Deimination of Arginine Residues in Nucleophosmin/B23 and Histones in HL-60 Granulocytes

Hagiwara, Tokio; Nakashima, Katsuhiko; Hirano, Hisashi; Senshu, Tatsuo; Yamada, Masao

doi:10.1006/bbrc.2001.6303

Cited by 168 publications

(147 citation statements)

References 30 publications

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“…Crystallographic data suggest that PAD4 does not recognize specific arginine residues in a unique sequence context but rather recognizes five successive residues with the consensus sequence 1 ϕXRXX 5 , in which ϕ denotes amino acids with small side-chain moieties, and X denotes any amino acid (21). The target sequence must also be accessible and flexible enough to fit into the active pocket of PAD4, much like the N-terminal sequence SGRGK of human histones H4 and H2A, whose arginine at position 3 (R3) is one of the better described targets of PAD4 (14,21,22). Because initiating methionines (M) often are removed during protein maturation, we searched the proteome for sequences beginning with an amino (N)-terminal MSGR motif using pBLAST and PredMod (23).…”

Section: Stable Knockdown Of Pad4 Induces Emt and Increases The Invasivementioning

confidence: 99%

Dysregulation of PAD4-mediated citrullination of nuclear GSK3β activates TGF-β signaling and induces epithelial-to-mesenchymal transition in breast cancer cells

Stadler

Vincent

Fedorov

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

117

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Peptidylarginine deiminase 4 (PAD4) is a Ca 2+ -dependent enzyme that converts arginine and methylarginine residues to citrulline, with histone proteins being among its best-described substrates to date. However, the biological function of this posttranslational modification, either in histones or in nonhistone proteins, is poorly understood. Here, we show that PAD4 recognizes, binds, and citrullinates glycogen synthase kinase-3β (GSK3β), both in vitro and in vivo. Among other functions, GSK3β is a key regulator of transcription factors involved in tumor progression, and its dysregulation has been associated with progression of human cancers. We demonstrate that silencing of PAD4 in breast cancer cells leads to a striking reduction of nuclear GSK3β protein levels, increased TGF-β signaling, induction of epithelial-to-mesenchymal transition, and production of more invasive tumors in xenograft assays. Moreover, in breast cancer patients, reduction of PAD4 and nuclear GSK3β is associated with increased tumor invasiveness. We propose that PAD4-mediated citrullination of GSK3β is a unique posttranslational modification that regulates its nuclear localization and thereby plays a critical role in maintaining an epithelial phenotype. We demonstrate a dynamic and previously unappreciated interplay between histone-modifying enzymes, citrullination of nonhistone proteins, and epithelial-to-mesenchymal transition.

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Section: Stable Knockdown Of Pad4 Induces Emt and Increases The Invasivementioning

confidence: 99%

Dysregulation of PAD4-mediated citrullination of nuclear GSK3β activates TGF-β signaling and induces epithelial-to-mesenchymal transition in breast cancer cells

Stadler

Vincent

Fedorov

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

117

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“…Synovial intracellular citrullinated proteins were detected using a well-described and validated mouse IgM monoclonal antibody (mAb), F95, which was raised against a decacitrullinated peptide linked to the carrier protein, keyhole limpet hemocyanin (33)(34)(35). For the detection of the citrullinating PAD enzymes, we focused on PAD-2, based on the presence of both PAD-2 and PAD-4, but not PAD-1 and PAD-3, subtypes in synovial extracts (36), the expression of PAD-4 in the nucleus rather than the cytoplasm (37)(38)(39), and the absence of correlation between PAD-4 polymorphisms and intracellular citrullinated proteins (40). A single-chain protein against human PAD-2, similar to a light chain of an IgG antibody, was developed by phage display technology.…”

Section: Methodsmentioning

confidence: 99%

Synovial intracellular citrullinated proteins colocalizing with peptidyl arginine deiminase as pathophysiologically relevant antigenic determinants of rheumatoid arthritis–specific humoral autoimmunity

