MECHANISM OF UV-INDUCED APOPTOSIS IN HUMAN LEUKEMIA-CELLS - ROLES OF CA2+ MG2+-DEPENDENT ENDONUCLEASE, CASPASE-3, AND STRESS-ACTIVATED PROTEIN-KINASES/

Ultraviolet light (UV) induced rapid apoptosis of U937 leukemia cells, concurrent with DNA fragmentation and cleavage of poly(ADP-ribose)pol ymerase (PARP) by activated caspase-3. The in vitro reconstitution of intact HeLa S3 nuclei and apoptotic U937 cytosolic extract (CE) reveal ed that (i) Ca2+/Mg2+-dependent, Zn2+-sensitive endonuclease activated in the apoptotic CE induced DNA ladder in HeLa nuclei at pH 6.8-7.4, (ii) activated caspase-3 cleaved PARP in HeLa nuclei, and (iii) when t he apoptotic CE was treated with the caspase-3 inhibitor (1 mu M Ac-DE VD-CHO) or the caspase-1 inhibitor (10 mu M Ac-YVAD-CHO), the former, but not the latter, caused a 50% inhibition of DNA fragmentation and t he complete inhibition of PARP cleavage in HeLa nuclei. Similarly, Ac- DEVD-CHO (100 mu M) inhibited apoptosis and DNA ladder by 50% and PARP cleavage completely in UV-irradiated U937 cells, but Ac-YVAD-CHO (100 mu M) did not. Thus, UV-induced apoptosis of U937 cells involves the Ca2+/Mg2+-dependent endonuclease pathway and the caspase-3-PARP cleava ge-Ca2+/Mg2+-dependent endonuclease pathway. The former pathway produc ed directly 50% of apoptotic DNA ladder, and the latter involved activ ated caspase-3 and PARP cleavage, followed by formation of the remaini ng 50% DNA ladder by the activated endonuclease. In UV-irradiated B-ce ll lines, further, p53-dependent increase of Bax resulted in a greater caspase-3 activation compared to its absence. However, UV-induced act ivation of JNK1 and p38 was not affected by the caspase-1 and -3 inhib itors in U937 cells, so that caspases-1 and -3 do not function upstrea m of JNK1 and p38. (C) 1998 Academic Press.