Eric Bennett

Assistant Professor
Section of Cell and Developmental Biology

e-mail: e1bennett@ucsd.edu
Lab Homepage: Bennett Lab

     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. 

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,

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)

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.

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.

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.

Kim, W.*, Bennett, E. J*., Huttlin, E. L., Guo, A., Li, J., Possemato, A., Sowa, M. E., Rad, R., Rush, J., Comb, M. J., Harper, J. W., and Gygi, S. P. Systematic and quantitative assessment of the ubiquitin modified proteome  Molecular Cell, Online-08 September 2011
*Authors contributed equally

     Dr. Bennett received his PhD from Stanford University. He then carried out postdoctoral work at the Harvard Medical School.  He will be joining U.C. San Diego in July, 2011.