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1.
Nat Immunol ; 21(9): 1070-1081, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32661361

RESUMO

Tissue-resident memory CD8+ T cells (TRM cells) are crucial in protecting against reinvading pathogens, but the impact of reinfection on their tissue confinement and contribution to recall responses is unclear. We developed a unique lineage tracer mouse model exploiting the TRM-defining transcription factor homolog of Blimp-1 in T cells (Hobit) to fate map the TRM progeny in secondary responses. After reinfection, a sizeable fraction of secondary memory T cells in the circulation developed downstream of TRM cells. These tissue-experienced ex-TRM cells shared phenotypic properties with the effector memory T cell population but were transcriptionally and functionally distinct from other secondary effector memory T cell cells. Adoptive transfer experiments of TRM cells corroborated their potential to form circulating effector and memory cells during recall responses. Moreover, specific ablation of primary TRM cell populations substantially impaired the secondary T cell response, both locally and systemically. Thus, TRM cells retain developmental plasticity and shape both local and systemic T cell responses on reinfection.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Memória Imunológica/imunologia , Fator 1 de Ligação ao Domínio I Regulador Positivo/metabolismo , Transferência Adotiva , Animais , Diferenciação Celular , Linhagem da Célula , Plasticidade Celular , Células Cultivadas , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Fator 1 de Ligação ao Domínio I Regulador Positivo/genética
2.
Brief Bioinform ; 22(3)2021 05 20.
Artigo em Inglês | MEDLINE | ID: mdl-32935832

RESUMO

Insights into the evolution of ancestral complexes and pathways are generally achieved through careful and time-intensive manual analysis often using phylogenetic profiles of the constituent proteins. This manual analysis limits the possibility of including more protein-complex components, repeating the analyses for updated genome sets or expanding the analyses to larger scales. Automated orthology inference should allow such large-scale analyses, but substantial differences between orthologous groups generated by different approaches are observed. We evaluate orthology methods for their ability to recapitulate a number of observations that have been made with regard to genome evolution in eukaryotes. Specifically, we investigate phylogenetic profile similarity (co-occurrence of complexes), the last eukaryotic common ancestor's gene content, pervasiveness of gene loss and the overlap with manually determined orthologous groups. Moreover, we compare the inferred orthologies to each other. We find that most orthology methods reconstruct a large last eukaryotic common ancestor, with substantial gene loss, and can predict interacting proteins reasonably well when applying phylogenetic co-occurrence. At the same time, derived orthologous groups show imperfect overlap with manually curated orthologous groups. There is no strong indication of which orthology method performs better than another on individual or all of these aspects. Counterintuitively, despite the orthology methods behaving similarly regarding large-scale evaluation, the obtained orthologous groups differ vastly from one another. Availability and implementation The data and code underlying this article are available in github and/or upon reasonable request to the corresponding author: https://github.com/ESDeutekom/ComparingOrthologies.


Assuntos
Benchmarking/métodos , Eucariotos/genética , Filogenia , Proteínas/genética , Proteoma/genética , Bases de Dados de Proteínas , Eucariotos/classificação , Evolução Molecular , Genoma/genética , Genômica/métodos , Internet , Proteínas/metabolismo , Proteoma/metabolismo , Reprodutibilidade dos Testes , Software
3.
Annu Rev Pharmacol Toxicol ; 58: 271-291, 2018 01 06.
Artigo em Inglês | MEDLINE | ID: mdl-28715978

RESUMO

Insight into drug transport mechanisms is highly relevant to the efficacious treatment of tuberculosis (TB). Major problems in TB treatment are related to the transport of antituberculosis (anti-TB) drugs across human and mycobacterial membranes, affecting the concentrations of these drugs systemically and locally. Firstly, transporters located in the intestines, liver, and kidneys all determine the pharmacokinetics and pharmacodynamics of anti-TB drugs, with a high risk of drug-drug interactions in the setting of concurrent use of antimycobacterial, antiretroviral, and antidiabetic agents. Secondly, human efflux transporters limit the penetration of anti-TB drugs into the brain and cerebrospinal fluid, which is especially important in the treatment of TB meningitis. Finally, efflux transporters located in the macrophage and Mycobacterium tuberculosis cell membranes play a pivotal role in the emergence of phenotypic tolerance and drug resistance, respectively. We review the role of efflux transporters in TB drug disposition and evaluate the promise of efflux pump inhibition from a novel holistic perspective.


