RESUMO
The killifish Nothobranchius furzeri is the shortest-lived vertebrate that can be bred in the laboratory. Its rapid growth, early sexual maturation, fast aging, and arrested embryonic development (diapause) make it an attractive model organism in biomedical research. Here, we report a draft sequence of its genome that allowed us to uncover an intra-species Y chromosome polymorphism representing-in real time-different stages of sex chromosome formation that display features of early mammalian XY evolution "in action." Our data suggest that gdf6Y, encoding a TGF-ß family growth factor, is the master sex-determining gene in N. furzeri. Moreover, we observed genomic clustering of aging-related genes, identified genes under positive selection, and revealed significant similarities of gene expression profiles between diapause and aging, particularly for genes controlling cell cycle and translation. The annotated genome sequence is provided as an online resource (http://www.nothobranchius.info/NFINgb).
Assuntos
Evolução Biológica , Peixes Listrados/genética , Cromossomos Sexuais , Envelhecimento , Animais , Feminino , Genoma , Peixes Listrados/fisiologia , Masculino , Dados de Sequência Molecular , Processos de Determinação SexualRESUMO
The use of alternative translation initiation sites enables production of more than one protein from a single gene, thereby expanding the cellular proteome. Although several such examples have been serendipitously found in bacteria, genome-wide mapping of alternative translation start sites has been unattainable. We found that the antibiotic retapamulin specifically arrests initiating ribosomes at start codons of the genes. Retapamulin-enhanced Ribo-seq analysis (Ribo-RET) not only allowed mapping of conventional initiation sites at the beginning of the genes, but strikingly, it also revealed putative internal start sites in a number of Escherichia coli genes. Experiments demonstrated that the internal start codons can be recognized by the ribosomes and direct translation initiation in vitro and in vivo. Proteins, whose synthesis is initiated at internal in-frame and out-of-frame start sites, can be functionally important and contribute to the "alternative" bacterial proteome. The internal start sites may also play regulatory roles in gene expression.
Assuntos
Genoma Bacteriano/genética , Iniciação Traducional da Cadeia Peptídica , Proteoma/genética , Proteômica , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Códon de Iniciação/genética , Diterpenos/farmacologia , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Regulação Bacteriana da Expressão Gênica/efeitos dos fármacos , Genoma Bacteriano/efeitos dos fármacos , RNA Mensageiro/genética , Ribossomos/efeitos dos fármacos , Ribossomos/genéticaRESUMO
Resting conventional T cells (Tconv) can be distinguished from T regulatory cells (Treg) by the canonical markers FOXP3, CD25 and CD127. However, the expression of these proteins alters after T-cell activation leading to overlap between Tconv and Treg. The objective of this study was to distinguish resting and antigen-responsive T effector (Tconv) and Treg using single-cell technologies. CD4+ Treg and Tconv cells were stimulated with antigen and responsive and non-responsive populations processed for targeted and non-targeted single-cell RNAseq. Machine learning was used to generate a limited set of genes that could distinguish responding and non-responding Treg and Tconv cells and which was used for single-cell multiplex qPCR and to design a flow cytometry panel. Targeted scRNAseq clearly distinguished the four-cell populations. A minimal set of 27 genes was identified by machine learning algorithms to provide discrimination of the four populations at >95% accuracy. In all, 15 of the genes were validated to be differentially expressed by single-cell multiplex qPCR. Discrimination of responding Treg from responding Tconv could be achieved by a flow cytometry strategy that included staining for CD25, CD127, FOXP3, IKZF2, ITGA4, and the novel marker TRIM which was strongly expressed in Tconv and weakly expressed in both responding and non-responding Treg. A minimal set of genes was identified that discriminates responding and non-responding CD4+ Treg and Tconv cells and, which have identified TRIM as a marker to distinguish Treg by flow cytometry.