Rycke¹,

Nicholas²,

Cantaert³

et al. 2005

Arthritis & Rheumatism

118

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Objective. To address the ongoing debate concerning the specificity of synovial citrullinated proteins for rheumatoid arthritis (RA) and to analyze their pathophysiologic relevance to the induction or perpetuation of the RA-specific anti-citrullinated protein antibodies (ACPAs).Methods. Synovium of 19 RA patients and 19 non-RA controls was immunostained for the presence of citrullinated proteins with a mouse monoclonal antibody (F95), for the citrullinating enzyme peptidyl arginine deiminase type 2 (PAD-2), and for the free citrulline-producing enzyme inducible nitric oxide synthase (iNOS). Extending the RA cohort to 61 patients, the findings of anticitrulline staining in synovium were related to serum and synovial fluid ACPA levels, as measured by enzyme-linked immunosorbent assay.Results. F95 staining indicated the presence of synovial intracellular citrullinated proteins in 53% of RA samples versus 5% of control samples, whereas extracellular staining was not RA specific. Immunoblotting and inhibition experiments confirmed that the antibody recognized citrullinated proteins but not free citrulline. Accordingly, iNOS was equally found in RA and control synovium and in intracellular citrullinated protein-positive and intracellular citrullinated proteinnegative samples. In contrast, intracellular citrullinated proteins colocalized with PAD-2, which was found in 59% of RA samples versus 17% of control samples. Independent of local disease activity, the presence of the RA-specific synovial intracellular citrullinated proteins was associated with significantly higher systemic and local ACPA levels and with local ACPA production in the joint.Conclusion. These data confirm the presence of RA-specific intracellular citrullinated proteins in synovium. The link with PAD-2 and local and systemic ACPA levels emphasizes their pathophysiologic relevance for RA-specific humoral autoimmunity.

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“…One intriguing histone posttranslational modification is the deimination of specific arginines in histones H2A, H3, and H4 (2)(3)(4). The conversion of arginine to citrulline, an atypical amino acid that lacks arginine's positive charge, is accomplished by a member of the peptidylarginine deiminase (PAD) 3 family of enzymes, PAD4 (5).…”

mentioning

confidence: 99%

Histone Deimination As a Response to Inflammatory Stimuli in Neutrophils

Neeli

Khan

Radic

2008

The Journal of Immunology

486

524

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Posttranslational modifications, such as the deimination of arginine to citrulline by peptidyl arginine deiminase (PAD4), change protein structure and function. For autoantigens, covalent modifications represent a mechanism to sidestep tolerance and stimulate autoimmunity. To examine conditions leading to histone deimination in neutrophils, we used Abs that detect citrullines in the N terminus of histone H3. Deimination was investigated in human neutrophils and HL-60 cells differentiated into granulocytes. We observed rapid and robust H3 deimination in HL-60 cells exposed to LPS, TNF, lipoteichoic acid, f-MLP, or hydrogen peroxide, which are stimuli that activate neutrophils. Importantly, we also observed H3 deimination in human neutrophils exposed to these stimuli. Citrullinated histones were identified as components of extracellular chromatin traps (NETs) produced by degranulating neutrophils. In contrast, apoptosis proceeded without detectable H3 deimination in HL-60 cells exposed to staurosporine or camptothecin. We conclude that histone deimination in neutrophils is induced in response to inflammatory stimuli and not by treatments that induce apoptosis. Our results further suggest that deiminated histone H3, a covalently modified form of a prominent nuclear autoantigen, is released to the extracellular space as part of the neutrophil response to infections. The possible association of a modified autoantigen with microbial components could, in predisposed individuals, increase the risk of autoimmunity.

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Deimination of Arginine Residues in Nucleophosmin/B23 and Histones in HL-60 Granulocytes

Cited by 168 publications

References 30 publications

Dysregulation of PAD4-mediated citrullination of nuclear GSK3β activates TGF-β signaling and induces epithelial-to-mesenchymal transition in breast cancer cells

Dysregulation of PAD4-mediated citrullination of nuclear GSK3β activates TGF-β signaling and induces epithelial-to-mesenchymal transition in breast cancer cells

Synovial intracellular citrullinated proteins colocalizing with peptidyl arginine deiminase as pathophysiologically relevant antigenic determinants of rheumatoid arthritis–specific humoral autoimmunity

Histone Deimination As a Response to Inflammatory Stimuli in Neutrophils

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