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Eric Bennett

Research

My lab is interested in understanding the fundamental principles of how proteins are destroyed within cells. The cellular environment must constantly adapt to changing conditions and one mechanistic strategy commonly employed is the targeted modification of proteins by ubiquitin chains and subsequent degradation by the proteasome. The ubiquitin-proteasome system (UPS) can be viewed as the cellular waste management system. The UPS is utilized as the cellular quality control system as it is tasked with the degradation of potentially toxic misfolded and misassembled proteins. The inability of the UPS to properly destroy these proteins has been linked to various human pathologies including amyotrophic lateral sclerosis (ALS, commonly referred to as Lou Gehrig's disease), Parkinson's disease, and many human cancers. Thus, understanding the mechanisms of how the UPS recognizes proteins to be destroyed and how the pathway becomes dysfunctional will be instrumental in understanding the etiology of these diseases. The lab uses integrated approaches to understand UPS activity coupling biochemistry, cell biology, and systems-level approaches.

Deubiquitylating enzyme regulation

The ubiquitylation of substrates occurs via a hierarchy of enzyme activity where ubiquitin activating enzymes (E1s) activate free ubiquitin and transfers the activated ubiquitin to ubiquitin conjugating enzymes, (E2s), who partner with ubiquitin ligases (E3s) to finally conjugate ubiquitin to substrates.

figure 1

Deubiquitylating enzymes (Dubs) remove ubiquitin chains from substrates thereby antagonizing ubiquitin ligase activity. While the basic enzymatic function of Dubs is understood, how their activity is regulated, what cellular pathways are regulated by Dub activity, and what cellular substrates are the targets for Dub regulationare not fully understood. We previously characterized 75 of the 100 human Dubs based on their cellular interaction partners using proteomic technologies. During this study we established an integrative proteomic platform, termed CompPASS, figure 2

to rapidly analyze interaction proteomics data. We found that Dubs not only frequently associate with ubiquitin ligases but are themselves often modified by ubiquitin. This suggests that Dub activity may be regulated by E3 ligases and vice versa. The lab is interested in understanding the mechanisms of Dub activity regulation via association with ubiquitin ligases using both biochemical and cellular techniques.

Cullin-RING ligase regulation

One family of ubiquitin ligases are the multi-subunit ligases consisting of Cullin-RING ligases (CRLs)

figure 3

and the anaphase promoting complex (APC). Cullin-RING ligases are modular assemblies that can target many cellular proteins for regulated degradation by dynamically remodeling its subunit architecture. CRLs are known to degrade many critical oncoproteins and tumor suppressors, and as such there has been recent interest in the development of therapeutics targeting this pathway as an anti-cancer strategy.

CRLs are also regulated through a switch like mechanism in which the ubiquitin like protein Nedd8 modifies and subsequently activates the cullin subunit. Thus, by simply adding and removing the Nedd8 modification, CRL activity can be switched on and off. We are utilizing quantitative proteomics to study the cellular mechanisms used to control CRL activity. Additionally, this allows us to probe the dynamic nature of CRL assemblies and to understand how their architecture is remodeled to adapt to changes in the cellular environment.

figure 4

Global quantitative proteomic analysis of UPS function

The cellular proteome is continually monitored by the protein folding and degradation machinery to prevent the accumulation of potentially toxic misfolded or misassembled proteins. Proteins that contain errors in their primary sequence that arise during the translation process need to be recognized and selectively degraded by the UPS. When this quality control function breaks down it can have dire consequences on cellular and organismal fitness. Thus, the ability to globally monitor the activity of the UPS would allow for the identification of conditions that lead to UPS dysfunction. We have developed technologies that allow for the interrogation of the UPS activity on a systematic level. From these studies it is clear that mistranslated proteins are constantly ubiquitylated and cleared from cells. We are interested in trying to answer several questions related to quality control mechanisms. For instance, what are the ubiquitin ligases responsible for ubiquitylation mistranslated substrates? How do changes in free amino acid levels within the cell effect protein degradation and UPS function? Can we monitor alterations in UPS function in human pathologies associated with a decrease in quality control function? How does changing the redox potential within cells effect the quality control machinery? By understanding how proteins are recognized as quality control substrates and how the cell responds to challenges in protein homeostasis we hope to develop therapeutics that modulate quality control function.

