RESUMO
Modern humans inhabit a variety of environments and are exposed to a plethora of selective pressures, leading to multiple genetic adaptations to local environmental conditions. These include adaptations to climate, UV exposure, disease, diet, altitude, or cultural practice and have generated important genetic and phenotypic differences amongst populations. In recent years, new methods to identify the genomic signatures of natural selection underlying these adaptations, combined with novel types of genetic data (e.g., ancient DNA), have provided unprecedented insights into the origin of adaptive alleles and the modes of adaptation. As a result, numerous instances of local adaptation have been identified in humans. Here, we review the most exciting recent developments and discuss, in our view, the future of this field.
Assuntos
Adaptação Fisiológica , Evolução Biológica , Variação Genética , Genômica/métodos , Seleção Genética , Animais , HumanosRESUMO
OBJECTIVES: To define the host mechanisms contributing to the pathological interferon (IFN) type 1 signature in Juvenile dermatomyositis (JDM). METHODS: RNA-sequencing was performed on CD4+, CD8+, CD14+ and CD19+ cells sorted from pretreatment and on-treatment JDM (pretreatment n=10, on-treatment n=11) and age/sex-matched child healthy-control (CHC n=4) peripheral blood mononuclear cell (PBMC). Mitochondrial morphology and superoxide were assessed by fluorescence microscopy, cellular metabolism by 13C glucose uptake assays, and oxidised mitochondrial DNA (oxmtDNA) content by dot-blot. Healthy-control PBMC and JDM pretreatment PBMC were cultured with IFN-α, oxmtDNA, cGAS-inhibitor, TLR-9 antagonist and/or n-acetyl cysteine (NAC). IFN-stimulated gene (ISGs) expression was measured by qPCR. Total numbers of patient and controls for functional experiments, JDM n=82, total CHC n=35. RESULTS: Dysregulated mitochondrial-associated gene expression correlated with increased ISG expression in JDM CD14+ monocytes. Altered mitochondrial-associated gene expression was paralleled by altered mitochondrial biology, including 'megamitochondria', cellular metabolism and a decrease in gene expression of superoxide dismutase (SOD)1. This was associated with enhanced production of oxidised mitochondrial (oxmt)DNA. OxmtDNA induced ISG expression in healthy PBMC, which was blocked by targeting oxidative stress and intracellular nucleic acid sensing pathways. Complementary experiments showed that, under in vitro experimental conditions, targeting these pathways via the antioxidant drug NAC, TLR9 antagonist and to a lesser extent cGAS-inhibitor, suppressed ISG expression in pretreatment JDM PBMC. CONCLUSIONS: These results describe a novel pathway where altered mitochondrial biology in JDM CD14+ monocytes lead to oxmtDNA production and stimulates ISG expression. Targeting this pathway has therapeutical potential in JDM and other IFN type 1-driven autoimmune diseases.
Assuntos
Dermatomiosite , Interferon Tipo I , Criança , Humanos , Leucócitos Mononucleares/metabolismo , Monócitos/metabolismo , DNA Mitocondrial , Interferon Tipo I/metabolismo , NucleotidiltransferasesRESUMO
It has been shown that Neanderthals contributed genetically to modern humans outside Africa 47,000-65,000 years ago. Here we analyse the genomes of a Neanderthal and a Denisovan from the Altai Mountains in Siberia together with the sequences of chromosome 21 of two Neanderthals from Spain and Croatia. We find that a population that diverged early from other modern humans in Africa contributed genetically to the ancestors of Neanderthals from the Altai Mountains roughly 100,000 years ago. By contrast, we do not detect such a genetic contribution in the Denisovan or the two European Neanderthals. We conclude that in addition to later interbreeding events, the ancestors of Neanderthals from the Altai Mountains and early modern humans met and interbred, possibly in the Near East, many thousands of years earlier than previously thought.