Assuntos
Antituberculosos/farmacologia , Antituberculosos/uso terapêutico , Proteínas de Membrana Transportadoras/metabolismo , Tuberculose/tratamento farmacológico , Tuberculose/metabolismo , Animais , Desenvolvimento de Medicamentos/métodos , Humanos , Mycobacterium tuberculosis/efeitos dos fármacos
4.
Bioessays ; 41(5): e1900006, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31026339

RESUMO

Comparative genomics has proven a fruitful approach to acquire many functional and evolutionary insights into core cellular processes. Here it is argued that in order to perform accurate and interesting comparative genomics, one first and foremost has to be able to recognize, postulate, and revise different evolutionary scenarios. After all, these studies lack a simple protocol, due to different proteins having different evolutionary dynamics and demanding different approaches. The authors here discuss this challenge from a practical (what are the observations?) and conceptual (how do these indicate a specific evolutionary scenario?) viewpoint, with the aim to guide investigators who want to analyze the evolution of their protein(s) of interest. By sharing how the authors draft, test, and update such a scenario and how it directs their investigations, the authors hope to illuminate how to execute molecular evolution studies and how to interpret them. Also see the video abstract here https://youtu.be/VCt3l2pbdbQ.


Assuntos
Biologia Computacional/métodos , Evolução Molecular , Proteínas/genética , Proteínas de Caenorhabditis elegans/genética , Bases de Dados de Proteínas , Células Eucarióticas , Genômica/métodos , Humanos , Filogenia , Domínios Proteicos , Proteínas/química
5.
PLoS Comput Biol ; 15(8): e1007301, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31461468

RESUMO

In recent years it became clear that in eukaryotic genome evolution gene loss is prevalent over gene gain. However, the absence of genes in an annotated genome is not always equivalent to the loss of genes. Due to sequencing issues, or incorrect gene prediction, genes can be falsely inferred as absent. This implies that loss estimates are overestimated and, more generally, that falsely inferred absences impact genomic comparative studies. However, reliable estimates of how prevalent this issue is are lacking. Here we quantified the impact of gene prediction on gene loss estimates in eukaryotes by analysing 209 phylogenetically diverse eukaryotic organisms and comparing their predicted proteomes to that of their respective six-frame translated genomes. We observe that 4.61% of domains per species were falsely inferred to be absent for Pfam domains predicted to have been present in the last eukaryotic common ancestor. Between phylogenetically different categories this estimate varies substantially: for clade-specific loss (ancestral loss) we found 1.30% and for species-specific loss 16.88% to be falsely inferred as absent. For BUSCO 1-to-1 orthologous families, 18.30% were falsely inferred to be absent. Finally, we showed that falsely inferred absences indeed impact loss estimates, with the number of losses decreasing by 11.78%. Our work strengthens the increasing number of studies showing that gene loss is an important factor in eukaryotic genome evolution. However, while we demonstrate that on average inferring gene absences from predicted proteomes is reliable, caution is warranted when inferring species-specific absences.


Assuntos
Eucariotos/genética , Evolução Molecular , Animais , Biologia Computacional , Deleção de Genes , Duplicação Gênica , Genoma , Humanos , Filogenia , Domínios Proteicos/genética , Proteoma , Especificidade da Espécie
6.
Nucleic Acids Res ; 45(18): 10634-10648, 2017 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-28977405

RESUMO

Hotspots of rapid genome evolution hold clues about human adaptation. We present a comparative analysis of nine whole-genome sequenced primates to identify high-confidence targets of positive selection. We find strong statistical evidence for positive selection in 331 protein-coding genes (3%), pinpointing 934 adaptively evolving codons (0.014%). Our new procedure is stringent and reveals substantial artefacts (20% of initial predictions) that have inflated previous estimates. The final 331 positively selected genes (PSG) are strongly enriched for innate and adaptive immunity, secreted and cell membrane proteins (e.g. pattern recognition, complement, cytokines, immune receptors, MHC, Siglecs). We also find evidence for positive selection in reproduction and chromosome segregation (e.g. centromere-associated CENPO, CENPT), apolipoproteins, smell/taste receptors and mitochondrial proteins. Focusing on the virus-host interaction, we retrieve most evolutionary conflicts known to influence antiviral activity (e.g. TRIM5, MAVS, SAMHD1, tetherin) and predict 70 novel cases through integration with virus-human interaction data. Protein structure analysis further identifies positive selection in the interaction interfaces between viruses and their cellular receptors (CD4-HIV; CD46-measles, adenoviruses; CD55-picornaviruses). Finally, primate PSG consistently show high sequence variation in human exomes, suggesting ongoing evolution. Our curated dataset of positive selection is a rich source for studying the genetics underlying human (antiviral) phenotypes. Procedures and data are available at https://github.com/robinvanderlee/positive-selection.


