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Fusing DEDD with ubiquitin changes its intracellular localization and apoptotic potential
J C Lee1, G X Wang, O Schickling
1The Ben May Institute for Cancer Research, University of Chicago, 924 E. 57th Street, Chicago, IL 60637, USA.
Apoptosis : an International Journal on Programmed Cell Death
|October 20, 2005
Summary
Monoubiquitination of the death effector domain-containing protein (DEDD) regulates its cytoplasmic localization and enhances its apoptosis-inducing potential. Cellular inhibitor of apoptosis proteins (cIAP-1/2) can modify DEDD ubiquitination status.
Area of Science:
- Cell Biology
- Molecular Biology
- Apoptosis Research
Background:
- The death effector domain-containing protein (DEDD) is a conserved protein involved in apoptosis.
- DEDD exists in non-, mono-, and diubiquitinated forms, with distinct cellular localizations.
- Previous work indicated unmodified DEDD in nucleoli and ubiquitinated forms in the cytosol associated with caspase-3.
Purpose of the Study:
- To investigate the role of DEDD ubiquitination in regulating its cellular localization and apoptosis-regulating functions.
- To identify the mechanisms and regulatory factors involved in DEDD ubiquitination.
- To explore the interaction between DEDD and cellular inhibitor of apoptosis proteins (cIAPs).
Main Methods:
- Analysis of DEDD ubiquitination sites, focusing on lysine residues.
- Functional studies using DEDD mutants and ubiquitin-fused DEDD constructs.
- Co-transfection experiments with DEDD and cIAP-1/2 to assess ubiquitination and localization changes.
Main Results:
- Multiple lysine residues serve as alternative ubiquitination sites, maintaining DEDD monoubiquitination.
- A central region (amino acids 109-305) is crucial for DEDD ubiquitination.
- Mimicking monoubiquitination by C-terminal ubiquitin fusion relocated DEDD to the cytosol and increased its apoptosis potential.
- Cellular inhibitor of apoptosis proteins 1 and 2 (cIAP-1/2) mediate both mono- and polyubiquitination of DEDD.
- Co-expression of DEDD with cIAP-1/2 caused relocalization of cIAPs to the nucleoli.
Conclusions:
- Monoubiquitination is a key regulatory mechanism for DEDD, controlling its cytoplasmic localization and proapoptotic activity.
- The ubiquitination machinery interacts with a central region of DEDD.
- cIAP-1/2 proteins are E3 ubiquitin ligases that can modify DEDD ubiquitination and influence its subcellular distribution, suggesting a role in apoptosis regulation.