Apoptosis induction in human leukemic cells by a novel protein Bengalin, isolated from Indian black scorpion venom: through mitochondrial pathway and inhibition of heat shock proteins.
Scorpion venom possesses protein toxins having numerous biological activities, some of which are potentially anticancerous. Previously we had reported antiproliferative activity of the venom of Indian black scorpion, Heterometrus bengalensis Koch. Here we have isolated and purified a novel protein named Bengalin (72kDa) from the venom, responsible for antiproliferative and apoptogenic activities against human leukemic cells U937 (histiocytic lymphoma) and K562 (chronic myelogenous leukemia). N-terminal sequence of first 20 amino acids of Bengalin was G-P-L-T-I-L-H-I-N-D-V-H-A-A/R-F-E-Q/G-F/G-N-T. Bengalin induced cell growth inhibition at IC(50) values of 3.7 and 4.1 microg/ml for U937 and K562 cells respectively did not significantly affect normal human lymphocytes. Inhibition of U937 and K562 cell proliferation occurred by apoptosis as evidenced from damaged nuclei, cell cycle arrest at sub G1 phase, increase of early apoptotic cells, augmentation of DNA fragmentation and also a reduction of telomerase activity. Further insights revealed that Bax:Bcl2 ratio was elevated after Bengalin treatment. Moreover Bengalin elicited loss of mitochondrial membrane potential (MMP) which commenced cytochrome c release in cytosol, decreased heat shock protein (HSP) 70 and 90 expression, activated caspase-9, caspase-3 and induced poly(ADP-ribose) polymerase (PARP) cleavage. We have also determined that HSP70 and 90 inhibitions correlated with Bengalin induced antiproliferation, caspase-3 upregulation, apoptogenesis and increased DNA fragmentation. These results hypothesize that Bengalin might provide a putative molecular mechanism for their anticancer effect on human leukemic cells which might be mediated by mitochondrial death cascade. Inhibition of HSPs might also play a crucial role in induction of apoptosis.
Gupta
Shubho Das
SD
Drug Development Division, Indian Institute of Chemical Biology, 4, Raja S.C. Mullick Road, Kolkata-700032, India.
Gomes
Antony
A
Debnath
Anindita
A
Saha
Archita
A
Gomes
Aparna
A
D000595:Amino Acid Sequence; D000818:Animals; D000970:Antineoplastic Agents / Q000302:isolation & purification / Q000633:toxicity*; D017209:Apoptosis; D053148:Caspase 3 / Q000378:metabolism; D053453:Caspase 9 / Q000378:metabolism; D045304:Cytochromes c / Q000378:metabolism; D016193:G1 Phase; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D006801:Humans; D007194:India; D020014:K562 Cells; D007938:Leukemia / Q000188:drug therapy; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D008969:Molecular Sequence Data; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D012604:Scorpion Venoms / Q000737:chemistry*; D020298:U937 Cells; D051028:bcl-2-Associated X Protein / Q000378:metabolism
D000970:Antineoplastic Agents; D018840:HSP70 Heat-Shock Proteins; D018841:HSP90 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012604:Scorpion Venoms; D051028:bcl-2-Associated X Protein; D045304:Cytochromes c; D011065:Poly(ADP-ribose) Polymerases; D053148:Caspase 3; D053453:Caspase 9