Assuntos
Evolução Molecular , Seleção Genética , Animais , Artefatos , Conversão Gênica , Variação Genética , Genômica , Interações Hospedeiro-Patógeno/genética , Humanos , Imunidade/genética , Família Multigênica , Primatas/genética , Proteínas/genética , Receptores Virais/química , Proteínas Virais/química , Viroses/genética
7.
Hum Mol Genet ; 25(3): 497-513, 2016 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-26643951

RESUMO

Oral-facial-digital (OFD) syndromes are rare heterogeneous disorders characterized by the association of abnormalities of the face, the oral cavity and the extremities, some due to mutations in proteins of the transition zone of the primary cilia or the closely associated distal end of centrioles. These two structures are essential for the formation of functional cilia, and for signaling events during development. We report here causal compound heterozygous mutations of KIAA0753/OFIP in a patient with an OFD VI syndrome. We show that the KIAA0753/OFIP protein, whose sequence is conserved in ciliated species, associates with centrosome/centriole and pericentriolar satellites in human cells and forms a complex with FOR20 and OFD1. The decreased expression of any component of this ternary complex in RPE1 cells causes a defective recruitment onto centrosomes and satellites. The OFD KIAA0753/OFIP mutant loses its capacity to interact with FOR20 and OFD1, which may be the molecular basis of the defect. We also show that KIAA0753/OFIP has microtubule-stabilizing activity. OFD1 and FOR20 are known to regulate the integrity of the centriole distal end, confirming that this structural element is a target of importance for pathogenic mutations in ciliopathies.


Assuntos
Centríolos/metabolismo , Centrossomo/metabolismo , Proteínas Associadas aos Microtúbulos/metabolismo , Síndromes Orofaciodigitais/metabolismo , Proteínas/metabolismo , Sequência de Aminoácidos , Sequência de Bases , Centríolos/ultraestrutura , Centrossomo/ultraestrutura , Cílios/genética , Cílios/metabolismo , Cílios/patologia , Sequência Conservada , Feminino , Expressão Gênica , Heterozigoto , Humanos , Recém-Nascido , Proteínas Associadas aos Microtúbulos/genética , Dados de Sequência Molecular , Mutação , Síndromes Orofaciodigitais/genética , Síndromes Orofaciodigitais/patologia , Ligação Proteica , Proteínas/genética , Alinhamento de Sequência
8.
Proc Natl Acad Sci U S A ; 110(17): 6943-8, 2013 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-23569277

RESUMO

The intraflagellar transport (IFT) complex is an integral component of the cilium, a quintessential organelle of the eukaryotic cell. The IFT system consists of three subcomplexes [i.e., intraflagellar transport (IFT)-A, IFT-B, and the BBSome], which together transport proteins and other molecules along the cilium. IFT dysfunction results in diseases collectively called ciliopathies. It has been proposed that the IFT complexes originated from vesicle coats similar to coat protein complex (COP) I, COPII, and clathrin. Here we provide phylogenetic evidence for common ancestry of IFT subunits and α, ß', and ε subunits of COPI, and trace the origins of the IFT-A, IFT-B, and the BBSome subcomplexes. We find that IFT-A and the BBSome likely arose from an IFT-B-like complex by intracomplex subunit duplication. The distribution of IFT proteins across eukaryotes identifies the BBSome as a frequently lost, modular component of the IFT. Significantly, loss of the BBSome from a taxon is a frequent precursor to complete cilium loss in related taxa. Given the inferred late origin of the BBSome in cilium evolution and its frequent loss, the IFT complex behaves as a "last-in, first-out" system. The protocoatomer origin of the IFT complex corroborates involvement of IFT components in vesicle transport. Expansion of IFT subunits by duplication and their subsequent independent loss supports the idea of modularity and structural independence of the IFT subcomplexes.