figure 5 figure 6

Publications

  • Harper JW, Bennett EJ. Proteome complexity and the forces that drive proteome imbalance. Nature. 2016 Sep 14;537(7620):328-38. PMID: 27629639
  • Arribas-Layton M, Dennis J, Bennett EJ, Damgaard CK, Lykke-Andersen J. The C-terminal RGG domain of human Lsm4 promotes processing body formation stimulated by arginine dimethylation. Mol Cell Biol. 2016 May 31 PMID: 27247266
  • Gendron JM, Webb K, Yang B, Rising L, Zuzow N, Bennett EJ. Using the ubiquitin-modified proteome to monitor distinct and spatially restricted protein homeostasis dysfunction. Mol Cell Proteomics. 2016 May 16. PMID: 27185884
  • Fu R, Olsen MT, Webb K, Bennett EJ, Lykke-Andersen J. Recruitment of the 4EHP-GYF2 cap-binding complex to tetraproline motifs of tristetraprolin promotes repression and degradation of mRNAs with AU-rich elements. RNA. 2016 Mar;22(3):373-82. PMID: 26763119
  • Higgins R, Gendron JM, Rising L, Mak R, Webb K, Kaiser SE, Zuzow N, Riviere P, Yang B, Fenech E, Tang X, Lindsay S, Christianson JC, Hampton RY, Wasserman SA, Bennett EJ. The unfolded protein response triggers site-specific regulatory ubiquitylation of 40S ribosomal proteins. Mol Cell. 2015 Jul 2;59(1):35-49. PMID: 26051182
  • Erickson SL, Corpuz EO, Maloy JP, Fillman C, Webb K, Bennett EJ, Lykke-Andersen J. A competition between decapping complex formation and ubiquitin-mediated proteasomal degradation controls human Dcp2 decapping activity. Mol Cell Biol. 2015 Apr 13.PMID: 25870104
  • Facette MR, Park Y, Sutimantanapi D, Luo A, Cartwright HN, Yang B, Bennett EJ, Sylvester AW, Smith LG. The SCAR/WAVE complex polarizes PAN receptors and promotes division asymmetry in maize. Nat Plants. 2015 Jan 26;1:14024. PMID: 27246760
  • Lyumkis D, Oliveira dos Passos D, Tahara EB, Webb K, Bennett EJ, Vinterbo S, Potter CS, Carragher B, Joazeiro CA. Structural basis for translational surveillance by the large ribosomal subunit-associated protein quality control complex. Proc Natl Acad Sci U S A. 2014 Nov 11;111(45):15981-6. PMID: 25349383
  • Ikeuchi Y, Dadakhujaev S, Chandhoke AS, Huynh MA, Oldenborg A, Ikeuchi M, Deng L, Bennett EJ, Harper JW, Bonni A, Bonni S. TIF1gamma Regulates Epithelial Mesenchymal Transition by Operating as a SUMO E3 Ligase for theTranscriptional Regulator SnoN1. J Biol Chem. 2014 Sep 5;289(36):25067-78. PMID: 25059663
  • Lykke-Andersen J, Bennett EJ. Protecting the proteome: Eukaryotic cotranslational quality control pathways.J Cell Biol. 2014 Feb 17;204(4):467-76. PMID: 24535822
  • Mejia LA, Litterman N, Ikeuchi Y, de la Torre-Ubieta L, Bennett EJ, Zhang C, Harper JW, Bonni A. A novel Hap1-Tsc1 interaction regulates neuronal mTORC1 signaling and morphogenesis in the brain. J Neurosci. 2013 Nov 13;33(46):18015-21. PMID: 24227713
  • Ziemba A, Hill S, Sandoval D, Webb K, Bennett EJ, Kleiger G. Multimodal mechanism of action for the Cdc34 acidic loop: a case study for why ubiquitin-conjugating enzymes have loops and tails. J Biol Chem. 2013 Nov 29;288(48):34882-96. PMID: 24129577
  • Tan MK, Lim HJ, Bennett EJ, Shi Y, Harper JW. Parallel SCF Adaptor Capture Proteomics Reveals a Role for SCFFBXL17 in NRF2 Activation via BACH1 Repressor Turnover. Molecular Cell 2013 Sep 10. PMID: 24035498
