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1.
Am J Hum Genet ; 108(7): 1283-1300, 2021 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-34214447

RESUMO

Most rare clinical missense variants cannot currently be classified as pathogenic or benign. Deficiency in human 5,10-methylenetetrahydrofolate reductase (MTHFR), the most common inherited disorder of folate metabolism, is caused primarily by rare missense variants. Further complicating variant interpretation, variant impacts often depend on environment. An important example of this phenomenon is the MTHFR variant p.Ala222Val (c.665C>T), which is carried by half of all humans and has a phenotypic impact that depends on dietary folate. Here we describe the results of 98,336 variant functional-impact assays, covering nearly all possible MTHFR amino acid substitutions in four folinate environments, each in the presence and absence of p.Ala222Val. The resulting atlas of MTHFR variant effects reveals many complex dependencies on both folinate and p.Ala222Val. MTHFR atlas scores can distinguish pathogenic from benign variants and, among individuals with severe MTHFR deficiency, correlate with age of disease onset. Providing a powerful tool for understanding structure-function relationships, the atlas suggests a role for a disordered loop in retaining cofactor at the active site and identifies variants that enable escape of inhibition by S-adenosylmethionine. Thus, a model based on eight MTHFR variant effect maps illustrates how shifting landscapes of environment- and genetic-background-dependent missense variation can inform our clinical, structural, and functional understanding of MTHFR deficiency.


Assuntos
Metilenotetra-Hidrofolato Redutase (NADPH2)/genética , Mutação de Sentido Incorreto , Substituição de Aminoácidos , Análise Mutacional de DNA , Diploide , Biblioteca Gênica , Genótipo , Humanos , Metilenotetra-Hidrofolato Redutase (NADPH2)/deficiência , Metilenotetra-Hidrofolato Redutase (NADPH2)/fisiologia , Saccharomyces cerevisiae/genética
2.
Bioinformatics ; 2021 Mar 27.
Artigo em Inglês | MEDLINE | ID: mdl-33774657

RESUMO

SUMMARY: Multiplexed assays of variant effect (MAVEs) are capable of experimentally testing all possible single nucleotide or amino acid variants in selected genomic regions, generating 'variant effect maps', which provide biochemical insight and functional evidence to enable more rapid and accurate clinical interpretation of human variation. Because the international community applying MAVE approaches is growing rapidly, we developed the online MaveRegistry platform to catalyze collaboration, reduce redundant efforts, allow stakeholders to nominate targets, and enable tracking and sharing of progress on ongoing MAVE projects. AVAILABILITY AND IMPLEMENTATION: MaveRegistry service: https://registry.varianteffect.org. MaveRegistry source code: https://github.com/kvnkuang/maveregistry-front-end. SUPPLEMENTARY INFORMATION: no Supplementary data.

3.
Microorganisms ; 8(8)2020 Aug 04.
Artigo em Inglês | MEDLINE | ID: mdl-32759834

RESUMO

The Neurospora crassa AOD1 protein is a mitochondrial alternative oxidase that passes electrons directly from ubiquinol to oxygen. The enzyme is encoded by the nuclear aod-1 gene and is produced when the standard electron transport chain is inhibited. We previously identified eleven strains in the N. crassa single gene deletion library that were severely deficient in their ability to produce AOD1 when grown in the presence of chloramphenicol, an inhibitor of mitochondrial translation that is known to induce the enzyme. Three mutants affected previously characterized genes. In this report we examined the remaining mutants and found that the deficiency of AOD1 was due to secondary mutations in all but two of the strains. One of the authentic mutants contained a deletion of the yvh1 gene and was found to have a deficiency of aod-1 transcripts. The YVH1 protein localized to the nucleus and a post mitochondrial pellet from the cytoplasm. A zinc binding domain in the protein was required for rescue of the AOD1 deficiency. In other organisms YVH1 is required for ribosome assembly and mutants have multiple phenotypes. Lack of YVH1 in N. crassa likely also affects ribosome assembly leading to phenotypes that include altered regulation of AOD1 production.

4.
Nature ; 580(7803): 402-408, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32296183

RESUMO

Global insights into cellular organization and genome function require comprehensive understanding of the interactome networks that mediate genotype-phenotype relationships1,2. Here we present a human 'all-by-all' reference interactome map of human binary protein interactions, or 'HuRI'. With approximately 53,000 protein-protein interactions, HuRI has approximately four times as many such interactions as there are high-quality curated interactions from small-scale studies. The integration of HuRI with genome3, transcriptome4 and proteome5 data enables cellular function to be studied within most physiological or pathological cellular contexts. We demonstrate the utility of HuRI in identifying the specific subcellular roles of protein-protein interactions. Inferred tissue-specific networks reveal general principles for the formation of cellular context-specific functions and elucidate potential molecular mechanisms that might underlie tissue-specific phenotypes of Mendelian diseases. HuRI is a systematic proteome-wide reference that links genomic variation to phenotypic outcomes.


