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@article{sourisseau_deep_2019,
title = {Deep {Mutational} {Scanning} {Comprehensively} {Maps} {How} {Zika} {Envelope} {Protein} {Mutations} {Affect} {Viral} {Growth} and {Antibody} {Escape}},
volume = {93},
issn = {0022-538X, 1098-5514},
url = {https://journals.asm.org/doi/10.1128/JVI.01291-19},
doi = {10.1128/JVI.01291-19},
abstract = {Zika virus has recently been shown to be associated with severe birth defects. The virus’s E protein mediates its ability to infect cells and is also the primary target of the antibodies that are elicited by natural infection and vaccines that are being developed against the virus. Therefore, determining the effects of mutations to this protein is important for understanding its function, its susceptibility to vaccine-mediated immunity, and its potential for future evolution. We completely mapped how amino acid mutations to the E protein affected the virus’s ability to grow in cells in the laboratory and escape from several antibodies. The resulting maps relate changes in the E protein’s sequence to changes in viral function and therefore provide a valuable complement to existing maps of the physical structure of the protein.
,
ABSTRACT
Functional constraints on viral proteins are often assessed by examining sequence conservation among natural strains, but this approach is relatively ineffective for Zika virus because all known sequences are highly similar. Here, we take an alternative approach to map functional constraints on Zika virus’s envelope (E) protein by using deep mutational scanning to measure how all amino acid mutations to the E protein affect viral growth in cell culture. The resulting sequence-function map is consistent with existing knowledge about E protein structure and function but also provides insight into mutation-level constraints in many regions of the protein that have not been well characterized in prior functional work. In addition, we extend our approach to completely map how mutations affect viral neutralization by two monoclonal antibodies, thereby precisely defining their functional epitopes. Overall, our study provides a valuable resource for understanding the effects of mutations to this important viral protein and also offers a roadmap for future work to map functional and antigenic selection to Zika virus at high resolution.
IMPORTANCE
Zika virus has recently been shown to be associated with severe birth defects. The virus’s E protein mediates its ability to infect cells and is also the primary target of the antibodies that are elicited by natural infection and vaccines that are being developed against the virus. Therefore, determining the effects of mutations to this protein is important for understanding its function, its susceptibility to vaccine-mediated immunity, and its potential for future evolution. We completely mapped how amino acid mutations to the E protein affected the virus’s ability to grow in cells in the laboratory and escape from several antibodies. The resulting maps relate changes in the E protein’s sequence to changes in viral function and therefore provide a valuable complement to existing maps of the physical structure of the protein.},
language = {en},
number = {23},
urldate = {2022-07-11},
journal = {Journal of Virology},
author = {Sourisseau, Marion and Lawrence, Daniel J. P. and Schwarz, Megan C. and Storrs, Carina H. and Veit, Ethan C. and Bloom, Jesse D. and Evans, Matthew J.},
editor = {Pfeiffer, Julie K.},
month = dec,
year = {2019},
pages = {e01291--19},
file = {Full Text:/Users/admin/Zotero/storage/D4BDUGQ6/Sourisseau et al. - 2019 - Deep Mutational Scanning Comprehensively Maps How .pdf:application/pdf},
}
@article{deng_deep_2012,
title = {Deep {Sequencing} of {Systematic} {Combinatorial} {Libraries} {Reveals} Beta-{Lactamase} {Sequence} {Constraints} at {High} {Resolution}},
volume = {424},
issn = {00222836},
url = {https://linkinghub.elsevier.com/retrieve/pii/S0022283612007711},
doi = {10.1016/j.jmb.2012.09.014},
language = {en},
number = {3-4},
urldate = {2022-07-11},
journal = {Journal of Molecular Biology},
author = {Deng, Zhifeng and Huang, Wanzhi and Bakkalbasi, Erol and Brown, Nicholas G. and Adamski, Carolyn J. and Rice, Kacie and Muzny, Donna and Gibbs, Richard A. and Palzkill, Timothy},
month = dec,
year = {2012},
pages = {150--167},
file = {Accepted Version:/Users/admin/Zotero/storage/X5RENMCM/Deng et al. - 2012 - Deep Sequencing of Systematic Combinatorial Librar.pdf:application/pdf},
}
@article{amorosi_massively_2021,
title = {Massively parallel characterization of {CYP2C9} variant enzyme activity and abundance},
volume = {108},
issn = {00029297},
url = {https://linkinghub.elsevier.com/retrieve/pii/S000292972100269X},
doi = {10.1016/j.ajhg.2021.07.001},
language = {en},
number = {9},
urldate = {2022-07-11},
journal = {The American Journal of Human Genetics},
author = {Amorosi, Clara J. and Chiasson, Melissa A. and McDonald, Matthew G. and Wong, Lai Hong and Sitko, Katherine A. and Boyle, Gabriel and Kowalski, John P. and Rettie, Allan E. and Fowler, Douglas M. and Dunham, Maitreya J.},
month = sep,
year = {2021},
pages = {1735--1751},
file = {Full Text:/Users/admin/Zotero/storage/8DWGHK9I/Amorosi et al. - 2021 - Massively parallel characterization of CYP2C9 vari.pdf:application/pdf},
}
@article{chiasson_multiplexed_2020,
title = {Multiplexed measurement of variant abundance and activity reveals {VKOR} topology, active site and human variant impact},
volume = {9},
issn = {2050-084X},
url = {https://elifesciences.org/articles/58026},
doi = {10.7554/eLife.58026},
abstract = {Vitamin K epoxide reductase (VKOR) drives the vitamin K cycle, activating vitamin K-dependent blood clotting factors. VKOR is also the target of the widely used anticoagulant drug, warfarin. Despite VKOR’s pivotal role in coagulation, its structure and active site remain poorly understood. In addition, VKOR variants can cause vitamin K-dependent clotting factor deficiency or alter warfarin response. Here, we used multiplexed, sequencing-based assays to measure the effects of 2,695 VKOR missense variants on abundance and 697 variants on activity in cultured human cells. The large-scale functional data, along with an evolutionary coupling analysis, supports a four transmembrane domain topology, with variants in transmembrane domains exhibiting strongly deleterious effects on abundance and activity. Functionally constrained regions of the protein define the active site, and we find that, of four conserved cysteines putatively critical for function, only three are absolutely required. Finally, 25\% of human VKOR missense variants show reduced abundance or activity, possibly conferring warfarin sensitivity or causing disease.},
language = {en},
urldate = {2022-07-11},
journal = {eLife},
author = {Chiasson, Melissa A and Rollins, Nathan J and Stephany, Jason J and Sitko, Katherine A and Matreyek, Kenneth A and Verby, Marta and Sun, Song and Roth, Frederick P and DeSloover, Daniel and Marks, Debora S and Rettie, Allan E and Fowler, Douglas M},
month = sep,
year = {2020},
pages = {e58026},
file = {Full Text:/Users/admin/Zotero/storage/GWWNSVDK/Chiasson et al. - 2020 - Multiplexed measurement of variant abundance and a.pdf:application/pdf},
}
@article{wrenbeck_single-mutation_2017,
title = {Single-mutation fitness landscapes for an enzyme on multiple substrates reveal specificity is globally encoded},
volume = {8},
issn = {2041-1723},
url = {http://www.nature.com/articles/ncomms15695},
doi = {10.1038/ncomms15695},
language = {en},
number = {1},
urldate = {2022-07-11},
journal = {Nature Communications},
author = {Wrenbeck, Emily E. and Azouz, Laura R. and Whitehead, Timothy A.},
month = aug,
year = {2017},
pages = {15695},
