Chemical proteomic map of dimethyl fumarate–sensitive cysteines in primary human T cells

Abstract

Dimethyl fumarate (DMF) is an electrophilic drug that is used to treat autoimmune conditions, including multiple sclerosis and psoriasis. The mechanism of action of DMF is unclear, but may involve the covalent modification of proteins or DMF serving as a pro-drug that is converted to monomethyl fumarate (MMF). Here, we found that DMF, but not MMF, blocked the activation of primary human and mouse T cells. Using a quantitative, site-specific chemical proteomic platform, we determined the DMF-sensitivity of > 2400 cysteine residues in human T cells. Cysteines sensitive to DMF, but not MMF, were identified in several proteins with established biochemical or genetic links to T cell function, including protein kinase C θ (PKCθ). Furthermore, DMF blocked the association of PKCθ with the costimulatory receptor CD28 by perturbing a CXXC motif in the C2 domain of this kinase. Mutation of these DMF-sensitive cysteines also impaired PKCθ-CD28 interactions and T cell activation, designating the C2 domain of PKCθ as a key functional, electrophile-sensing module important for T cell biology.

Effects of fumarates on inflammatory human astrocyte responses and oligodendrocyte differentiation

“…Taken together, these results support the hypothesis that in addition to MMF, DMF can also directly contribute to the anti‐inflammatory and regenerative effects of this DMT. These results align with recent findings that have been previously observed in peripheral immune cells9, 10, 15 and brain‐resident microglia 9, 16…”

“…Furthermore, it has been demonstrated that the anti‐inflammatory effects of fumarates may be restricted to DMF, with other fumarate metabolites having little influence on inflammation 9, 10, 13, 14. A recent report has demonstrated that DMF, but not metabolite monomethyl fumarate (MMF), is a potent inhibitor of NF‐ κ B signaling within immune cells through the modification in cysteine residues 15. This is consistent with findings that DMF promotes an anti‐inflammatory shift in activated microglia 9, 16.…”

Global profiling of lysine reactivity and ligandability in the human proteome

“…We also believe that our studies, despite having uncovered more than 100 lysines targeted by fragment electrophiles, almost certainly still underestimate the global ligandability of lysines in the human proteome. The development and evaluation of larger compound libraries displaying more diversified recognition and amine-reactive elements, including covalent-reversible electrophiles (e.g., aldehydes), in combination with surveying complementary cell types (e.g., primary immune cells 62 , metabolic organs 63 ), should greatly enrich our understanding of functional and ligandable lysines in the human proteome and, through doing so, extend its druggable landscape for basic and translational research objectives…”

Transcription Factor NRF2 as a Therapeutic Target for Chronic Diseases: A Systems Medicine Approach

“…Thus, DMF may interact with cysteine residues in several proteins that regulate NF-kB signaling (Blewett et al, 2016). In addition, DMF inhibits ubiquitin-conjugating enzymes and thus prevents the degradation of the IkB repressor of NF-kB in response to IL-1b or Toll-like receptor agonists (McGuire et al, 2016).…”

“…In addition, DMF inhibits ubiquitin-conjugating enzymes and thus prevents the degradation of the IkB repressor of NF-kB in response to IL-1b or Toll-like receptor agonists (McGuire et al, 2016). Moreover, DMF binds directly to specific cysteine residues in protein kinase C-u, a key kinase involved in signaling by the T cell receptor (Blewett et al, 2016). In addition, MMF and DMF activate the hydroxycarboxylic acid receptor-2, resulting in inhibition of NF-kB and downregulation of proinflammatory cytokines and adhesion molecules Gillard et al, 2015) and leading to decreased neutrophil infiltration .…”