infección por AV-6, mejor que el polyl:polyC y de forma similar al IFN-Ó. Ninguna de las tres sustancias afectó significativamente la viabilidad celular. En cambio, la síntesis de DNA sí disminuyó de manera directa en relación con la concentración de los inductores, no así con el IFN-Ó. En los plasmas de los sujetos tratados con el tRNA fúngico se encontró un aumento en la concentración de IFN-n (de 84.2 ñ 107.6 a 171.42 ñ 129.5 UI/mL) a los cinco días, aunque la diferencia no fue estadísticamente significativa. Conclusión. El tRNA fúngico mostró actividad antiviral contra el AV-6, no afectó la viabilidad pero sí la síntesis celular de DNA, y con una capacidad de mantener elevada hasta por cinco días la concentración plasmática de IFN-n en humanos

Los inductores de la síntesis de interferón endógeno ¿una posible opción terapéutica? / Endogenous interferon synthesis inducers. A possible therapeutic option?

La linfocina conocida como interferón (IFN) existe en tres tipos principales (-Ó, -ß y -ç), cada uno de ellos con diferente capacidad como agente antiviral, antitumoral o como inmunomodulador, esta características motivaron su aplicción exógena desde su descubrimiento como terapia para el tratamiento de diversos padecimientos, esto sin embargo, trajo consigo severos efectos colaterales del IFN exógeno, y de estimular la síntesis endógena en el organismo para aprovechar sus propiedades en la eliminación de alguna enfermedad (viral, tumoral, etc.), se han probado una enorme cantidad de compuestos, tanto de origen natural como sintético (inductores), lamentablemente hasta el momento no se ha encontrado al inductor idóneo, debido principalmente a factores, como: el tipo y grado de la infección o proceso tumoral, población estudiada, toxicidad de la molécula, etc., por estos motivos son necesarios la realización de un mayor número de trabajos para encontrar al mejor inductor, capaz de utilizarse con fines terapéuticos y que además muestre un mínimo de efectos secundarios al paciente al cual se aplique. En el presente trabajo se revisan algunos estudios que hablan de la utilidad, así como de los efectos secundarios de los inductores de la síntesis de IFN como el polyA:polyU, el PolyI:polyC, PolyI:polyC12U, Ridostine, Bropirimine y de un ARNt de orgien fúngico utilizados en el tratamiento de procesos infecciosos y tumorales

Incremento en la concentración de IFN-ç y TNF-Ó en el plasma de sujetos sanos a los que se administró un RNAt de origen fúngico. Reporte preliminar / Increase in the concentrations of IFN-ç and TNF-Ó in the plasm of healthy subjects treated with fungal tRNA. Preliminary report

Antecedentes: Es importante el uso con fines terapéuticos de inmunomoduladores que funcionen como inductores de la sintesis de interferón natural (IFN-n), principalmente si se considera el costo y toxicidad del interferón recombinante. En este trabajo, se estudia el efecto que la aplicación de uno de estos inductores, el ácido ribonucleico de transferencia (RNAt), tuvo sobre los niveles séricos del interferón gamma (IFN-ç) y del factor de necrosis tumoral alfa (TNF-Ó). Objetivos: Determinar la concentración del INF-ç y el TNF-Ó en el plasma de sujetos a quienes se les aplicó un RNAt de origen fúngico como inductor de la síntesis de IFN-n. Material y Métodos: A nueve voluntarios sanos se les administró por vía IM 200 mg del RNAt, y se valoró en plasma de sangre venosa antes y dos horas después de la apliación del RNAt, la concentración del IFN-ç y del TNF-Ó por la técnica de ELISA. Resultados: Se observó que los niveles de IFN-ç y del TNF-Ó se incrementaron de 267.83 ñ 208.03 a 2230 ñ 2088.73 pg/mL, y de 48.92 ñ 88.07 a 261.65 ñ 80.61 pg/mL respectivamante (Prom. ñ D.S, p < 0.05), a las dos horas de la administración de RNAt. Conclusión: Con la administración del RNAt se observa una tendencia de incremento en la concentración de IFN-ç y del TNF-Ó, en el plasma de sujetos normales

tRNA rRNA sequence matches from inter- and intraspecies comparisons suggest common origins for the two RNAs

1. Comparisons were made of the results of searches within and among different species of organisms for sequence matches between transfer RNAs and ribosomal RNAs. The purpose was to determine whether the matching sequences might result form selection acting on the two RNAs within a common cellular environment. 2. The results indicate that most matches do not reflect such selection. The matches described we more frequent than those found in searches among randomized sequences and the frequency of intraspecific matches was not significantly higher than that of interspecific matches. 3. The matches ara thought to identify conserved vestiges of a molecule or molecules ancestral to both classes of RNAs (Bloch, D.P., McArthur, B. and Mirrop, S (1985), BioSystems, 17:209-225). The matching sequences are interpreted as homologies