Cdk1/Cyclin B1 Controls Fas-Mediated Apoptosis by Regulating Caspase-8 Activity

Abstract

Caspase activation is a hallmark of apoptosis. However, the molecular mechanisms underlying the regulation of caspase-8 activation within the extrinsic death pathway are not well understood. In this study, we demonstrate that procaspase-8 is phosphorylated in mitotic cells by Cdk1/cyclin B1 on Ser-387, which is located at the N terminus of the catalytic subunit p10. This phosphorylation of procaspase-8 on Ser-387 occurs in cancer cell lines, as well as in primary breast tissues and lymphocytes. Furthermore, RNA interferencemediated silencing of cyclin B1 or treatment with the Cdk1 inhibitor RO-3306 enhances the Fas-mediated activation and processing of procaspase-8 in mitotic cells. A nonphosphorylatable procaspase-8 (S387A) facilitates Fas-induced apoptosis during mitosis. Our findings suggest that Cdk1/cyclin B1 activity shields human cells against extrinsic death stimuli and unravel the molecular details of the cross talk between cell cycle and extrinsic apoptotic pathways. Finally, this new mechanism may also contribute to tumorigenesis.Intact apoptotic machinery is essential to maintain the integrity and homeostasis of multicellular organisms (8). The evasion of apoptosis is a hallmark of cancer, as demonstrated by the inability of cancer cells to respond appropriately to signals that normally control unrestricted growth (11). In mammalian cells, the apoptotic response is mediated by either the intrinsic or the extrinsic pathway, depending on the origin of the death stimulus. After stimulation of the death receptor Fas (APO-1/CD95) the death-inducing signaling complex (DISC) assembles, which contains the oligomerized receptor, the adaptor molecule FADD, two isoforms of procaspase-8 (procaspase-8a and -8b), procaspase-10, and c-Flip L/S/R (28, 37). In accordance with the induced proximity model, immediately after DISC formation, procaspase-8, which consists of two death effector domains (DED) and a protease domain containing the p18 and p10 subunits, is proteolytically processed (28). This autoprocessing follows a sequential order of events: while the first cleavage step occurs at Asp-374 and results in the formation of the subunits p43/p41 and p12, the second cleavage at Asp-216 and Asp-384 produces the enzymatically active subunits p18, p10, and the prodomain p26/p24 (5, 14, 27, 40). The mature caspase-8 heterotetramer p18 2 -p10 2 then translocates from the DISC to the cytosol, where it cleaves several substrates, such as Bid, and effector caspases to initiate the apoptotic cascade (22).Increasing evidence highlights the functional importance of procaspase-8 for carcinogenesis; several findings suggest that the impairment of procaspase-8 function by genetic and epigenetic mechanisms correlates with the malignant potential of different types of cancer (6,12,43,44). In the present study, we investigated the functional correlation of the cell cycle with the extrinsic death pathway. The cyclin-dependent kinase 1 (Cdk1) in complex with cyclin B1 (Cdk1/cyclin B1) is one of the key mitotic kinases. The kinase ac...

BimEL is phosphorylated at mitosis by Aurora A and targeted for degradation by βTrCP1

“…Caspase-2, -8, and -9 are phosphorylated by Cdk1/Cyclin B1 during mitosis and these modifications inhibit pro-apoptotic activity. [40][41][42] A general feature of mitotic cells is the loss of attachment to extracellular matrix as the cytoskeleton is remodeled in preparation for spindle formation. When anchorage-dependent cells remain in a detached state for extended periods, they undergo a process termed anoikis.…”

Killing cells by targeting mitosis

“…82 In contrast, Cdk1 may also phosphorylate caspase-9 and caspase-8, hampering their ability to trigger the intrinsic or extrinsic apoptotic pathways, respectively. 83,84 Altogether, these data suggest that cyclin B1-Cdk1 complexes have a key role in balancing survival and cell death pathways that dictate the fate of cells during mitotic arrest. How the proapoptotic and prosurvival activities of Cdk1 are controlled is not clear at present.…”

Regulation of CD95/Fas signaling at the DISC

“…Phosphorylation of procaspase-8 at the catalytic subunits has been shown to have an important role in the activity of procaspase-8, likely by interfering with processing of procaspase-8. 40 Phosphorylation at Y380 of procaspase-8 has been shown to inhibit CD95-mediated apoptosis. 41 In addition, polyubiquitination of procaspase-8 at the DISC has been reported recently to stabilize the active caspase-8 heterotetramer and, in this way, to have a pro-apoptotic role.…”