Chymotrypsin B cached in rat liver lysosomes and involved in apoptotic regulation through a mitochondrial pathway.
Lysosomes can trigger the mitochondrial apoptotic pathway by releasing proteases. Here we report that a 25-kDa protein purified from rat liver lysosomes possesses a long standing potent Bid cleavage activity at neutral pH, and the truncated Bid can in turn induce rapid mitochondrial release of cytochrome c. This protease was revealed as chymotrypsin B by biochemical and mass spectrometric analysis. Although it was long recognized as a digestive protease exclusively secreted by the exocrine pancreas, our data support that it also expresses and intracellularly resides in rat liver lysosomes. Translocation of lysosomal chymotrypsin B into cytosol was triggered by apoptotic stimuli such as tumor necrosis factor-alpha, and intracellular delivery of chymotrypsin B protein induced apoptotic cell death with a potency comparable with cathepsin B, suggestive of a lysosomal-mitochondrial pathway to apoptosis regulated by chymotrypsin B following its release. Noteworthily, either knockdown of chymotrypsin B expression by RNA interference or pretreatment with chymotrypsin B inhibitor N-p-tosyl-L-phenylalanine chloromethyl ketone significantly reduced tumor necrosis factor-alpha-induce apoptosis. These results demonstrate for the first time that chymotrypsin B is not only restricted to the pancreas but can function intracellularly as a pro-apoptotic protease.
Miao
Qi
Q
National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Chaoyang District, Beijing, China.
Sun
Yang
Y
Wei
Taotao
T
Zhao
Xingyu
X
Zhao
Kai
K
Yan
Ling
L
Zhang
Xujia
X
Shu
Hongjun
H
Yang
Fuyu
F
D000818:Animals; D017209:Apoptosis; D002460:Cell Line; D002918:Chymotrypsin / Q000235:genetics / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D022781:Hepatocytes / Q000201:enzymology; D008099:Liver / Q000201:enzymology*; D008247:Lysosomes / Q000201:enzymology*; D008928:Mitochondria / Q000201:enzymology*; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction
D012333:RNA, Messenger; D002918:Chymotrypsin; C117609:chymotrypsin B