RESUMEN

The standard genetic code determines that in most species, including viruses, there are 20 amino acids that are coded by 61 codons, while the other three codons are stop triplets. Considering the whole proteome each species features its own amino acid frequencies, given the slow rate of change, closely related species display similar GC content and amino acids usage. In contrast, distantly related species display different amino acid frequencies. Furthermore, within certain multicellular species, as mammals, intragenomic differences in the usage of amino acids are evident. In this communication, we shall summarize some of the most prominent and well-established factors that determine the differences found in the amino acid usage, both across evolution and intragenomically.

Asunto(s)

Aminoácidos , Código Genético , Animales , Aminoácidos/genética , Codón/genética , Composición de Base , Proteoma/genética , Evolución Molecular , Mamíferos/genética

RESUMEN

Current available information on reptile genomes provides great potential for the study of unique adaptations from a genomic perspective. We compared differences in base composition and codon usage patterns across 400 reptile mitochondrial genomes assessing AT and GC skew, GC frequency, codon usage, effective number of codons, and codon adaptation index. We identified poor GC content in reptile mitochondrial genomes, with a predominant bias toward Adenine. We determined a compositional asymmetry between different taxonomic groups, which are inversely correlated to the rates of rearrangements in the mitogenome. We found that the most common codons in reptile mitochondrion are CTA (L), ATA (M) and ACA (T), which relates with have been found in birds, meaning that these patterns are shared across sauropsid mitogenomes. Codon usage bias clustering and effective codon number analyses revelated compositional asymmetry based on RSCU as well as that reptile mitogenomes are translationally efficient and are under selection pressure. Codon adaptation index revealed highest values in turtles indicating higher translational efficiency of mitochondrial genes among all reptiles, which could be related to metabolic adaptations (i.e., tolerance to anoxic conditions). This was also seen in other groups such as crocodiles (i.e., acclimation to cold) and snakes (phylogenetic origin of toxin-secreting oral glands and the evolutionary redesign of cytochrome c oxidase complex genes). We discuss our findings in the context of potential adaptations and evolutionary implications that these genomic differences provide to reptiles.

Asunto(s)

Uso de Codones , Genoma Mitocondrial , Animales , Genoma Mitocondrial/genética , Filogenia , Codón/genética , Reptiles/genética

RESUMEN

Codon usage is the outcome of different evolutionary processes and can inform us about the conditions in which organisms live and evolve. Here, we present R_ENC', which is an improvement to the original S index developed by dos Reis et al. (2004). Our index is less sensitive to G+C content, which greatly affects synonymous codon usage in prokaryotes, making it better suited to detect selection acting on codon usage. We used R_ENC' to estimate the extent of selected codon usage bias in 1800 genomes representing 26 prokaryotic phyla. We found that Gammaproteobacteria, Betaproteobacteria, Actinobacteria, and Firmicutes are the phyla/subphyla showing more genomes with selected codon usage bias. In particular, we found that several lineages within Gammaproteobacteria and Firmicutes show a similar set of functional terms enriched in genes under selected codon usage bias, indicating convergent evolution. We also show that selected codon usage bias tends to evolve in genes coding for the translation machinery before other functional GO terms. Finally, we discuss the possibility to use R_ENC' to predict whether lineages evolved in copiotrophic or oligotrophic environments.

Asunto(s)

Bacterias , Uso de Codones , Uso de Codones/genética , Codón/genética , Composición de Base , Bacterias/genética , Selección Genética , Evolución Molecular

RESUMEN

The Culicidae family is distributed worldwide and comprises about 3587 species subdivided into the subfamilies Anophelinae and Culicinae. This is the first description of complete mitochondrial DNA sequences from Aedes fluviatilis, Aedeomyia squamipennis, Coquillettidia nigricans, Psorophora albipes, and Psorophora ferox. The mitogenomes showed an average length of 15,046 pb and 78.02% AT content, comprising 37 functional subunits (13 protein coding genes, 22 tRNAs, and two rRNAs). The most common start codons were ATT/ATG, and TAA was the stop codon for all PCGs. The tRNAs had the typical leaf clover structure, except tRNASer1. Phylogeny was inferred by analyzing the 13 PCGs concatenated nucleotide sequences of 48 mitogenomes. Maximum likelihood and Bayesian inference analysis placed Ps. albipes and Ps. ferox in the Janthinosoma group, like the accepted classification of Psorophora genus. Ae. fluviatilis was placed in the Aedini tribe, but was revealed to be more related to the Haemagogus genus, a result that may have been hampered by the poor sampling of Aedes sequences. Cq. nigricans clustered with Cq. chrysonotum, both related to Mansonia. Ae. squamipennis was placed as the most external lineage of the Culicinae subfamily. The yielded topology supports the concept of monophyly of all groups and ratifies the current taxonomic classification.