Assuntos
Proteínas de Transporte/genética , Cílios/fisiologia , Evolução Molecular , Flagelos/fisiologia , Modelos Moleculares , Complexos Multiproteicos/genética , Filogenia , Sequência de Aminoácidos , Sequência de Bases , Transporte Biológico/genética , Análise por Conglomerados , Complexo I de Proteína do Envoltório/genética , Humanos , Dados de Sequência Molecular , Complexos Multiproteicos/metabolismo , Dobramento de Proteína , Subunidades Proteicas/genética , Alinhamento de Sequência , Análise de Sequência de DNA , Trypanosoma brucei brucei
9.
PLoS One ; 17(4): e0251833, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35421089

RESUMO

Phylogenetic profiling in eukaryotes is of continued interest to study and predict the functional relationships between proteins. This interest is likely driven by the increased number of available diverse genomes and computational methods to infer orthologies. The evaluation of phylogenetic profiles has mainly focussed on reference genome selection in prokaryotes. However, it has been proven to be challenging to obtain high prediction accuracies in eukaryotes. As part of our recent comparison of orthology inference methods for eukaryotic genomes, we observed a surprisingly high performance for predicting interacting orthologous groups. This high performance, in turn, prompted the question of what factors influence the success of phylogenetic profiling when applied to eukaryotic genomes. Here we analyse the effect of species, orthologous group and interactome selection on protein interaction prediction using phylogenetic profiles. We select species based on the diversity and quality of the genomes and compare this supervised selection with randomly generated genome subsets. We also analyse the effect on the performance of orthologous groups defined to be in the last eukaryotic common ancestor of eukaryotes to that of orthologous groups that are not. Finally, we consider the effects of reference interactome set filtering and reference interactome species. In agreement with other studies, we find an effect of genome selection based on quality, less of an effect based on genome diversity, but a more notable effect based on the amount of information contained within the genomes. Most importantly, we find it is not merely selecting the correct genomes that is important for high prediction performance. Other choices in meta parameters such as orthologous group selection, the reference species of the interaction set, and the quality of the interaction set have a much larger impact on the performance when predicting protein interactions using phylogenetic profiles. These findings shed light on the differences in reported performance amongst phylogenetic profiles approaches, and reveal on a more fundamental level for which types of protein interactions this method has most promise when applied to eukaryotes.


Assuntos
Eucariotos , Genoma , Eucariotos/genética , Células Eucarióticas , Evolução Molecular , Filogenia , Células Procarióticas
10.
J Mol Evol ; 73(3-4): 209-20, 2011 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-22057117

RESUMO

The TOR kinase is a major regulator of growth in eukaryotes. Many components of the TOR pathway are implicated in cancer and metabolic diseases in humans. Analysis of the evolution of TOR and its pathway may provide fundamental insight into the evolution of growth regulation in eukaryotes and provide a practical framework on which experimental evidence can be compared between species. Here we performed phylogenetic analyses on the components of the TOR pathway and determined their point of invention. We find that the two TOR complexes and a large part of the TOR pathway originated before the Last Eukaryotic Common Ancestor and form a core to which new inputs have been added during animal evolution. In addition, we provide insight into how duplications and sub-functionalization of the S6K, RSK, SGK and PKB kinases shaped the complexity of the TOR pathway. In yeast we identify novel AGC kinases that are orthologous to the S6 kinase. These results demonstrate how a vital signaling pathway can be both highly conserved and flexible in eukaryotes.


Assuntos
Evolução Molecular , Transdução de Sinais/genética , Serina-Treonina Quinases TOR/genética , Sequência de Aminoácidos , Animais , Sequência Conservada , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Duplicação Gênica , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Humanos , Modelos Genéticos , Complexos Multiproteicos/genética , Filogenia , Proteínas de Protozoários/química , Proteínas de Protozoários/genética , Homologia de Sequência de Aminoácidos , Serina-Treonina Quinases TOR/química , Fatores de Transcrição/química , Fatores de Transcrição/genética
11.
Sci Immunol ; 6(62)2021 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-34417257

