Citrullination of CXCL8 by peptidylarginine deiminase alters receptor usage, prevents proteolysis, and dampens tissue inflammation

Abstract

Biological functions of proteins are influenced by posttranslational modifications such as on/off switching by phosphorylation and modulation by glycosylation. Proteolytic processing regulates cytokine and chemokine activities. In this study, we report that natural posttranslational citrullination or deimination alters the biological activities of the neutrophil chemoattractant and angiogenic cytokine CXCL8/interleukin-8 (IL-8). Citrullination of arginine in position 5 was discovered on 14% of natural leukocyte-derived CXCL8(1–77), generating CXCL8(1–77)Cit5. Peptidylarginine deiminase (PAD) is known to citrullinate structural proteins, and it may initiate autoimmune diseases. PAD efficiently and site-specifically citrullinated CXCL5, CXCL8, CCL17, CCL26, but not IL-1β. In comparison with CXCL8(1–77), CXCL8(1–77)Cit5 had reduced affinity for glycosaminoglycans and induced less CXCR2-dependent calcium signaling and extracellular signal-regulated kinase 1/2 phosphorylation. In contrast to CXCL8(1–77), CXCL8(1–77)Cit5 was resistant to thrombin- or plasmin-dependent potentiation into CXCL8(6–77). Upon intraperitoneal injection, CXCL8(6–77) was a more potent inducer of neutrophil extravasation compared with CXCL8(1–77). Despite its retained chemotactic activity in vitro, CXCL8(1–77)Cit5 was unable to attract neutrophils to the peritoneum. Finally, in the rabbit cornea angiogenesis assay, the equally potent CXCL8(1–77) and CXCL8(1–77)Cit5 were less efficient angiogenic molecules than CXCL8(6–77). This study shows that PAD citrullinates the chemokine CXCL8, and thus may dampen neutrophil extravasation during acute or chronic inflammation.

Recognition Versus Adaptive Up-regulation and Degradation of CC Chemokines by the Chemokine Decoy Receptor D6 Are Determined by Their N-terminal Sequence

“…Alternatively, they may also be classified on the basis of their expression patterns and functions as "inflammatory" (produced only upon activation) and "homeostatic" (expressed in discrete locations in the absence of apparent activating stimuli) (4). In addition to gene expression regulation, post-transcriptional regulatory mechanisms, including chemokine degradation by non-signaling chemokine decoy receptors (5,6), binding to glycosaminoglycans (7), and posttranslational processing by endogenous peptidases or peptidyl arginine deiminases (8,9), are emerging mechanisms that fine tune chemokine biological activities and leukocyte recruitment.…”

Citrullination and Proteolytic Processing of Chemokines by Porphyromonas gingivalis

“…3). At first, an obvious explanation for this could be the NH 2 -terminal cleavage of CXCL8 at Lys8 by Kgp, which potentially removes the first 8 amino acids, including the Arg in position 5, i.e., the Arg which is most susceptible to citrullination by human PAD2 and PAD4 (8). However, the citrullination levels detected upon incubation with PPAD in vitro were also very low.…”

“…PTMs affect the biological activity of cytokines and chemokines (2,4,58). Recently, citrullination, a new PTM of chemokines, has been discovered (8,21). The conversion of peptidylarginine to peptidylcitrulline is performed by the mammalian PAD enzymes (25).…”

“…Also, naturally occurring N-and O-glycosylated forms of chemokines with altered lectin binding properties have been identified (18)(19)(20). In addition to NH 2 -and COOH-terminal proteolytic processing and glycosylation, citrullination, i.e., deimination of arginine (Arg) to citrulline (Cit), is a recently discovered PTM on the natural chemokines interleukin-8 (IL-8; CXCL8) and gamma interferon-inducible protein 10 (IP-10; CXCL10) (8,13,(21)(22)(23)(24). CXCL8, a ligand for CXC chemokine receptors CXCR1 and CXCR2, is known to be the major human neutrophil chemoattractant.…”

“…The biological effect of such proteolytic processing varies depending on the chemokine and protease involved. First, limited NH 2 -terminal proteolytic processing of chemokines by proteases can result in enhanced biological activity, e.g., CXCL8 processing by thrombin (8,9) or matrix metalloproteinase 9 (MMP-9)/gelatinase B (10) and CCL3L1 processing by DPPIV/CD26 (11), or decreased biological activity, e.g., CCL7 processing by MMP-2/gelatinase A (12) and CXCL10 processing by CD26 (13). For some chemokines, like CXCL7 and CCL14, more extensive NH 2 -terminal truncation is even mandatory to obtain receptor signaling and chemotactic properties (14)(15)(16).…”