title: Apoptosis induction-related cytosolic calcium responses revealed by the dual FRET imaging of calcium signals and caspase-3 activation in a single cell.
abstract: Stimulus-induced changes in the intracellular Ca(2+) concentration control cell fate decision, including apoptosis. However, the precise patterns of the cytosolic Ca(2+) signals that are associated with apoptotic induction remain unknown. We have developed a novel genetically encoded sensor of activated caspase-3 that can be applied in combination with a genetically encoded sensor of the Ca(2+) concentration and have established a dual imaging system that enables the imaging of both cytosolic Ca(2+) signals and caspase-3 activation, which is an indicator of apoptosis, in the same cell. Using this system, we identified differences in the cytosolic Ca(2+) signals of apoptotic and surviving DT40 B lymphocytes after B cell receptor (BCR) stimulation. In surviving cells, BCR stimulation evoked larger initial Ca(2+) spikes followed by a larger sustained elevation of the Ca(2+) concentration than those in apoptotic cells; BCR stimulation also resulted in repetitive transient Ca(2+) spikes, which were mediated by the influx of Ca(2+) from the extracellular space. Our results indicate that the observation of both Ca(2+) signals and cells fate in same cell is crucial to gain an accurate understanding of the function of intracellular Ca(2+) signals in apoptotic induction.
authors.affiliation: Laboratory for Developmental Neurobiology, RIKEN Brain Science Institute, Wako, Saitama 351-0198, Japan; Division of Neuronal Network, Department of Basic Medical Sciences, Institute of Medical Science, The University of Tokyo, Minato-ku, Tokyo 108-8639, Japan.
authors.forename: Akitoshi
authors.initials: A
authors.lastname: Miyamoto
authors.affiliation: Saitama Medical University Hospital, Iruma, Saitama 350-0495, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Miyauchi
authors.affiliation: Laboratory for Cell Function Dynamics, RIKEN Brain Science Institute, Wako, Saitama 351-0198, Japan.
authors.forename: Takako
authors.initials: T
authors.lastname: Kogure
authors.affiliation: Laboratory for Cell Function Dynamics, RIKEN Brain Science Institute, Wako, Saitama 351-0198, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Miyawaki
authors.affiliation: Laboratory for Developmental Neurobiology, RIKEN Brain Science Institute, Wako, Saitama 351-0198, Japan; Brain Science Institute, Saitama University, 255 Shimo-Okubo, Sakura-ku, Saitama 338-8570, Japan; Laboratory for Behavioral Genetics, RIKEN Brain Science Institute, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan. Electronic address: t-michikawa@brain.riken.jp.
authors.forename: Takayuki
authors.initials: T
authors.lastname: Michikawa
authors.affiliation: Laboratory for Developmental Neurobiology, RIKEN Brain Science Institute, Wako, Saitama 351-0198, Japan. Electronic address: mikosiba@brain.riken.jp.
authors.forename: Katsuhiko
authors.initials: K
authors.lastname: Mikoshiba
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000502:physiology*; D053148:Caspase 3 / Q000378:metabolism*; D002460:Cell Line; D002645:Chickens; D003593:Cytoplasm / Q000378:metabolism; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D036641:Microscopy, Fluorescence, Multiphoton / Q000379:methods*
chemical_list: D051017:Apoptosis Regulatory Proteins; D053148:Caspase 3; D002118:Calcium
keywords: Apoptosis; Biosensors; Calcium imaging; Caspase; FRET