RESUMO

Tissue-resident memory CD8+ T cells (TRM) constitute a noncirculating memory T cell subset that provides early protection against reinfection. However, how TRM arise from antigen-triggered T cells has remained unclear. Exploiting the TRM-restricted expression of Hobit, we used TRM reporter/deleter mice to study TRM differentiation. We found that Hobit was up-regulated in a subset of LCMV-specific CD8+ T cells located within peripheral tissues during the effector phase of the immune response. These Hobit+ effector T cells were identified as TRM precursors, given that their depletion substantially decreased TRM development but not the formation of circulating memory T cells. Adoptive transfer experiments of Hobit+ effector T cells corroborated their biased contribution to the TRM lineage. Transcriptional profiling of Hobit+ effector T cells underlined the early establishment of TRM properties including down-regulation of tissue exit receptors and up-regulation of TRM-associated molecules. We identified Eomes as a key factor instructing the early bifurcation of circulating and resident lineages. These findings establish that commitment of TRM occurs early in antigen-driven T cell differentiation and reveal the molecular mechanisms underlying this differentiation pathway.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Células T de Memória/imunologia , Proteínas com Domínio T/imunologia , Animais , Diferenciação Celular , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos
12.
J Cell Biol ; 219(1)2020 01 06.
Artigo em Inglês | MEDLINE | ID: mdl-31740506

RESUMO

Multi-ciliary arrays promote fluid flow and cellular motility using the polarized and coordinated beating of hundreds of motile cilia. Tetrahymena basal bodies (BBs) nucleate and position cilia, whereby BB-associated striated fibers (SFs) promote BB anchorage and orientation into ciliary rows. Mutants that shorten SFs cause disoriented BBs. In contrast to the cytotaxis model, we show that disoriented BBs with short SFs can regain normal orientation if SF length is restored. In addition, SFs adopt unique lengths by their shrinkage and growth to establish and maintain BB connections and cortical interactions in a ciliary force-dependent mechanism. Tetrahymena SFs comprise at least eight uniquely localizing proteins belonging to the SF-assemblin family. Loss of different proteins that localize to the SF base disrupts either SF steady-state length or ciliary force-induced SF elongation. Thus, the dynamic regulation of SFs promotes BB connections and cortical interactions to organize ciliary arrays.


Assuntos
Corpos Basais/fisiologia , Cílios/fisiologia , Proteínas Associadas aos Microtúbulos/metabolismo , Microtúbulos/metabolismo , Proteínas de Protozoários/metabolismo , Tetrahymena thermophila/crescimento & desenvolvimento , Tetrahymena thermophila/metabolismo , Fenômenos Mecânicos , Proteínas Associadas aos Microtúbulos/genética , Proteínas de Protozoários/genética , Tetrahymena thermophila/genética
13.
Biochim Biophys Acta Bioenerg ; 1861(8): 148202, 2020 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-32275929

RESUMO

Protein complexes from the oxidative phosphorylation (OXPHOS) system are assembled with the help of proteins called assembly factors. We here delineate the function of the inner mitochondrial membrane protein TMEM70, in which mutations have been linked to OXPHOS deficiencies, using a combination of BioID, complexome profiling and coevolution analyses. TMEM70 interacts with complex I and V and for both complexes the loss of TMEM70 results in the accumulation of an assembly intermediate followed by a reduction of the next assembly intermediate in the pathway. This indicates that TMEM70 has a role in the stability of membrane-bound subassemblies or in the membrane recruitment of subunits into the forming complex. Independent evidence for a role of TMEM70 in OXPHOS assembly comes from evolutionary analyses. The TMEM70/TMEM186/TMEM223 protein family, of which we show that TMEM186 and TMEM223 are mitochondrial in human as well, only occurs in species with OXPHOS complexes. Our results validate the use of combining complexome profiling with BioID and evolutionary analyses in elucidating congenital defects in protein complex assembly.


Assuntos
Complexo I de Transporte de Elétrons/metabolismo , Proteínas de Membrana/metabolismo , Proteínas Mitocondriais/metabolismo , ATPases Mitocondriais Próton-Translocadoras/metabolismo , Biotinilação , Evolução Molecular , Técnicas de Inativação de Genes , Células HEK293 , Humanos , Proteínas de Membrana/deficiência , Proteínas de Membrana/genética , Proteínas Mitocondriais/deficiência , Proteínas Mitocondriais/genética , Fosforilação Oxidativa , Ligação Proteica
14.
PLoS Comput Biol ; 4(7): e1000132, 2008 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-18711636

RESUMO

The formation of proteins into stable protein complexes plays a fundamental role in the operation of the cell. The study of the degree of evolutionary conservation of protein complexes between species and the evolution of protein-protein interactions has been hampered by lack of comprehensive coverage of the high-throughput (HTP) technologies that measure the interactome. We show that new high-throughput datasets on protein co-purification in yeast have a substantially lower false negative rate than previous datasets when compared to known complexes. These datasets are therefore more suitable to estimate the conservation of protein complex membership than hitherto possible. We perform comparative genomics between curated protein complexes from human and the HTP data in Saccharomyces cerevisiae to study the evolution of co-complex memberships. This analysis revealed that out of the 5,960 protein pairs that are part of the same complex in human, 2,216 are absent because both proteins lack an ortholog in S. cerevisiae, while for 1,828 the co-complex membership is disrupted because one of the two proteins lacks an ortholog. For the remaining 1,916 protein pairs, only 10% were never co-purified in the large-scale experiments. This implies a conservation level of co-complex membership of 90% when the genes coding for the protein pairs that participate in the same protein complex are also conserved. We conclude that the evolutionary dynamics of protein complexes are, by and large, not the result of network rewiring (i.e. acquisition or loss of co-complex memberships), but mainly due to genomic acquisition or loss of genes coding for subunits. We thus reveal evidence for the tight interrelation of genomic and network evolution.


Assuntos
Evolução Molecular , Proteínas/química , Animais , Cromatografia de Afinidade , Drosophila melanogaster/química , Humanos , Proteínas/isolamento & purificação , Saccharomyces cerevisiae/química , Espectrometria de Massas em Tandem
15.
PLoS One ; 14(5): e0216705, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31095607

RESUMO

The cilium is an essential organelle at the surface of mammalian cells whose dysfunction causes a wide range of genetic diseases collectively called ciliopathies. The current rate at which new ciliopathy genes are identified suggests that many ciliary components remain undiscovered. We generated and rigorously analyzed genomic, proteomic, transcriptomic and evolutionary data and systematically integrated these using Bayesian statistics into a predictive score for ciliary function. This resulted in 285 candidate ciliary genes. We generated independent experimental evidence of ciliary associations for 24 out of 36 analyzed candidate proteins using multiple cell and animal model systems (mouse, zebrafish and nematode) and techniques. For example, we show that OSCP1, which has previously been implicated in two distinct non-ciliary processes, causes ciliogenic and ciliopathy-associated tissue phenotypes when depleted in zebrafish. The candidate list forms the basis of CiliaCarta, a comprehensive ciliary compendium covering 956 genes. The resource can be used to objectively prioritize candidate genes in whole exome or genome sequencing of ciliopathy patients and can be accessed at http://bioinformatics.bio.uu.nl/john/syscilia/ciliacarta/.


Assuntos
Cílios/genética , Genômica , Animais , Teorema de Bayes , Caenorhabditis elegans/citologia , Caenorhabditis elegans/genética , Anotação de Sequência Molecular , Fenótipo , Reprodutibilidade dos Testes , Células Receptoras Sensoriais/metabolismo , Peixe-Zebra/genética
16.
Sci Rep ; 8(1): 410, 2018 01 11.
Artigo em Inglês | MEDLINE | ID: mdl-29323249

RESUMO

Plasmodium gametocytes are the sexual forms of the malaria parasite essential for transmission to mosquitoes. To better understand how gametocytes differ from asexual blood-stage parasites, we performed a systematic analysis of available 'omics data for P. falciparum and other Plasmodium species. 18 transcriptomic and proteomic data sets were evaluated for the presence of curated "gold standards" of 41 gametocyte-specific versus 46 non-gametocyte genes and integrated using Bayesian probabilities, resulting in gametocyte-specificity scores for all P. falciparum genes. To illustrate the utility of the gametocyte score, we explored newly predicted gametocyte-specific genes as potential biomarkers of gametocyte carriage and exposure. We analyzed the humoral immune response in field samples against 30 novel gametocyte-specific antigens and found five antigens to be differentially recognized by gametocyte carriers as compared to malaria-infected individuals without detectable gametocytes. We also validated the gametocyte-specificity of 15 identified gametocyte transcripts on culture material and samples from naturally infected individuals, resulting in eight transcripts that were >1000-fold higher expressed in gametocytes compared to asexual parasites and whose transcript abundance allowed gametocyte detection in naturally infected individuals. Our integrated genome-wide gametocyte-specificity scores provide a comprehensive resource to identify targets and monitor P. falciparum gametocytemia.