Assuntos
Fluxo Gênico/genética , Homem de Neandertal/genética , Altitude , Animais , Teorema de Bayes , Cromossomos Humanos Par 21/genética , Croácia/etnologia , Genoma Humano/genética , Genômica , Haplótipos/genética , Heterozigoto , Humanos , Hibridização Genética/genética , Filogenia , Densidade Demográfica , Sibéria , Espanha/etnologia , Fatores de TempoRESUMO
Chimpanzees, humans' closest relatives, are in danger of extinction. Aside from direct human impacts such as hunting and habitat destruction, a key threat is transmissible disease. As humans continue to encroach upon their habitats, which shrink in size and grow in density, the risk of inter-population and cross-species viral transmission increases, a point dramatically made in the reverse with the global HIV/AIDS pandemic. Inhabiting central Africa, the four subspecies of chimpanzees differ in demographic history and geographical range, and are likely differentially adapted to their particular local environments. To quantitatively explore genetic adaptation, we investigated the genic enrichment for SNPs highly differentiated between chimpanzee subspecies. Previous analyses of such patterns in human populations exhibited limited evidence of adaptation. In contrast, chimpanzees show evidence of recent positive selection, with differences among subspecies. Specifically, we observe strong evidence of recent selection in eastern chimpanzees, with highly differentiated SNPs being uniquely enriched in genic sites in a way that is expected under recent adaptation but not under neutral evolution or background selection. These sites are enriched for genes involved in immune responses to pathogens, and for genes inferred to differentiate the immune response to infection by simian immunodeficiency virus (SIV) in natural vs. non-natural host species. Conversely, central chimpanzees exhibit an enrichment of signatures of positive selection only at cytokine receptors, due to selective sweeps in CCR3, CCR9 and CXCR6 -paralogs of CCR5 and CXCR4, the two major receptors utilized by HIV to enter human cells. Thus, our results suggest that positive selection has contributed to the genetic and phenotypic differentiation of chimpanzee subspecies, and that viruses likely play a predominate role in this differentiation, with SIV being a likely selective agent. Interestingly, our results suggest that SIV has elicited distinctive adaptive responses in these two chimpanzee subspecies.
Assuntos
Adaptação Fisiológica/genética , Imunidade Inata/genética , Pan troglodytes/genética , Seleção Genética/genética , Adaptação Fisiológica/imunologia , Animais , Demografia , Deriva Genética , Especiação Genética , HIV/genética , HIV/imunologia , HIV/patogenicidade , Humanos , Pan troglodytes/imunologia , Pan troglodytes/virologia , Polimorfismo de Nucleotídeo Único/genética , Receptores CCR/genética , Receptores CCR3/genética , Receptores CCR5/genética , Receptores CXCR4/genética , Receptores CXCR6/imunologia , Vírus da Imunodeficiência Símia/genética , Vírus da Imunodeficiência Símia/imunologia , Vírus da Imunodeficiência Símia/patogenicidadeRESUMO
We sequenced the genomes of a â¼7,000-year-old farmer from Germany and eight â¼8,000-year-old hunter-gatherers from Luxembourg and Sweden. We analysed these and other ancient genomes with 2,345 contemporary humans to show that most present-day Europeans derive from at least three highly differentiated populations: west European hunter-gatherers, who contributed ancestry to all Europeans but not to Near Easterners; ancient north Eurasians related to Upper Palaeolithic Siberians, who contributed to both Europeans and Near Easterners; and early European farmers, who were mainly of Near Eastern origin but also harboured west European hunter-gatherer related ancestry. We model these populations' deep relationships and show that early European farmers had â¼44% ancestry from a 'basal Eurasian' population that split before the diversification of other non-African lineages.
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Genoma Humano/genética , População Branca/classificação , População Branca/genética , Agricultura/história , Ásia/etnologia , Europa (Continente) , História Antiga , Humanos , Dinâmica Populacional , Análise de Componente Principal , Recursos HumanosRESUMO
Selenium (Se), a sparse element on earth, is an essential micronutrient in the vertebrate diet and its intake depends on its content in soils and waters worldwide. Selenium is required due to its function in selenoproteins, which contain selenocysteine (Sec), the 21st amino acid in the genetic code, as one of their constituent residues. Selenocysteine is analogous to the amino acid cysteine (Cys), which uses the abounding element sulfur instead. Despite the irregular distribution of Se worldwide, its distinct biochemical properties have made the substitution of Sec for Cys rare in vertebrate proteins. Still, vertebrates inhabited environments with different amounts of Se and may have distinctly adapted to it. To address this question, we compared the evolutionary forces acting on the coding sequences of selenoprotein genes and genes that regulate Se between vertebrate clades and between the Se-dependent genes and their paralogs with Cys. We find that the strength of natural selection in genes that use or regulate Se is distinct between land vertebrates and teleost fishes and more variable than in the Cys paralogs, particularly in genes involved in the preferential supply of Se to some organs and the tissue-specific expression of selenoproteins. This is compatible with vertebrates adapting to Se scarcity in land and its abundance in waters. In agreement, teleost fishes duplicated and subfunctionalized or neofunctionalized selenoprotein genes and maintained their capacity for Se transport in the body, which declined (under neutrality) for millions of years in terrestrial vertebrates. Dietary Se has thus distinctly shaped vertebrate evolution.