Asunto(s)

Culicidae/genética , Genoma Mitocondrial/genética , Aedes/genética , Animales , Composición de Base/genética , Brasil , Codón/genética , Culicidae/metabolismo , ADN Mitocondrial/genética , Conformación de Ácido Nucleico , Sistemas de Lectura Abierta/genética , Filogenia , ARN Ribosómico/genética , ARN de Transferencia/genética

RESUMEN

Since the genetic code is degenerate, several codons are translated to the same amino acid. Although these triplets were historically considered to be "synonymous" and therefore expected to be used at rather equal frequencies in all genomes, we now know that this is not the case. Indeed, since several coding sequences were obtained in the late '70s and early '80s in the last century, coming from either the same or different species, it was evident that (a) each genome, taken globally, displayed different codon usage patterns, which means that different genomes display a particular global codon usage table when all genes are considered together, and (b) there is a strong intragenomic diversity: in other words, within a given species the codon usage pattern can (and usually do) differ greatly among genes in the same genome. These different patterns were attributed to two main factors: first, the mutational bias characteristic of each genome, which determines that GC- poor species display a general bias towards A/T codons while the reverse is true for GC- rich species. Second, the differences in codon usage among genes from the same species are due to natural selection acting at the level of translation, in such a way that highly expressed genes tend to use codons that match with the most abundant isoacceptor tRNAs. Thus, these genes are translated at a highest rate, which in turn leads to avoid the limiting factor in translation which is the number of available ribosomes per cell. Although these explanations are still valid, new factors are almost constantly postulated to affect codon usage. In this mini review, we shall try to summarize them.

Asunto(s)

Uso de Codones , Código Genético , Codón/genética , ARN de Transferencia/genética , Selección Genética

RESUMEN

In December 2019, rising pneumonia cases caused by a novel ß-coronavirus (SARS-CoV-2) occurred in Wuhan, China, which has rapidly spread worldwide, causing thousands of deaths. The WHO declared the SARS-CoV-2 outbreak as a public health emergency of international concern, since then several scientists are dedicated to its study. It has been observed that many human viruses have codon usage biases that match highly expressed proteins in the tissues they infect and depend on the host cell machinery for the replication and co-evolution. In this work, we analysed 91 molecular features and codon usage patterns for 339 viral genes and 463 human genes that consisted of 677,873 codon positions. Hereby, we selected the highly expressed genes from human lung tissue to perform computational studies that permit to compare their molecular features with those of SARS, SARS-CoV-2 and MERS genes. The integrated analysis of all the features revealed that certain viral genes and overexpressed human genes have similar codon usage patterns. The main pattern was the A/T bias that together with other features could propitiate the viral infection, enhanced by a host dependant specialization of the translation machinery of only some of the overexpressed genes. The envelope protein E, the membrane glycoprotein M and ORF7 could be further benefited. This could be the key for a facilitated translation and viral replication conducting to different comorbidities depending on the genetic variability of population due to the host translation machinery. This is the first codon usage approach that reveals which human genes could be potentially deregulated due to the codon usage similarities between the host and the viral genes when the virus is already inside the human cells of the lung tissues. Our work leaded to the identification of additional highly expressed human genes which are not the usual suspects but might play a role in the viral infection and settle the basis for further research in the field of human genetics associated with new viral infections. To identify the genes that could be deregulated under a viral infection is important to predict the collateral effects and determine which individuals would be more susceptible based on their genetic features and comorbidities associated.