Assuntos
Perfilação da Expressão Gênica/métodos , Malária/imunologia , Plasmodium/fisiologia , Proteômica/métodos , Animais , Antígenos de Protozoários/genética , Antígenos de Protozoários/metabolismo , Teorema de Bayes , Bases de Dados Genéticas , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Imunidade Humoral , Malária/parasitologia , Plasmodium/imunologia , Análise Serial de Proteínas/métodos , Proteínas de Protozoários/genética , Proteínas de Protozoários/imunologia , Proteínas de Protozoários/metabolismo
17.
Cilia ; 6: 10, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29177046

RESUMO

BACKGROUND: Recent research into ciliary structure and function provides important insights into inherited diseases termed ciliopathies and other cilia-related disorders. This wealth of knowledge needs to be translated into a computational representation to be fully exploitable by the research community. To this end, members of the Gene Ontology (GO) and SYSCILIA Consortia have worked together to improve representation of ciliary substructures and processes in GO. METHODS: Members of the SYSCILIA and Gene Ontology Consortia suggested additions and changes to GO, to reflect new knowledge in the field. The project initially aimed to improve coverage of ciliary parts, and was then broadened to cilia-related biological processes. Discussions were documented in a public tracker. We engaged the broader cilia community via direct consultation and by referring to the literature. Ontology updates were implemented via ontology editing tools. RESULTS: So far, we have created or modified 127 GO terms representing parts and processes related to eukaryotic cilia/flagella or prokaryotic flagella. A growing number of biological pathways are known to involve cilia, and we continue to incorporate this knowledge in GO. The resulting expansion in GO allows more precise representation of experimentally derived knowledge, and SYSCILIA and GO biocurators have created 199 annotations to 50 human ciliary proteins. The revised ontology was also used to curate mouse proteins in a collaborative project. The revised GO and annotations, used in comparative 'before and after' analyses of representative ciliary datasets, improve enrichment results significantly. CONCLUSIONS: Our work has resulted in a broader and deeper coverage of ciliary composition and function. These improvements in ontology and protein annotation will benefit all users of GO enrichment analysis tools, as well as the ciliary research community, in areas ranging from microscopy image annotation to interpretation of high-throughput studies. We welcome feedback to further enhance the representation of cilia biology in GO.

18.
Nat Cell Biol ; 18(1): 122-31, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26595381

RESUMO

The transition zone (TZ) ciliary subcompartment is thought to control cilium composition and signalling by facilitating a protein diffusion barrier at the ciliary base. TZ defects cause ciliopathies such as Meckel-Gruber syndrome (MKS), nephronophthisis (NPHP) and Joubert syndrome (JBTS). However, the molecular composition and mechanisms underpinning TZ organization and barrier regulation are poorly understood. To uncover candidate TZ genes, we employed bioinformatics (coexpression and co-evolution) and identified TMEM107 as a TZ protein mutated in oral-facial-digital syndrome and JBTS patients. Mechanistic studies in Caenorhabditis elegans showed that TMEM-107 controls ciliary composition and functions redundantly with NPHP-4 to regulate cilium integrity, TZ docking and assembly of membrane to microtubule Y-link connectors. Furthermore, nematode TMEM-107 occupies an intermediate layer of the TZ-localized MKS module by organizing recruitment of the ciliopathy proteins MKS-1, TMEM-231 (JBTS20) and JBTS-14 (TMEM237). Finally, MKS module membrane proteins are immobile and super-resolution microscopy in worms and mammalian cells reveals periodic localizations within the TZ. This work expands the MKS module of ciliopathy-causing TZ proteins associated with diffusion barrier formation and provides insight into TZ subdomain architecture.


Assuntos
Cerebelo/anormalidades , Cílios/metabolismo , Proteínas de Membrana/metabolismo , Retina/anormalidades , Anormalidades Múltiplas/genética , Anormalidades Múltiplas/metabolismo , Animais , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Cerebelo/metabolismo , Anormalidades do Olho/genética , Anormalidades do Olho/metabolismo , Humanos , Doenças Renais Císticas/genética , Doenças Renais Císticas/metabolismo , Proteínas de Membrana/genética , Retina/metabolismo
19.
Nat Commun ; 7: 11491, 2016 05 13.
Artigo em Inglês | MEDLINE | ID: mdl-27173435

RESUMO

Cellular organelles provide opportunities to relate biological mechanisms to disease. Here we use affinity proteomics, genetics and cell biology to interrogate cilia: poorly understood organelles, where defects cause genetic diseases. Two hundred and seventeen tagged human ciliary proteins create a final landscape of 1,319 proteins, 4,905 interactions and 52 complexes. Reverse tagging, repetition of purifications and statistical analyses, produce a high-resolution network that reveals organelle-specific interactions and complexes not apparent in larger studies, and links vesicle transport, the cytoskeleton, signalling and ubiquitination to ciliary signalling and proteostasis. We observe sub-complexes in exocyst and intraflagellar transport complexes, which we validate biochemically, and by probing structurally predicted, disruptive, genetic variants from ciliary disease patients. The landscape suggests other genetic diseases could be ciliary including 3M syndrome. We show that 3M genes are involved in ciliogenesis, and that patient fibroblasts lack cilia. Overall, this organelle-specific targeting strategy shows considerable promise for Systems Medicine.