  • Zhang C, Mejia LA, Huang J, Valnegri P, Bennett EJ, Anckar J, Jahani-Asl A, Gallardo G, Ikeuchi Y, Yamada T, Rudnicki M, Harper JW, Bonni A. The X-linked intellectual disability protein PHF6 associates with the PAF1 complex and regulates neuronal migration in the mammalian brain. Neuron 2013 Jun 19;78(6):986-93. PMID: 23791194
  • Carrano AC, Bennett EJ. Using the ubiquitin modified proteome to monitor protein homeostasis function. Mol Cell Proteomics 2013 May 23. PMID: 23704779
  • Armour SM, Bennett EJ, Braun CR, Zhang XY, McMahon SB, Gygi SP, Harper JW, Sinclair DA. A high-confidence interaction map identifies SIRT1 as a mediator of acetylation of USP22 and the SAGA coactivator complex. Mol Cell Biol. 2013 Apr;33(8):1487-502. PMID: 23382074
  • Monda JK, Scott DC, Miller DJ, Lydeard J, King D, Harper JW, Bennett EJ, Schulman BA. Structural Conservation of Distinctive N-terminal Acetylation-Dependent Interactions across a Family of Mammalian NEDD8 Ligation Enzymes. Structure 2013 Jan 8;21(1):42-53. PMID: 23201271
  • Christianson JC*, Olzmann JA*, Shaler TA, Sowa ME, Bennett EJ, Richter CM, Tyler RE, Greenblatt EJ, Harper JW, Kopito RR. Defining Human ERAD Networks through an Integrative Mapping Strategy. Nature Cell Biology 2011 Nov 27;14(1):93-105. PMID: 22119785
    *Authors contributed equally
  • Scott DC, Monda JK, Bennett EJ, Harper JW, Schulman BA. N-Terminal Acetylation Acts as an Avidity Enhancer Within an Interconnected Multiprotein Complex. Science 2011 Nov 4;334(6056): 674-8. PMID: 21940857
  • Kim W*, Bennett EJ*, Huttlin EL, Guo A, Li J, Possemato A, Sowa ME, Rad R, Rush J, Comb MJ, Harper JW, and Gygi SP. Systematic and quantitative assessment of the ubiquitin modified proteome Molecular Cell 2011 Oct 1;44(2):325-40. PMID: 21906983
    *Authors contributed equally
  • Bennett EJ, Rush J, Gygi SP, Harper JW. Dynamics of cullin-RING ubiquitin ligase assembly revealed by systematic quantitative proteomics. Cell 2010 Dec 10;143(6):951-965.
  • Bennett EJ, Harper JW. Ubiquitin gets CARDed. Cell. 2010 Apr 16;141(2):220-222.
  • Bennett EJ, Sowa ME, Harper JW. Cellular regulation by deubiquitinating enzymes. Nat. Rev. Mol Cell Bio. 2010 Feb Poster insert.
  • Sowa ME*, Bennett EJ*, Gygi SP, Harper JW. Defining the Human Deubiquitinating Enzyme Interaction Landscape. Cell. 2009 Jul 23;138(2):389-403. *Authors contributed equally
  • Bennett EJ, Harper JW. DNA damage: ubiquitin marks the spot. Nat Struct Mol Biol. 2008 Jan;15(1):20-22.
  • Bennett EJ, Shaler TA, Woodman B, Ryu KY, Zaitseva TS, Becker CH, Bates GP, Schulman H, Kopito RR. Global changes to the ubiquitin system in Huntington’s disease. Nature 2007 Aug 9;448(7154):704-708
  • Bennett EJ, Bence NF, Jayakumar R, Kopito RR. Global impairment of the ubiquitinproteasome system by nuclear or cytoplasmic protein aggregates precedes inclusion body formation. Molecular Cell 2005 Feb 4;17(3):351-365.
  • Bence NF, Bennett EJ, Kopito RR. Application and Analysis of the GFPu Family of Ubiquitin-Proteasome System Reporters. Methods in Enzymology 2005;399:481-490.
  • Bennett EJ, Bjerregaard J, Knapp JE, Chavous DA, Friedman AM, Royer WE, and O’Connor CM. Catalytic Implications from the Drosophila Protein L-Isoaspartyl Methyltransferase Structure and Site-Directed Mutagenesis. Biochemistry 2003 Nov 11;42(44):12844-12853.

Biography

Dr. Bennett received his PhD from Stanford University. He then carried out postdoctoral work at the Harvard Medical School.