Asunto(s)

Betacoronavirus/genética , Infecciones por Coronavirus/genética , Infecciones por Coronavirus/virología , Codón/genética , Uso de Codones , Biología Computacional/métodos , Coronavirus/genética , Infecciones por Coronavirus/metabolismo , Genes Virales , Genoma Viral , Humanos , Coronavirus del Síndrome Respiratorio de Oriente Medio/genética , Filogenia , Coronavirus Relacionado al Síndrome Respiratorio Agudo Severo/genética , SARS-CoV-2/genética

RESUMEN

PURPOSE: To determine the presence of a group of mutations, and establish the prognostic value for recurrence and progression. MATERIALS AND METHODS: Prospective observational study. Intermediate-to-high-risk non-muscle invasive bladder cancer (NMIBC) was evaluated. Data from genetic analyses were included in a database along with clinicopathological variables of interest. RESULTS: Seventy-four patients. Twenty-five (33.8%) recurred and 3 (4.1%) progressed. Median time to recurrence: 8 months (5.7-12.7). Median time to progression: 14 months (P75: 12). Mutation distribution: KRAS codon 12: one patient (1.4%), BAT25: five patients (6.8%), BAT-26: four patients (5.4%), and D2S123: 6 patients (8.1%). Arg72Pro polymorphism: 50 patients (67.6%) exhibited homozygous mutations, 23 (31.1%) were heterozygous, and 1 patient (1.4%) did not present the mutation. We found an association between presence of MSI at BAT26 and female sex (p < 0.05) and tumor stage and the TP53 Arg72Pro polymorphism. Recurrence-free survival (RFS) was significantly associated with presence of MSI at D2S123, with a HR of 5.44 for patients presenting the mutation (95% CI 1.83-16.16). On multivariate analysis, we found a statistically significant increase in risk of recurrence among patients with MSI at D2S123 (HR 5.15; p < 0.05) and more than 2 previous transurethral bladder resections (TURBs) (HR 5.07; p < 0.05) adjusted for tumor stage and grade. Harrell's concordance index revealed an accuracy of 0.74 (p < 0.05). CONCLUSION: An association was found between presence BAT26 MSI and female sex, Arg72Pro polymorphism with tumor stage and D2S123 and more than 2 TUR procedures were associated with RFS adjusted to tumor stage and grade.

Asunto(s)

Biomarcadores de Tumor/genética , Progresión de la Enfermedad , Recurrencia Local de Neoplasia/genética , Mutación Puntual , Neoplasias de la Vejiga Urinaria/genética , Anciano , Análisis de Varianza , Codón/genética , Dipéptidos/genética , Femenino , Genes p53/genética , Genes ras , Marcadores Genéticos/genética , Humanos , Masculino , Inestabilidad de Microsatélites , Repeticiones de Microsatélite/genética , Pronóstico , Estudios Prospectivos , Factores Sexuales , Factores de Tiempo , Neoplasias de la Vejiga Urinaria/patología , Neoplasias de la Vejiga Urinaria/cirugía

RESUMEN

Circumsporozoite protein (CSP) is the primary pre-erythrocytic vaccine target in Plasmodium species. Knowledge about their genetic diversity can help predict vaccine efficacy and the spread of novel parasite variants. Thus, we investigated pvcsp gene polymorphisms in 219 isolates (136 from Brazilian Amazon [BA], 71 from Rio de Janeiro Atlantic Forest [AF], and 12 from non-Brazilian countries [NB]). Forty-eight polymorphic sites were detected, 46 in the central repeat region (CR), and two in the C-terminal region. Also, the CR presents InDels and a variable number of repeats. All samples correspond to the VK210 variant, and 24 VK210 subtypes based on CR. Nucleotide diversity (π = 0.0135) generated a significant number of haplotypes (168) with low genetic differentiation between the Brazilian regions (Fst = 0.208). The haplotype network revealed similar distances among the BA and AF regions. The linkage disequilibrium indicates that recombination does not seem to be acting in diversity, reinforcing natural selection's role in accelerating adaptive evolution. The high diversity (low Fst) and polymorphism frequencies could be indicators of balancing selection. Although malaria in BA and AF have distinct vector species and different host immune pressures, consistent genetic signature was found in two regions. The immunodominant B-cell epitope mapped in the CR varies from seven to 19 repeats. The CR T-cell epitope is conserved only in 39 samples. Concerning to C-terminal region, the Th2R epitope presented nonsynonymous SNP only in 6% of Brazilian samples, and the Th3R epitope remained conserved in all studied regions. We conclude that, although the uneven distribution of alleles may jeopardize the deployment of vaccines directed to a specific variable locus, a unique vaccine formulation could protect populations in all Brazilian regions.

