Generation of an indestructible cyclin B1 by caspase-6-dependent cleavage during mitotic catastrophe

Abstract

Overriding the G 2 DNA damage checkpoint permits precocious entry into mitosis that ultimately leads to mitotic catastrophe. Mitotic catastrophe is manifested by an unscheduled activation of CDK1, caspase activation and apoptotic cell death. We found that although cyclin B1 was required for mitotic catastrophe, it was cleaved into a B35 kDa protein by a caspase-dependent mechanism during the process. Cyclin B1 cleavage occurred after Asp123 in the motif ILVD 123 k, and mutation of this motif attenuated the cleavage. Cleavage was abolished by a pan-caspase inhibitor as well as by specific inhibitors for the effector caspase-6 and the initiator caspase-8. Cleavage created a truncated cyclin B1 lacking part of the NH 2 -terminal regulatory domain that included the destruction box sequence. Although cleavage of cyclin B1 itself was not absolutely required for mitotic catastrophe, expression of the truncated product enhanced cell death. In support of this, ectopic expression of this truncated cyclin B1 was not only sufficient to induce mitotic block and apoptosis but also enhanced mitotic catastrophe induced by ionizing radiation and caffeine. These data underscore a possible linkage between mitotic and apoptotic functions by caspase-dependent processing of mitotic activators.

Adenovirus Replaces Mitotic Checkpoint Controls

“…Cells infected with E4orf3-mutant viruses also contain a cyclin B-related protein (Fig. 6) similar in size to a cleaved form of cyclin B1 that sustains the mitotic block (34). In the context of an E1B-55K deletion, E4orf3 may prevent accumulation of this inhibitory product.…”

“…This smaller form of cyclin B1 appears to correspond to a cleavage product found during mitotic catastrophe, termed cyclin B1⌬ (34). It was suggested that cyclin B1⌬ acts as a dominant-negative inhibitor of cyclin B1 function and sustains the mitotic block in cells that would otherwise exit mitosis (34). Both the elevated levels of cyclin B1 and the presence of a dominant-negative inhibitor could retard E1B-55K/E4orf3 double mutant virus-infected cells in mitosis.…”

“…A 35-kDa protein recognized by cyclin B1 antibodies was detected in lysates from cells infected with the E1B-55K/E4orf3 double mutant virus and, to a lesser extent, with the E4orf3-mutant virus. This smaller form of cyclin B1 appears to correspond to a cleavage product found during mitotic catastrophe, termed cyclin B1⌬ (34). It was suggested that cyclin B1⌬ acts as a dominant-negative inhibitor of cyclin B1 function and sustains the mitotic block in cells that would otherwise exit mitosis (34).…”

“…It has been proposed that mitotic catastrophe be considered an oncosuppressive mechanism leading to cell death resulting from the disorder of mitotic machinery that is typified by a period of aberrant mitotic arrest (32). Additional hallmarks of mitotic catastrophe include the formation of multipolar spindles and the appearance of a cleaved form of cyclin B1 that is unable to be degraded and is therefore presumed to maintain activity of the mitotic kinase (33,34). Aberrant mitotic arrest requires that cells enter mitosis but halt prior to anaphase.…”

Determinants of Mitotic Catastrophe on Abrogation of the G2 DNA Damage Checkpoint by UCN-01

“…Cyclin B-CDK1 activity is known to be associated with cell death. For example, mitotic catastrophe can be promoted by cyclin B1-CDK1 overexpression (37,38) and inhibited by cyclin B1 knockdown (39). Although the underlying principles of CDK1-mediated toxicity remain largely unresolved, a few targets that affect survival, including members of the BCL-2 family have been identified (11).…”