file = {Full Text:/Users/admin/Zotero/storage/7WF3NED2/Wrenbeck et al. - 2017 - Single-mutation fitness landscapes for an enzyme o.pdf:application/pdf},
}
@article{brenan_phenotypic_2016,
title = {Phenotypic {Characterization} of a {Comprehensive} {Set} of {MAPK1} /{ERK2} {Missense} {Mutants}},
volume = {17},
issn = {22111247},
url = {https://linkinghub.elsevier.com/retrieve/pii/S2211124716313171},
doi = {10.1016/j.celrep.2016.09.061},
language = {en},
number = {4},
urldate = {2022-07-11},
journal = {Cell Reports},
author = {Brenan, Lisa and Andreev, Aleksandr and Cohen, Ofir and Pantel, Sasha and Kamburov, Atanas and Cacchiarelli, Davide and Persky, Nicole S. and Zhu, Cong and Bagul, Mukta and Goetz, Eva M. and Burgin, Alex B. and Garraway, Levi A. and Getz, Gad and Mikkelsen, Tarjei S. and Piccioni, Federica and Root, David E. and Johannessen, Cory M.},
month = oct,
year = {2016},
pages = {1171--1183},
file = {Full Text:/Users/admin/Zotero/storage/W3B46C64/Brenan et al. - 2016 - Phenotypic Characterization of a Comprehensive Set.pdf:application/pdf},
}
@article{kozek_high-throughput_2020,
title = {High-throughput discovery of trafficking-deficient variants in the cardiac potassium channel {KV11}.1},
volume = {17},
issn = {15475271},
url = {https://linkinghub.elsevier.com/retrieve/pii/S1547527120305427},
doi = {10.1016/j.hrthm.2020.05.041},
language = {en},
number = {12},
urldate = {2022-07-11},
journal = {Heart Rhythm},
author = {Kozek, Krystian A. and Glazer, Andrew M. and Ng, Chai-Ann and Blackwell, Daniel and Egly, Christian L. and Vanags, Loren R. and Blair, Marcia and Mitchell, Devyn and Matreyek, Kenneth A. and Fowler, Douglas M. and Knollmann, Bjorn C. and Vandenberg, Jamie I. and Roden, Dan M. and Kroncke, Brett M.},
month = dec,
year = {2020},
pages = {2180--2189},
file = {Accepted Version:/Users/admin/Zotero/storage/JETSTCHG/Kozek et al. - 2020 - High-throughput discovery of trafficking-deficient.pdf:application/pdf},
}
@article{davidi_highly_2020,
title = {Highly active rubiscos discovered by systematic interrogation of natural sequence diversity},
volume = {39},
issn = {0261-4189, 1460-2075},
url = {https://onlinelibrary.wiley.com/doi/10.15252/embj.2019104081},
doi = {10.15252/embj.2019104081},
language = {en},
number = {18},
urldate = {2022-07-11},
journal = {The EMBO Journal},
author = {Davidi, Dan and Shamshoum, Melina and Guo, Zhijun and Bar‐On, Yinon M and Prywes, Noam and Oz, Aia and Jablonska, Jagoda and Flamholz, Avi and Wernick, David G and Antonovsky, Niv and Pins, Benoit and Shachar, Lior and Hochhauser, Dina and Peleg, Yoav and Albeck, Shira and Sharon, Itai and Mueller‐Cajar, Oliver and Milo, Ron},
month = sep,
year = {2020},
file = {Full Text:/Users/admin/Zotero/storage/PYUPVUZK/Davidi et al. - 2020 - Highly active rubiscos discovered by systematic in.pdf:application/pdf},
}
@article{sarkisyan_local_2016,
title = {Local fitness landscape of the green fluorescent protein},
volume = {533},
issn = {0028-0836, 1476-4687},
url = {http://www.nature.com/articles/nature17995},
doi = {10.1038/nature17995},
language = {en},
number = {7603},
urldate = {2022-07-11},
journal = {Nature},
author = {Sarkisyan, Karen S. and Bolotin, Dmitry A. and Meer, Margarita V. and Usmanova, Dinara R. and Mishin, Alexander S. and Sharonov, George V. and Ivankov, Dmitry N. and Bozhanova, Nina G. and Baranov, Mikhail S. and Soylemez, Onuralp and Bogatyreva, Natalya S. and Vlasov, Peter K. and Egorov, Evgeny S. and Logacheva, Maria D. and Kondrashov, Alexey S. and Chudakov, Dmitry M. and Putintseva, Ekaterina V. and Mamedov, Ilgar Z. and Tawfik, Dan S. and Lukyanov, Konstantin A. and Kondrashov, Fyodor A.},
month = may,
year = {2016},
pages = {397--401},
file = {Accepted Version:/Users/admin/Zotero/storage/KPQ2DLXG/Sarkisyan et al. - 2016 - Local fitness landscape of the green fluorescent p.pdf:application/pdf},
}
@article{olson_comprehensive_2014,
title = {A {Comprehensive} {Biophysical} {Description} of {Pairwise} {Epistasis} throughout an {Entire} {Protein} {Domain}},
volume = {24},
issn = {09609822},
url = {https://linkinghub.elsevier.com/retrieve/pii/S0960982214012688},
doi = {10.1016/j.cub.2014.09.072},
language = {en},
number = {22},
urldate = {2022-07-11},
journal = {Current Biology},
author = {Olson, C. Anders and Wu, Nicholas C. and Sun, Ren},
month = nov,
year = {2014},
pages = {2643--2651},
file = {Full Text:/Users/admin/Zotero/storage/XHQR6TLB/Olson et al. - 2014 - A Comprehensive Biophysical Description of Pairwis.pdf:application/pdf},
}
@article{duenas-decamp_saturation_2016,
title = {Saturation {Mutagenesis} of the {HIV}-1 {Envelope} {CD4} {Binding} {Loop} {Reveals} {Residues} {Controlling} {Distinct} {Trimer} {Conformations}},
volume = {12},
issn = {1553-7374},
url = {https://dx.plos.org/10.1371/journal.ppat.1005988},
doi = {10.1371/journal.ppat.1005988},
language = {en},
number = {11},
urldate = {2022-07-11},
journal = {PLOS Pathogens},
author = {Duenas-Decamp, Maria and Jiang, Li and Bolon, Daniel and Clapham, Paul R.},
editor = {Desrosiers, Ronald C.},
month = nov,
year = {2016},
pages = {e1005988},
file = {Full Text:/Users/admin/Zotero/storage/B948VI9X/Duenas-Decamp et al. - 2016 - Saturation Mutagenesis of the HIV-1 Envelope CD4 B.pdf:application/pdf},
}
@article{thompson_altered_2020,
title = {Altered expression of a quality control protease in {E}. coli reshapes the in vivo mutational landscape of a model enzyme},
volume = {9},
issn = {2050-084X},
url = {https://elifesciences.org/articles/53476},
doi = {10.7554/eLife.53476},
abstract = {Protein mutational landscapes are shaped by the cellular environment, but key factors and their quantitative effects are often unknown. Here we show that Lon, a quality control protease naturally absent in common
E. coli
expression strains, drastically reshapes the mutational landscape of the metabolic enzyme dihydrofolate reductase (DHFR). Selection under conditions that resolve highly active mutants reveals that 23.3\% of all single point mutations in DHFR are advantageous in the absence of Lon, but advantageous mutations are largely suppressed when Lon is reintroduced. Protein stability measurements demonstrate extensive activity-stability tradeoffs for the advantageous mutants and provide a mechanistic explanation for Lon’s widespread impact. Our findings suggest possibilities for tuning mutational landscapes by modulating the cellular environment, with implications for protein design and combatting antibiotic resistance.},
language = {en},
urldate = {2022-07-11},
journal = {eLife},
author = {Thompson, Samuel and Zhang, Yang and Ingle, Christine and Reynolds, Kimberly A and Kortemme, Tanja},
month = jul,
year = {2020},
pages = {e53476},
file = {Full Text:/Users/admin/Zotero/storage/4U2CHYQR/Thompson et al. - 2020 - Altered expression of a quality control protease i.pdf:application/pdf},
}
@article{starita_activity-enhancing_2013,
title = {Activity-enhancing mutations in an {E3} ubiquitin ligase identified by high-throughput mutagenesis},
volume = {110},
issn = {0027-8424, 1091-6490},
url = {https://pnas.org/doi/full/10.1073/pnas.1303309110},
doi = {10.1073/pnas.1303309110},
abstract = {Significance
Ubiquitin is a 76 residue protein that is attached to target proteins as a posttranslational modification. This modification is dependent on the successive activity of three enzymes, designated E1, E2, and E3. We developed a high-throughput mutagenesis strategy to probe the mechanism of E3-catalyzed transfer of ubiquitin from the E2 to the target protein. By scoring the effect of nearly 100,000 mutations in an E3, we identified mutations that affect direct and allosteric interactions between the E3 and the E2. These results highlight the general utility of high-throughput mutagenesis in delineating the molecular basis of enzyme activity.
