Mechanism of beta 2-microglobulin-induced apoptosis in the K562 leukemia cell line, defective in major histocompatibility class 1.
BACKGROUND
It has been shown that exogenous beta 2-microglobulin (beta 2m) triggers significant apoptosis in several cell lines, but the molecular mechanism of beta 2m-induced apoptosis remains to be found.


METHODS
To understand the mechanism of beta 2m-induced apoptosis, we added purified human beta 2m to cultures of K562 human chronic myelocytic leukemia cells, detected apoptosis by DNA fragmentation and annexin V binding assays, measured mitochondrial membrane potential (delta psi m), and used Z-VAD-fmk, a general inhibitor of caspases, inhibitors of caspases-1 and-3, as well as Western blot analysis to detect activated caspases.


RESULTS
beta 2m-induced apoptosis was associated with decreased delta psi m K562 cells. Treatment with the general caspase inhibitor Z-VAD-fmk, as well as the caspase-1 inhibitor YVAD-CHO, significantly blocked beta 2m-induced apoptosis. However, Western blot analysis revealed that caspase-1 was intrinsically activated in untreated as well as beta 2m-treated K562 cells. Furthermore, beta 2m-induced apoptosis was not associated with the cleavage of caspase-3 as revealed by Western blot analysis, but was inhibited by an inhibitor of caspase-3, Z-DEVD-CHO, suggesting that not caspase-3, but a caspase-3-like enzyme may be involved in beta 2m-induced apoptosis. Western blot analysis also revealed that caspases-4, -8 and -9 were not activated during beta 2m-induced apoptosis in these cells.


CONCLUSIONS
These results reveal that beta 2m-induced apoptosis in K562 cells occurs by a mechanism dependent on decreased mitochondrial delta psi m. Moreover, while caspase-1 activity may be one of the factors involved in beta 2m-induced apoptosis, activation of this caspase alone does not cause apoptosis, and other proapoptotic factors including activation of a caspase-3-like enzyme, independently of caspases-4, -8 and -9 activity, may be required to trigger apoptosis.
Wu
Ching-Huang
CH
Department of Pharmacology and Toxicology and Indiana University Cancer Center, 1044 West Walnut R4-119, Indianapolis, IN 46202, USA.
Gordon
John D
JD
Zhong
Xiaoling
X
Safa
Ahmad R
AR
D017209:Apoptosis / Q000187:drug effects* / Q000276:immunology / Q000502:physiology; D020170:Caspase 1 / Q000378:metabolism; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects; D003574:Cytochrome c Group / Q000378:metabolism; D006680:HLA Antigens / Q000096:biosynthesis* / Q000276:immunology; D006801:Humans; D007425:Intracellular Membranes / Q000187:drug effects / Q000502:physiology; D020014:K562 Cells / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology / Q000276:immunology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008928:Mitochondria / Q000187:drug effects / Q000502:physiology; D001613:beta 2-Microglobulin / Q000037:antagonists & inhibitors / Q000276:immunology / Q000494:pharmacology* / Q000502:physiology
D061945:Caspase Inhibitors; D003574:Cytochrome c Group; D006680:HLA Antigens; D001613:beta 2-Microglobulin; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D020170:Caspase 1