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Steven P. Gygi

Bio: Steven P. Gygi is an academic researcher from Harvard University. The author has contributed to research in topics: Phosphorylation & Proteome. The author has an hindex of 172, co-authored 704 publications receiving 129173 citations. Previous affiliations of Steven P. Gygi include University of Rochester Medical Center & Cell Signaling Technology.


Papers
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Journal ArticleDOI
TL;DR: It is demonstrated that the selection for specific KRAS mutants in human cancers from different tissues is due to their distinct signaling properties, demonstrating that context-dependent variations in signaling downstream of differentKRAS mutants drive the KRAS mutational pattern seen in cancer.
Abstract: KRAS is the most frequently mutated oncogene. The incidence of specific KRAS alleles varies between cancers from different sites, but it is unclear whether allelic selection results from biological selection for specific mutant KRAS proteins. We used a cross-disciplinary approach to compare KRASG12D, a common mutant form, and KRASA146T, a mutant that occurs only in selected cancers. Biochemical and structural studies demonstrated that KRASA146T exhibits a marked extension of switch 1 away from the protein body and nucleotide binding site, which activates KRAS by promoting a high rate of intrinsic and guanine nucleotide exchange factor-induced nucleotide exchange. Using mice genetically engineered to express either allele, we found that KRASG12D and KRASA146T exhibit distinct tissue-specific effects on homeostasis that mirror mutational frequencies in human cancers. These tissue-specific phenotypes result from allele-specific signaling properties, demonstrating that context-dependent variations in signaling downstream of different KRAS mutants drive the KRAS mutational pattern seen in cancer. SIGNIFICANCE: Although epidemiologic and clinical studies have suggested allele-specific behaviors for KRAS, experimental evidence for allele-specific biological properties is limited. We combined structural biology, mass spectrometry, and mouse modeling to demonstrate that the selection for specific KRAS mutants in human cancers from different tissues is due to their distinct signaling properties.See related commentary by Hobbs and Der, p. 696.This article is highlighted in the In This Issue feature, p. 681.

120 citations

Journal ArticleDOI
TL;DR: The results suggest that Tb MP52 and TbMP48 are components of the RNA editing complex.
Abstract: RNA editing in kinetoplastid mitochondria inserts and deletes uridylates at multiple sites in pre-mRNAs as directed by guide RNAs. This occurs by a series of steps that are catalyzed by endoribonuclease, 3*-terminal uridylyl transferase, 3*-exouridylylase, and RNA ligase activities. A multiprotein complex that contains these activities and catalyzes deletion editing in vitro was enriched from Trypanosoma brucei mitochondria by sequential ion-exchange and gel filtration chromatography, followed by glycerol gradient sedimentation. The complex size is approximately 1,600 kDa, and the purified fraction contains 20 major polypeptides. A monoclonal antibody that was generated against the enriched complex reacts with an ;49-kDa protein and specifically immunoprecipitates in vitro deletion RNA editing activity. The protein recognized by the antibody was identified by mass spectrometry, and the corresponding gene, designated TbMP52, was cloned. Recombinant TbMP52 reacts with the monoclonal antibody. Another novel protein, TbMP48, which is similar to TbMP52, and its gene were also identified in the enriched complex. These results suggest that TbMP52 and TbMP48 are components of the RNA editing complex.

120 citations

Journal ArticleDOI
TL;DR: The results suggest that PGC-1β acetylation by GCN5 might be an important step in the control of glucose and lipid pathways and its dysregulation could contribute to metabolic diseases.

118 citations

Journal ArticleDOI
TL;DR: This work demonstrates that Mek1 is activated in response to DSBs by autophosphorylation of two conserved threonines, T327 and T331, in the Mek1 activation loop.
Abstract: Mek1 is a meiosis-specific kinase in budding yeast which promotes recombination between homologous chromosomes by suppressing double-strand break (DSB) repair between sister chromatids. Previous work has shown that in the absence of the meiosis-specific recombinase gene, DMC1, cells arrest in prophase due to unrepaired DSBs and that Mek1 kinase activity is required in this situation to prevent repair of the breaks using sister chromatids. This work demonstrates that Mek1 is activated in response to DSBs by autophosphorylation of two conserved threonines, T327 and T331, in the Mek1 activation loop. Using a version of Mek1 that can be conditionally dimerized during meiosis, Mek1 function was shown to be promoted by dimerization, perhaps as a way of enabling autophosphorylation of the activation loop in trans. A putative HOP1-dependent dimerization domain within the C terminus of Mek1 has been identified. Dimerization alone, however, is insufficient for activation, as DSBs and Mek1 recruitment to the meiosis-specific chromosomal core protein Red1 are also necessary. Phosphorylation of S320 in the activation loop inhibits sister chromatid repair specifically in dmc1-arrested cells. Ectopic dimerization of Mek1 bypasses the requirement for S320 phosphorylation, suggesting this phosphorylation is necessary for maintenance of Mek1 dimers during checkpointinduced arrest.

