Bcl-2 binds to and inhibits ryanodine receptors.
The anti-apoptotic B-cell lymphoma-2 (Bcl-2) protein not only counteracts apoptosis at the mitochondria by scaffolding pro-apoptotic Bcl-2-family members, but also acts at the endoplasmic reticulum, thereby controlling intracellular Ca(2+) dynamics. Bcl-2 inhibits Ca(2+) release by targeting the inositol 1,4,5-trisphosphate receptor (IP3R). Sequence analysis has revealed that the Bcl-2-binding site on the IP3R displays strong similarity with a conserved sequence present in all three ryanodine receptor (RyR) isoforms. We now report that Bcl-2 co-immunoprecipitated with RyRs in ectopic expression systems and in native rat hippocampi, indicating that endogenous RyR-Bcl-2 complexes exist. Purified RyR domains containing the putative Bcl-2-binding site bound full-length Bcl-2 in pulldown experiments and interacted with the BH4 domain of Bcl-2 in surface plasmon resonance (SPR) experiments, suggesting a direct interaction. Exogenous expression of full-length Bcl-2 or electroporation loading of the BH4 domain of Bcl-2 dampened RyR-mediated Ca(2+) release in HEK293 cell models. Finally, introducing the BH4-domain peptide into hippocampal neurons through a patch pipette decreased RyR-mediated Ca(2+) release. In conclusion, this study identifies Bcl-2 as a new inhibitor of RyR-based intracellular Ca(2+)-release channels.
KU Leuven, Laboratory of Molecular and Cellular Signaling, Department of Cellular and Molecular Medicine, B-3000 Leuven, Belgium.
Tim
T
Vervliet
University of Ghent, Physiology Group, Department Basic Medical Sciences, B-9000 Ghent, Belgium.
Elke
E
Decrock
Institut de Neurobiologie Alfred Fessard, CNRS, Laboratoire de Neurobiologie et Développement, 91198 Gif sur Yvette cedex, France.
Jordi
J
Molgó
University of Siena, Molecular Medicine Section, Department of Molecular and Developmental Medicine, and Interuniversitary Institute of Myology, 53100 Siena, Italy.
Vincenzo
V
Sorrentino
KU Leuven, Laboratory of Molecular and Cellular Signaling, Department of Cellular and Molecular Medicine, B-3000 Leuven, Belgium.
Ludwig
L
Missiaen
University of Ghent, Physiology Group, Department Basic Medical Sciences, B-9000 Ghent, Belgium.
Luc
L
Leybaert
KU Leuven, Laboratory of Molecular and Cellular Signaling, Department of Cellular and Molecular Medicine, B-3000 Leuven, Belgium.
Humbert
H
De Smedt
Radboud University Medical Center, Donders Institute for Brain, Cognition and Behaviour, Department of Cognitive Neuroscience, 6500HB Nijmegen, The Netherlands.
Nael Nadif
NN
Kasri
KU Leuven, Laboratory of Molecular and Cellular Signaling, Department of Cellular and Molecular Medicine, B-3000 Leuven, Belgium.
Jan B
JB
Parys
KU Leuven, Laboratory of Molecular and Cellular Signaling, Department of Cellular and Molecular Medicine, B-3000 Leuven, Belgium geert.bultynck@med.kuleuven.be.
Geert
G
Bultynck
D000595:Amino Acid Sequence; D000818:Animals; D015220:Calcium Channels / Q000378:metabolism; D020013:Calcium Signaling; D057809:HEK293 Cells; D006624:Hippocampus / Q000166:cytology; D006801:Humans; D051379:Mice; D008907:Mink; D008969:Molecular Sequence Data; D009474:Neurons / Q000378:metabolism; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism*; D011817:Rabbits; D051381:Rats; D019837:Ryanodine Receptor Calcium Release Channel / Q000737:chemistry / Q000378:metabolism*
D015220:Calcium Channels; D019253:Proto-Oncogene Proteins c-bcl-2; D019837:Ryanodine Receptor Calcium Release Channel
Bcl-2; Ca2+ signaling; Hippocampus; Ryanodine receptor