Asunto(s)

Variación Genética , Parásitos/genética , Plasmodium vivax/genética , Proteínas Protozoarias/genética , Selección Genética , Secuencia de Aminoácidos , Sustitución de Aminoácidos , Animales , Océano Atlántico , Brasil , Codón/genética , Epítopos de Linfocito B/química , Epítopos de Linfocito B/genética , Epítopos de Linfocito T/química , Epítopos de Linfocito T/genética , Geografía , Haplotipos/genética , Mutación INDEL/genética , Desequilibrio de Ligamiento/genética , Nucleótidos/genética , Péptidos/química , Filogenia , Plasmodium vivax/aislamiento & purificación , Polimorfismo Genético , Proteínas Protozoarias/química

RESUMEN

The R337H is a TP53 germline pathogenic variant that has been associated with several types of cancers, including breast cancer. Our main objective was to determine the frequency of the R337H variant in sporadic breast cancer patients from Paraná state, South Brazil, its association with prognosis and its impact in genomic instability. The genotyping of 805 breast cancer tissues revealed a genotypic and allelic frequency of the R337H variant of 2.36% and 1.18%, respectively. In these R337H+ cases a lower mean age at diagnosis was observed when compared to the R337H-cases. Array-CGH analysis showed that R337H+ patients presented a higher number of copy number alterations (CNAs), compared to the R337H-. These CNAs affected genes and miRNAs that regulate critical cancer signaling pathways; a number of these genes were associated with survival after querying the KMplot database. Furthermore, homozygous (R337H+/R337H+) fibroblasts presented increased levels of copy number variants when compared to heterozygous or R337H- cells. In conclusion, the R337H variant may contribute to 2.36% of the breast cancer cases without family cancer history in Paraná. Among other mechanisms, R337H increases the level of genomic instability, as evidenced by a higher number of CNAs in the R337H+ cases compared to the R337H-.

Asunto(s)

Neoplasias de la Mama/genética , Inestabilidad Genómica/genética , Mutación de Línea Germinal/genética , Proteína p53 Supresora de Tumor/genética , Factores de Edad , Anciano , Brasil , Neoplasias de la Mama/mortalidad , Codón/genética , Exones/genética , Femenino , Dosificación de Gen/genética , Frecuencia de los Genes , Humanos , Persona de Mediana Edad , Tasa de Supervivencia

RESUMEN

Sea turtles are the only extant chelonian representatives that inhabit the marine environment. One key to successful colonization of this habitat is the adaptation to different energetic demands. Such energetic requirement is intrinsically related to the mitochondrial ability to generate energy through oxidative phosphorylation (OXPHOS) process. Here, we estimated Testudines phylogenetic relationships from 90 complete chelonian mitochondrial genomes and tested the adaptive evolution of 13 mitochondrial protein-coding genes of sea turtles to determine how natural selection shaped mitochondrial genes of the Chelonioidea clade. Complete mitogenomes showed strong support and resolution, differing at the position of the Chelonioidea clade in comparison to the turtle phylogeny based on nuclear genomic data. Codon models retrieved a relatively increased dN/dS (ω) on three OXPHOS genes for sea turtle lineages. Also, we found evidence of positive selection on at least three codon positions, encoded by NADH dehydrogenase genes (ND4 and ND5). The accelerated evolutionary rates found for sea turtles on COX2, ND1 and CYTB and the molecular footprints of positive selection found on ND4 and ND5 genes may be related to mitochondrial molecular adaptation to stress likely resulted from a more active lifestyle in sea turtles. Our study provides insight into the adaptive evolution of the mtDNA genome in sea turtles and its implications for the molecular mechanism of oxidative phosphorylation.

Asunto(s)

ADN Mitocondrial/genética , Ecosistema , Evolución Molecular , Genoma Mitocondrial/genética , Proteínas Mitocondriales/genética , Océanos y Mares , Filogenia , Selección Genética/genética , Tortugas/genética , Adaptación Fisiológica/genética , Animales , Codón/genética , Ciclooxigenasa 2 , Metabolismo Energético/genética , NADH Deshidrogenasa/genética , Fosforilación Oxidativa