,
Although ubiquitination plays a critical role in virtually all cellular processes, mechanistic details of ubiquitin (Ub) transfer are still being defined. To identify the molecular determinants within E3 ligases that modulate activity, we scored each member of a library of nearly 100,000 protein variants of the murine ubiquitination factor E4B (Ube4b) U-box domain for auto-ubiquitination activity in the presence of the E2 UbcH5c. This assay identified mutations that enhance activity both in vitro and in cellular p53 degradation assays. The activity-enhancing mutations fall into two distinct mechanistic classes: One increases the U-box:E2-binding affinity, and the other allosterically stimulates the formation of catalytically active conformations of the E2∼Ub conjugate. The same mutations enhance E3 activity in the presence of another E2, Ube2w, implying a common allosteric mechanism, and therefore the general applicability of our observations to other E3s. A comparison of the E3 activity with the two different E2s identified an additional variant that exhibits E3:E2 specificity. Our results highlight the general utility of high-throughput mutagenesis in delineating the molecular basis of enzyme activity.},
language = {en},
number = {14},
urldate = {2022-07-11},
journal = {Proceedings of the National Academy of Sciences},
author = {Starita, Lea M. and Pruneda, Jonathan N. and Lo, Russell S. and Fowler, Douglas M. and Kim, Helen J. and Hiatt, Joseph B. and Shendure, Jay and Brzovic, Peter S. and Fields, Stanley and Klevit, Rachel E.},
month = apr,
year = {2013},
file = {Full Text:/Users/admin/Zotero/storage/FBVQ2HYR/Starita et al. - 2013 - Activity-enhancing mutations in an E3 ubiquitin li.pdf:application/pdf;Full Text:/Users/admin/Zotero/storage/SF4WML4G/Starita et al. - 2013 - Activity-enhancing mutations in an E3 ubiquitin li.pdf:application/pdf},
}
@article{araya_fundamental_2012,
title = {A fundamental protein property, thermodynamic stability, revealed solely from large-scale measurements of protein function},
volume = {109},
issn = {0027-8424, 1091-6490},
url = {https://pnas.org/doi/full/10.1073/pnas.1209751109},
doi = {10.1073/pnas.1209751109},
abstract = {The ability of a protein to carry out a given function results from fundamental physicochemical properties that include the protein’s structure, mechanism of action, and thermodynamic stability. Traditional approaches to study these properties have typically required the direct measurement of the property of interest, oftentimes a laborious undertaking. Although protein properties can be probed by mutagenesis, this approach has been limited by its low throughput. Recent technological developments have enabled the rapid quantification of a protein’s function, such as binding to a ligand, for numerous variants of that protein. Here, we measure the ability of 47,000 variants of a WW domain to bind to a peptide ligand and use these functional measurements to identify stabilizing mutations without directly assaying stability. Our approach is rooted in the well-established concept that protein function is closely related to stability. Protein function is generally reduced by destabilizing mutations, but this decrease can be rescued by stabilizing mutations. Based on this observation, we introduce partner potentiation, a metric that uses this rescue ability to identify stabilizing mutations, and identify 15 candidate stabilizing mutations in the WW domain. We tested six candidates by thermal denaturation and found two highly stabilizing mutations, one more stabilizing than any previously known mutation. Thus, physicochemical properties such as stability are latent within these large-scale protein functional data and can be revealed by systematic analysis. This approach should allow other protein properties to be discovered.},
language = {en},
number = {42},
urldate = {2022-07-11},
journal = {Proceedings of the National Academy of Sciences},
author = {Araya, Carlos L. and Fowler, Douglas M. and Chen, Wentao and Muniez, Ike and Kelly, Jeffery W. and Fields, Stanley},
month = oct,
year = {2012},
pages = {16858--16863},
file = {Full Text:/Users/admin/Zotero/storage/KR46AVWQ/Araya et al. - 2012 - A fundamental protein property, thermodynamic stab.pdf:application/pdf},
}
@article{wu_high-throughput_2014,
title = {High-throughput profiling of influenza {A} virus hemagglutinin gene at single-nucleotide resolution},
volume = {4},
issn = {2045-2322},
url = {https://www.nature.com/articles/srep04942},
doi = {10.1038/srep04942},
abstract = {Abstract
Genetic research on influenza virus biology has been informed in large part by nucleotide variants present in seasonal or pandemic samples, or individual mutants generated in the laboratory, leaving a substantial part of the genome uncharacterized. Here, we have developed a single-nucleotide resolution genetic approach to interrogate the fitness effect of point mutations in 98\% of the amino acid positions in the influenza A virus hemagglutinin (HA) gene. Our HA fitness map provides a reference to identify indispensable regions to aid in drug and vaccine design as targeting these regions will increase the genetic barrier for the emergence of escape mutations. This study offers a new platform for studying genome dynamics, structure-function relationships, virus-host interactions and can further rational drug and vaccine design. Our approach can also be applied to any virus that can be genetically manipulated.},
language = {en},
number = {1},
urldate = {2022-07-11},
journal = {Scientific Reports},
author = {Wu, Nicholas C. and Young, Arthur P. and Al-Mawsawi, Laith Q. and Olson, C. Anders and Feng, Jun and Qi, Hangfei and Chen, Shu-Hwa and Lu, I.-Hsuan and Lin, Chung-Yen and Chin, Robert G. and Luan, Harding H. and Nguyen, Nguyen and Nelson, Stanley F. and Li, Xinmin and Wu, Ting-Ting and Sun, Ren},
month = dec,
year = {2014},
pages = {4942},
file = {Full Text:/Users/admin/Zotero/storage/IPZQQNJQ/Wu et al. - 2014 - High-throughput profiling of influenza A virus hem.pdf:application/pdf},
}
@techreport{young_deep_2021,
type = {preprint},
title = {Deep {Mutagenesis} of a {Transporter} for {Uptake} of a {Non}-{Native} {Substrate} {Identifies} {Conformationally} {Dynamic} {Regions}},
url = {http://biorxiv.org/lookup/doi/10.1101/2021.04.19.440442},
abstract = {Abstract
The serotonin transporter, SERT, catalyzes serotonin reuptake at the synapse to terminate neurotransmission via an alternating access mechanism, and SERT inhibitors are the most widely prescribed antidepressants. Here, deep mutagenesis is used to determine the effects of nearly all amino acid substitutions on human SERT surface expression and transport of the fluorescent substrate analogue APP+, identifying many mutations that enhance APP+ import. Comprehensive simulations of the entire ion-coupled import process reveal that while binding of the native substrate, serotonin, reduces free energy barriers between conformational states to promote SERT dynamics, the conformational free energy landscape in the presence of APP+ instead resembles Na
+
bound-SERT, with a higher free energy barrier for transitioning to an inward-facing state. The deep mutational scan for SERT-catalyzed import of APP+ finds mutations that promote the necessary conformational changes that would otherwise be facilitated by the native substrate. Indeed, hundreds of gain-of-function mutations for APP+ import are found along the permeation pathway, most notably mutations that favor opening of a solvent-exposed intracellular vestibule. The mutagenesis data support the simulated mechanism in which the neurotransmitter and a symported sodium share a common cytosolic exit pathway to achieve coupling. Furthermore, the mutational landscape for SERT surface trafficking, which likely filters out misfolded sequences, reveals that residues along the permeation pathway are mutationally tolerant, providing plausible evolutionary pathways for changes in transporter properties while maintaining folded structure.},
language = {en},
urldate = {2022-07-11},
institution = {Biochemistry},
author = {Young, Heather J. and Chan, Matthew and Selvam, Balaji and Szymanski, Steven K. and Shukla, Diwakar and Procko, Erik},
month = apr,
year = {2021},
doi = {10.1101/2021.04.19.440442},
file = {Submitted Version:/Users/admin/Zotero/storage/T69VH4DC/Young et al. - 2021 - Deep Mutagenesis of a Transporter for Uptake of a .pdf:application/pdf},
}
@article{jiang_balance_2016,
title = {A {Balance} between {Inhibitor} {Binding} and {Substrate} {Processing} {Confers} {Influenza} {Drug} {Resistance}},
volume = {428},
issn = {00222836},
url = {https://linkinghub.elsevier.com/retrieve/pii/S0022283615006907},
doi = {10.1016/j.jmb.2015.11.027},
language = {en},
number = {3},
urldate = {2022-07-11},
journal = {Journal of Molecular Biology},
author = {Jiang, Li and Liu, Ping and Bank, Claudia and Renzette, Nicholas and Prachanronarong, Kristina and Yilmaz, Lutfu S. and Caffrey, Daniel R. and Zeldovich, Konstantin B. and Schiffer, Celia A. and Kowalik, Timothy F. and Jensen, Jeffrey D. and Finberg, Robert W. and Wang, Jennifer P. and Bolon, Daniel N.A.},
month = feb,
year = {2016},
pages = {538--553},
file = {Accepted Version:/Users/admin/Zotero/storage/TJGBH3YJ/Jiang et al. - 2016 - A Balance between Inhibitor Binding and Substrate .pdf:application/pdf},
}
@article{melamed_deep_2013,
title = {Deep mutational scanning of an {RRM} domain of the \textit{{Saccharomyces} cerevisiae} poly({A})-binding protein},
volume = {19},
issn = {1355-8382, 1469-9001},
url = {http://rnajournal.cshlp.org/lookup/doi/10.1261/rna.040709.113},
doi = {10.1261/rna.040709.113},
abstract = {The RNA recognition motif (RRM) is the most common RNA-binding domain in eukaryotes. Differences in RRM sequences dictate, in part, both RNA and protein-binding specificities and affinities. We used a deep mutational scanning approach to study the sequence-function relationship of the RRM2 domain of the
Saccharomyces cerevisiae
poly(A)-binding protein (Pab1). By scoring the activity of more than 100,000 unique Pab1 variants, including 1246 with single amino acid substitutions, we delineated the mutational constraints on each residue. Clustering of residues with similar mutational patterns reveals three major classes, composed principally of RNA-binding residues, of hydrophobic core residues, and of the remaining residues. The first class also includes a highly conserved residue not involved in RNA binding, G150, which can be mutated to destabilize Pab1. A comparison of the mutational sensitivity of yeast Pab1 residues to their evolutionary conservation reveals that most residues tolerate more substitutions than are present in the natural sequences, although other residues that tolerate fewer substitutions may point to specialized functions in yeast. An analysis of ∼40,000 double mutants indicates a preference for a short distance between two mutations that display an epistatic interaction. As examples of interactions, the mutations N139T, N139S, and I157L suppress other mutations that interfere with RNA binding and protein stability. Overall, this study demonstrates that living cells can be subjected to a single assay to analyze hundreds of thousands of protein variants in parallel.},
language = {en},
number = {11},
urldate = {2022-07-11},
journal = {RNA},
author = {Melamed, Daniel and Young, David L. and Gamble, Caitlin E. and Miller, Christina R. and Fields, Stanley},
month = nov,
year = {2013},
pages = {1537--1551},
file = {Full Text:/Users/admin/Zotero/storage/AFZEI4G6/Melamed et al. - 2013 - Deep mutational scanning of an RRM domain of the .pdf:application/pdf},
}
@article{jia_massively_2021,
title = {Massively parallel functional testing of {MSH2} missense variants conferring {Lynch} syndrome risk},
volume = {108},
issn = {00029297},
url = {https://linkinghub.elsevier.com/retrieve/pii/S0002929720304390},
doi = {10.1016/j.ajhg.2020.12.003},
language = {en},
number = {1},
urldate = {2022-07-11},
journal = {The American Journal of Human Genetics},
author = {Jia, Xiaoyan and Burugula, Bala Bharathi and Chen, Victor and Lemons, Rosemary M. and Jayakody, Sajini and Maksutova, Mariam and Kitzman, Jacob O.},
month = jan,
year = {2021},
pages = {163--175},
file = {Full Text:/Users/admin/Zotero/storage/4J7YRE9U/Jia et al. - 2021 - Massively parallel functional testing of MSH2 miss.pdf:application/pdf},
}
@article{mavor_determination_2016,
title = {Determination of ubiquitin fitness landscapes under different chemical stresses in a classroom setting},
volume = {5},
issn = {2050-084X},
url = {https://elifesciences.org/articles/15802},
doi = {10.7554/eLife.15802},
abstract = {Ubiquitin is essential for eukaryotic life and varies in only 3 amino acid positions between yeast and humans. However, recent deep sequencing studies indicate that ubiquitin is highly tolerant to single mutations. We hypothesized that this tolerance would be reduced by chemically induced physiologic perturbations. To test this hypothesis, a class of first year UCSF graduate students employed deep mutational scanning to determine the fitness landscape of all possible single residue mutations in the presence of five different small molecule perturbations. These perturbations uncover 'shared sensitized positions' localized to areas around the hydrophobic patch and the C-terminus. In addition, we identified perturbation specific effects such as a sensitization of His68 in HU and a tolerance to mutation at Lys63 in DTT. Our data show how chemical stresses can reduce buffering effects in the ubiquitin proteasome system. Finally, this study demonstrates the potential of lab-based interdisciplinary graduate curriculum.
