We are upgrading the repository! The content freeze has been extended to December 11, 2024, when we expect the new repository to become available. New submissions or changes to existing items will not be allowed until after the new website goes live. All content already published will remain publicly available for searching and downloading. Updates will be posted in the Website Upgrade 2024 FAQ in the sidebar Help menu. Reach out to escholarship@umassmed.edu with any questions.
Determination of ubiquitin fitness landscapes under different chemical stresses in a classroom setting
UMass Chan Affiliations
Department of Biochemistry and Molecular PharmacologyDocument Type
Journal ArticlePublication Date
2016-04-25Keywords
S. cerevisiaebiophysics
chemical biology
computational biology
deep mutational scanning
evolutionary biology
genomics
proteasome
proteostasis
systems biologyubiquitinBiochemistryBiophysicsComputational BiologyGenomicsOther Ecology and Evolutionary BiologySystems Biology
Metadata
Show full item recordAbstract
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.Source
Elife. 2016 Apr 25;5. pii: e15802. doi: 10.7554/eLife.15802. Link to article on publisher's site
DOI
10.7554/eLife.15802Permanent Link to this Item
http://hdl.handle.net/20.500.14038/40049PubMed ID
27111525Notes
Full author list omitted for brevity. For full list of authors see article.
Related Resources
Rights
Copyright © 2016, Mavor et al. This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.Distribution License
http://creativecommons.org/licenses/by/4.0/Scopus Count
Collections
Except where otherwise noted, this item's license is described as Copyright © 2016, Mavor et al. This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.
Related items
Showing items related by title, author, creator and subject.
-
The genome-wide multi-layered architecture of chromosome pairing in early Drosophila embryosGenome organization involves cis and trans chromosomal interactions, both implicated in gene regulation, development, and disease. Here, we focus on trans interactions in Drosophila, where homologous chromosomes are paired in somatic cells from embryogenesis through adulthood. We first address long-standing questions regarding the structure of embryonic homolog pairing and, to this end, develop a haplotype-resolved Hi-C approach to minimize homolog misassignment and thus robustly distinguish trans-homolog from cis contacts. This computational approach, which we call Ohm, reveals pairing to be surprisingly structured genome-wide, with trans-homolog domains, compartments, and interaction peaks, many coinciding with analogous cis features. We also find a significant genome-wide correlation between pairing, transcription during zygotic genome activation, and binding of the pioneer factor Zelda. Our findings reveal a complex, highly structured organization underlying homolog pairing, first discovered a century ago in Drosophila. Finally, we demonstrate the versatility of our haplotype-resolved approach by applying it to mammalian embryos.
-
Combined experimental and computational analysis of DNA damage signaling reveals context-dependent roles for Erk in apoptosis and G1/S arrest after genotoxic stressFollowing DNA damage, cells display complex multi-pathway signaling dynamics that connect cell-cycle arrest and DNA repair in G1, S, or G2/M phase with phenotypic fate decisions made between survival, cell-cycle re-entry and proliferation, permanent cell-cycle arrest, or cell death. How these phenotypic fate decisions are determined remains poorly understood, but must derive from integrating genotoxic stress signals together with inputs from the local microenvironment. To investigate this in a systematic manner, we undertook a quantitative time-resolved cell signaling and phenotypic response study in U2OS cells receiving doxorubicin-induced DNA damage in the presence or absence of TNFalpha co-treatment; we measured key nodes in a broad set of DNA damage signal transduction pathways along with apoptotic death and cell-cycle regulatory responses. Two relational modeling approaches were then used to identify network-level relationships between signals and cell phenotypic events: a partial least squares regression approach and a complementary new technique which we term 'time-interval stepwise regression.' Taken together, the results from these analysis methods revealed complex, cytokine-modulated inter-relationships among multiple signaling pathways following DNA damage, and identified an unexpected context-dependent role for Erk in both G1/S arrest and apoptotic cell death following treatment with this commonly used clinical chemotherapeutic drug.
-
Mapping and analysis of Caenorhabditis elegans transcription factor sequence specificitiesCaenorhabditis elegans is a powerful model for studying gene regulation, as it has a compact genome and a wealth of genomic tools. However, identification of regulatory elements has been limited, as DNA-binding motifs are known for only 71 of the estimated 763 sequence-specific transcription factors (TFs). To address this problem, we performed protein binding microarray experiments on representatives of canonical TF families in C. elegans, obtaining motifs for 129 TFs. Additionally, we predict motifs for many TFs that have DNA-binding domains similar to those already characterized, increasing coverage of binding specificities to 292 C. elegans TFs (~40%). These data highlight the diversification of binding motifs for the nuclear hormone receptor and C2H2 zinc finger families, and reveal unexpected diversity of motifs for T-box and DM families. Motif enrichment in promoters of functionally related genes is consistent with known biology, and also identifies putative regulatory roles for unstudied TFs.