RESUMEN

Prokaryote genomes exhibit a wide range of GC contents and codon usages, both resulting from an interaction between mutational bias and natural selection. In order to investigate the basis underlying specific codon changes, we performed a comprehensive analysis of 29 different prokaryote families. The analysis of core gene sets with increasing ancestries in each family lineage revealed that the codon usages became progressively more adapted to the tRNA pools. While, as previously reported, highly expressed genes presented the most optimized codon usage, the singletons contained the less selectively favored codons. The results showed that usually codons with the highest translational adaptation were preferentially enriched. In agreement with previous reports, a C bias in 2- to 3-fold pyrimidine-ending codons, and a U bias in 4-fold codons occurred in all families, irrespective of the global genomic GC content. Furthermore, the U biases suggested that U3-mRNA-U34-tRNA interactions were responsible for a prominent codon optimization in both the most ancestral core and the highly expressed genes. A comparative analysis of sequences that encode conserved (cr) or variable (vr) translated products, with each one being under high (HEP) and low (LEP) expression levels, demonstrated that the efficiency was more relevant (by a factor of 2) than accuracy to modeling codon usage. Finally, analysis of the third position of codons (GC3) revealed that in genomes with global GC contents higher than 35 to 40%, selection favored a GC3 increase, whereas in genomes with very low GC contents, a decrease in GC3 occurred. A comprehensive final model is presented in which all patterns of codon usage variations are condensed in four distinct behavioral groups.IMPORTANCE The prokaryotic genomes-the current heritage of the most ancient life forms on earth-are comprised of diverse gene sets, all characterized by varied origins, ancestries, and spatial-temporal expression patterns. Such genetic diversity has for a long time raised the question of how cells shape their coding strategies to optimize protein demands (i.e., product abundance) and accuracy (i.e., translation fidelity) through the use of the same genetic code in genomes with GC contents that range from less than 20 to more than 80%. Here, we present evidence on how codon usage is adjusted in the prokaryotic tree of life and on how specific biases have operated to improve translation. Through the use of proteome data, we characterized conserved and variable sequence domains in genes of either high or low expression level and quantitated the relative weight of efficiency and accuracy-as well as their interaction-in shaping codon usage in prokaryotes.

Asunto(s)

Archaea/genética , Bacterias/genética , Uso de Codones , Codón/genética , Código Genético , ARN de Transferencia/genética , Archaea/clasificación , Bacterias/clasificación , Composición de Base , Mutación , Biosíntesis de Proteínas , Proteoma

RESUMEN

To evaluate the impact of hypermutation on the HIV-1 dissemination at the population level we studied 7072 sequences HIV-1 gene vif retrieved from the public databank. From this dataset 854 sequences were selected because they had associated values of CD4+ T lymphocytes counts and viral loads and they were used to assess the correlation between clinical parameters and hypermutation. We found that the frequency of stop codons at sites 5, 11 and 79 ranged from 2.8x10-4 to 4.2x10-4. On the other hand, at codons 21, 38, 70, 89 and 174 the frequency of stop codons ranged from 1.4x10-3 to 2.5x10-3. We also found a correlation between clinical parameters and hypermutation where patients harboring proviruses with one or more stop codons at the tryptophan sites of the gene vif had higher CD4+ T lymphocytes counts and lower viral loads compared to the population. Our findings indicate that A3 activity potentially restrains HIV-1 replication because individuals with hypermutated proviruses tend to have lower numbers of RNA copies. However, owing to the low frequency of hypermutated sequences observed in the databank (44 out of 7072), it is unlikely that A3 has a significant impact to curb HIV-1 dissemination at the population level.

Asunto(s)

Codón/genética , VIH-1/genética , Triptófano , Productos del Gen vif del Virus de la Inmunodeficiencia Humana/genética , Recuento de Linfocito CD4 , Codón de Terminación/genética , VIH-1/fisiología , Mutación , Carga Viral/genética

RESUMEN

Recombinant protein expression is widely used to produce large quantities of protein for diverse uses including functional characterization of selected sequences and vaccination trials. In the postgenomic era, high-throughput techniques that allow us to manipulate several sequences are needed. Cloning by in vivo recombination is a technique that consists in the insertion of a linear DNA into a linearized plasmid DNA by in vivo recombination using a recA+ E. coli strain. This methodology provides high-throughput cloning with high efficiency without the need for restriction enzyme digestion. In this chapter, we describe two protocols for DNA cloning: one using in vivo recombination and the other by using restriction enzymes. We also describe the application of different conditions to produce functional proteins that needs the incorporation of the amino acid selenocysteine (Sec), like thioredoxin-glutathione reductase enzyme.