,
The ability of an organism to grow and reproduce, that is, it’s “fitness”, is determined by how its genes interact with the environment. Yeast is a model organism in which researchers can control the exact mutations present in the yeast’s genes (its genotype) and the conditions in which the yeast cells live (their environment). This allows researchers to measure how a yeast cell’s genotype and environment affect its fitness.
Ubiquitin is a protein that many organisms depend on to manage cell stress by acting as a tag that targets other proteins for degradation. Essential proteins such as ubiquitin often remain unchanged by mutation over long periods of time. As a result, these proteins evolve very slowly. Like all proteins, ubiquitin is built from a chain of amino acid molecules linked together, and the ubiquitin proteins of yeast and humans are made of almost identical sequences of amino acids.
Although ubiquitin has barely changed its sequence over evolution, previous studies have shown that – under normal growth conditions in the laboratory – most amino acids in ubiquitin can be mutated without any loss of cell fitness. This led Mavor et al. to hypothesize that treating the yeast cells with chemicals that cause cell stress might lead to amino acids in ubiquitin becoming more sensitive to mutation.
To test this idea, a class of graduate students at the University of California, San Francisco grew yeast cells with different ubiquitin mutations together, and with different chemicals that induce cell stress, and measured their growth rates. Sequencing the ubiquitin gene in the thousands of tested yeast cells revealed that three of the chemicals cause a shared set of amino acids in ubiquitin to become more sensitive to mutation.
This result suggests that these amino acids are important for the stress response, possibly by altering the ability of yeast cells to target certain proteins for degradation. Conversely, another chemical causes yeast to become more tolerant to changes in the ubiquitin sequence. The experiments also link changes in particular amino acids in ubiquitin to specific stress responses.
Mavor et al. show that many of ubquitin’s amino acids are sensitive to mutation under different stress conditions, while others can be mutated to form different amino acids without effecting fitness. By testing the effects of other chemicals, future experiments could further characterize how the yeast’s genotype and environment interact.},
language = {en},
urldate = {2022-07-11},
journal = {eLife},
author = {Mavor, David and Barlow, Kyle and Thompson, Samuel and Barad, Benjamin A and Bonny, Alain R and Cario, Clinton L and Gaskins, Garrett and Liu, Zairan and Deming, Laura and Axen, Seth D and Caceres, Elena and Chen, Weilin and Cuesta, Adolfo and Gate, Rachel E and Green, Evan M and Hulce, Kaitlin R and Ji, Weiyue and Kenner, Lillian R and Mensa, Bruk and Morinishi, Leanna S and Moss, Steven M and Mravic, Marco and Muir, Ryan K and Niekamp, Stefan and Nnadi, Chimno I and Palovcak, Eugene and Poss, Erin M and Ross, Tyler D and Salcedo, Eugenia C and See, Stephanie K and Subramaniam, Meena and Wong, Allison W and Li, Jennifer and Thorn, Kurt S and Conchúir, Shane Ó and Roscoe, Benjamin P and Chow, Eric D and DeRisi, Joseph L and Kortemme, Tanja and Bolon, Daniel N and Fraser, James S},
month = apr,
year = {2016},
pages = {e15802},
file = {Full Text:/Users/admin/Zotero/storage/H2575JWN/Mavor et al. - 2016 - Determination of ubiquitin fitness landscapes unde.pdf:application/pdf},
}
@article{glazer_deep_2020,
title = {Deep {Mutational} {Scan} of an \textit{{SCN5A}} {Voltage} {Sensor}},
volume = {13},
issn = {2574-8300},
url = {https://www.ahajournals.org/doi/10.1161/CIRCGEN.119.002786},
doi = {10.1161/CIRCGEN.119.002786},
abstract = {Background:
Variants in ion channel genes have classically been studied in low throughput by patch clamping. Deep mutational scanning is a complementary approach that can simultaneously assess function of thousands of variants.
Methods:
We have developed and validated a method to perform a deep mutational scan of variants in
SCN5A
, which encodes the major voltage-gated sodium channel in the heart. We created a library of nearly all possible variants in a 36 base region of
SCN5A
in the S4 voltage sensor of domain IV and stably integrated the library into HEK293T cells.
Results:
In preliminary experiments, challenge with 3 drugs (veratridine, brevetoxin, and ouabain) could discriminate wild-type channels from gain- and loss-of-function pathogenic variants. High-throughput sequencing of the pre- and postdrug challenge pools was used to count the prevalence of each variant and identify variants with abnormal function. The deep mutational scan scores identified 40 putative gain-of-function and 33 putative loss-of-function variants. For 8 of 9 variants, patch clamping data were consistent with the scores.
Conclusions:
These experiments demonstrate the accuracy of a high-throughput in vitro scan of
SCN5A
variant function, which can be used to identify deleterious variants in
SCN5A
and other ion channel genes.},
language = {en},
number = {1},
urldate = {2022-07-11},
journal = {Circulation: Genomic and Precision Medicine},
author = {Glazer, Andrew M. and Kroncke, Brett M. and Matreyek, Kenneth A. and Yang, Tao and Wada, Yuko and Shields, Tiffany and Salem, Joe-Elie and Fowler, Douglas M. and Roden, Dan M.},
month = feb,
year = {2020},
pages = {e002786},
file = {Full Text:/Users/admin/Zotero/storage/AHYB9LRW/Glazer et al. - 2020 - Deep Mutational Scan of an SCN5A Voltage Se.pdf:application/pdf},
}
@article{kelsic_rna_2016,
title = {{RNA} {Structural} {Determinants} of {Optimal} {Codons} {Revealed} by {MAGE}-{Seq}},
volume = {3},
issn = {24054712},
url = {https://linkinghub.elsevier.com/retrieve/pii/S2405471216303684},
doi = {10.1016/j.cels.2016.11.004},
language = {en},
number = {6},
urldate = {2022-07-11},
journal = {Cell Systems},
author = {Kelsic, Eric D. and Chung, Hattie and Cohen, Niv and Park, Jimin and Wang, Harris H. and Kishony, Roy},
month = dec,
year = {2016},
pages = {563--571.e6},
file = {Full Text:/Users/admin/Zotero/storage/IF39P5MD/Kelsic et al. - 2016 - RNA Structural Determinants of Optimal Codons Reve.pdf:application/pdf},
}
@article{rockah-shmuel_systematic_2015,
title = {Systematic {Mapping} of {Protein} {Mutational} {Space} by {Prolonged} {Drift} {Reveals} the {Deleterious} {Effects} of {Seemingly} {Neutral} {Mutations}},
volume = {11},
issn = {1553-7358},
url = {https://dx.plos.org/10.1371/journal.pcbi.1004421},
doi = {10.1371/journal.pcbi.1004421},
language = {en},
number = {8},
urldate = {2022-07-11},
journal = {PLOS Computational Biology},
author = {Rockah-Shmuel, Liat and Tóth-Petróczy, Ágnes and Tawfik, Dan S.},
editor = {Orengo, Christine A.},
month = aug,
year = {2015},
pages = {e1004421},
file = {Full Text:/Users/admin/Zotero/storage/QVEEKDBN/Rockah-Shmuel et al. - 2015 - Systematic Mapping of Protein Mutational Space by .pdf:application/pdf},
}
@article{kitzman_massively_2015,
title = {Massively parallel single-amino-acid mutagenesis},
volume = {12},
issn = {1548-7091, 1548-7105},
url = {http://www.nature.com/articles/nmeth.3223},
doi = {10.1038/nmeth.3223},
language = {en},
number = {3},
urldate = {2022-07-11},
journal = {Nature Methods},
author = {Kitzman, Jacob O and Starita, Lea M and Lo, Russell S and Fields, Stanley and Shendure, Jay},
month = mar,
year = {2015},
pages = {203--206},
file = {Accepted Version:/Users/admin/Zotero/storage/SSUEVZ32/Kitzman et al. - 2015 - Massively parallel single-amino-acid mutagenesis.pdf:application/pdf},
}
@article{aakre_evolving_2015,
title = {Evolving {New} {Protein}-{Protein} {Interaction} {Specificity} through {Promiscuous} {Intermediates}},
volume = {163},
issn = {00928674},
url = {https://linkinghub.elsevier.com/retrieve/pii/S0092867415012726},
doi = {10.1016/j.cell.2015.09.055},
language = {en},
number = {3},
urldate = {2022-07-11},
journal = {Cell},
author = {Aakre, Christopher D. and Herrou, Julien and Phung, Tuyen N. and Perchuk, Barrett S. and Crosson, Sean and Laub, Michael T.},
month = oct,
year = {2015},
pages = {594--606},
file = {Full Text:/Users/admin/Zotero/storage/4IIQTP93/Aakre et al. - 2015 - Evolving New Protein-Protein Interaction Specifici.pdf:application/pdf},
}
@article{bolognesi_mutational_2019,
title = {The mutational landscape of a prion-like domain},
volume = {10},
issn = {2041-1723},
url = {http://www.nature.com/articles/s41467-019-12101-z},
doi = {10.1038/s41467-019-12101-z},
abstract = {Abstract
Insoluble protein aggregates are the hallmarks of many neurodegenerative diseases. For example, aggregates of TDP-43 occur in nearly all cases of amyotrophic lateral sclerosis (ALS). However, whether aggregates cause cellular toxicity is still not clear, even in simpler cellular systems. We reasoned that deep mutagenesis might be a powerful approach to disentangle the relationship between aggregation and toxicity. We generated {\textgreater}50,000 mutations in the prion-like domain (PRD) of TDP-43 and quantified their toxicity in yeast cells. Surprisingly, mutations that increase hydrophobicity and aggregation strongly decrease toxicity. In contrast, toxic variants promote the formation of dynamic liquid-like condensates. Mutations have their strongest effects in a hotspot that genetic interactions reveal to be structured in vivo, illustrating how mutagenesis can probe the in vivo structures of unstructured proteins. Our results show that aggregation of TDP-43 is not harmful but protects cells, most likely by titrating the protein away from a toxic liquid-like phase.},
language = {en},
number = {1},
urldate = {2022-07-11},
journal = {Nature Communications},
