Shirley R. Kronheim

Viral inhibition of inflammation: Cowpox virus encodes an inhibitor of the interleukin-1β converting enzyme

Cowpox virus effectively inhibits inflammatory responses against viral infection in the chick embryo. This study demonstrates that one of the viral genes necessary for this inhibition, the crmA gene (a cytokine response modifier gene), encodes a serpin that is a specific inhibitor of the interleukin-1 beta converting enzyme. This serpin can prevent the proteolytic activation of interleukin-1 beta, thereby suppressing an interleukin-1 beta response to infection. However, the modification of this single cytokine response is not sufficient to inhibit inflammatory responses. This suggests that cowpox virus encodes several cytokine response modifiers that act together to inhibit the release of pro-inflammatory cytokines in response to infection. These viral countermeasures to host defenses against infection may contribute significantly to the pathology associated with poxvirus infections.

Activation of interleukin‐ 1β by a co‐induced protease

The proteolytic generation of mature interleukin‐ 1β (IL‐1β) from its inactive precursor does not proceed by a conventional pathway for hormonal processing. Pro‐IL‐ 1β is found dispersed in the cytoplasm, and there are no basic amino acid residues or other commonly recognized processing sites adjoining the mature N‐terminus. Processing appears to occur during release of the hormone. In the present study, we have identified a specific protease that generates mature IL‐ 1β from the precursor. This enzyme is co‐induced with the hormone, and it differs in its cleavage specificity and inhibitor sensitivity from all known proteases.

Purification of interleukin-1β converting enzyme, the protease that cleaves the interleukin-1β precursor

We have purified the IL-1β converting enzyme from the THP-1 cell line using standard chromatographic techniques and obtained the N-terminal amino acid sequence of this novel protein. After stimulation of THP-1 cells with lipopolysaccharide, hydroxyurea, and silica, the protease was solubilized by multiple freeze/thawing. The protein was purified by ion-exchange chromatography, affinity chromatography on blue agarose, gel filtration, and chromatofocusing. The molecular weight of the protein is approximately 22,000 Da and the pI is between 7.1 and 6.8. The overall yield for this procedure was 16% of the activity found in the initial cell lysates. An antiserum raised against a peptide based on the N-terminus was used to precipitate the protease, confirming our identification of the 22,000-Da protein as the IL-1β converting enzyme.