118 citations

Journal ArticleDOI
04 Aug 2017-Science
TL;DR: Results indicate that UBE2O may function as a hybrid enzyme with both E2 and E3 (ubiquitin-ligating) activities in the reticulocyte–red blood cell transition, and that a major component of the specificity underlying differentiation-linked proteome remodeling appears to be carried by UBE1O itself.
Abstract: INTRODUCTION The reticulocyte–red blood cell transition is a canonical example of terminal differentiation. The mature red blood cell has one of the simplest cellular proteomes known, with hemoglobin remarkably concentrated to ~98% of soluble protein. During reticulocyte maturation, the proteome is remodeled through the programmed elimination of most generic constituents of the cell, in parallel with abundant synthesis of cell type–specific proteins such as hemoglobin. The mechanisms that drive rapid turnover of soluble and normally stable proteins in terminally differentiating cells remain largely unclear. RATIONALE The ubiquitin-proteasome system (UPS) was discovered in reticulocytes, where it is highly active. However, its function in this developmental context has not been established. UBE2O is an E2 (ubiquitin-conjugating) enzyme that is co-induced with globin and expressed at elevated levels late in the erythroid lineage. We identified an anemic mouse line with a null mutation in Ube2o , and used multiplexed quantitative proteomics to identify candidate substrates of UBE2O in an unbiased and global manner. We found that the protein compositions of mutant and wild-type reticulocytes differed markedly, suggesting that UBE2O-dependent ubiquitination might target its substrates for degradation to effect remodeling of the proteome. To test whether UBE2O was sufficient for proteome remodeling, we engineered a non-erythroid cell line to inducibly express UBE2O above its basal level. Upon induction, we observed the decline of hundreds of proteins from these cells, in many cases the same proteins as those eliminated from reticulocytes. Overexpression of an active-site mutant of UBE2O did not show these effects. Therefore, a major component of the specificity underlying differentiation-linked proteome remodeling appears to be carried by UBE2O itself. These results also indicate that UBE2O may function as a hybrid enzyme with both E2 and E3 (ubiquitin-ligating) activities. In support of this model, candidate substrates identified by proteomics were ubiquitinated by purified UBE2O without the assistance of additional specificity factors. RESULTS The most prominent phenotypes of the Ube2o mutant are an anemia characterized by small cells with low hemoglobin content (microcytic hypochromic anemia), and a defect in the elimination of ribosomes, the latter being a key aspect of reticulocyte maturation. When we added recombinant UBE2O protein to reticulocyte lysates from the null mutant, ubiquitin was conjugated primarily to ribosomal proteins. Moreover, immunoblot analysis and quantitative proteomics revealed elevated levels of multiple ribosomal proteins in mutant reticulocytes. Sucrose gradient analysis indicated the persistence not only of ribosomal proteins but of ribosomes themselves during ex vivo differentiation of mutant reticulocytes. Accordingly, ribosomes were eliminated upon induction of UBE2O in non-erythroid cells. The elimination of organelles from reticulocytes, as exemplified by that of mitochondria, was not affected in the Ube2o mutant, indicating the specificity of its effects on programmed protein turnover. Free ribosomal proteins were ubiquitinated by purified UBE2O, which suggests that these proteins are true substrates of the enzyme. However, UBE2O substrates are diverse in nature and not limited to ribosomal proteins. Individual domains of UBE2O bound substrates with distinct specificities. Thus, the broad specificity of UBE2O reflects the presence of multiple substrate recognition domains within the enzyme. Proteasome inhibitors blocked the degradation of UBE2O-dependent substrates in reticulocytes, although UBE2O does not form polyubiquitin chains. Rather, UBE2O adds single ubiquitin groups to substrates at multiple sites. Proteasome inhibitor treatment ex vivo led to depletion of the pools of many amino acids; this result implies that the flux of ubiquitinated substrates through the reticulocyte proteasome is sufficient to supply amino acids needed for late-stage translation of mRNA. In late erythropoiesis, several ubiquitin-conjugating enzymes and ligases are induced together with Ube2o while most components of the UPS disappear. We propose that the UPS is not simply amplified during erythroid maturation, but is instead broadly reconfigured to promote remodeling of the proteome. CONCLUSION A highly specialized UPS is expressed in the reticulocyte and is used to remodel the proteome of these cells on a global scale. UBE2O, a hybrid E2-E3 enzyme, functions as a major specificity factor in this process. In reticulocytes, and perhaps in other differentiated cells such as in the lens, the induction of ubiquitinating factors may drive the transition from a complex to a simple proteome.

