Mining of caspase-7 substrates using a degradomic approach.
Caspases play critical roles in the execution of apoptosis. Caspase-3 and caspase-7 are closely related in sequence as well as in substrate specificity. The two caspases have overlapping substrate specificities with special preference for the DEVD motif. However, they are targeted to different subcellular locations during apoptosis, implying the existence of substrates specific for one or other caspase. To identify new caspase-7 substrates, we digested cell lysates obtained from the caspase-3-deficient MCF-7 cell line with purified recombinant caspase-7, and analyzed spots that disappeared or decreased by 2-DE (we refer to this as the caspase-7 degradome). Several proteins with various cellular functions underwent caspase-7- dependent proteolysis. The substrates of capase-7 identified by the degradomic approach were rather different from those of caspase-3 (Proteomics, 4, 3429-3435, 2004). Among the candidate substrates, we confirmed that Valosin-containing protein (VCP) was cleaved by both capspase-7 and caspase-3 in vitro and during apoptosis. Cleavage occurred at both DELD(307) and DELD(580). The degradomic study yielded several candidate caspase-7 substrates and their further analysis should provide valuables clues to the functions of caspase-7 during apoptosis.
Jang
Mi
M
Translational Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon 305-806, Korea.
Park
Byoung Chul
BC
Kang
Sunghyun
S
Lee
Do Hee
DH
Cho
Sayeon
S
Lee
Sang Chul
SC
Bae
Kwang-Hee
KH
Park
Sung Goo
SG
D000251:Adenosine Triphosphatases / Q000378:metabolism; D017209:Apoptosis; D053148:Caspase 3 / Q000378:metabolism; D053179:Caspase 7 / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000378:metabolism; D045744:Cell Line, Tumor; D015180:Electrophoresis, Gel, Two-Dimensional; D018922:HL-60 Cells; D006801:Humans; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013379:Substrate Specificity; D000074405:Valosin Containing Protein
D018797:Cell Cycle Proteins; D053148:Caspase 3; D053179:Caspase 7; D000251:Adenosine Triphosphatases; C000614178:VCP protein, human; D000074405:Valosin Containing Protein