Assuntos
Ativação Linfocitária , Linfócitos T Reguladores , Biomarcadores/metabolismo , Citometria de Fluxo , Fatores de Transcrição Forkhead/metabolismo , Contagem de LinfócitosRESUMO
The repeated evolution of dietary specialization represents a hallmark of mammalian ecology. To detect genomic changes that are associated with dietary adaptations, we performed a systematic screen for convergent gene losses associated with an obligate herbivorous or carnivorous diet in 31 placental mammals. For herbivores, our screen discovered the repeated loss of the triglyceride lipase inhibitor PNLIPRP1, suggesting enhanced triglyceride digestion efficiency. Furthermore, several herbivores lost the pancreatic exocytosis factor SYCN, providing an explanation for continuous pancreatic zymogen secretion in these species. For carnivores, we discovered the repeated loss of the hormone-receptor pair INSL5-RXFP4 that regulates appetite and glucose homeostasis, which likely relates to irregular feeding patterns and constant gluconeogenesis. Furthermore, reflecting the reduced need to metabolize plant-derived xenobiotics, several carnivores lost the xenobiotic receptors NR1I3 and NR1I2 Finally, the carnivore-associated loss of the gastrointestinal host defense gene NOX1 could be related to a reduced gut microbiome diversity. By revealing convergent gene losses associated with differences in dietary composition, feeding patterns, and gut microbiomes, our study contributes to understanding how similar dietary specializations evolved repeatedly in mammals.
Assuntos
Carnivoridade/fisiologia , Trato Gastrointestinal/microbiologia , Herbivoria/genética , Filogenia , Animais , Dieta , Feminino , Microbioma Gastrointestinal/genética , Trato Gastrointestinal/metabolismo , Genoma/genética , Herbivoria/fisiologia , Mamíferos/microbiologia , Plantas , Gravidez , RNA Ribossômico 16S/genéticaRESUMO
Toll-like receptors (TLRs) play an important role for the innate immune system by detecting pathogen-associated molecular patterns. TLR5 encodes the major extracellular receptor for bacterial flagellin and frequently evolves under positive selection, consistent with coevolutionary arms races between the host and pathogens. Furthermore, TLR5 is inactivated in several vertebrates and a TLR5 stop codon polymorphism is widespread in human populations. Here, we analyzed the genomes of 120 mammals and discovered that TLR5 is convergently lost in four independent lineages, comprising guinea pigs, Yangtze river dolphin, pinnipeds, and pangolins. Validated inactivating mutations, absence of protein-coding transcript expression, and relaxed selection on the TLR5 remnants confirm these losses. PCR analysis further confirmed the loss of TLR5 in the pinniped stem lineage. Finally, we show that TLR11, encoding a second extracellular flagellin receptor, is also absent in these four lineages. Independent losses of TLR5 and TLR11 suggest that a major pathway for detecting flagellated bacteria is not essential for different mammals and predicts an impaired capacity to sense extracellular flagellin.
Assuntos
Evolução Biológica , Flagelina/imunologia , Mamíferos/genética , Receptor 5 Toll-Like/genética , Animais , Genoma , Cobaias , Humanos , CoelhosRESUMO
Descent of testes from a position near the kidneys into the lower abdomen or into the scrotum is an important developmental process that occurs in all placental mammals, with the exception of five afrotherian lineages. Since soft-tissue structures like testes are not preserved in the fossil record and since key parts of the placental mammal phylogeny remain controversial, it has been debated whether testicular descent is the ancestral or derived condition in placental mammals. To resolve this debate, we used genomic data of 71 mammalian species and analyzed the evolution of two key genes (relaxin/insulin-like family peptide receptor 2 [RXFP2] and insulin-like 3 [INSL3]) that induce the development of the gubernaculum, the ligament that is crucial for testicular descent. We show that both RXFP2 and INSL3 are lost or nonfunctional exclusively in four afrotherians (tenrec, cape elephant shrew, cape golden mole, and manatee) that completely lack testicular descent. The presence of remnants of once functional orthologs of both genes in these afrotherian species shows that these gene losses happened after the split from the placental mammal ancestor. These "molecular vestiges" provide strong evidence that testicular descent is the ancestral condition, irrespective of persisting phylogenetic discrepancies. Furthermore, the absence of shared gene-inactivating mutations and our estimates that the loss of RXFP2 happened at different time points strongly suggest that testicular descent was lost independently in Afrotheria. Our results provide a molecular mechanism that explains the loss of testicular descent in afrotherians and, more generally, highlight how molecular vestiges can provide insights into the evolution of soft-tissue characters.