author = {Bolognesi, Benedetta and Faure, Andre J. and Seuma, Mireia and Schmiedel, Jörn M. and Tartaglia, Gian Gaetano and Lehner, Ben},
month = dec,
year = {2019},
pages = {4162},
file = {Full Text:/Users/admin/Zotero/storage/3J8H73B4/Bolognesi et al. - 2019 - The mutational landscape of a prion-like domain.pdf:application/pdf},
}
@techreport{flynn_comprehensive_2022,
type = {preprint},
title = {Comprehensive fitness landscape of {SARS}-{CoV}-2 {M} $^{\textrm{pro}}$ reveals insights into viral resistance mechanisms},
url = {http://biorxiv.org/lookup/doi/10.1101/2022.01.26.477860},
abstract = {Abstract
With the continual evolution of new strains of SARS-CoV-2 that are more virulent, transmissible, and able to evade current vaccines, there is an urgent need for effective anti-viral drugs. SARS-CoV-2 main protease (M
pro
) is a leading target for drug design due to its conserved and indispensable role in the viral life cycle. Drugs targeting M
pro
appear promising but will elicit selection pressure for resistance. To understand resistance potential in M
pro
, we performed a comprehensive mutational scan of the protease that analyzed the function of all possible single amino acid changes. We developed three separate high-throughput assays of M
pro
function in yeast, based on either the ability of M
pro
variants to cleave at a defined cut-site or on the toxicity of their expression to yeast. We used deep sequencing to quantify the functional effects of each variant in each screen. The protein fitness landscapes from all three screens were strongly correlated, indicating that they captured the biophysical properties critical to M
pro
function. The fitness landscapes revealed a non-active site location on the surface that is extremely sensitive to mutation making it a favorable location to target with inhibitors. In addition, we found a network of critical amino acids that physically bridge the two active sites of the M
pro
dimer. The clinical variants of M
pro
were predominantly functional in our screens, indicating that M
pro
is under strong selection pressure in the human population. Our results provide predictions of mutations that will be readily accessible to M
pro
evolution and that are likely to contribute to drug resistance. This complete mutational guide of M
pro
can be used in the design of inhibitors with reduced potential of evolving viral resistance.},
language = {en},
urldate = {2022-07-11},
institution = {Molecular Biology},
author = {Flynn, Julia M. and Samant, Neha and Schneider-Nachum, Gily and Barkan, David T. and Yilmaz, Nese Kurt and Schiffer, Celia A. and Moquin, Stephanie A. and Dovala, Dustin and Bolon, Daniel N.A.},
month = jan,
year = {2022},
doi = {10.1101/2022.01.26.477860},
file = {Submitted Version:/Users/admin/Zotero/storage/MR6KIKAK/Flynn et al. - 2022 - Comprehensive fitness landscape of SARS-CoV-2 M s.pdf:application/pdf},
}
@article{haddox_mapping_2018,
title = {Mapping mutational effects along the evolutionary landscape of {HIV} envelope},
volume = {7},
issn = {2050-084X},
url = {https://elifesciences.org/articles/34420},
doi = {10.7554/eLife.34420},
abstract = {The immediate evolutionary space accessible to HIV is largely determined by how single amino acid mutations affect fitness. These mutational effects can shift as the virus evolves. However, the prevalence of such shifts in mutational effects remains unclear. Here, we quantify the effects on viral growth of all amino acid mutations to two HIV envelope (Env) proteins that differ at
{\textgreater}
100 residues. Most mutations similarly affect both Envs, but the amino acid preferences of a minority of sites have clearly shifted. These shifted sites usually prefer a specific amino acid in one Env, but tolerate many amino acids in the other. Surprisingly, shifts are only slightly enriched at sites that have substituted between the Envs—and many occur at residues that do not even contact substitutions. Therefore, long-range epistasis can unpredictably shift Env’s mutational tolerance during HIV evolution, although the amino acid preferences of most sites are conserved between moderately diverged viral strains.
,
The virus that causes AIDS, or HIV, has a protein called Env on its surface, which is essential for the virus to infect cells. Env can also be recognized by the immune system, which then targets the virus for destruction or blocks it from infecting cells. Unfortunately, Env evolves very quickly, which means that HIV can evade our defenses. However, there are limits to how much this protein can change, since it still needs to perform its essential role in helping viruses enter cells.
In the century since HIV first appeared in human populations, the virus has evolved considerably. There are now many HIV strains that infect people, and they bear Env proteins with substantially different sequences. However, it is not clear if these changes in sequence have resulted in Envs from distinct strains being able to tolerate different mutations.
To examine this question, Haddox et al. compared how the Envs from two strains of HIV react to modifications in their sequences. They created all possible individual mutations in the proteins, and the resulting collections of mutated viruses were then tested for their ability to infect cells in the laboratory.
Most mutations had similar effects in both Env proteins. This allowed Haddox et al. to identify portions of the protein that easily accommodate changes, and portions that must remain unchanged for viruses to remain infectious—at least in the laboratory. Some of these mutations are under different types of pressures when the virus faces the immune system, and those were identified using computational approaches.
However, some mutations were tolerated differently by the two Env proteins. Therefore, viral strains differ in how their Env proteins can evolve. The parts of Env that showed differences in mutational tolerance between the strains were not necessarily the parts that differ in sequence. This shows that changes in sequence in one part of the protein can modify how other portions evolve.
It remains to be determined whether changes in tolerance to mutations translate into differences in how the virus can escape immunity. This is an important question given that the rapid evolution of Env is a major obstacle to creating a vaccine for HIV.},
language = {en},
urldate = {2022-07-11},
journal = {eLife},
author = {Haddox, Hugh K and Dingens, Adam S and Hilton, Sarah K and Overbaugh, Julie and Bloom, Jesse D},
month = mar,
year = {2018},
pages = {e34420},
file = {Full Text:/Users/admin/Zotero/storage/2JX4KQTR/Haddox et al. - 2018 - Mapping mutational effects along the evolutionary .pdf:application/pdf},
}
@article{stiffler_evolvability_2015,
title = {Evolvability as a {Function} of {Purifying} {Selection} in {TEM}-1 Beta-{Lactamase}},
volume = {160},
issn = {00928674},
url = {https://linkinghub.elsevier.com/retrieve/pii/S0092867415000781},
doi = {10.1016/j.cell.2015.01.035},
language = {en},
number = {5},
urldate = {2022-07-11},
journal = {Cell},
author = {Stiffler, Michael A. and Hekstra, Doeke R. and Ranganathan, Rama},
month = feb,
year = {2015},
pages = {882--892},
file = {Full Text:/Users/admin/Zotero/storage/D95TSRL8/Stiffler et al. - 2015 - Evolvability as a Function of Purifying Selection .pdf:application/pdf},
}
@article{tripathi_molecular_2016,
title = {Molecular {Determinants} of {Mutant} {Phenotypes}, {Inferred} from {Saturation} {Mutagenesis} {Data}},
volume = {33},
issn = {0737-4038, 1537-1719},
url = {https://academic.oup.com/mbe/article-lookup/doi/10.1093/molbev/msw182},
doi = {10.1093/molbev/msw182},
language = {en},
number = {11},
urldate = {2022-07-11},
journal = {Molecular Biology and Evolution},
author = {Tripathi, Arti and Gupta, Kritika and Khare, Shruti and Jain, Pankaj C. and Patel, Siddharth and Kumar, Prasanth and Pulianmackal, Ajai J. and Aghera, Nilesh and Varadarajan, Raghavan},
month = nov,
year = {2016},
pages = {2960--2975},
file = {Full Text:/Users/admin/Zotero/storage/UFKDKVYH/Tripathi et al. - 2016 - Molecular Determinants of Mutant Phenotypes, Infer.pdf:application/pdf},
}
@article{findlay_accurate_2018,
title = {Accurate classification of {BRCA1} variants with saturation genome editing},
volume = {562},
issn = {0028-0836, 1476-4687},
url = {http://www.nature.com/articles/s41586-018-0461-z},
doi = {10.1038/s41586-018-0461-z},
language = {en},
number = {7726},
urldate = {2022-07-11},
journal = {Nature},
author = {Findlay, Gregory M. and Daza, Riza M. and Martin, Beth and Zhang, Melissa D. and Leith, Anh P. and Gasperini, Molly and Janizek, Joseph D. and Huang, Xingfan and Starita, Lea M. and Shendure, Jay},
month = oct,
year = {2018},
pages = {217--222},
file = {Accepted Version:/Users/admin/Zotero/storage/QFJF3L2Y/Findlay et al. - 2018 - Accurate classification of BRCA1 variants with sat.pdf:application/pdf},
}
@article{lee_deep_2018,
title = {Deep mutational scanning of hemagglutinin helps predict evolutionary fates of human {H3N2} influenza variants},
volume = {115},
issn = {0027-8424, 1091-6490},
url = {https://pnas.org/doi/full/10.1073/pnas.1806133115},
doi = {10.1073/pnas.1806133115},
abstract = {Significance
A key goal in the study of influenza virus evolution is to forecast which viral strains will persist and which ones will die out. Here we experimentally measure the effects of all amino acid mutations to the hemagglutinin protein from a human H3N2 influenza strain on viral growth in cell culture. We show that these measurements have utility for distinguishing among viral strains that do and do not succeed in nature. Overall, our work suggests that new high-throughput experimental approaches may be useful for understanding virus evolution in nature.