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Organismos Aquáticos/genética , Seleção Genética , Selenoproteínas/genética , Vertebrados/genética , Animais , Duplicação Gênica , Selênio/metabolismo , Selenoproteínas/metabolismoRESUMO
Sequencing the genomes of extinct hominids has reshaped our understanding of modern human origins. Here, we analyze â¼120 kb of exome-captured Y-chromosome DNA from a Neandertal individual from El Sidrón, Spain. We investigate its divergence from orthologous chimpanzee and modern human sequences and find strong support for a model that places the Neandertal lineage as an outgroup to modern human Y chromosomes-including A00, the highly divergent basal haplogroup. We estimate that the time to the most recent common ancestor (TMRCA) of Neandertal and modern human Y chromosomes is â¼588 thousand years ago (kya) (95% confidence interval [CI]: 447-806 kya). This is â¼2.1 (95% CI: 1.7-2.9) times longer than the TMRCA of A00 and other extant modern human Y-chromosome lineages. This estimate suggests that the Y-chromosome divergence mirrors the population divergence of Neandertals and modern human ancestors, and it refutes alternative scenarios of a relatively recent or super-archaic origin of Neandertal Y chromosomes. The fact that the Neandertal Y we describe has never been observed in modern humans suggests that the lineage is most likely extinct. We identify protein-coding differences between Neandertal and modern human Y chromosomes, including potentially damaging changes to PCDH11Y, TMSB4Y, USP9Y, and KDM5D. Three of these changes are missense mutations in genes that produce male-specific minor histocompatibility (H-Y) antigens. Antigens derived from KDM5D, for example, are thought to elicit a maternal immune response during gestation. It is possible that incompatibilities at one or more of these genes played a role in the reproductive isolation of the two groups.
Assuntos
Cromossomos Humanos Y/genética , Homem de Neandertal/genética , Animais , Caderinas/genética , DNA/genética , Variação Genética , Histona Desmetilases/genética , Humanos , Masculino , Antígenos de Histocompatibilidade Menor/genética , Mutação de Sentido Incorreto , Pan troglodytes/genética , Protocaderinas , Análise de Sequência de DNA , Espanha , Ubiquitina Tiolesterase/genéticaRESUMO
The human genome contains 25 genes coding for selenocysteine-containing proteins (selenoproteins). These proteins are involved in a variety of functions, most notably redox homeostasis. Selenoprotein enzymes with known functions are designated according to these functions: TXNRD1, TXNRD2, and TXNRD3 (thioredoxin reductases), GPX1, GPX2, GPX3, GPX4, and GPX6 (glutathione peroxidases), DIO1, DIO2, and DIO3 (iodothyronine deiodinases), MSRB1 (methionine sulfoxide reductase B1), and SEPHS2 (selenophosphate synthetase 2). Selenoproteins without known functions have traditionally been denoted by SEL or SEP symbols. However, these symbols are sometimes ambiguous and conflict with the approved nomenclature for several other genes. Therefore, there is a need to implement a rational and coherent nomenclature system for selenoprotein-encoding genes. Our solution is to use the root symbol SELENO followed by a letter. This nomenclature applies to SELENOF (selenoprotein F, the 15-kDa selenoprotein, SEP15), SELENOH (selenoprotein H, SELH, C11orf31), SELENOI (selenoprotein I, SELI, EPT1), SELENOK (selenoprotein K, SELK), SELENOM (selenoprotein M, SELM), SELENON (selenoprotein N, SEPN1, SELN), SELENOO (selenoprotein O, SELO), SELENOP (selenoprotein P, SeP, SEPP1, SELP), SELENOS (selenoprotein S, SELS, SEPS1, VIMP), SELENOT (selenoprotein T, SELT), SELENOV (selenoprotein V, SELV), and SELENOW (selenoprotein W, SELW, SEPW1). This system, approved by the HUGO Gene Nomenclature Committee, also resolves conflicting, missing, and ambiguous designations for selenoprotein genes and is applicable to selenoproteins across vertebrates.