Asunto(s)

Codón/genética , Escherichia coli/genética , Proteínas Recombinantes/genética , Clonación Molecular/métodos , ADN/genética , Enzimas de Restricción del ADN/genética , Plásmidos/genética , Selenocisteína/genética

RESUMEN

PURPOSE: Factors influencing fallopian tube occlusion in women with a lower genital tract infection remain incompletely elucidated. We evaluated whether a polymorphism in the mannose-binding lectin (MBL) gene at codon 54 influences the occurrence of fallopian tube blockage in relation to exposure to Chlamydia trachomatis. METHODS: In a case-control study at The Hospital das Clínicas, University of São Paulo, Brazil, 75 women with hysterosalpingography-documented tubal occlusion and 75 women with patent fallopian tubes were analyzed for detection of single-nucleotide polymorphism in codon 54 of the MBL gene and for IgG anti-C. trachomatis antibodies in their sera. Both groups were matched for age, race, and sexual variables. RESULTS: Prior exposure to C. trachomatis, as evidenced by the presence of IgG antibodies, was comparable in both groups. Detection of the polymorphic MBL allele was more prevalent in women with blocked tubes (p < 0.01), regardless of whether or not there was evidence of prior chlamydial exposure. CONCLUSION: The level of MBL-related innate immunity influences the consequences of infection by C. trachomatis or other microbes.

Asunto(s)

Infecciones por Chlamydia/genética , Chlamydia trachomatis/aislamiento & purificación , Enfermedades de las Trompas Uterinas/diagnóstico por imagen , Trompas Uterinas/diagnóstico por imagen , Infertilidad Femenina/diagnóstico por imagen , Infertilidad Femenina/genética , Lectina de Unión a Manosa/genética , Adulto , Brasil , Estudios de Casos y Controles , Infecciones por Chlamydia/microbiología , Chlamydia trachomatis/genética , Chlamydia trachomatis/inmunología , Codón/genética , Enfermedades de las Trompas Uterinas/microbiología , Trompas Uterinas/microbiología , Femenino , Predisposición Genética a la Enfermedad , Humanos , Histerosalpingografía , Inmunoglobulina G/sangre , Infertilidad Femenina/microbiología , Polimorfismo Genético , Polimorfismo de Nucleótido Simple

RESUMEN

Mycobacteriophages are viruses -mostly temperates- that infect Mycobacterium smegmatis and sometimes Mycobacterium tuberculosis. Mycobacteriophages are grouped in clusters on the basis of the overall nucleotide sequence homology, being further divided in subclusters as more mycobacteriophage genomes are sequenced and annotated. As part of our on-going screening for novel isolates, we herein report the bioinformatics analysis of CRB2, a mycobacteriophage belonging into the Siphoviridae family that propagates at 30°C. CRB2 has a 72,217 bp genome with a 69.78% GC content that belongs to Cluster B; nucleotide comparison with other B cluster members positions CRB2 as the sole member of a new subcluster, B9, being mycobacteriophage Saguaro (belonging into subcluster B7) its closest relative. Sequencing and annotation of 14 mycobacteriophages isolated by our group has yielded six cluster A members, a singleton, four of the five members of subcluster B6, one of the three reported members of subcluster G4, and CRB2 which defines subcluster B9. Considering the massive mycobacteriophage search performed in USA and the relatively rarity of our phages, we propose that factors other than size of the sampling determine the variability of mycobacteriophage distribution, and thus a world-wide concerted mining would most likely bring extremely rare and yet undiscovered mycobacteriophages.

Asunto(s)

Biodiversidad , Codón/genética , ADN Viral/genética , Genoma Viral , Micobacteriófagos/clasificación , Micobacteriófagos/genética , Mycobacterium tuberculosis/virología , Micobacteriófagos/aislamiento & purificación , Filogenia

