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Actin cleavage by CPP-32/apopain during the development of apoptosis

T Mashima1, M Naito, K Noguchi

  • 1Laboratory of Biomedical Research, Institute of Molecular and Cellular Biosciences, University of Tokyo, Japan.

Oncogene
|March 6, 1997
PubMed

Insights

The study identifies activated CPP-32/apopain as the protease that cleaves actin during apoptosis. This actin cleavage, inhibited by Z-EVD-CH2-DCB, is crucial for programmed cell death.

Area of Science:

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Background:

  • Interleukin-1beta-converting enzyme (ICE)/ced-3 family proteases are critical for apoptosis.
  • The specific cellular substrates targeted by these proteases during apoptosis remain largely unidentified.
  • Previous work indicated a distinct ICE family protease cleaves actin during VP-16-induced apoptosis.

Purpose of the Study:

  • To identify the specific ICE family protease responsible for actin cleavage during apoptosis.
  • To investigate the role of actin cleavage in the mechanism of apoptosis.
  • To determine if inhibiting this protease affects cellular apoptosis.

Main Methods:

  • In vitro cleavage assays using apoptotic cell extracts and purified actin.
  • Western blot analysis using antibodies against specific actin fragments.
  • In vivo studies using VP-16-treated U937 cells and a selective CPP-32(-like) protease inhibitor (Z-EVD-CH2-DCB).

Main Results:

  • Activated CPP-32/apopain was identified as the protease cleaving actin in vitro, generating 15 kDa and 31 kDa fragments.
  • In vivo cleavage of actin during apoptosis was confirmed using an antibody specific to the N-terminus of the 15 kDa fragment.
  • Inhibition of CPP-32(-like) protease activity with Z-EVD-CH2-DCB effectively blocked actin cleavage and VP-16-induced apoptosis in U937 cells.

Conclusions:

  • Actin serves as a direct substrate for CPP-32/apopain(-like) protease in both in vitro and in vivo settings.
  • The cleavage of actin by CPP-32/apopain is a significant event during apoptosis.
  • These findings suggest a role for actin in the regulation of cell growth and programmed cell death.

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