A specific isozyme of 2'-5' oligoadenylate synthetase is a dual function proapoptotic protein of the Bcl-2 family.
2-5(A) synthetases are a family of interferon-induced enzymes that polymerize ATP into 2'-5' linked oligoadenylates that activate RNase L and cause mRNA degradation. Because they all can synthesize 2-5(A), the reason for the existence of so many synthetase isozymes is unclear. Here we report that the 9-2 isozyme of 2-5(A) synthetase has an additional activity: it promotes apoptosis in mammalian cells. The proapoptotic activity of 9-2 was isozyme-specific and enzyme activity-independent. The 9-2-expressing cells exhibited many properties of cells undergoing apoptosis, such as DNA fragmentation, caspase activation, and poly ADP-ribose polymerase and lamin B cleavage. The isozyme-specific carboxyl-terminal tail of the 9-2 protein was shown, by molecular modeling, to contain a Bcl-2 homology 3 (BH3) domain, suggesting that it may be able to interact with members of the Bcl-2 family that contain BH1 and BH2 domains. Co-immunoprecipitate assays and confocal microscopy showed that 9-2 can indeed interact with the anti-apoptotic proteins Bcl-2 and Bclx(L) in vivo and in vitro. Mutations in the BH3 domain that eliminated the 9-2-Bcl-2 amd 9-2-Bclx(L) interactions also eliminated the apoptotic activity of 9-2. Thus, we have identified an interferon-induced dual function protein of the Bcl-2 family that can synthesize 2-5(A) and promote cellular apoptosis independently. Moreover, the cellular abundance of this protein is regulated by alternative splicing; the other isozymes encoded by the same gene are not proapoptotic.
Ghosh
A
A
Department of Molecular Biology, The Lerner Research Institute, The Cleveland Clinic Foundation, 9500 Euclid Ave., Cleveland, OH 44195, USA.
Sarkar
S N
SN
Rowe
T M
TM
Sen
G C
GC
D015088:2',5'-Oligoadenylate Synthetase / Q000235:genetics / Q000502:physiology*; D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D017209:Apoptosis; D002478:Cells, Cultured; D005456:Fluorescent Dyes; D006801:Humans; D020287:In Situ Nick-End Labeling; D007211:Indoles; D007527:Isoenzymes / Q000502:physiology*; D051379:Mice; D008856:Microscopy, Fluorescence; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000502:physiology*; D014162:Transfection
D000911:Antibodies, Monoclonal; D005456:Fluorescent Dyes; D007211:Indoles; D007527:Isoenzymes; D019253:Proto-Oncogene Proteins c-bcl-2; C007293:DAPI; D015088:2',5'-Oligoadenylate Synthetase