Nonenzymatic Conversion of ADP-Ribosylated Arginines to Ornithine Alters the Biological Activities of Human Neutrophil Peptide-1

Abstract

Activated neutrophils, recruited to the airway of diseased lung, release human neutrophil peptides (HNP1–4) that are cytotoxic to airway cells as well as microbes. Airway epithelial cells express arginine-specific ADP-ribosyltransferase (ART)-1, a glycophosphatidylinositol-anchored ART that transfers ADP-ribose from NAD to arginines 14 and 24 of HNP-1.We previously reported that ADP-ribosyl-arginine is converted non-enzymatically to ornithine and that ADP-ribosylated HNP-1 and ADP-ribosyl-HNP-(ornithine) were isolated from bronchoalveolar lavage fluid of a patient with idiopathic pulmonary fibrosis, indicating that these reactions occur in vivo. To determine effects of HNP-ornithine on the airway, three analogs of HNP-1, HNP-(R14orn), HNP-(R24orn) and HNP-(R14,24orn) were tested for their activity against Pseudomonas aeruginosa, Escherichia coli and Staphylococcus aureus, their cytotoxic effects on A549, NCI-H441, SAEC epithelial-like cells and normal human lung fibroblasts (NHLF) and their ability to stimulate IL-8 and TGF-β1 release from A549 cells, and to serve as ART1 substrates. HNP and the three analogs had similar effects on IL-8 and TGF-β1 release from A549 cells and were all cytotoxic for SAEC, NCI-H441 and NHLF. HNP-(R14,24orn) when compared to HNP-1 and HNP-1 with a single ornithine substitution for arginine 14 or 24 exhibited reduced cytotoxicity, but it enhanced proliferation of A549 cells and had antibacterial activity. Thus, arginines 14 and 24, which can be ADP-ribosylated by ART1, are critical to the regulation of the cytotoxic and antibacterial effects of HNP-1. The HNP analog, HNP-(R14,24orn) lacks the epithelial cell cytotoxicity of HNP-1 but partially retains its antibacterial activity and thus may have clinical applications in airway disease.

Presence of anti-acetylated peptide antibodies (AAPA) in inflammatory arthritis and other rheumatic diseases suggests discriminative diagnostic capacity towards early rheumatoid arthritis

“… 10 , 32 Ornithine-like citrulline is an unusual amino acid that is not encoded in the genome but created by non-enzymatic post-translational modification as described by several authors. 38 , 39 Interestingly however, this does not appear sufficient to create a neo-epitope since only the peptide carrying acetylated ornithine was targeted by autoantibodies. Thus, it would be tempting to speculate that ac-ornithine containing neo-epitopes generated by aberrant post-translational modification form one of the primary targets of the AAPA response, which then may partially cross-react with ac-lysine containing epitopes but also with citrullinated or carbamylated epitopes.…”

Role of a TRIM72 ADP-ribosylation cycle in myocardial injury and membrane repair

“…In previous reports, however, other proteins, e.g., integrin α7 (48, 49) and FGF-2 (11), have been shown to be ADP-ribosylated in vitro. The only confirmed substrate of ART1 in vivo is human neutrophil peptide-1 (12,50). Cholera toxin catalyzes ADP-ribosylation of Gαs, a substrate of ARH1 in ARH1-KO mice in vivo (51).…”

ARH1 in Health and Disease

“…Six ADP-ribosylated proteins overlapping with mouse heart, mouse skeletal muscle, and C2C12 myotubes were identified (Figure 2), e.g., Golgi apparatus protein 1 (Glg1), basement membrane-specific heparan sulfate proteoglycan core protein (Hspg2), integrin α-7, nidogen-1, protein disulfide-isomerase A3, and TRIM72 (Table 1). HNP-1, integrin α-7, and TRIM72 represent the only previously reported ART1 target proteins [5,27,36,80,81]. Among these 354 proteins, only 6 proteins were identified in Art1-deficient mouse skeletal muscle (Table 2) [39].…”

“…HNP-1 ADP-ribosylated on arginines 14 and 24 shows reduced antimicrobial and cytotoxic activities [35]. Non-enzymatic replacement of the ADP-ribosylated arginines of HNP-1 with ornithine resulted in a peptide with less cytotoxicity than unmodified HNP-1 but with retention of its antibacterial activity [36]. Thus, the ornithine-containing HNP-1, which is no longer a substrate for ARH1, may have therapeutic potential as an ARH1-resistant molecule.…”