Mcl-1 is required for Akata6 B-lymphoma cell survival and is converted to a cell death molecule by efficient caspase-mediated cleavage

Abstract

Enforced expression of the antiapoptotic Bcl-2 family protein Mcl-1 promotes lymphomagenesis in the mouse; however, the functional role of Mcl-1 in human B-cell lymphoma remains unclear. We demonstrate that Mcl-1 is widely expressed in malignant B-cells, and high-level expression of Mcl-1 is required for B-lymphoma cell survival, since transfection of Mcl-1-specific antisense oligodeoxynucleotides was sufficient to promote apoptosis in Akata6 lymphoma cells. Mcl-1 was efficiently cleaved by caspases at evolutionarily conserved aspartic acid residues in vitro, and during cisplatin-induced apoptosis in B-lymphoma cell lines and spontaneous apoptosis of primary malignant B-cells. Overexpression of the Mcl-1 cleavage product that accumulated during apoptosis was sufficient to kill cells. Therefore, Mcl-1 is an essential survival molecule for B-lymphoma cells and is cleaved by caspases to a death-promoting molecule during apoptosis. In contrast to Mcl-1, Bcl-2 and Bcl-X L were relatively resistant to caspase cleavage in vitro and in intact cells. Interfering with Mcl-1 function appears to be an effective means of inducing apoptosis in Mcl-1-positive B-cell lymphoma, and the unique sensitivity of Mcl-1 to caspasemediated cleavage suggests an attractive strategy for converting it to a proapoptotic molecule.

Unraveling MCL-1 degradation

“…8 In dying cells, MCL-1 can be eliminated by caspase-mediated cleavage generating a potent proapoptotic protein that can enhance the cell death response. 9 These findings demonstrate that MCL-1 is tightly regulated at several levels, but it remains to be established what the physiological regulation of MCL-1 is during normal development and homeostasis.…”

Effective elimination of fludarabine-resistant CLL cells by PEITC through a redox-mediated mechanism

“…Because a high level of MCL1 was shown to mediate resistance to fludarabine in vivo and in vitro, 28,29 abrogation of MCL1 by PEITC likely facilitated the killing of F-ara-A-refractory CLL cells. As MCL1 may be a better substrate for caspase than BCL2, 30 we used the pan-caspase inhibitor Z-VAD-fmk to test if it could prevent PEITC-induced MCL1 degradation, and showed that the 20 M Z-VAD-fmk largely suppressed MCL1 degradation in PEITC-treated cells ( Figure 6C). Similar effect was observed by using the specific inhibitor to caspase-3, Z-DEVD (data not shown), suggesting that caspase-3 might be a major protease that cleaved MCL1.…”

Sequence and partial functional analysis of canine Bcl-2 family proteins

“…Several members of the Bcl-2 protein family display protease cleavage sites, and their cleavage products have been shown in general to enhance apoptotic activity (Cheng et al, 1997;Clem et al, 1998;Fujita et al, 1998;Wood et al, 1998;Condorelli et al, 2001;Michels et al, 2004) and in some cases to mediate apoptotic activity (Ofengeim et al, 2012). Sequence comparison between human, canine and murine Bcl-2 family members has revealed a number of fully conserved cleavage sites.…”

“…Figure 1 shows the comparative tetramer sequences preceding cleavage sites reported in the literature. Sequences 100% conserved in all three species include both caspase-3 cleavage sites of Bcl-xL, the major caspase-3 cleavage sites of Mcl-1 and of Bad, and the calpain recognition sequence of Bax (Cheng et al, 1997;Clem et al, 1998;Fujita et al, 1998;Wood et al, 1998;Condorelli et al, 2001;Michels et al, 2004).…”