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Journal of Histochemistry and Cytochemistry, Vol. 50, 961-972, July 2002, Copyright © 2002, The Histochemical Society, Inc.

ARTICLE

c-Jun-like Immunoreactivity in Apoptosis Is the Result of a Crossreaction with Neoantigenic Sites Exposed by Caspase-3-mediated Proteolysis

Joan Riberaa, Victoria Ayalaa, and Josep E. Esquerdaa
a Unitat de Neurobiologia Cellular, Departament de Ciències Mèdiques Bàsiques, Facultat de Medicina, Lleida, Catalunya, Spain

Correspondence to: Josep E. Esquerda, Unitat de Neurobiologia Cellular, Departament de Ciències Mèdiques Bàsiques, Facultat de Medicina, Avd Rovira Roure 44, E25198 Lleida, Catalunya, Spain. E-mail: josep.esquerda@cmb.udl.es

Previous reports in various cells and species have shown that apoptotic cells are specifically and strongly labeled by certain c-Jun/N-terminal antibodies, such as c-Jun/sc45. This kind of immunoreactivity is confined to the cytoplasm. It is not due to c-Jun but appears to be related to c-Jun-like neoepitopes generated during apoptosis. This study was planned to gain further information about c-Jun-like immunostaining during apoptosis and to evaluate these antibodies as possible tools for characterizing cell death. Most of the experiments were performed in chick embryo spinal cord. When the apoptotic c-Jun-like immunoreactivity and caspase-3 immunostaining patterns were compared, we found that both antibodies immunostained the same dying cells in a similar pattern. In contrast to TUNEL staining, which reveals a positive reaction in both apoptotic and necrotic dying cells, active caspase-3 and c-Jun/sc45 antibodies are more selective because they stained only apoptotic cells. When cytosolic extracts from normal tissues were digested in vitro with caspase-3, c-Jun/sc45 immunoreactivity was strongly induced in several proteins, as demonstrated by Western blotting. Similar results were found when normal tissue sections were treated with caspase-3. Our results show that c-Jun/sc45 antibodies react with neoepitopes generated from cell proteins cleaved by activated caspases during apoptosis. We conclude that c-Jun/sc45 antibodies may be useful for detecting apoptosis. They can even be used in archival paraffin-embedded tissue samples. (J Histochem Cytochem 50:961–972, 2002)

Key Words: apoptosis, c-Jun, caspase, immunocytochemistry, cell death, necrosis, chick embryo spinal cord, c-Jun/sc45


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