Assuntos
Selenoproteínas/classificação , Selenoproteínas/genética , Humanos , Terminologia como AssuntoRESUMO
We develop and evaluate methods for inferring relatedness among individuals from low-coverage DNA sequences of their genomes, with particular emphasis on sequences obtained from fossil remains. We suggest the major factors complicating the determination of relatedness among ancient individuals are sequencing depth, the number of overlapping sites, the sequencing error rate and the presence of contamination from present-day genetic sources. We develop a theoretical model that facilitates the exploration of these factors and their relative effects, via measurement of pairwise genetic distances, without calling genotypes, and determine the power to infer relatedness under various scenarios of varying sequencing depth, present-day contamination and sequencing error. The model is validated by a simulation study as well as the analysis of aligned sequences from present-day human genomes. We then apply the method to the recently published genome sequences of ancient Europeans, developing a statistical treatment to determine confidence in assigned relatedness that is, in some cases, more precise than previously reported. As the majority of ancient specimens are from animals, this method would be applicable to investigate kinship in nonhuman remains. The developed software grups (Genetic Relatedness Using Pedigree Simulations) is implemented in Python and freely available.
Assuntos
Simulação por Computador , Genoma Humano , Modelos Genéticos , Linhagem , Genótipo , Humanos , Polimorfismo de Nucleotídeo Único , SoftwareRESUMO
We present the DNA sequence of 17,367 protein-coding genes in two Neandertals from Spain and Croatia and analyze them together with the genome sequence recently determined from a Neandertal from southern Siberia. Comparisons with present-day humans from Africa, Europe, and Asia reveal that genetic diversity among Neandertals was remarkably low, and that they carried a higher proportion of amino acid-changing (nonsynonymous) alleles inferred to alter protein structure or function than present-day humans. Thus, Neandertals across Eurasia had a smaller long-term effective population than present-day humans. We also identify amino acid substitutions in Neandertals and present-day humans that may underlie phenotypic differences between the two groups. We find that genes involved in skeletal morphology have changed more in the lineage leading to Neandertals than in the ancestral lineage common to archaic and modern humans, whereas genes involved in behavior and pigmentation have changed more on the modern human lineage.
Assuntos
Exoma , Variação Genética , Homem de Neandertal/genética , Substituição de Aminoácidos , Animais , Croácia , DNA/genética , Frequência do Gene , Humanos , Paleontologia , Filogenia , Polimorfismo de Nucleotídeo Único , Sibéria , EspanhaRESUMO
As humans migrated around the world, they came to inhabit environments that differ widely in the soil levels of certain micronutrients, including selenium (Se). Coupled with cultural variation in dietary practices, these migrations have led to a wide range of Se intake levels in populations around the world. Both excess and deficiency of Se in the diet can have adverse health consequences in humans, with severe Se deficiency resulting in diseases of the bone and heart. Se is required by humans mainly due to its function in selenoproteins, which contain the amino acid selenocysteine as one of their constituent residues. To understand the evolution of the use of this micronutrient in humans, we surveyed the patterns of polymorphism in all selenoprotein genes and genes involved in their regulation in 50 human populations. We find that single nucleotide polymorphisms from populations in Asia, particularly in populations living in the extreme Se-deficient regions of China, have experienced concerted shifts in their allele frequencies. Such differentiation in allele frequencies across genes is not observed in other regions of the world and is not expected under neutral evolution, being better explained by the action of recent positive selection. Thus, recent changes in the use and regulation of Se may harbor the genetic adaptations that helped humans inhabit environments that do not provide adequate levels of Se in the diet.