,
Human influenza virus rapidly accumulates mutations in its major surface protein hemagglutinin (HA). The evolutionary success of influenza virus lineages depends on how these mutations affect HA’s functionality and antigenicity. Here we experimentally measure the effects on viral growth in cell culture of all single amino acid mutations to the HA from a recent human H3N2 influenza virus strain. We show that mutations that are measured to be more favorable for viral growth are enriched in evolutionarily successful H3N2 viral lineages relative to mutations that are measured to be less favorable for viral growth. Therefore, despite the well-known caveats about cell-culture measurements of viral fitness, such measurements can still be informative for understanding evolution in nature. We also compare our measurements for H3 HA to similar data previously generated for a distantly related H1 HA and find substantial differences in which amino acids are preferred at many sites. For instance, the H3 HA has less disparity in mutational tolerance between the head and stalk domains than the H1 HA. Overall, our work suggests that experimental measurements of mutational effects can be leveraged to help understand the evolutionary fates of viral lineages in nature—but only when the measurements are made on a viral strain similar to the ones being studied in nature.},
language = {en},
number = {35},
urldate = {2022-07-11},
journal = {Proceedings of the National Academy of Sciences},
author = {Lee, Juhye M. and Huddleston, John and Doud, Michael B. and Hooper, Kathryn A. and Wu, Nicholas C. and Bedford, Trevor and Bloom, Jesse D.},
month = aug,
year = {2018},
file = {Full Text:/Users/admin/Zotero/storage/HG3QHAPN/Lee et al. - 2018 - Deep mutational scanning of hemagglutinin helps pr.pdf:application/pdf},
}
@article{weile_framework_2017,
title = {A framework for exhaustively mapping functional missense variants},
volume = {13},
issn = {1744-4292, 1744-4292},
url = {https://onlinelibrary.wiley.com/doi/10.15252/msb.20177908},
doi = {10.15252/msb.20177908},
language = {en},
number = {12},
urldate = {2022-07-11},
journal = {Molecular Systems Biology},
author = {Weile, Jochen and Sun, Song and Cote, Atina G and Knapp, Jennifer and Verby, Marta and Mellor, Joseph C and Wu, Yingzhou and Pons, Carles and Wong, Cassandra and Lieshout, Natascha and Yang, Fan and Tasan, Murat and Tan, Guihong and Yang, Shan and Fowler, Douglas M and Nussbaum, Robert and Bloom, Jesse D and Vidal, Marc and Hill, David E and Aloy, Patrick and Roth, Frederick P},
month = dec,
year = {2017},
pages = {957},
file = {Full Text:/Users/admin/Zotero/storage/YYUXTHFT/Weile et al. - 2017 - A framework for exhaustively mapping functional mi.pdf:application/pdf;Full Text:/Users/admin/Zotero/storage/HCRUH6RQ/Weile et al. - 2017 - A framework for exhaustively mapping functional mi.pdf:application/pdf},
}
@article{qi_quantitative_2014,
title = {A {Quantitative} {High}-{Resolution} {Genetic} {Profile} {Rapidly} {Identifies} {Sequence} {Determinants} of {Hepatitis} {C} {Viral} {Fitness} and {Drug} {Sensitivity}},
volume = {10},
issn = {1553-7374},
url = {https://dx.plos.org/10.1371/journal.ppat.1004064},
doi = {10.1371/journal.ppat.1004064},
language = {en},
number = {4},
urldate = {2022-07-11},
journal = {PLoS Pathogens},
author = {Qi, Hangfei and Olson, C. Anders and Wu, Nicholas C. and Ke, Ruian and Loverdo, Claude and Chu, Virginia and Truong, Shawna and Remenyi, Roland and Chen, Zugen and Du, Yushen and Su, Sheng-Yao and Al-Mawsawi, Laith Q. and Wu, Ting-Ting and Chen, Shu-Hua and Lin, Chung-Yen and Zhong, Weidong and Lloyd-Smith, James O. and Sun, Ren},
editor = {Wilke, Claus O.},
month = apr,
year = {2014},
pages = {e1004064},
file = {Full Text:/Users/admin/Zotero/storage/ZXNH72PK/Qi et al. - 2014 - A Quantitative High-Resolution Genetic Profile Rap.pdf:application/pdf},
}
@article{chan_correlation_2017,
title = {Correlation of fitness landscapes from three orthologous {TIM} barrels originates from sequence and structure constraints},
volume = {8},
issn = {2041-1723},
url = {http://www.nature.com/articles/ncomms14614},
doi = {10.1038/ncomms14614},
language = {en},
number = {1},
urldate = {2022-07-11},
journal = {Nature Communications},
author = {Chan, Yvonne H. and Venev, Sergey V. and Zeldovich, Konstantin B. and Matthews, C. Robert},
month = apr,
year = {2017},
pages = {14614},
file = {Full Text:/Users/admin/Zotero/storage/A6E42JVX/Chan et al. - 2017 - Correlation of fitness landscapes from three ortho.pdf:application/pdf},
}
@article{melnikov_comprehensive_2014,
title = {Comprehensive mutational scanning of a kinase \textit{in vivo} reveals substrate-dependent fitness landscapes},
volume = {42},
issn = {0305-1048, 1362-4962},
url = {https://academic.oup.com/nar/article-lookup/doi/10.1093/nar/gku511},
doi = {10.1093/nar/gku511},
language = {en},
number = {14},
urldate = {2022-07-11},
journal = {Nucleic Acids Research},
author = {Melnikov, Alexandre and Rogov, Peter and Wang, Li and Gnirke, Andreas and Mikkelsen, Tarjei S.},
month = aug,
year = {2014},
pages = {e112--e112},
file = {Full Text:/Users/admin/Zotero/storage/U454BG33/Melnikov et al. - 2014 - Comprehensive mutational scanning of a kinase i.pdf:application/pdf},
}
@article{nutschel_systematically_2020,
title = {Systematically {Scrutinizing} the {Impact} of {Substitution} {Sites} on {Thermostability} and {Detergent} {Tolerance} for \textit{{Bacillus} subtilis} {Lipase} {A}},
volume = {60},
issn = {1549-9596, 1549-960X},
url = {https://pubs.acs.org/doi/10.1021/acs.jcim.9b00954},
doi = {10.1021/acs.jcim.9b00954},
language = {en},
number = {3},
urldate = {2022-07-11},
journal = {Journal of Chemical Information and Modeling},
author = {Nutschel, Christina and Fulton, Alexander and Zimmermann, Olav and Schwaneberg, Ulrich and Jaeger, Karl-Erich and Gohlke, Holger},
month = mar,
year = {2020},
pages = {1568--1584},
}
@article{jacquier_capturing_2013,
title = {Capturing the mutational landscape of the beta-lactamase {TEM}-1},
volume = {110},
issn = {0027-8424, 1091-6490},
url = {https://pnas.org/doi/full/10.1073/pnas.1215206110},
doi = {10.1073/pnas.1215206110},
abstract = {Adaptation proceeds through the selection of mutations. The distribution of mutant fitness effect and the forces shaping this distribution are therefore keys to predict the evolutionary fate of organisms and their constituents such as enzymes. Here, by producing and sequencing a comprehensive collection of 10,000 mutants, we explore the mutational landscape of one enzyme involved in the spread of antibiotic resistance, the beta-lactamase TEM-1. We measured mutation impact on the enzyme activity through the estimation of amoxicillin minimum inhibitory concentration on a subset of 990 mutants carrying a unique missense mutation, representing 64\% of possible amino acid changes in that protein reachable by point mutation. We established that mutation type, solvent accessibility of residues, and the predicted effect of mutations on protein stability primarily determined alone or in combination changes in minimum inhibitory concentration of mutants. Moreover, we were able to capture the drastic modification of the mutational landscape induced by a single stabilizing point mutation (M182T) by a simple model of protein stability. This work thereby provides an integrated framework to study mutation effects and a tool to understand/define better the epistatic interactions.},