Assuntos
Eutérios/embriologia , Eutérios/genética , Evolução Molecular , Insulina/genética , Proteínas/genética , Receptores Acoplados a Proteínas G/genética , Testículo/embriologia , Sequência de Aminoácidos , Animais , Sequência de Bases , DNA/genética , Eutérios/classificação , Gubernáculo/crescimento & desenvolvimento , Masculino , Mutação , Filogenia , Talidomida/análogos & derivadosRESUMO
Genome alignments provide a powerful basis to transfer gene annotations from a well-annotated reference genome to many other aligned genomes. The completeness of these annotations crucially depends on the sensitivity of the underlying genome alignment. Here, we investigated the impact of the genome alignment parameters and found that parameters with a higher sensitivity allow the detection of thousands of novel alignments between orthologous exons that have been missed before. In particular, comparisons between species separated by an evolutionary distance of >0.75 substitutions per neutral site, like human and other non-placental vertebrates, benefit from increased sensitivity. To systematically test if increased sensitivity improves comparative gene annotations, we built a multiple alignment of 144 vertebrate genomes and used this alignment to map human genes to the other 143 vertebrates with CESAR. We found that higher alignment sensitivity substantially improves the completeness of comparative gene annotations by adding on average 2382 and 7440 novel exons and 117 and 317 novel genes for mammalian and non-mammalian species, respectively. Our results suggest a more sensitive alignment strategy that should generally be used for genome alignments between distantly-related species. Our 144-vertebrate genome alignment and the comparative gene annotations (https://bds.mpi-cbg.de/hillerlab/144VertebrateAlignment_CESAR/) are a valuable resource for comparative genomics.
Assuntos
Biologia Computacional/métodos , Éxons/genética , Genoma/genética , Anotação de Sequência Molecular/métodos , Alinhamento de Sequência/métodos , Animais , Sequência de Bases , Genômica/métodos , Humanos , Internet , Reprodutibilidade dos Testes , Homologia de Sequência do Ácido Nucleico , Especificidade da Espécie , Vertebrados/classificação , Vertebrados/genéticaRESUMO
MOTIVATION: Homology-based gene prediction is a powerful concept to annotate newly sequenced genomes. We have previously demonstrated that whole genome alignments can be utilized for accurate comparative coding gene annotation. RESULTS: Here we present CESAR 2.0 that utilizes genome alignments to transfer coding gene annotations from one reference to many other aligned genomes. We show that CESAR 2.0 is 77 times faster and requires 31 times less memory compared to its predecessor. CESAR 2.0 substantially improves the ability to align splice sites that have shifted over larger distances, allowing for precise identification of the exon boundaries in the aligned genome. Finally, CESAR 2.0 supports entire genes, which enables the annotation of joined exons that arose by complete intron deletions. CESAR 2.0 can readily be applied to new genome alignments to annotate coding genes in many other genomes at improved accuracy and without necessitating large-computational resources. AVAILABILITY AND IMPLEMENTATION: Source code is freely available at https://github.com/hillerlab/CESAR2.0. CONTACT: hiller@mpi-cbg.de. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
Assuntos
Biologia Computacional/métodos , Anotação de Sequência Molecular , Software , Éxons , Genoma , Genômica , Íntrons , Alinhamento de SequênciaRESUMO
Identifying coding genes is an essential step in genome annotation. Here, we utilize existing whole genome alignments to detect conserved coding exons and then map gene annotations from one genome to many aligned genomes. We show that genome alignments contain thousands of spurious frameshifts and splice site mutations in exons that are truly conserved. To overcome these limitations, we have developed CESAR (Coding Exon-Structure Aware Realigner) that realigns coding exons, while considering reading frame and splice sites of each exon. CESAR effectively avoids spurious frameshifts in conserved genes and detects 91% of shifted splice sites. This results in the identification of thousands of additional conserved exons and 99% of the exons that lack inactivating mutations match real exons. Finally, to demonstrate the potential of using CESAR for comparative gene annotation, we applied it to 188 788 exons of 19 865 human genes to annotate human genes in 99 other vertebrates. These comparative gene annotations are available as a resource (http://bds.mpi-cbg.de/hillerlab/CESAR/). CESAR (https://github.com/hillerlab/CESAR/) can readily be applied to other alignments to accurately annotate coding genes in many other vertebrate and invertebrate genomes.
Assuntos
Éxons , Genoma , Genômica/métodos , Anotação de Sequência Molecular , Software , Animais , Bovinos , Códon , Biologia Computacional/métodos , Cães , Evolução Molecular , Humanos , Íntrons , Camundongos , Mutação , Fases de Leitura Aberta , Filogenia , Sítios de Splice de RNA , Ratos , Fases de Leitura , Reprodutibilidade dos Testes , NavegadorRESUMO
Escherichia coli and yeast DNA-dependent RNA polymerases are shown to mediate efficient nascent transcript stem loop formation-dependent RNA-DNA hybrid realignment. The realignment was discovered on the heteropolymeric sequence T5C5 and yields transcripts lacking a C residue within a corresponding U5C4. The sequence studied is derived from a Roseiflexus insertion sequence (IS) element where the resulting transcriptional slippage is required for transposase synthesis. The stability of the RNA structure, the proximity of the stem loop to the slippage site, the length and composition of the slippage site motif, and the identity of its 3' adjacent nucleotides (nt) are crucial for transcripts lacking a single C. In many respects, the RNA structure requirements for this slippage resemble those for hairpin-dependent transcription termination. In a purified in vitro system, the slippage efficiency ranges from 5% to 75% depending on the concentration ratios of the nucleotides specified by the slippage sequence and the 3' nt context. The only previous proposal of stem loop mediated slippage, which was in Ebola virus expression, was based on incorrect data interpretation. We propose a mechanical slippage model involving the RNAP translocation state as the main motor in slippage directionality and efficiency. It is distinct from previously described models, including the one proposed for paramyxovirus, where following random movement efficiency is mainly dependent on the stability of the new realigned hybrid. In broadening the scope for utilization of transcription slippage for gene expression, the stimulatory structure provides parallels with programmed ribosomal frameshifting at the translation level.
Assuntos
Conformação de Ácido Nucleico , RNA Mensageiro/química , Regiões Terminadoras Genéticas , Transcrição Gênica , Sequência de Aminoácidos , Sequência de Bases , Chloroflexi/genética , RNA Polimerases Dirigidas por DNA/metabolismo , Dados de Sequência Molecular , Motivos de Nucleotídeos/genética , RNA Mensageiro/genética , Saccharomyces cerevisiae/genética , Inversão de SequênciaRESUMO
Programmed ribosomal -1 frameshifting is a non-standard decoding process occurring when ribosomes encounter a signal embedded in the mRNA of certain eukaryotic and prokaryotic genes. This signal has a mandatory component, the frameshift motif: it is either a Z_ZZN tetramer or a X_XXZ_ZZN heptamer (where ZZZ and XXX are three identical nucleotides) allowing cognate or near-cognate repairing to the -1 frame of the A site or A and P sites tRNAs. Depending on the signal, the frameshifting frequency can vary over a wide range, from less than 1% to more than 50%. The present study combines experimental and bioinformatics approaches to carry out (i) a systematic analysis of the frameshift propensity of all possible motifs (16 Z_ZZN tetramers and 64 X_XXZ_ZZN heptamers) in Escherichia coli and (ii) the identification of genes potentially using this mode of expression amongst 36 Enterobacteriaceae genomes. While motif efficiency varies widely, a major distinctive rule of bacterial -1 frameshifting is that the most efficient motifs are those allowing cognate re-pairing of the A site tRNA from ZZN to ZZZ. The outcome of the genomic search is a set of 69 gene clusters, 59 of which constitute new candidates for functional utilization of -1 frameshifting.