Assuntos
Adaptação Fisiológica/genética , Dieta , Evolução Molecular , Selênio , Selenoproteínas/genética , China , Frequência do Gene , Humanos , Anotação de Sequência Molecular , Polimorfismo de Nucleotídeo Único , RNA Mensageiro/genética , Seleção Genética , Selênio/deficiência , Selenocisteína/genéticaRESUMO
SelenoDB (http://www.selenodb.org) aims to provide high-quality annotations of selenoprotein genes, proteins and SECIS elements. Selenoproteins are proteins that contain the amino acid selenocysteine (Sec) and the first release of the database included annotations for eight species. Since the release of SelenoDB 1.0 many new animal genomes have been sequenced. The annotations of selenoproteins in new genomes usually contain many errors in major databases. For this reason, we have now fully annotated selenoprotein genes in 58 animal genomes. We provide manually curated annotations for human selenoproteins, whereas we use an automatic annotation pipeline to annotate selenoprotein genes in other animal genomes. In addition, we annotate the homologous genes containing cysteine (Cys) instead of Sec. Finally, we have surveyed genetic variation in the annotated genes in humans. We use exon capture and resequencing approaches to identify single-nucleotide polymorphisms in more than 50 human populations around the world. We thus present a detailed view of the genetic divergence of Sec- and Cys-containing genes in animals and their diversity in humans. The addition of these datasets into the second release of the database provides a valuable resource for addressing medical and evolutionary questions in selenium biology.
Assuntos
Bases de Dados de Proteínas , Variação Genética , Anotação de Sequência Molecular , Selenoproteínas/genética , Animais , Genes , Genoma , Humanos , Internet , Selenoproteínas/classificaçãoRESUMO
Selenocysteine, the 21st amino acid specified by the genetic code, is a rare selenium-containing residue found in the catalytic site of selenoprotein oxidoreductases. Selenocysteine is analogous to the common cysteine amino acid, but its selenium atom offers physical-chemical properties not provided by the corresponding sulfur atom in cysteine. Catalytic sites with selenocysteine in selenoproteins of vertebrates are under strong purifying selection, but one enzyme, glutathione peroxidase 6 (GPX6), independently exchanged selenocysteine for cysteine <100 million years ago in several mammalian lineages. We reconstructed and assayed these ancient enzymes before and after selenocysteine was lost and up to today and found them to have lost their classic ability to reduce hydroperoxides using glutathione. This loss of function, however, was accompanied by additional amino acid changes in the catalytic domain, with protein sites concertedly changing under positive selection across distant lineages abandoning selenocysteine in glutathione peroxidase 6. This demonstrates a narrow evolutionary range in maintaining fitness when sulfur in cysteine impairs the catalytic activity of this protein, with pleiotropy and epistasis likely driving the observed convergent evolution. We propose that the mutations shared across distinct lineages may trigger enzymatic properties beyond those in classic glutathione peroxidases, rather than simply recovering catalytic rate. These findings are an unusual example of adaptive convergence across mammalian selenoproteins, with the evolutionary signatures possibly representing the evolution of novel oxidoreductase functions.
Assuntos
Selênio , Selenocisteína , Animais , Selenocisteína/genética , Selenocisteína/química , Selenocisteína/metabolismo , Cisteína/genética , Cisteína/metabolismo , Selênio/metabolismo , Selenoproteínas/genética , Selenoproteínas/química , Selenoproteínas/metabolismo , Glutationa Peroxidase/genética , Glutationa Peroxidase/metabolismo , Aminoácidos , Glutationa , Enxofre , Mamíferos/genética , Mamíferos/metabolismoRESUMO
The two types of craniopharyngioma, adamantinomatous (ACP) and papillary (PCP), are clinically relevant tumours in children and adults. Although the biology of primary craniopharyngioma is starting to be unravelled, little is known about the biology of recurrence. To fill this gap in knowledge, we have analysed through methylation array, RNA sequencing and pERK1/2 immunohistochemistry a cohort of paired primary and recurrent samples (32 samples from 14 cases of ACP and 4 cases of PCP). We show the presence of copy number alterations and clonal evolution across recurrence in 6 cases of ACP, and analysis of additional whole genome sequencing data from the Children's Brain Tumour Network confirms chromosomal arm copy number changes in at least 7/67 ACP cases. The activation of the MAPK/ERK pathway, a feature previously shown in primary ACP, is observed in all but one recurrent cases of ACP. The only ACP without MAPK activation is an aggressive case of recurrent malignant human craniopharyngioma harbouring a CTNNB1 mutation and loss of TP53. Providing support for a functional role of this TP53 mutation, we show that Trp53 loss in a murine model of ACP results in aggressive tumours and reduced mouse survival. Finally, we characterise the tumour immune infiltrate showing differences in the cellular composition and spatial distribution between ACP and PCP. Together, these analyses have revealed novel insights into recurrent craniopharyngioma and provided preclinical evidence supporting the evaluation of MAPK pathway inhibitors and immunomodulatory approaches in clinical trials in against recurrent ACP.