RESUMEN

Decades of unmanaged insecticide use and routine exposure to agrochemicals have left many populations of malaria vectors in the Americas resistant to multiple classes of insecticides, including pyrethroids. The molecular basis of pyrethroid resistance is relatively uncharacterised in American malaria vectors, preventing the design of suitable resistance management strategies. Using whole transcriptome sequencing, we characterized the mechanisms of pyrethroid resistance in Anopheles albimanus from Peru and Guatemala. An. albimanus were phenotyped as either deltamethrin or alpha-cypermethrin resistant. RNA from 1) resistant, 2) unexposed, and 3) a susceptible laboratory strain of An. albimanus was sequenced and analyzed using RNA-Seq. Expression profiles of the three groups were compared based on the current annotation of the An. albimanus reference genome. Several candidate genes associated with pyrethroid resistance in other malaria vectors were found to be overexpressed in resistant An. albimanus. In addition, gene ontology terms related to serine-type endopeptidase activity, extracellular activity and chitin metabolic process were also commonly overexpressed in the field caught resistant and unexposed samples from both Peru and Guatemala when compared to the susceptible strain. The cytochrome P450 CYP9K1 was overexpressed 14x in deltamethrin and 8x in alpha-cypermethrin-resistant samples from Peru and 2x in deltamethrin-resistant samples from Guatemala, relative to the susceptible laboratory strain. CYP6P5 was overexpressed 68x in deltamethrin-resistant samples from Peru but not in deltamethrin-resistant samples from Guatemala. When comparing overexpressed genes between deltamethrin-resistant and alpha-cypermethrin-resistant samples from Peru, a single P450 gene, CYP4C26, was overexpressed 9.8x (p<0.05) in alpha-cypermethrin-resistant samples. In Peruvian deltamethrin-resistant samples, the knockdown resistance mutation (kdr) variant alleles at position 1014 were rare, with approximately 5% frequency, but in the alpha-cypermethrin-resistant samples, the frequency of these alleles was approximately 15-30%. Functional validation of the candidate genes and the kdr mutation as a resistance marker for alpha-cypermethrin will confirm the role of these mechanisms in conferring pyrethroid resistance.

Asunto(s)

Anopheles/genética , Perfilación de la Expresión Génica , Resistencia a los Insecticidas/genética , Malaria/parasitología , Mosquitos Vectores/genética , Piretrinas/toxicidad , Animales , Anopheles/efectos de los fármacos , Codón/genética , Complejo IV de Transporte de Electrones/genética , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Genes de Insecto , Geografía , Guatemala , Haplotipos/genética , Inactivación Metabólica/efectos de los fármacos , Resistencia a los Insecticidas/efectos de los fármacos , Mosquitos Vectores/efectos de los fármacos , Mutación/genética , Nitrilos/toxicidad , Perú , Reproducibilidad de los Resultados , Análisis de Secuencia de ARN

RESUMEN

The codon stabilization coefficient (CSC) is derived from the correlation between each codon frequency in transcripts and mRNA half-life experimental data. In this work, we used this metric as a reference to compare previously published Saccharomyces cerevisiae mRNA half-life datasets and investigate how codon composition related to protein levels. We generated CSCs derived from nine studies. Four datasets produced similar CSCs, which also correlated with other independent parameters that reflected codon optimality, such as the tRNA abundance and ribosome residence time. By calculating the average CSC for each gene, we found that most mRNAs tended to have more non-optimal codons. Conversely, a high proportion of optimal codons was found for genes coding highly abundant proteins, including proteins that were only transiently overexpressed in response to stress conditions. We also used CSCs to identify and locate mRNA regions enriched in non-optimal codons. We found that these stretches were usually located close to the initiation codon and were sufficient to slow ribosome movement. However, in contrast to observations from reporter systems, we found no position-dependent effect on the mRNA half-life. These analyses underscore the value of CSCs in studies of mRNA stability and codon bias and their relationships with protein expression.

Asunto(s)

Codón/genética , Biosíntesis de Proteínas , Estabilidad del ARN , ARN Mensajero/genética , ARN Mensajero/metabolismo , Saccharomyces cerevisiae/genética , Secuencia de Bases , Conjuntos de Datos como Asunto , Genes Fúngicos/genética , Genoma Fúngico/genética , Semivida , Biosíntesis de Proteínas/genética , Ribosomas/metabolismo , Proteínas de Saccharomyces cerevisiae/análisis , Proteínas de Saccharomyces cerevisiae/genética

