The 26S proteasome is the primary site for protein degradation in mammalian cells and is critical in cell regulation, protein quality control, and antigen presentation. Although proteasome inhibitors are widely used as research tools and in treatment of certain cancers, the mechanisms by which proteasomes catalyze the ATP- dependent degradation of ubiquitinated proteins are still poorly understood. Our recent studies defined several new ATP-dependent steps in this process. In coming years, we hope to learn how these various steps are regulated and coordinated to ensure efficient degradation of different types of proteins. After binding to the 26S, ubiquitin (Ub) conjugates activate their own degradation by stimulating ATP hydrolysis and gate opening via interactions with either of the 19S-associated deubiquitinating enzymes, Usp14 or Uch37. It will be important to understand further their regulatory roles, and how they coordinate substrate deubiquitination and proteolysis. Protein breakdown by the 26S is tightly linked to ATP hydrolysis, and we have shown that the six 19S ATPase subunits hydrolyze ATP in an ordered manner. We plan to critically test this novel cyclic mechanism and to learn if other AAA ATPases operate in a similar manner. We also recently found that prions (PrPSc) aggregates disrupt proteasome function by inhibiting gate opening and substrate entry into the 20S particle. We shall explore whether the soluble protein aggregates that appear to cause other neurodegenerative diseases cause a similar inhibition of proteasome gating. Related studies will explore how three newly discovered modifications of proteasome structure regulate 19S activity: 1) we recently found that the Ub-binding subunit Rpn13 is continually poly-ubiquitinated by 26S- associated Ub ligases, and that this modification inhibits conjugate degradation. 2) Upon excitatory stimulation of neurons, the ATPase subunit Rpt6 becomes phosphorylated, and this modification is important in synaptic plasticity. 3) The 19S caps on singly and doubly capped forms of the 26S proteasomes appear to differ in content of certain subunits. We hope to clarify the functional consequences of these intriguing proteasome modifications and learn how they influence different steps in conjugate degradation. Unlike mammalian cells, bacteria catalyze protein degradation by ubiquitin-independent, ATP-requiring protease complexes. Mycobacterium tuberculosis (Mtb) has two such ClpP genes and two regulatory AAA ATPases, ClpC1 and ClpX, which are all attractive new drug targets. We discovered that ClpP1 and ClpP2 encode a novel 2-ring complex, ClpP1P2, with a unique assembly mechanism. We now hope to solve its structure and define the detailed mechanisms of these two types of AAA ATPases and of the promising new antibiotic, Novo23, that we found uncouples ATP hydrolysis by ClpC1 from proteolysis by ClpP1P2. !
The ubiquitin-proteasome system is the major pathway for protein degradation in eukaryotic cells and plays a critical role in eliminating misfolded and key regulatory proteins. The overall goal of our studies is to clarify the biochemical mechanisms by which the 26S proteasome catalyzes the rapid and efficient degradation of proteins tagged for degradation by the linkage to a chain of ubiquitin molecules. These studies are focusing on the roles of the proteasome-regulatory ATPases that bind protein substrates and translocate them into the 20S proteasome for degradation, but in addition, we are investigating a novel ATP-dependent proteolytic complex from Mycobacteria tuberculosis, ClpP1P2, which has unique biochemical properties and is a very attractive drug target.
|Myeku, Natura; Clelland, Catherine L; Emrani, Sheina et al. (2016) Tau-driven 26S proteasome impairment and cognitive dysfunction can be prevented early in disease by activating cAMP-PKA signaling. Nat Med 22:46-53|
|Sha, Zhe; Goldberg, Alfred L (2016) Reply to Vangala et al.: Complete inhibition of the proteasome reduces new proteasome production by causing Nrf1 aggregation. Curr Biol 26:R836-7|
|Zhao, Jinghui; Garcia, Gonzalo A; Goldberg, Alfred L (2016) Control of proteasomal proteolysis by mTOR. Nature 529:E1-2|
|Zhao, Jinghui; Zhai, Bo; Gygi, Steven P et al. (2015) mTOR inhibition activates overall protein degradation by the ubiquitin proteasome system as well as by autophagy. Proc Natl Acad Sci U S A 112:15790-7|
|Lokireddy, Sudarsanareddy; Kukushkin, Nikolay Vadimovich; Goldberg, Alfred Lewis (2015) cAMP-induced phosphorylation of 26S proteasomes on Rpn6/PSMD11 enhances their activity and the degradation of misfolded proteins. Proc Natl Acad Sci U S A 112:E7176-85|
|Tsvetkov, Peter; Mendillo, Marc L; Zhao, Jinghui et al. (2015) Compromising the 19S proteasome complex protects cells from reduced flux through the proteasome. Elife 4:|
|Akopian, Tatos; Kandror, Olga; Tsu, Christopher et al. (2015) Cleavage Specificity of Mycobacterium tuberculosis ClpP1P2 Protease and Identification of Novel Peptide Substrates and Boronate Inhibitors with Anti-bacterial Activity. J Biol Chem 290:11008-20|
|Goldberg, Alfred L; Zhao, Jinghui; Collins, Galen A (2015) Blocking Cancer Growth with Less POMP or Proteasomes. Mol Cell 59:143-5|
|Rock, Kenneth L; FarfÃ¡n-Arribas, Diego J; Colbert, Jeff D et al. (2014) Re-examining class-I presentation and the DRiP hypothesis. Trends Immunol 35:144-52|
|Gavrish, Ekaterina; Sit, Clarissa S; Cao, Shugeng et al. (2014) Lassomycin, a ribosomally synthesized cyclic peptide, kills mycobacterium tuberculosis by targeting the ATP-dependent protease ClpC1P1P2. Chem Biol 21:509-18|
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