Related Experiment Video
Updated: Aug 7, 2026
Flow Cytometric Analysis of Apoptotic Biomarkers in Actinomycin D-Treated SiHa Cervical Cancer Cells
Published on: August 26, 2021
D4-GDI is cleaved by caspase-3 during daunorubicin-induced apoptosis in HL-60 cells
Kang-Beom Kwon1, Eun-Kyung Park, Do-Gon Ryu
1Department of Physiology, School of Oriental Medicine, Won-Kwang University, Chonbuk, Korea.
Abstract:
Daunorubicin, an anti-cancer drug, is known to induce apoptosis in HL-60 cells in a dose-dependent manner through the activation of caspase-3 (CPP32). Caspase-3 selective inhibitor, Ac-DEVD-CHO, prevented both the activation of caspase-3 and cleavage of poly(ADP-ribose) polymerase (PARP). D4-GDI is a GDP dissociation inhibitor for the Ras-related Rho family GTPase in hematopoietic cells. Here we report that D4-GDI is a substrate for the caspase-3. D4-GDI was cleaved to a 23 kDa fragment by daunorubicin treatment in HL-60 cells with kinetics that parallel the onset of apoptosis. D4-GDI cleavage as well as DNA fragmentation was inhibited by treatment with Ac-DEVD-CHO but not with Ac-YVAD-CHO, a caspase-1 inhibitor. These data suggest that D4-GDI of Rho family GTPase may be regulated during apoptosis through the caspase-3 mediated cleavage of the GDI protein.
Insights
Daunorubicin induces apoptosis by activating caspase-3, which cleaves D4-GDI, a Rho family GTPase regulator. This cleavage is a key event in the apoptotic process in HL-60 cells.
Area of Science:
- Molecular Biology
- Cell Biology
- Biochemistry
Background:
- Daunorubicin is an anti-cancer drug that triggers apoptosis in HL-60 cells.
- Apoptosis involves the activation of caspase-3 (CPP32) and cleavage of substrates like poly(ADP-ribose) polymerase (PARP).
- D4-GDI is a GDP dissociation inhibitor for Ras-related Rho family GTPases in hematopoietic cells.
Purpose of the Study:
- To investigate whether D4-GDI is a substrate for caspase-3.
- To determine the role of D4-GDI cleavage in daunorubicin-induced apoptosis.
Main Methods:
- HL-60 cells were treated with daunorubicin.
- Caspase-3 activity was assessed using a selective inhibitor (Ac-DEVD-CHO).
- Cleavage of D4-GDI and PARP was analyzed via Western blotting.
- DNA fragmentation was measured to confirm apoptosis.
Main Results:
- Daunorubicin treatment led to the cleavage of D4-GDI into a 23 kDa fragment in HL-60 cells, paralleling apoptosis onset.
- The caspase-3 inhibitor Ac-DEVD-CHO blocked D4-GDI cleavage and DNA fragmentation.
- A caspase-1 inhibitor (Ac-YVAD-CHO) did not affect D4-GDI cleavage.
Conclusions:
- D4-GDI is a novel substrate for caspase-3.
- Caspase-3 mediated cleavage of D4-GDI is involved in the regulation of Rho family GTPases during daunorubicin-induced apoptosis.
- This cleavage event contributes to the apoptotic pathway in hematopoietic cells.
