Identification of caspase-3 degradome by two-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization-time of flight analysis.
The activation of caspases is a critical event for the execution phase of programmed cell death. Caspases are highly specific in their ability to activate or inhibit many crucial proteins in the cell via site-specific cleavage. To date, more than 60 proteins have been shown to be substrates of one or more caspases in mammalian cells, and the list is still growing. In this study, to identify human caspase-3 substrates, we digested lysates obtained from a caspase-3-deficient MCF-7 cell line with purified caspase-3 and analyzed eliminated or decreased spots by 2-DE. Proteins degraded by caspase-3, termed as caspase-3 degradome, are involved in a variety of cellular functions, such as stress-responsive proteins, signaling molecules, structural proteins, and unclassified proteins. Interestingly, the cellular level of vinculin, a caspase-3 substrate, was dramatically reduced during the apoptotic process, where the expression level of caspase-3 was increased. This degradomic approach could provide a powerful tool in finding physiological substrates of many proteolytic enzymes whose functions remain to be determined.
Systemic Proteomics Research Center, Korea Research Institute of Bioscience and Biotechnology, Yusung, Taejon, South Korea.
Ah Young
AY
Lee
Byoung Chul
BC
Park
Mi
M
Jang
Sayeon
S
Cho
Do Hee
DH
Lee
Sang Chul
SC
Lee
Pyung Keun
PK
Myung
Sung Goo
SG
Park
D017209:Apoptosis / Q000502:physiology*; D053148:Caspase 3; D020169:Caspases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013379:Substrate Specificity; D016596:Vinculin / Q000378:metabolism
D016596:Vinculin; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases