Transient expression of the Bcl-2 family member, A1-a, results in nuclear localization and resistance to staurosporine-induced apoptosis

• Published in: Cell death and differentiation Vol. 8; no. 8; pp. 785 - 793
• Main Authors: Somogyi, R D, Wu, Y, Orlofsky, A, Prystowsky, M B
• Format: Journal Article
• Language: English
• Published: England Nature Publishing Group 01.08.2001
• Subjects: Animals; Apoptosis; Apoptosis - drug effects; Apoptosis - physiology; Binding Sites - physiology; Cell Compartmentation - drug effects; Cell Compartmentation - physiology; Cell death; Cell Nucleus - drug effects; Cell Nucleus - metabolism; COS Cells; Cytochrome; Enzyme Inhibitors - pharmacology; Fluorescent Antibody Technique; Gene Expression Regulation - physiology; Genetic Vectors - physiology; Localization; Microscopy; Monoclonal antibodies; Polymers - metabolism; Proteins; Proto-Oncogene Proteins c-bcl-2 - drug effects; Proto-Oncogene Proteins c-bcl-2 - metabolism; Staurosporine - pharmacology; Subcellular Fractions - drug effects; Subcellular Fractions - metabolism; United States > US
• ISSN: 1350-9047; 1476-5403
• DOI: 10.1038/sj.cdd.4400879

The Bcl-2 family of proteins has been characterized by either anti-apoptotic or pro-apoptotic activity. Insight into how Bcl-2 family members function has been gained by determining their intracellular localization. We have generated a monoclonal anti-A1-a antibody and used a COS-7 overexpression system to study the localization of the murine anti-apoptotic Bcl-2 family member, A1-a. A1-a overexpressed in COS-7 cells localized to the nucleus as determined by subcellular fractionation and immunofluorescent microscopy. A1-a in the COS-7 nucleus bound tightly to the nuclear matrix as evidenced by resistance to treatment with DNAse I and RNAse A and sequential extraction with 1.0% Triton X-100, 0.15 M NaCl, 0.25 M HCl, 0.5 M Tris pH 7.4 and 6 M urea. HPLC analysis of A1-a, subsequent to SDS extraction, produced fractions that gave multiple bands when analyzed by Western blot analysis suggesting a propensity to form multimers. COS-7 cells transfected with A1-a were protected from apoptotic induction by staurosporine treatment.