Assuntos
Evolução Clonal , Craniofaringioma , Sistema de Sinalização das MAP Quinases , Recidiva Local de Neoplasia , Neoplasias Hipofisárias , Proteína Supressora de Tumor p53 , Animais , Feminino , Humanos , Masculino , Camundongos , beta Catenina/genética , beta Catenina/metabolismo , Evolução Clonal/genética , Craniofaringioma/genética , Craniofaringioma/patologia , Craniofaringioma/metabolismo , Progressão da Doença , Sistema de Sinalização das MAP Quinases/genética , Sistema de Sinalização das MAP Quinases/fisiologia , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/patologia , Neoplasias Hipofisárias/genética , Neoplasias Hipofisárias/patologia , Neoplasias Hipofisárias/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismoRESUMO
The use of induced pluripotent stem cells (iPSC) as models for development and human disease has enabled the study of otherwise inaccessible tissues. A remaining challenge in developing reliable models is our limited understanding of the factors driving irregular differentiation of iPSCs, particularly the impact of acquired somatic mutations. We leveraged data from a pooled dopaminergic neuron differentiation experiment of 238 iPSC lines profiled with single-cell RNA and whole-exome sequencing to study how somatic mutations affect differentiation outcomes. We found that deleterious somatic mutations in key developmental genes, notably the BCOR gene, are strongly associated with failure in dopaminergic neuron differentiation and a larger proliferation rate in culture. We further identified broad differences in cell type composition between incorrectly and successfully differentiating lines, as well as significant changes in gene expression contributing to the inhibition of neurogenesis. Our work calls for caution in interpreting differentiation-related phenotypes in disease-modeling experiments.
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Primary keratinocytes including keratinocyte stem cells (KSCs) can be cultured as epidermal sheets in vitro and are attractive for cell and gene therapies for genetic skin disorders. However, the initial slow growth of freshly isolated keratinocytes hinders clinical applications. Rho-associated kinase inhibitor (ROCKi) has been used to overcome this obstacle, but its influence on the characteristics of KSC and its safety for clinical application remains unknown. In this study, primary keratinocytes were treated with ROCKi Y-27632 for six days (short-term). Significant increases in colony formation and cell proliferation during the six-day ROCKi treatment were observed and confirmed by related protein markers and single-cell transcriptomic analysis. In addition, short-term ROCKi-treated cells maintained their differentiation ability as examined by 3D-organotypic culture. However, these changes could be reversed and became indistinguishable between treated and untreated cells once ROCKi treatment was withdrawn. Further, the short-term ROCKi treatment did not reduce the number of KSCs. In addition, AKT and ERK pathways were rapidly activated upon ROCKi treatment. In conclusion, short-term ROCKi treatment can transiently and reversibly accelerate initial primary keratinocyte expansion while preserving the holoclone-forming cell population (KSCs), providing a safe avenue for clinical applications.
Assuntos
Queratinócitos , Quinases Associadas a rho , Células Cultivadas , Células-Tronco , Epiderme , Inibidores de Proteínas Quinases/farmacologiaRESUMO
BACKGROUND: Alström syndrome (ALMS) is a rare autosomal recessive disease that is associated with mutations in ALMS1 gene. The main clinical manifestations of ALMS are retinal dystrophy, obesity, type 2 diabetes mellitus, dilated cardiomyopathy and multi-organ fibrosis, characteristic in kidneys and liver. Depletion of the protein encoded by ALMS1 has been associated with the alteration of different processes regulated via the primary cilium, such as the NOTCH or TGF-ß signalling pathways. However, the cellular impact of these deregulated pathways in the absence of ALMS1 remains unknown. METHODS: In this study, we integrated RNA-seq and proteomic analysis to determine the gene expression profile of hTERT-BJ-5ta ALMS1 knockout fibroblasts after TGF-ß stimulation. In addition, we studied alterations in cross-signalling between the TGF-ß pathway and the AKT pathway in this cell line. RESULTS: We found that ALMS1 depletion affects the TGF-ß pathway and its cross-signalling with other pathways such as PI3K/AKT, EGFR1 or p53. In addition, alterations associated with ALMS1 depletion clustered around the processes of extracellular matrix regulation and lipid metabolism in both the transcriptome and proteome. By studying the enriched pathways of common genes differentially expressed in the transcriptome and proteome, collagen fibril organisation, ß-oxidation of fatty acids and eicosanoid metabolism emerged as key processes altered by the absence of ALMS1. Finally, an overactivation of the AKT pathway was determined in the absence of ALMS1 that could be explained by a decrease in PTEN gene expression. CONCLUSION: ALMS1 deficiency disrupts cross-signalling between the TGF-ß pathway and other dependent pathways in hTERT-BJ-5ta cells. Furthermore, altered cross-signalling impacts the regulation of extracellular matrix-related processes and fatty acid metabolism, and leads to over-activation of the AKT pathway.
