Selective cleavage of nucleolar autoantigen B23 by granzyme B in differentiated vascular smooth muscle cells: Insights into the association of specific autoantibodies with distinct disease phenotypes

Abstract

Objective. To investigate the association of specific autoantibodies with distinct disease phenotypes. The association of autoantibodies to nucleophosmin/ B23 with pulmonary hypertension in scleroderma, and the susceptibility of autoantigens to cleavage by granzyme B (GB), provided a focus for these studies.Methods. Intact cells were subjected to cytotoxic lymphocyte granule-induced death, and the susceptibility of autoantigens to cleavage by GB was addressed by immunoblotting and/or by a novel immunofluorescence assay.Results. B23 was cleaved efficiently by GB in vitro, but was highly resistant to cleavage by GB during cytotoxic lymphocyte granule-mediated death of many intact cell types. In contrast, this molecule was highly susceptible to GB-mediated proteolysis exclusively in differentiated vascular smooth muscle cells. Topoisomerase I and several other GB substrates did not show this striking change in cleavage susceptibility in different cell types.Conclusion. These data demonstrate that the cleavage of B23 by GB in intact cells is dependent upon both cell type and phenotype. The susceptibility of this autoantigen (which is associated with a distinct pulmonary vascular phenotype in scleroderma) to GBmediated proteolysis selectively in vascular smooth muscle cells suggests that the GB-cleavable conformation of autoantigens may occur selectively in the target tissue, and may play a role in shaping the phenotypespecific autoimmune response.

Antiangiogenic plasma activity in patients with systemic sclerosis

“…Among those reported are various MMPs (17)(18)(19) and plasmin reductase (20,21). Granzyme B, a serine protease found in T cell granules, is strikingly associated with cleavage of autoantigens targeted by autoantibodies in patients with SSc (33,56). In agreement with this hypothesis, we demonstrated that both granzyme B and the full granule content could indeed cleave plasminogen into varioussized fragments containing angiostatin K [1][2][3] .…”

“…Although several proteases can potentially cleave plasminogen in a manner that would lead to angiostatin production, granzyme B is a particularly interesting candidate given its high levels in the peripheral blood cells of some patients with SSc (32,33). To determine whether granzyme B can cleave plasminogen, SSc and control plasma samples were allowed to interact with granzyme B or the entire granule content (the source of granzyme B) in vitro ( Figure 2C).…”

Granzyme B-induced and Caspase 3-dependent Cleavage of Gelsolin by Mouse Cytotoxic T Cells Modifies Cytoskeleton Dynamics

“…We cannot exclude the possibility that gzmB directly processes c-gelsolin in specific subtypes of cells. For example, the B23 autoantigen is most susceptible to cleavage by gzmB in differentiated smooth muscle cells (42).…”

Molecular pathogenesis of pulmonary arterial hypertension

“…Autoantibodies against B23, a cleavage product of a nuclear protein produced by the T cell enzyme granzyme B, can distinguish the subset of patients with scleroderma who also have PAH from patients with scleroderma without PAH (111). The mechanistic significance of this biomarker is not known.…”