language = {en},
number = {32},
urldate = {2022-07-11},
journal = {Proceedings of the National Academy of Sciences},
author = {Jacquier, Hervé and Birgy, André and Le Nagard, Hervé and Mechulam, Yves and Schmitt, Emmanuelle and Glodt, Jérémy and Bercot, Beatrice and Petit, Emmanuelle and Poulain, Julie and Barnaud, Guilène and Gros, Pierre-Alexis and Tenaillon, Olivier},
month = aug,
year = {2013},
pages = {13067--13072},
file = {Full Text:/Users/admin/Zotero/storage/KCXP8PUL/Jacquier et al. - 2013 - Capturing the mutational landscape of the beta-lac.pdf:application/pdf},
}
@article{mishra_systematic_2016,
title = {Systematic {Mutant} {Analyses} {Elucidate} {General} and {Client}-{Specific} {Aspects} of {Hsp90} {Function}},
volume = {15},
issn = {22111247},
url = {https://linkinghub.elsevier.com/retrieve/pii/S2211124716303175},
doi = {10.1016/j.celrep.2016.03.046},
language = {en},
number = {3},
urldate = {2022-07-11},
journal = {Cell Reports},
author = {Mishra, Parul and Flynn, Julia M. and Starr, Tyler N. and Bolon, Daniel N.A.},
month = apr,
year = {2016},
pages = {588--598},
file = {Full Text:/Users/admin/Zotero/storage/N3VLLXTY/Mishra et al. - 2016 - Systematic Mutant Analyses Elucidate General and C.pdf:application/pdf},
}
@article{pokusaeva_experimental_2019,
title = {An experimental assay of the interactions of amino acids from orthologous sequences shaping a complex fitness landscape},
volume = {15},
issn = {1553-7404},
url = {https://dx.plos.org/10.1371/journal.pgen.1008079},
doi = {10.1371/journal.pgen.1008079},
language = {en},
number = {4},
urldate = {2022-07-11},
journal = {PLOS Genetics},
author = {Pokusaeva, Victoria O. and Usmanova, Dinara R. and Putintseva, Ekaterina V. and Espinar, Lorena and Sarkisyan, Karen S. and Mishin, Alexander S. and Bogatyreva, Natalya S. and Ivankov, Dmitry N. and Akopyan, Arseniy V. and Avvakumov, Sergey Ya. and Povolotskaya, Inna S. and Filion, Guillaume J. and Carey, Lucas B. and Kondrashov, Fyodor A.},
editor = {Petrov, Dmitri A.},
month = apr,
year = {2019},
pages = {e1008079},
file = {Full Text:/Users/admin/Zotero/storage/39VZ7AQN/Pokusaeva et al. - 2019 - An experimental assay of the interactions of amino.pdf:application/pdf},
}
@article{fernandes_functional_2016,
title = {Functional {Segregation} of {Overlapping} {Genes} in {HIV}},
volume = {167},
issn = {00928674},
url = {https://linkinghub.elsevier.com/retrieve/pii/S0092867416316038},
doi = {10.1016/j.cell.2016.11.031},
language = {en},
number = {7},
urldate = {2022-07-11},
journal = {Cell},
author = {Fernandes, Jason D. and Faust, Tyler B. and Strauli, Nicolas B. and Smith, Cynthia and Crosby, David C. and Nakamura, Robert L. and Hernandez, Ryan D. and Frankel, Alan D.},
month = dec,
year = {2016},
pages = {1762--1773.e12},
file = {Full Text:/Users/admin/Zotero/storage/M2JCUQLM/Fernandes et al. - 2016 - Functional Segregation of Overlapping Genes in HIV.pdf:application/pdf},
}
@techreport{sinai_generative_2021,
type = {preprint},
title = {Generative {AAV} capsid diversification by latent interpolation},
url = {http://biorxiv.org/lookup/doi/10.1101/2021.04.16.440236},
abstract = {Summary
Adeno-associated virus (AAV) capsids have shown clinical promise as delivery vectors for gene therapy. However, the high prevalence of pre-existing immunity against natural capsids poses a challenge for widespread treatment. The generation of diverse capsids that are potentially more capable of immune evasion is challenging because introducing multiple mutations often breaks capsid assembly. Here we target a representative, immunologically relevant 28-amino-acid segment of the AAV2 capsid and show that a low-complexity Variational Auto-encoder (VAE) can interpolate in sequence space to produce diverse and novel capsids capable of packaging their own genomes. We first train the VAE on a 564-sample Multiple-Sequence Alignment (MSA) of dependo-parvoviruses, and then further augment this dataset by adding 22,704 samples from a deep mutational exploration (DME) on the target region. In both cases the VAE generated viable variants with many mutations, which we validated experimentally. We propose that this simple approach can be used to optimize and diversify other proteins, as well as other capsid traits of interest for gene delivery.},
language = {en},
urldate = {2022-07-11},
institution = {Synthetic Biology},
author = {Sinai, Sam and Jain, Nina and Church, George M and Kelsic, Eric D},
month = apr,
year = {2021},
doi = {10.1101/2021.04.16.440236},
file = {Submitted Version:/Users/admin/Zotero/storage/TEMTVFT7/Sinai et al. - 2021 - Generative AAV capsid diversification by latent in.pdf:application/pdf},
}
@article{wu_functional_2015,
title = {Functional {Constraint} {Profiling} of a {Viral} {Protein} {Reveals} {Discordance} of {Evolutionary} {Conservation} and {Functionality}},
volume = {11},
issn = {1553-7404},
url = {https://dx.plos.org/10.1371/journal.pgen.1005310},
doi = {10.1371/journal.pgen.1005310},
language = {en},
number = {7},
urldate = {2022-07-11},
journal = {PLOS Genetics},
author = {Wu, Nicholas C. and Olson, C. Anders and Du, Yushen and Le, Shuai and Tran, Kevin and Remenyi, Roland and Gong, Danyang and Al-Mawsawi, Laith Q. and Qi, Hangfei and Wu, Ting-Ting and Sun, Ren},
editor = {Worobey, Michael},
month = jul,
year = {2015},
pages = {e1005310},
file = {Full Text:/Users/admin/Zotero/storage/NNQX65KR/Wu et al. - 2015 - Functional Constraint Profiling of a Viral Protein.pdf:application/pdf},
}
@article{matreyek_integrating_2021,
title = {Integrating thousands of {PTEN} variant activity and abundance measurements reveals variant subgroups and new dominant negatives in cancers},
volume = {13},
issn = {1756-994X},
url = {https://genomemedicine.biomedcentral.com/articles/10.1186/s13073-021-00984-x},
doi = {10.1186/s13073-021-00984-x},
abstract = {Abstract
Background
PTEN is a multi-functional tumor suppressor protein regulating cell growth, immune signaling, neuronal function, and genome stability. Experimental characterization can help guide the clinical interpretation of the thousands of germline or somatic PTEN variants observed in patients. Two large-scale mutational datasets, one for PTEN variant intracellular abundance encompassing 4112 missense variants and one for lipid phosphatase activity encompassing 7244 variants, were recently published. The combined information from these datasets can reveal variant-specific phenotypes that may underlie various clinical presentations, but this has not been comprehensively examined, particularly for somatic PTEN variants observed in cancers.
Methods
Here, we add to these efforts by measuring the intracellular abundance of 764 new PTEN variants and refining abundance measurements for 3351 previously studied variants. We use this expanded and refined PTEN abundance dataset to explore the mutational patterns governing PTEN intracellular abundance, and then incorporate the phosphatase activity data to subdivide PTEN variants into four functionally distinct groups.
Results
This analysis revealed a set of highly abundant but lipid phosphatase defective variants that could act in a dominant-negative fashion to suppress PTEN activity. Two of these variants were, indeed, capable of dysregulating Akt signaling in cells harboring a WT PTEN allele. Both variants were observed in multiple breast or uterine tumors, demonstrating the disease relevance of these high abundance, inactive variants.