Assuntos
Escherichia coli/genética , Mudança da Fase de Leitura do Gene Ribossômico , RNA Mensageiro/química , Genes Bacterianos , Motivos de Nucleotídeos , Nucleotídeos/análiseRESUMO
Replacement of beta cells through transplantation is a potential therapeutic approach for individuals with pancreas removal or poorly controllable type 1 diabetes. However, stress and death of beta cells pose significant challenges. Circulating miRNA has emerged as potential biomarkers reflecting early beta cell stress and death, allowing for timely intervention. The aim of this study was to identify miRNAs as potential biomarkers for beta cell health. Literature review combined with small RNA sequencing was employed to select islet-enriched miRNA. The release of those miRNA was assessed by RT-qPCR in vivo, using a streptozotocin induced diabetes mouse model and in vitro, through mouse and human islets exposed to varying degrees of hypoxic and cytokine stressors. Utilizing the streptozotocin induced model, we identified 18 miRNAs out of 39 candidate islet-enriched miRNA to be released upon islet stress in vivo. In vitro analysis of culture supernatants from cytokine and/or hypoxia stressed islets identified the release of 45 miRNAs from mouse and 8 miRNAs from human islets. Investigation into the biological pathways targeted by the cytokine- and/or hypoxia-induced miRNA suggested the involvement of MAPK and PI3K-Akt signaling pathways in both mouse and human islets. We have identified miRNAs associated with beta cell health and stress. The findings allowed us to propose a panel of 47 islet-related human miRNA that is potentially valuable for application in clinical contexts of beta cell transplantation and presymptomatic early-stage type 1 diabetes.
Assuntos
Diabetes Mellitus Experimental , Ilhotas Pancreáticas , MicroRNAs , Animais , MicroRNAs/genética , MicroRNAs/metabolismo , Humanos , Camundongos , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/metabolismo , Ilhotas Pancreáticas/metabolismo , Células Secretoras de Insulina/metabolismo , Reação em Cadeia da Polimerase em Tempo Real/métodos , Estresse Fisiológico/genética , Masculino , RNA-Seq/métodos , Camundongos Endogâmicos C57BL , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/metabolismoRESUMO
MOTIVATION: Conserved patterns across a multiple sequence alignment can be visualized by generating sequence logos. Sequence logos show each column in the alignment as stacks of symbol(s) where the height of a stack is proportional to its informational content, whereas the height of each symbol within the stack is proportional to its frequency in the column. Sequence logos use symbols of either nucleotide or amino acid alphabets. However, certain regulatory signals in messenger RNA (mRNA) act as combinations of codons. Yet no tool is available for visualization of conserved codon patterns. RESULTS: We present the first application which allows visualization of conserved regions in a multiple sequence alignment in the context of codons. CodonLogo is based on WebLogo3 and uses the same heuristics but treats codons as inseparable units of a 64-letter alphabet. CodonLogo can discriminate patterns of codon conservation from patterns of nucleotide conservation that appear indistinguishable in standard sequence logos. AVAILABILITY: The CodonLogo source code and its implementation (in a local version of the Galaxy Browser) are available at http://recode.ucc.ie/CodonLogo and through the Galaxy Tool Shed at http://toolshed.g2.bx.psu.edu/.
Assuntos
Códon , Alinhamento de Sequência/métodos , Software , Algoritmos , Sequência de Bases , Biologia Computacional/métodos , Reconhecimento Automatizado de Padrão , Matrizes de Pontuação de Posição Específica , Linguagens de ProgramaçãoRESUMO
Annotating coding genes and inferring orthologs are two classical challenges in genomics and evolutionary biology that have traditionally been approached separately, limiting scalability. We present TOGA (Tool to infer Orthologs from Genome Alignments), a method that integrates structural gene annotation and orthology inference. TOGA implements a different paradigm to infer orthologous loci, improves ortholog detection and annotation of conserved genes compared with state-of-the-art methods, and handles even highly fragmented assemblies. TOGA scales to hundreds of genomes, which we demonstrate by applying it to 488 placental mammal and 501 bird assemblies, creating the largest comparative gene resources so far. Additionally, TOGA detects gene losses, enables selection screens, and automatically provides a superior measure of mammalian genome quality. TOGA is a powerful and scalable method to annotate and compare genes in the genomic era.
