ArticlePLoS genetics2014
A massively parallel pipeline to clone DNA variants and examine molecular phenotypes of human disease mutations.
Article in PLoS genetics, 2014. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 36 papers.
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Who cites it
36 citing papers in PubMed, 57 citations in OpenAlex.
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- Uncovering cell-type-specific immunomodulatory variants and molecular phenotypes in COVID-19 using structurally resolved protein networks.Cell reports · 2024Article
- The genetic architecture of protein stability.Nature · 2024Article
- A multiscale functional map of somatic mutations in cancer integrating protein structure and network topology.bioRxiv : the preprint server for biology · 2024Article
- Structurally-informed human interactome reveals proteome-wide perturbations by disease mutations.bioRxiv : the preprint server for biology · 2024Article
- In vivo versus in silico assessment of potentially pathogenic missense variants in human reproductive genes.Proceedings of the National Academy of Sciences of the United States of America · 2023Article
- HAP1, a new revolutionary cell model for gene editing using CRISPR-Cas9.Frontiers in cell and developmental biology · 2023Review
- Predicting functional consequences of mutations using molecular interaction network features.Human genetics · 2022Article
- Article
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- A 3D structural SARS-CoV-2-human interactome to explore genetic and drug perturbations.Nature methods · 2021Article
- Human MLH1/3 variants causing aneuploidy, pregnancy loss, and premature reproductive aging.Nature communications · 2021Article
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- Strategies to Identify Genetic Variants Causing Infertility.Trends in molecular medicine · 2021Review
- In-depth and 3-dimensional exploration of the budding yeast phosphoproteome.EMBO reports · 2021Article
- Transcription imparts architecture, function and logic to enhancer units.Nature genetics · 2020Article
Corrections and comments
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Authors and funding
13 authors at 2 institutions in 2 countries.
Funding
Abstract
Understanding the functional relevance of DNA variants is essential for all exome and genome sequencing projects. However, current mutagenesis cloning protocols require Sanger sequencing, and thus are prohibitively costly and labor-intensive. We describe a massively-parallel site-directed mutagenesis approach, "Clone-seq", leveraging next-generation sequencing to rapidly and cost-effectively generate a large number of mutant alleles. Using Clone-seq, we further develop a comparative interactome-scanning pipeline integrating high-throughput GFP, yeast two-hybrid (Y2H), and mass spectrometry assays to systematically evaluate the functional impact of mutations on protein stability and interactions. We use this pipeline to show that disease mutations on protein-protein interaction interfaces are significantly more likely than those away from interfaces to disrupt corresponding interactions. We also find that mutation pairs with similar molecular phenotypes in terms of both protein stability and interactions are significantly more likely to cause the same disease than those with different molecular phenotypes, validating the in vivo biological relevance of our high-throughput GFP and Y2H assays, and indicating that both assays can be used to determine candidate disease mutations in the future. The general scheme of our experimental pipeline can be readily expanded to other types of interactome-mapping methods to comprehensively evaluate the functional relevance of all DNA variants, including those in non-coding regions.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.