RESUMEN

The aim of this study was to characterize the most frequent mutations associated with rifampicin (RIF) and isoniazid (INH) resistance of Mycobacterium tuberculosis isolates from Ecuador. Sequence analysis of 40 strains, resistant for the tuberculosis drugs INH, RIF, or for both showed that of the 31 strains with resistance to INH, 20 strains (64.5%) carried a mutation in the katG gene (codon 315). Eight INH-resistant strains carried a mutation in the katG gene at codon 463. This katG463 mutation, considered a phylogenetic marker, was exclusively found in INH-resistant strains and not in 121 INH-susceptible strains. Of the 35 strains resistant to RIF, 33 (93.9%) had mutations in the hot spot region of the rpoB gene, predominantly in codons 531, 516, and 526. Our results show that sequence-based detection for drug resistance of the katG will identify, respectively, 64.5% or, considering katG463 as a marker, 90.3% of the INH-resistant strains. Sequencing of the hot spot region of the rpoB gene will detect 94.3% of the RIF drug-resistant isolates in Ecuador. This is appropriate for fast screening for drug resistance with the GeneXpert MTB/RIF assay or by direct sequencing of a part of the genes katG and rpoB of PCR products obtained from DNA isolation from primary cultures.

Asunto(s)

Proteínas Bacterianas/genética , Farmacorresistencia Bacteriana/genética , Mutación/genética , Mycobacterium tuberculosis/genética , Tuberculosis Resistente a Múltiples Medicamentos/genética , Antituberculosos/farmacología , Codón/genética , ADN Bacteriano/genética , Ecuador , Humanos , Pruebas de Sensibilidad Microbiana/métodos , Tasa de Mutación , Mycobacterium tuberculosis/efectos de los fármacos , Filogenia , Tuberculosis Resistente a Múltiples Medicamentos/tratamiento farmacológico

RESUMEN

The definition of a genomic signature (GS) is "the total net response to selective pressure". Recent isolation and sequencing of naturally occurring organisms, hereby named entoorganisms, within Acanthamoeba polyphaga, raised the hypothesis of a common genomic signature despite their diverse and unrelated evolutionary origin. Widely accepted and implemented tests for GS detection are oligonucleotide relative frequencies (OnRF) and relative codon usage (RCU) surveys. A common pattern and strong correlations were unveiled from OnRFs among A. polyphaga's Mimivirus and virophage Sputnik. RCU showed a common A-T bias at third codon position. We expanded tests to the amoebal mitochondrial genome and amoeba-resistant bacteria, achieving strikingly coherent results to the aforementioned viral analyses. The GSs in these entoorganisms of diverse evolutionary origin are coevolutionarily conserved within an intracellular environment that provides sanctuary for species of ecological and biomedical relevance.

Asunto(s)

Acanthamoeba/genética , Coevolución Biológica/genética , Mimiviridae/genética , Amoeba/genética , Animales , Bacterias/genética , Codón/genética , Evolución Molecular , Genoma Viral , Genómica , Mitocondrias/genética , Parásitos/genética , Proteínas Virales/genética , Virófagos/genética

RESUMEN

BACKGROUND: Small RNAs (sRNAs) are key regulators of gene expression in bacteria. In addition to modulating translation initiation, sRNAs can interact with mRNA coding regions to regulate mRNA stability and translation efficiency, enhancing or impeding progression of the ribosome along the mRNA. Since most amino acids are decoded by more than one codon (synonymous) we asked as to whether there is a codon bias in the interaction of sRNAs with coding regions of mRNAs. Therefore, we explored whether there are differences in codon usage or tRNA availability according to whether an mRNA is regulated by sRNAs or not. We also explored these parameters in the coding interaction regions in mRNAs. We focused our analysis on sRNAs that regulate multiple mRNAs. RESULTS: We found differences in codon adaptation index and tRNA adaptation index between sRNA-regulated and non-sRNA-regulated mRNAs. Interestingly, the sRNA-mRNA interacting regions tended to be enriched in unpreferred codons decoded by scarce tRNAs. We also found that sRNAs with multiple targets often contained modular segments capable of recognizing conserved motifs among these mRNAs. CONCLUSIONS: Our results show that sRNAs in E. coli tend to recognize mRNA coding regions in which the ribosome is predicted to advance at low speeds. Identified motifs in interacting regions are conserved among mRNAs that are recognized by the same sRNA.

Asunto(s)

Codón/genética , Escherichia coli/genética , ARN Bacteriano/genética , ARN Pequeño no Traducido/genética , ARN Mensajero/genética , Análisis de Secuencia de ARN