117 citations


Cited by
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Journal ArticleDOI
TL;DR: Several case studies of Cytoscape plug-ins are surveyed, including a search for interaction pathways correlating with changes in gene expression, a study of protein complexes involved in cellular recovery to DNA damage, inference of a combined physical/functional interaction network for Halobacterium, and an interface to detailed stochastic/kinetic gene regulatory models.
Abstract: Cytoscape is an open source software project for integrating biomolecular interaction networks with high-throughput expression data and other molecular states into a unified conceptual framework. Although applicable to any system of molecular components and interactions, Cytoscape is most powerful when used in conjunction with large databases of protein-protein, protein-DNA, and genetic interactions that are increasingly available for humans and model organisms. Cytoscape's software Core provides basic functionality to layout and query the network; to visually integrate the network with expression profiles, phenotypes, and other molecular states; and to link the network to databases of functional annotations. The Core is extensible through a straightforward plug-in architecture, allowing rapid development of additional computational analyses and features. Several case studies of Cytoscape plug-ins are surveyed, including a search for interaction pathways correlating with changes in gene expression, a study of protein complexes involved in cellular recovery to DNA damage, inference of a combined physical/functional interaction network for Halobacterium, and an interface to detailed stochastic/kinetic gene regulatory models.

32,980 citations

28 Jul 2005
TL;DR: PfPMP1)与感染红细胞、树突状组胞以及胎盘的单个或多个受体作用,在黏附及免疫逃避中起关键的作�ly.
Abstract: 抗原变异可使得多种致病微生物易于逃避宿主免疫应答。表达在感染红细胞表面的恶性疟原虫红细胞表面蛋白1(PfPMP1)与感染红细胞、内皮细胞、树突状细胞以及胎盘的单个或多个受体作用,在黏附及免疫逃避中起关键的作用。每个单倍体基因组var基因家族编码约60种成员,通过启动转录不同的var基因变异体为抗原变异提供了分子基础。

18,940 citations

Journal ArticleDOI
23 Jan 2009-Cell
TL;DR: The current understanding of miRNA target recognition in animals is outlined and the widespread impact of miRNAs on both the expression and evolution of protein-coding genes is discussed.

18,036 citations

Journal ArticleDOI
TL;DR: Machine learning addresses many of the same research questions as the fields of statistics, data mining, and psychology, but with differences of emphasis.
Abstract: Machine Learning is the study of methods for programming computers to learn. Computers are applied to a wide range of tasks, and for most of these it is relatively easy for programmers to design and implement the necessary software. However, there are many tasks for which this is difficult or impossible. These can be divided into four general categories. First, there are problems for which there exist no human experts. For example, in modern automated manufacturing facilities, there is a need to predict machine failures before they occur by analyzing sensor readings. Because the machines are new, there are no human experts who can be interviewed by a programmer to provide the knowledge necessary to build a computer system. A machine learning system can study recorded data and subsequent machine failures and learn prediction rules. Second, there are problems where human experts exist, but where they are unable to explain their expertise. This is the case in many perceptual tasks, such as speech recognition, hand-writing recognition, and natural language understanding. Virtually all humans exhibit expert-level abilities on these tasks, but none of them can describe the detailed steps that they follow as they perform them. Fortunately, humans can provide machines with examples of the inputs and correct outputs for these tasks, so machine learning algorithms can learn to map the inputs to the outputs. Third, there are problems where phenomena are changing rapidly. In finance, for example, people would like to predict the future behavior of the stock market, of consumer purchases, or of exchange rates. These behaviors change frequently, so that even if a programmer could construct a good predictive computer program, it would need to be rewritten frequently. A learning program can relieve the programmer of this burden by constantly modifying and tuning a set of learned prediction rules. Fourth, there are applications that need to be customized for each computer user separately. Consider, for example, a program to filter unwanted electronic mail messages. Different users will need different filters. It is unreasonable to expect each user to program his or her own rules, and it is infeasible to provide every user with a software engineer to keep the rules up-to-date. A machine learning system can learn which mail messages the user rejects and maintain the filtering rules automatically. Machine learning addresses many of the same research questions as the fields of statistics, data mining, and psychology, but with differences of emphasis. Statistics focuses on understanding the phenomena that have generated the data, often with the goal of testing different hypotheses about those phenomena. Data mining seeks to find patterns in the data that are understandable by people. Psychological studies of human learning aspire to understand the mechanisms underlying the various learning behaviors exhibited by people (concept learning, skill acquisition, strategy change, etc.).

13,246 citations

Journal ArticleDOI
TL;DR: The Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines target the reliability of results to help ensure the integrity of the scientific literature, promote consistency between laboratories, and increase experimental transparency.
Abstract: Background: Currently, a lack of consensus exists on how best to perform and interpret quantitative real-time PCR (qPCR) experiments. The problem is exacerbated by a lack of sufficient experimental detail in many publications, which impedes a reader’s ability to evaluate critically the quality of the results presented or to repeat the experiments. Content: The Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines target the reliability of results to help ensure the integrity of the scientific literature, promote consistency between laboratories, and increase experimental transparency. MIQE is a set of guidelines that describe the minimum information necessary for evaluating qPCR experiments. Included is a checklist to accompany the initial submission of a manuscript to the publisher. By providing all relevant experimental conditions and assay characteristics, reviewers can assess the validity of the protocols used. Full disclosure of all reagents, sequences, and analysis methods is necessary to enable other investigators to reproduce results. MIQE details should be published either in abbreviated form or as an online supplement. Summary: Following these guidelines will encourage better experimental practice, allowing more reliable and unequivocal interpretation of qPCR results.

12,469 citations