Increased Citrullination of Histone H3 in Multiple Sclerosis Brain and Animal Models of Demyelination: A Role for Tumor Necrosis Factor-Induced Peptidylarginine Deiminase 4 Translocation

Abstract

Modification of arginine residues by citrullination is catalyzed by peptidylarginine deiminases (PADs), of which five are known, generating irreversible protein structural modifications. We have shown previously that enhanced citrullination of myelin basic protein contributed to destabilization of the myelin membrane in the CNS of multiple sclerosis (MS) patients. We now report increased citrullination of nucleosomal histones by PAD4 in normal-appearing white matter (NAWM) of MS patients and in animal models of demyelination. Histone citrullination was attributable to increased levels and activity of nuclear PAD4. PAD4 translocation into the nucleus was attributable to elevated tumor necrosis factor-␣ (TNF-␣) protein. The elevated TNF-␣ in MS NAWM was not associated with CD3 ϩ or CD8 ϩ lymphocytes, nor was it associated with CD68 ϩ microglia/macrophages. GFAP, a measure of astrocytosis, was the only cytological marker that was consistently elevated in the MS NAWM, suggesting that TNF-␣ may have been derived from astrocytes. In cell cultures of mouse and human oligodendroglial cell lines, PAD4 was predominantly cytosolic but TNF-␣ treatment induced its nuclear translocation. To address the involvement of TNF-␣ in targeting PAD4 to the nucleus, we found that transgenic mice overexpressing TNF-␣ also had increased levels of citrullinated histones and elevated nuclear PAD4 before demyelination. In conclusion, high citrullination of histones consequent to PAD4 nuclear translocation is part of the process that leads to irreversible changes in oligodendrocytes and may contribute to apoptosis of oligodendrocytes in MS.

Myelin localization of peptidylarginine deiminases 2 and 4: comparison of PAD2 and PAD4 activities

“…These observations have led us to postulate that citrullinated MBP represents an important component in the pathogenesis of MS. 4 The mechanism by which arginine is converted to citrulline in proteins involves a family of enzymes, the peptidylarginine deiminases, of which five isoforms are known. 1 Although they are widely distributed, PAD2 is the predominant brain enzyme, but PAD4 is also present, as we have shown previously 14 and in this paper. In order to understand the important role of PAD2 in deimination of MBP in demyelinating disease, we have performed immunoelectron microscopy on intact mouse optic nerve with anti-PAD2 antibody, to localize it in the myelin sheath.…”

“…This is consistent with our immunoslot blot measurements showing a threefold increase in the amount of PAD2 enzyme in normal-appearing white matter from MS tissue. 14 …”

“…The number of gold particles in myelin from MS brain was increased threefold over normal, in agreement with a previous study in which we reported a threefold increase in the amount of PAD 2 by immunoblots. 14 In a study of the deiminated sites, no restrictions were observed, that is, all arginyl residues except one were targets for deimination in aqueous media. Since citrullinated MBP is generated by PAD, these studies suggest that decreased PAD enzyme activity may be a therapeutic target with PAD-specific inhibitors, as suggested for rheumatoid arthritis.…”

Post-Translational Modifications of Nucleosomal Histones in Oligodendrocyte Lineage Cells in Development and Disease

“…Increased histone citrullination on arginine residues was reported to be increased in the normal-appearing white matter of MS patients and in animal models of demyelination (Mastronardi et al 2006). Citrullination of arginine residues on histone tails is catalyzed by peptidylarginine deiminase 4 (PAD4).…”

“…Under normal conditions, PAD4 is cytosolically localized. However, under pathological conditions, such as in the presence of abnormally increased level of tumor necrosis factor α (TNFα), PAD4 is translocated to the nucleus, and its increased nuclear expression and activity catalyzes the citrullination of nucleosomal histones (Mastronardi et al 2006). Importantly, these molecular changes occur before the onset of symptoms in animal models of demyelination (Mastronardi et al 2006), indicating that TNFα-induced PAD4 nuclear localization and subsequent histone citrullination may be part of the etiopathogenesis of the disease.…”