Granzyme B is dispensable for immunologic tolerance to self in a murine model of systemic lupus erythematosus
✍ Scribed by Kareem L. Graham; Donna L. Thibault; Jonathan B. Steinman; Lance Okeke; Peter N. Kao; Paul J. Utz
- Publisher
- John Wiley and Sons
- Year
- 2005
- Tongue
- English
- Weight
- 246 KB
- Volume
- 52
- Category
- Article
- ISSN
- 0004-3591
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✦ Synopsis
Abstract
Objective
Proteolytic autoantigen cleavage by the serine protease granzyme B has been implicated in the development of systemic autoimmune disease; however, there has been no conclusive demonstration of a pathogenic role for granzyme B in autoimmunity. In this study, we evaluated the role of granzyme B in a murine model of autoimmunity.
Methods
To identify potential novel granzyme B substrates, complementary DNAs encoding nuclear factor 45 (NF45) and NF90 were used to generate ^35^S‐methionine–labeled proteins by coupled in vitro transcription/translation. Radiolabeled proteins were then incubated with purified recombinant granzyme B or caspases, and the cleavage products were analyzed by autoradiography. We also immunized granzyme B–deficient and granzyme B–intact mice with the mineral oil pristane. Production of autoantibodies directed against granzyme B substrates in response to pristane was evaluated by Western blotting, immunoprecipitation, and enzyme‐linked immunosorbent assay.
Results
The double‐stranded RNA–binding protein NF90 was identified as a novel substrate for caspases and granzyme B, both in vitro and in vivo. NF90 is uniquely cleaved by granzyme B in vitro; however, pristane immunization still induced anti‐NF90 antibodies in granzyme B–deficient mice. Pristane‐treated granzyme B–deficient mice also produced antibodies directed against the U1–70‐kd antigen, a previously identified granzyme B substrate. Last, antibodies directed against U1–70 kd arose spontaneously in granzyme B–deficient mice.
Conclusion
These results demonstrate that granzyme B is not required for the production of autoantibodies directed against antigens that are granzyme B substrates in vitro. The data also suggest a protective role for this proapoptotic protease in systemic autoimmunity.