Assuntos
Eutérios , Genômica , Anotação de Sequência Molecular , Animais , Feminino , Camundongos , Eutérios/genética , Genoma , Genômica/métodos , Anotação de Sequência Molecular/métodos , Aves/genéticaRESUMO
Bacterial genome annotations contain a number of coding sequences (CDSs) that, in spite of reading frame disruptions, encode a single continuous polypeptide. Such disruptions have different origins: sequencing errors, frameshift, or stop codon mutations, as well as instances of utilization of nontriplet decoding. We have extracted over 1,000 CDSs with annotated disruptions and found that about 75% of them can be clustered into 64 groups based on sequence similarity. Analysis of the clusters revealed deep phylogenetic conservation of open reading frame organization as well as the presence of conserved sequence patterns that indicate likely utilization of the nonstandard decoding mechanisms: programmed ribosomal frameshifting (PRF) and programmed transcriptional realignment (PTR). Further enrichment of these clusters with additional homologous nucleotide sequences revealed over 6,000 candidate genes utilizing PRF or PTR. Analysis of the patterns of conservation apparently associated with nontriplet decoding revealed the presence of both previously characterized frameshift-prone sequences and a few novel ones. Since the starting point of our analysis was a set of genes with already annotated disruptions, it is highly plausible that in this study, we have identified only a fraction of all bacterial genes that utilize PRF or PTR. In addition to the identification of a large number of recoded genes, a surprising observation is that nearly half of them are expressed via PTR-a mechanism that, in contrast to PRF, has not yet received substantial attention.
Assuntos
Mudança da Fase de Leitura do Gene Ribossômico/genética , Genes Bacterianos/genética , Variação Genética , Fases de Leitura Aberta/genética , Seleção Genética , Transcrição Gênica/genética , Análise por Conglomerados , Biologia Computacional , Sequência Conservada/genética , Filogenia , Projetos PilotoRESUMO
Despite the great diversity of vertebrate limb proportion and our deep understanding of the genetic mechanisms that drive skeletal elongation, little is known about how individual bones reach different lengths in any species. Here, we directly compare the transcriptomes of homologous growth cartilages of the mouse (Mus musculus) and bipedal jerboa (Jaculus jaculus), the latter of which has "mouse-like" arms but extremely long metatarsals of the feet. Intersecting gene-expression differences in metatarsals and forearms of the two species revealed that about 10% of orthologous genes are associated with the disproportionately rapid elongation of neonatal jerboa feet. These include genes and enriched pathways not previously associated with endochondral elongation as well as those that might diversify skeletal proportion in addition to their known requirements for bone growth throughout the skeleton. We also identified transcription regulators that might act as "nodes" for sweeping differences in genome expression between species. Among these, Shox2, which is necessary for proximal limb elongation, has gained expression in jerboa metatarsals where it has not been detected in other vertebrates. We show that Shox2 is sufficient to increase mouse distal limb length, and a nearby putative cis-regulatory region is preferentially accessible in jerboa metatarsals. In addition to mechanisms that might directly promote growth, we found evidence that jerboa foot elongation may occur in part by de-repressing latent growth potential. The genes and pathways that we identified here provide a framework to understand the modular genetic control of skeletal growth and the remarkable malleability of vertebrate limb proportion.
Assuntos
Roedores , Transcriptoma , Animais , Extremidades , Pé , Camundongos , Fatores de Transcrição/metabolismoRESUMO
The human gut microbiome, of which the genus Bifidobacterium is a prevalent and abundant member, is thought to sustain and enhance human health. Several surface-exposed structures, including so-called sortase-dependent pili, represent important bifidobacterial gut colonization factors. Here we show that expression of two sortase-dependent pilus clusters of the prototype Bifidobacterium breve UCC2003 depends on replication slippage at an intragenic G-tract, equivalents of which are present in various members of the Bifidobacterium genus. The nature and extent of this slippage is modulated by the host environment. Involvement of such sortase-dependent pilus clusters in microbe-host interactions, including bacterial attachment to the gut epithelial cells, has been shown previously and is corroborated here for one case. Using a Maximum Depth Sequencing strategy aimed at excluding PCR and sequencing errors introduced by DNA polymerase reagents, specific G-tract sequences in B. breve UCC2003 reveal a range of G-tract lengths whose plasticity within the population is functionally utilized. Interestingly, replication slippage is shown to be modulated under in vivo conditions in a murine model. This in vivo modulation causes an enrichment of a G-tract length which appears to allow biosynthesis of these sortase-dependent pili. This work provides the first example of productive replication slippage influenced by in vivo conditions. It highlights the potential for microdiversity generation in "beneficial" gut commensals.
