N-terminal Truncation of Antiapoptotic MCL1, but Not G2/M-induced Phosphorylation, Is Associated with Stabilization and Abundant Expression in Tumor Cells

Abstract

The antiapoptotic BCL2 family member MCL1 is normally upand down-modulated in response to environmental signals and conditions, but is constitutively expressed in cancer where it promotes cell survival and drug resistance. A post-translational modification identified here, truncation at the N terminus, was found to act along with previously described ERK-and GSK3-induced phosphorylation events to regulate the turnover of the MCL1 protein and thus its availability for antiapoptotic effects. Although both N-terminally truncated and full-length MCL1 contain sequences enriched in proline, glutamic acid, serine, and threonine and were susceptible to proteasomal degradation, the truncated form decayed less rapidly and was maintained for an extended period in the presence of ERK activation. This was associated with extended cell survival because the truncated form of MCL1 (unlike those of BCL2 and BCLX) retained antiapoptotic activity. N-terminal truncation slightly increased the electrophoretic mobility of MCL1 and differed from the phosphorylation/band shift to decreased mobility, which occurs in the G 2 /M phase and was not found to affect MCL1 turnover. The N-terminally truncated form of MCL1 was expressed to varying extents in normal lymphoid tissues and was the predominant form present in lymphomas from transgenic mice and human tumor lines of B-lymphoid origin. The degradation versus stabilized expression of antiapoptotic MCL1 is thus controlled by N-terminal truncation as well as by ERK-and GSK3 (but not G 2 /M)-induced phosphorylation. These modifications may contribute to dysregulated MCL1 expression in cancer and represent targets for promoting its degradation to enhance tumor cell death.The antiapoptotic mediator MCL1 was discovered based on rapid, transient up-regulation in ML-1 human myeloblastic leukemia cells initiating differentiation in the presence of 12-Otetradecanoylphorbol 13-acetate (TPA) 7 (1). MCL1 has since been found to be induced in cells at various stages in differentiation, in response to specific growth, differentiation, and survival factors. Induction of MCL1 expression serves to promote viability in cells undergoing changes in proliferation or differentiation or responding to stress, infection, or other signals. Thus, when MCL1 is conditionally knocked out, hematopoietic stem cells do not survive, and early stage B-or T-lymphocytes die instead of continuing their differentiation (2, 3); mature lymphoid cells likewise exhibit impaired survival despite the presence of growth factors (4, 5). Just as induction of MCL1 expression serves to promote viability, its down-regulation is an early, pivotal event in apoptosis initiated by DNA damage and other death stimuli (6, 7). Overall, in response to changing signals, increases and decreases in MCL1 expression regulate cell viability, maintaining cells and lineages that are needed while eliminating those that are no longer needed or are damaged or overabundant (1).Although MCL1 is normally expressed in particular cells in response to specific sti...

Ubiquitin-Independent Degradation of Antiapoptotic MCL-1

“…However, mutagenesis of the putative noncanonical splice donor/acceptor pair does not affect the generation of all three species of MCL-1 (data not shown). Our data are consistent with the observation that human MCL-1 undergoes aminoterminal truncation resulting in multiple protein species (18). Treatment of wild-type MEFs with MG-132, lactacystin, epoxomicin, and proteasome inhibitor-1 (PI-1) did not alter the level of the MCL-1 40-kDa band but significantly resulted in an increase in the levels of the 38-kDa and 36-kDa bands compared to those of untreated samples (Fig.…”

Enhanced stability of Mcl1, a prosurvival Bcl2 relative, blunts stress-induced apoptosis, causes male sterility, and promotes tumorigenesis

“…At a mechanistic level, precisely how the additional N-terminal amino acids block basal Mcl1 turnover is unclear. The N-terminal region of Mcl-1 may contain a poorly understood degron, because Mcl-1 turnover can be enhanced not only by lengthening the N terminus, as observed here, but by shortening it (34).…”

“…A recently recognized but puzzling aspect of Mcl1 regulation is that a proportion of the Mcl-1 protein molecules is processed to remove the N-terminal 33 amino acid residues and the truncated protein enters the mitochondrial matrix (34)(35)(36). One study has reported that the matrix form influences mitochondrial fission/ fusion and energetics (36 Fig.…”

Elevated Mcl-1 inhibits thymocyte apoptosis and alters thymic selection

“…For WT thymocytes, each of the cytotoxic agents provoked a major shift in the mobility of endogenous Mcl-1 by 24 h, presumably reflecting modification and/or degradation. 27 In the Mcl-1tg thymocytes, however, the level of intact Mcl-1 stayed high over the course of treatment with most agents, presumably accounting for the improved viability despite the increase in BH3-only proteins. Exposure to dexamethasone, in contrast, resulted in a decrease in Mcl-1 within 24 h, explaining the greater sensitivity of the tg cells to this agent.…”