Assuntos
Síndrome de Alstrom , Diabetes Mellitus Tipo 2 , Humanos , Metabolismo dos Lipídeos , Diabetes Mellitus Tipo 2/metabolismo , Proteoma/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteômica , Proteínas Proto-Oncogênicas c-akt , Proteínas de Ciclo Celular/metabolismo , Síndrome de Alstrom/genética , Síndrome de Alstrom/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Matriz Extracelular/metabolismoRESUMO
We recently described a low-affinity second-generation CD19 chimeric antigen receptor (CAR) CAT that showed enhanced expansion, cytotoxicity, and antitumor efficacy compared with the high-affinity (FMC63-based) CAR used in tisagenlecleucel, in preclinical models. Furthermore, CAT demonstrated an excellent toxicity profile, enhanced in vivo expansion, and long-term persistence in a phase 1 clinical study. To understand the molecular mechanisms behind these properties of CAT CAR T cells, we performed a systematic in vitro characterization of the transcriptomic (RNA sequencing) and protein (cytometry by time of flight) changes occurring in T cells expressing low-affinity vs high-affinity CD19 CARs following stimulation with CD19-expressing cells. Our results show that CAT CAR T cells exhibit enhanced activation to CD19 stimulation and a distinct transcriptomic and protein profile, with increased activation and cytokine polyfunctionality compared with FMC63 CAR T cells. We demonstrate that the enhanced functionality of low-affinity CAT CAR T cells is a consequence of an antigen-dependent priming induced by residual CD19-expressing B cells present in the manufacture.
Assuntos
Citocinas , Receptores de Antígenos Quiméricos , Citocinas/metabolismo , Imunoterapia Adotiva/métodos , Linfócitos T , Receptores de Antígenos Quiméricos/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Antígenos CD19RESUMO
Acute Myeloid Leukaemia (AML) is a phenotypically and genetically heterogenous blood cancer characterised by very poor prognosis, with disease relapse being the primary cause of treatment failure. AML heterogeneity arise from different genetic and non-genetic sources, including its proposed hierarchical structure, with leukemic stem cells (LSCs) and progenitors giving origin to a variety of more mature leukemic subsets. Recent advances in single-cell molecular and phenotypic profiling have highlighted the intra and inter-patient heterogeneous nature of AML, which has so far limited the success of cell-based immunotherapy approaches against single targets. Machine Learning (ML) can be uniquely used to find non-trivial patterns from high-dimensional datasets and identify rare sub-populations. Here we review some recent ML tools that applied to single-cell data could help disentangle cell heterogeneity in AML by identifying distinct core molecular signatures of leukemic cell subsets. We discuss the advantages and limitations of unsupervised and supervised ML approaches to cluster and classify cell populations in AML, for the identification of biomarkers and the design of personalised therapies.
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The process of natural selection leaves signatures in our genome that can be used to identify functionally important amino acid changes in proteins. In natural populations, amino acids that are better adapted to local conditions might increase in frequency, whereas moderately to severely deleterious protein mutations tend to be eliminated and do not contribute to protein differences between species. Amino acid mutations with no fitness consequences are, however, lost or fixed without regard to natural selection. Looking for evidence of natural selection is, therefore, an attractive strategy for characterizing the contribution of a residue to protein function. Because the majority of identified selenoproteins have now been found in Cys-form, the extent of exchangeability between Sec and Cys residues can be measured in proteins over long periods of time. The statistical analysis of the pattern of Sec/Cys exchanges in diversity (within species) and divergence (between species) data, provides robust inferences of the strength and mode of natural selection acting on these protein sites. Such inferences inform us not only of the long-term exchangeability between Sec and Cys residues, but also of the nature of the selective factors shaping Sec usage in proteins.