Assuntos
Cílios/metabolismo , Ciliopatias/genética , Nanismo/genética , Hipotonia Muscular/genética , Mapas de Interação de Proteínas , Proteínas/metabolismo , Coluna Vertebral/anormalidades , Transporte Biológico/fisiologia , Cromatografia de Afinidade/métodos , Ciliopatias/patologia , Ciliopatias/terapia , Análise Mutacional de DNA , Conjuntos de Dados como Assunto , Nanismo/patologia , Nanismo/terapia , Fibroblastos , Células HEK293 , Humanos , Espectrometria de Massas , Terapia de Alvo Molecular/métodos , Hipotonia Muscular/patologia , Hipotonia Muscular/terapia , Mapeamento de Interação de Proteínas/métodos , Proteínas/genética , Proteínas/isolamento & purificação , Proteômica/métodos , Coluna Vertebral/patologia , Análise de Sistemas
20.
Genome Biol ; 16: 293, 2015 Dec 29.
Artigo em Inglês | MEDLINE | ID: mdl-26714646

RESUMO

BACKGROUND: Joubert syndrome (JBTS) and related disorders are defined by cerebellar malformation (molar tooth sign), together with neurological symptoms of variable expressivity. The ciliary basis of Joubert syndrome related disorders frequently extends the phenotype to tissues such as the eye, kidney, skeleton and craniofacial structures. RESULTS: Using autozygome and exome analyses, we identified a null mutation in KIAA0556 in a multiplex consanguineous family with hallmark features of mild Joubert syndrome. Patient-derived fibroblasts displayed reduced ciliogenesis potential and abnormally elongated cilia. Investigation of disease pathophysiology revealed that Kiaa0556 (-/-) null mice possess a Joubert syndrome-associated brain-restricted phenotype. Functional studies in Caenorhabditis elegans nematodes and cultured human cells support a conserved ciliary role for KIAA0556 linked to microtubule regulation. First, nematode KIAA0556 is expressed almost exclusively in ciliated cells, and the worm and human KIAA0556 proteins are enriched at the ciliary base. Second, C. elegans KIAA0056 regulates ciliary A-tubule number and genetically interacts with an ARL13B (JBTS8) orthologue to control cilium integrity. Third, human KIAA0556 binds to microtubules in vitro and appears to stabilise microtubule networks when overexpressed. Finally, human KIAA0556 biochemically interacts with ciliary proteins and p60/p80 katanins. The latter form a microtubule-severing enzyme complex that regulates microtubule dynamics as well as ciliary functions. CONCLUSIONS: We have identified KIAA0556 as a novel microtubule-associated ciliary base protein mutated in Joubert syndrome. Consistent with the mild patient phenotype, our nematode, mice and human cell data support the notion that KIAA0556 has a relatively subtle and variable cilia-related function, which we propose is related to microtubule regulation.


Assuntos
Corpos Basais/metabolismo , Cerebelo/anormalidades , Proteínas Associadas aos Microtúbulos/genética , Mutação , Retina/anormalidades , Fatores de Ribosilação do ADP/metabolismo , Anormalidades Múltiplas/genética , Anormalidades Múltiplas/patologia , Adenosina Trifosfatases/metabolismo , Adulto , Animais , Corpos Basais/patologia , Encéfalo/metabolismo , Encéfalo/patologia , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Células Cultivadas , Cerebelo/patologia , Criança , Pré-Escolar , Cílios/genética , Cílios/patologia , Exoma , Anormalidades do Olho/genética , Anormalidades do Olho/patologia , Feminino , Humanos , Katanina , Doenças Renais Císticas/genética , Doenças Renais Císticas/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Associadas aos Microtúbulos/metabolismo , Microtúbulos/metabolismo , Linhagem , Ligação Proteica , Retina/patologia
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