Assuntos
Bifidobacterium breve , Microbioma Gastrointestinal , Animais , Bifidobacterium/genética , Bifidobacterium breve/metabolismo , Fímbrias Bacterianas/genética , Microbioma Gastrointestinal/genética , Interações entre Hospedeiro e Microrganismos , Humanos , CamundongosRESUMO
Human CD4+ T cells are essential mediators of immune responses. By altering the mitochondrial and metabolic states, we defined metabolic requirements of human CD4+ T cells for in vitro activation, expansion, and effector function. T-cell activation and proliferation were reduced by inhibiting oxidative phosphorylation, whereas early cytokine production was maintained by either OXPHOS or glycolytic activity. Glucose deprivation in the presence of mild mitochondrial stress markedly reduced all three T-cell functions, contrasting the exposure to resveratrol, an antioxidant and sirtuin-1 activator, which specifically inhibited cytokine production and T-cell proliferation, but not T-cell activation. Conditions that inhibited T-cell activation were associated with the down-regulation of 2',5'-oligoadenylate synthetase genes via interferon response pathways. Our findings indicate that T-cell function is grossly impaired by stressors combined with nutrient deprivation, suggesting that correcting nutrient availability, metabolic stress, and/or the function of T cells in these conditions will improve the efficacy of T-cell-based therapies.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Citocinas/metabolismo , Glucose/farmacologia , Glicólise/efeitos dos fármacos , Ativação Linfocitária/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Estresse Fisiológico/efeitos dos fármacos , 2',5'-Oligoadenilato Sintetase/genética , Adulto , Antioxidantes/farmacologia , Doadores de Sangue , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Células Cultivadas , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/genética , Glucose/metabolismo , Glicólise/genética , Humanos , Ativação Linfocitária/genética , Masculino , Mitocôndrias/metabolismo , Fosforilação Oxidativa/efeitos dos fármacos , Resveratrol/farmacologia , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Estresse Fisiológico/genética , Estresse Fisiológico/imunologiaRESUMO
We systematically investigate whether losses of human disease-associated genes occurred in other mammals during evolution. We first show that genes lost in any of 62 non-human mammals generally have a lower degree of pleiotropy, and are highly depleted in essential and disease-associated genes. Despite this under-representation, we discovered multiple genes implicated in human disease that are truly lost in non-human mammals. In most cases, traits resembling human disease symptoms are present but not deleterious in gene-loss species, exemplified by losses of genes causing human eye or teeth disorders in poor-vision or enamel-less mammals. We also found widespread losses of PCSK9 and CETP genes, where loss-of-function mutations in humans protect from atherosclerosis. Unexpectedly, we discovered losses of disease genes (TYMP, TBX22, ABCG5, ABCG8, MEFV, CTSE) where deleterious phenotypes do not manifest in the respective species. A remarkable example is the uric acid-degrading enzyme UOX, which we found to be inactivated in elephants and manatees. While UOX loss in hominoids led to high serum uric acid levels and a predisposition for gout, elephants and manatees exhibit low uric acid levels, suggesting alternative ways of metabolizing uric acid. Together, our results highlight numerous mammals that are 'natural knockouts' of human disease genes.
RESUMO
Alignment-based gene identification methods utilize sequence conservation between orthologous protein-coding genes to annotate genes in newly sequenced genomes. CESAR is an approach that makes use of existing genome alignments to transfer genes from one genome to other aligned genomes, and thus generates comparative gene annotations. To accurately detect conserved exons that exhibit an intact reading frame and consensus splice sites, CESAR produces a new alignment between orthologous exons, taking information about the exon's reading frame and splice site positions into account. Furthermore, CESAR is able to detect most evolutionary splice site shifts, which helps to annotate exon boundaries at high precision. Here, we describe how to apply CESAR to generate comparative gene annotations for one or many species, and discuss the strengths and limitations of this approach. CESAR is available at https://github.com/hillerlab/CESAR2.0 .