Protein Arginine Deiminase (PAD) activity is aberrantly upregulated in multiple human diseases, including rheumatoid arthritis, colitis, and cancer. Thus, these enzymes are potential therapeutic targets. One isozyme, PAD4, helps control a number of physiological processes, including gene transcription, apoptosis, cell growth, and neutrophil extracellular trap formation. However, the specific roles of PAD4 in each of these processes are incompletely defined. Additionally, its mode of regulation and its contribution kinase signaling are only beginning to be understood and appreciated. The physiological roles of the other human PADs (i.e., PADs 1, 2, 3, and 6) are even less well understood. Building on previous work that described the development of PAD- targeted irreversible inhibitors and activity based proteomic probes (ABPPs), we propose to develop isozyme specific PAD inactivators. Additionally, we will use these compounds to identify and characterize the factors that regulate PAD activity, focusing initially on PAD4. Additionally, we will examine crosstalk between serine phosphorylation and arginine citrullination at both the molecular and cell biology levels.
Specific aims are: (1) The first aim of this proposal is focused on developing isozyme specific PAD inhibitors with improved potency and bioavailability.
This aim builds on our exciting discovery that haloacetamidine bearing compounds act as irreversible PAD inhibitors. Two strategies are proposed: (i) the replacement of the backbone amides in Cl-amidine, an inhibitor developed by the PI's lab, with libraries of triazoles and tetrazoles;and (i) a cyclic peptoid library approach. We will also identify novel warheads with altered reactivity tha overcome the limitations of the fluoro- and chloroacetamidine warheads. These compounds will ultimately serve as useful chemical probes of PAD function both directly and when converted into ABPPs to discover the factors that regulate the activity of a particular isozyme. (2) The second aim focuses on characterizing the post-translational modifications (PTMs) that regulate PAD4 activity. This work builds on our prior efforts showing that PAD4 is proteolyzed, acetylated, and ubiquitinated in vivo and these PTMs correlate with different activity states. Specifically, we describe an integrated chemical biology approach to examine the effects of these PTMs on PAD4 activity both in vitro and in vivo. (3) The third aim will study crosstalk between citrullination and serine phosphorylation. We are focused on these studies because we hypothesize that such crosstalk exists to 'fine-tune'cell signaling. Specifically, we demonstrate that crosstalk plays a role in regulating the phosphorylation of ELK1. Additionally, we will take candidate and proteomic approaches to determine the scope of crosstalk between these two PTMs. Once complete, the proposed studies will not only increase our understanding of PAD biology but will provide a suite of chemical probes that can be used to study protein citrullination.
The project focuses on the development and use of chemical probes targeting the Protein Arginine Deiminases (PADs), a group of enzymes whose activity is aberrantly increased in rheumatoid arthritis, colitis, and cancer. These probes will be used in an integrated chemical biology approach to characterize the factors that regulate PAD activity and explore crosstalk between protein phosphorylation and protein citrullination. In total, these studies will increase our understanding of PAD biology, which is critical for understanding how these enzymes contribute to human disease.
|Mohamed, Bashir M; Boyle, Noreen T; Schinwald, Anja et al. (2018) Induction of protein citrullination and auto-antibodies production in murine exposed to nickel nanomaterials. Sci Rep 8:679|
|Carmona-Rivera, Carmelo; Carlucci, Philip M; Moore, Erica et al. (2017) Synovial fibroblast-neutrophil interactions promote pathogenic adaptive immunity in rheumatoid arthritis. Sci Immunol 2:|
|Muth, Aaron; Subramanian, Venkataraman; Beaumont, Edward et al. (2017) Development of a Selective Inhibitor of Protein Arginine Deiminase 2. J Med Chem 60:3198-3211|
|Sun, Bo; Dwivedi, Nishant; Bechtel, Tyler J et al. (2017) Citrullination of NF-?B p65 promotes its nuclear localization and TLR-induced expression of IL-1? and TNF?. Sci Immunol 2:|
|Young, Coleman H; Rothfuss, Heather M; Gard, Philip F et al. (2017) Citrullination regulates the expression of insulin-like growth factor-binding protein 1 (IGFBP1) in ovine uterine luminal epithelial cells. Reproduction 153:1-10|
|Kosgodage, Uchini S; Trindade, Rita P; Thompson, Paul R et al. (2017) Chloramidine/Bisindolylmaleimide-I-Mediated Inhibition of Exosome and Microvesicle Release and Enhanced Efficacy of Cancer Chemotherapy. Int J Mol Sci 18:|
|Clancy, Kathleen W; Russell, Anna-Maria; Subramanian, Venkataraman et al. (2017) Citrullination/Methylation Crosstalk on Histone H3 Regulates ER-Target Gene Transcription. ACS Chem Biol 12:1691-1702|
|Chang, Hui-Hsin; Liu, Guang-Yaw; Dwivedi, Nishant et al. (2016) A molecular signature of preclinical rheumatoid arthritis triggered by dysregulated PTPN22. JCI Insight 1:e90045|
|Hosseinzadeh, Ava; Thompson, Paul R; Segal, Brahm H et al. (2016) Nicotine induces neutrophil extracellular traps. J Leukoc Biol 100:1105-1112|
|Ghari, Fatemeh; Quirke, Anne-Marie; Munro, Shonagh et al. (2016) Citrullination-acetylation interplay guides E2F-1 activity during the inflammatory response. Sci Adv 2:e1501257|
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