Assuntos
Proteoma/metabolismo , Espaço Extracelular/metabolismo , Humanos , Especificidade de Órgãos , Mapeamento de Interação de Proteínas
5.
Cell Syst ; 10(1): 25-38.e10, 2020 01 22.
Artigo em Inglês | MEDLINE | ID: mdl-31668799

RESUMO

Many traits are complex, depending non-additively on variant combinations. Even in model systems, such as the yeast S. cerevisiae, carrying out the high-order variant-combination testing needed to dissect complex traits remains a daunting challenge. Here, we describe "X-gene" genetic analysis (XGA), a strategy for engineering and profiling highly combinatorial gene perturbations. We demonstrate XGA on yeast ABC transporters by engineering 5,353 strains, each deleted for a random subset of 16 transporters, and profiling each strain's resistance to 16 compounds. XGA yielded 85,648 genotype-to-resistance observations, revealing high-order genetic interactions for 13 of the 16 transporters studied. Neural networks yielded intuitive functional models and guided exploration of fluconazole resistance, which was influenced non-additively by five genes. Together, our results showed that highly combinatorial genetic perturbation can functionally dissect complex traits, supporting pursuit of analogous strategies in human cells and other model systems.


Assuntos
Transporte Biológico/genética , Proteínas de Membrana Transportadoras/genética , Humanos
6.
G3 (Bethesda) ; 9(10): 3453-3465, 2019 10 07.
Artigo em Inglês | MEDLINE | ID: mdl-31444295

RESUMO

The Neurospora crassa nuclear aod-1 gene encodes an alternative oxidase that functions in mitochondria. The enzyme provides a branch from the standard electron transport chain by transferring electrons directly from ubiquinol to oxygen. In standard laboratory strains, aod-1 is transcribed at very low levels under normal growth conditions. However, if the standard electron transport chain is disrupted, aod-1 mRNA expression is induced and the AOD1 protein is produced. We previously identified a strain of N. crassa, that produces high levels of aod-1 transcript under non-inducing conditions. Here we have crossed this strain to a standard lab strain and determined the genomic sequences of the parents and several progeny. Analysis of the sequence data and the levels of aod-1 mRNA in uninduced cultures revealed that a frameshift mutation in the flbA gene results in the high uninduced expression of aod-1 The flbA gene encodes a regulator of G protein signaling that decreases the activity of the Gα subunit of heterotrimeric G proteins. Our data suggest that strains with a functional flbA gene prevent uninduced expression of aod-1 by inactivating a G protein signaling pathway, and that this pathway is activated in cells grown under conditions that induce aod-1 Induced cells with a deletion of the gene encoding the Gα protein still have a partial increase in aod-1 mRNA levels, suggesting a second pathway for inducing transcription of the gene in N. crassa We also present evidence that a translational control mechanism prevents production of AOD1 protein in uninduced cultures.


Assuntos
Proteínas de Ligação ao GTP/genética , Regulação Fúngica da Expressão Gênica , Proteínas Mitocondriais/biossíntese , Neurospora crassa/genética , Neurospora crassa/metabolismo , Oxirredutases/biossíntese , Proteínas de Plantas/biossíntese , Mutação , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
7.
Nat Commun ; 10(1): 1240, 2019 03 18.
Artigo em Inglês | MEDLINE | ID: mdl-30886144

RESUMO

Despite exceptional experimental efforts to map out the human interactome, the continued data incompleteness limits our ability to understand the molecular roots of human disease. Computational tools offer a promising alternative, helping identify biologically significant, yet unmapped protein-protein interactions (PPIs). While link prediction methods connect proteins on the basis of biological or network-based similarity, interacting proteins are not necessarily similar and similar proteins do not necessarily interact. Here, we offer structural and evolutionary evidence that proteins interact not if they are similar to each other, but if one of them is similar to the other's partners. This approach, that mathematically relies on network paths of length three (L3), significantly outperforms all existing link prediction methods. Given its high accuracy, we show that L3 can offer mechanistic insights into disease mechanisms and can complement future experimental efforts to complete the human interactome.


Assuntos
Modelos Biológicos , Mapeamento de Interação de Proteínas/métodos , Mapas de Interação de Proteínas , Algoritmos , Animais , Proteínas de Arabidopsis/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Biologia Computacional/métodos , Conjuntos de Dados como Assunto , Proteínas de Drosophila/metabolismo , Humanos , Camundongos , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Schizosaccharomyces pombe/metabolismo , Software
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