Direct measurement of cathepsin B activity in the cytosol of apoptotic cells by an activity-based probe.
Cells control their own death through a program termed apoptosis, which is indispensable for development and homeostasis in all metazoans. Lysosomal cysteine proteases are not normally thought of as participating in apoptosis; however, recent reports have shown that the cathepsin proteases can be released from the lysosome during apoptosis, where they can participate in cell death. We report here the development of an activity-based probe that, under optimized conditions, reports on cathepsin B activity only in apoptotic cells by reading out the release of cathepsin B from the lysosomes. Biochemical characterization of apoptosis in cells from cathepsin B null mice shows delayed and suboptimal activation of caspases. Our data further supports a role for cathepsin B in the cytosol as a positive regulator of a cell death feed-forward loop and provides a chemical tool for future investigations.
Pratt
Matthew R
MR
Laboratory of Synthetic Protein Chemistry, The Rockefeller University, New York, NY 10065, USA. matthew.pratt@usc.edu
Sekedat
Matthew D
MD
Chiang
Kyle P
KP
Muir
Tom W
TW
D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D001710:Biotin / Q000737:chemistry / Q000378:metabolism; D002219:Carbamates / Q000737:chemistry / Q000494:pharmacology*; D020169:Caspases / Q000737:chemistry / Q000378:metabolism; D002401:Cathepsin B / Q000032:analysis / Q000378:metabolism*; D045744:Cell Line, Tumor; D003600:Cytosol / Q000378:metabolism; D006801:Humans; D008247:Lysosomes / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D041681:NIH 3T3 Cells; D009842:Oligopeptides / Q000737:chemistry / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
D002219:Carbamates; D009842:Oligopeptides; D011994:Recombinant Proteins; D001710:Biotin; D020169:Caspases; D002401:Cathepsin B