Conclusions
We show that multidimensional, large-scale variant functional data, when paired with public cancer genomics datasets and follow-up assays, can improve understanding of uncharacterized cancer-associated variants, and provide better insights into how they contribute to oncogenesis.},
language = {en},
number = {1},
urldate = {2022-07-11},
journal = {Genome Medicine},
author = {Matreyek, Kenneth A. and Stephany, Jason J. and Ahler, Ethan and Fowler, Douglas M.},
month = dec,
year = {2021},
pages = {165},
file = {Full Text:/Users/admin/Zotero/storage/8VXIZT8Q/Matreyek et al. - 2021 - Integrating thousands of PTEN variant activity and.pdf:application/pdf},
}
@article{mighell_saturation_2018,
title = {A {Saturation} {Mutagenesis} {Approach} to {Understanding} {PTEN} {Lipid} {Phosphatase} {Activity} and {Genotype}-{Phenotype} {Relationships}},
volume = {102},
issn = {00029297},
url = {https://linkinghub.elsevier.com/retrieve/pii/S0002929718301071},
doi = {10.1016/j.ajhg.2018.03.018},
language = {en},
number = {5},
urldate = {2023-06-14},
journal = {The American Journal of Human Genetics},
author = {Mighell, Taylor L. and Evans-Dutson, Sara and O’Roak, Brian J.},
month = may,
year = {2018},
pages = {943--955},
file = {Full Text:/Users/admin/Zotero/storage/9UEQ49C4/Mighell et al. - 2018 - A Saturation Mutagenesis Approach to Understanding.pdf:application/pdf},
}
@article{russ_evolution-based_2020,
title = {An evolution-based model for designing chorismate mutase enzymes},
volume = {369},
issn = {0036-8075, 1095-9203},
url = {https://www.sciencemag.org/lookup/doi/10.1126/science.aba3304},
doi = {10.1126/science.aba3304},
abstract = {The rational design of enzymes is an important goal for both fundamental and practical reasons. Here, we describe a process to learn the constraints for specifying proteins purely from evolutionary sequence data, design and build libraries of synthetic genes, and test them for activity in vivo using a quantitative complementation assay. For chorismate mutase, a key enzyme in the biosynthesis of aromatic amino acids, we demonstrate the design of natural-like catalytic function with substantial sequence diversity. Further optimization focuses the generative model toward function in a specific genomic context. The data show that sequence-based statistical models suffice to specify proteins and provide access to an enormous space of functional sequences. This result provides a foundation for a general process for evolution-based design of artificial proteins.},
language = {en},
number = {6502},
urldate = {2023-06-14},
journal = {Science},
author = {Russ, William P. and Figliuzzi, Matteo and Stocker, Christian and Barrat-Charlaix, Pierre and Socolich, Michael and Kast, Peter and Hilvert, Donald and Monasson, Remi and Cocco, Simona and Weigt, Martin and Ranganathan, Rama},
month = jul,
year = {2020},
pages = {440--445},
}
@article{gonzalez_fitness_2019,
title = {Fitness {Effects} of {Single} {Amino} {Acid} {Insertions} and {Deletions} in {TEM}-1 Beta-{Lactamase}},
volume = {431},
issn = {00222836},
url = {https://linkinghub.elsevier.com/retrieve/pii/S0022283619302372},
doi = {10.1016/j.jmb.2019.04.030},
language = {en},
number = {12},
urldate = {2023-06-14},
journal = {Journal of Molecular Biology},
author = {Gonzalez, Courtney E. and Roberts, Paul and Ostermeier, Marc},
month = may,
year = {2019},
pages = {2320--2330},
file = {Accepted Version:/Users/admin/Zotero/storage/ZWGQXGBD/Gonzalez et al. - 2019 - Fitness Effects of Single Amino Acid Insertions an.pdf:application/pdf},
}
@article{macdonald_dimple_2023,
title = {{DIMPLE}: deep insertion, deletion, and missense mutation libraries for exploring protein variation in evolution, disease, and biology},
volume = {24},
issn = {1474-760X},
shorttitle = {{DIMPLE}},
url = {https://genomebiology.biomedcentral.com/articles/10.1186/s13059-023-02880-6},
doi = {10.1186/s13059-023-02880-6},
abstract = {Abstract
Insertions and deletions (indels) enable evolution and cause disease. Due to technical challenges, indels are left out of most mutational scans, limiting our understanding of them in disease, biology, and evolution. We develop a low cost and bias method, DIMPLE, for systematically generating deletions, insertions, and missense mutations in genes, which we test on a range of targets, including Kir2.1. We use DIMPLE to study how indels impact potassium channel structure, disease, and evolution. We find deletions are most disruptive overall, beta sheets are most sensitive to indels, and flexible loops are sensitive to deletions yet tolerate insertions.},
language = {en},
number = {1},
urldate = {2023-06-14},
journal = {Genome Biology},
author = {Macdonald, Christian B. and Nedrud, David and Grimes, Patrick Rockefeller and Trinidad, Donovan and Fraser, James S. and Coyote-Maestas, Willow},
month = feb,
year = {2023},
pages = {36},
file = {Full Text:/Users/admin/Zotero/storage/3MMXFUJ4/Macdonald et al. - 2023 - DIMPLE deep insertion, deletion, and missense mut.pdf:application/pdf},
}
@article{jones_structural_2020,
title = {Structural and functional characterization of {G} protein–coupled receptors with deep mutational scanning},
volume = {9},
issn = {2050-084X},
url = {https://elifesciences.org/articles/54895},
doi = {10.7554/eLife.54895},
abstract = {The {\textgreater}800 human G protein–coupled receptors (GPCRs) are responsible for transducing diverse chemical stimuli to alter cell state- and are the largest class of drug targets. Their myriad structural conformations and various modes of signaling make it challenging to understand their structure and function. Here, we developed a platform to characterize large libraries of GPCR variants in human cell lines with a barcoded transcriptional reporter of G protein signal transduction. We tested 7800 of 7828 possible single amino acid substitutions to the beta-2 adrenergic receptor (Beta
2
AR) at four concentrations of the agonist isoproterenol. We identified residues specifically important for Beta
2
AR signaling, mutations in the human population that are potentially loss of function, and residues that modulate basal activity. Using unsupervised learning, we identify residues critical for signaling, including all major structural motifs and molecular interfaces. We also find a previously uncharacterized structural latch spanning the first two extracellular loops that is highly conserved across Class A GPCRs and is conformationally rigid in both the inactive and active states of the receptor. More broadly, by linking deep mutational scanning with engineered transcriptional reporters, we establish a generalizable method for exploring pharmacogenomics, structure and function across broad classes of drug receptors.},
language = {en},
urldate = {2023-06-14},
journal = {eLife},
author = {Jones, Eric M and Lubock, Nathan B and Venkatakrishnan, Aj and Wang, Jeffrey and Tseng, Alex M and Paggi, Joseph M and Latorraca, Naomi R and Cancilla, Daniel and Satyadi, Megan and Davis, Jessica E and Babu, M Madan and Dror, Ron O and Kosuri, Sriram},
month = oct,
year = {2020},
pages = {e54895},
file = {Full Text:/Users/admin/Zotero/storage/T2SD9K3D/Jones et al. - 2020 - Structural and functional characterization of G pr.pdf:application/pdf;Submitted Version:/Users/admin/Zotero/storage/WYWCXHH9/Jones et al. - 2020 - Structural and functional characterization of G pr.pdf:application/pdf},
}
@article{chen_comprehensive_2020,
title = {Comprehensive exploration of the translocation, stability and substrate recognition requirements in {VIM}-2 lactamase},
volume = {9},
issn = {2050-084X},
url = {https://elifesciences.org/articles/56707},
doi = {10.7554/eLife.56707},
abstract = {Metallo-Beta-lactamases (MBLs) degrade a broad spectrum of Beta-lactam antibiotics, and are a major disseminating source for multidrug resistant bacteria. Despite many biochemical studies in diverse MBLs, molecular understanding of the roles of residues in the enzyme’s stability and function, and especially substrate specificity, is lacking. Here, we employ deep mutational scanning (DMS) to generate comprehensive single amino acid variant data on a major clinical MBL, VIM-2, by measuring the effect of thousands of VIM-2 mutants on the degradation of three representative classes of Beta-lactams (ampicillin, cefotaxime, and meropenem) and at two different temperatures (25°C and 37°C). We revealed residues responsible for expression and translocation, and mutations that increase resistance and/or alter substrate specificity. The distribution of specificity-altering mutations unveiled distinct molecular recognition of the three substrates. Moreover, these function-altering mutations are frequently observed among naturally occurring variants, suggesting that the enzymes have continuously evolved to become more potent resistance genes.},
language = {en},
urldate = {2023-06-14},
journal = {eLife},
author = {Chen, John Z and Fowler, Douglas M and Tokuriki, Nobuhiko},
month = jun,
year = {2020},
pages = {e56707},
file = {Full Text:/Users/admin/Zotero/storage/LBP8NZRP/Chen et al. - 2020 - Comprehensive exploration of the translocation, st.pdf:application/pdf},
}
@article{kennouche_deep_2019,
title = {Deep mutational scanning of the \textit{{Neisseria} meningitidis} major pilin reveals the importance of pilus tip‐mediated adhesion},
volume = {38},
issn = {0261-4189, 1460-2075},
url = {https://www.embopress.org/doi/10.15252/embj.2019102145},
doi = {10.15252/embj.2019102145},
language = {en},
number = {22},
urldate = {2023-06-14},
journal = {The EMBO Journal},
author = {Kennouche, Paul and Charles‐Orszag, Arthur and Nishiguchi, Daiki and Goussard, Sylvie and Imhaus, Anne‐Flore and Dupré, Mathieu and Chamot‐Rooke, Julia and Duménil, Guillaume},
month = nov,
year = {2019},
pages = {e102145},
file = {Full Text:/Users/admin/Zotero/storage/K9LIAMBH/Kennouche et al. - 2019 - Deep mutational scanning of the Neisseria menin.pdf:application/pdf;Submitted Version:/Users/admin/Zotero/storage/9587BW8A/Kennouche et al. - 2019 - Deep mutational scanning of the Neisseria menin.pdf:application/pdf},
}
@article{haddox_experimental_2016,
title = {Experimental {Estimation} of the {Effects} of {All} {Amino}-{Acid} {Mutations} to {HIV}’s {Envelope} {Protein} on {Viral} {Replication} in {Cell} {Culture}},
volume = {12},
issn = {1553-7374},
url = {https://dx.plos.org/10.1371/journal.ppat.1006114},
doi = {10.1371/journal.ppat.1006114},
language = {en},
number = {12},
urldate = {2023-06-14},
journal = {PLOS Pathogens},
author = {Haddox, Hugh K. and Dingens, Adam S. and Bloom, Jesse D.},
editor = {Swanstrom, Ronald},
month = dec,
year = {2016},
pages = {e1006114},