Activation of cyclin-dependent kinases by Myc mediates induction of cyclin A, but not apoptosis.

Abstract

The activation of conditional alleles of Myc induces both cell proliferation and apoptosis in serum‐deprived RAT1 fibroblasts. Entry into S phase and apoptosis are both preceded by increased levels of cyclin E‐ and cyclin D1‐dependent kinase activities. To assess which, if any, cellular responses to Myc depend on active cyclin‐dependent kinases (cdks), we have microinjected expression plasmids encoding the cdk inhibitors p16, p21 or p27, and have used a specific inhibitor of cdk2, roscovitine. Expression of cyclin A, which starts late in G1 phase, served as a marker for cell cycle progression. Our data show that active G1 cyclin/cdk complexes are both necessary and sufficient for induction of cyclin A by Myc. In contrast, neither microinjection of cdk inhibitors nor chemical inhibition of cdk2 affected the ability of Myc to induce apoptosis in serum‐starved cells. Further, in isoleucine‐deprived cells, Myc induces apoptosis without altering cdk activity. We conclude that Myc acts upstream of cdks in stimulating cell proliferation and also that activation of cdks and induction of apoptosis are largely independent events that occur in response to induction of Myc.

Quiescence versus apoptosis: Myc abundance determines pathway of exit from the cell cycle

“…We ®nd that the overexpression of native c-Myc greatly enhances apoptosis when cells are serum starved. In addition, this apoptosis is associated with striking elevations in cdk2 activity as previously described (Rudolph et al, 1996;Steiner et al, 1995) and the transient activation of cdk2 is preceded by increases in cdc25A phosphatase (Galaktionov et al, 1996) and followed by increased p27 Kip1 kinase inhibitor levels (Steiner et al, 1995). Thus, our results con®rm that perturbations in cell cycle regulatory pathways are prominent in myc-overexpressing cells undergoing apoptosis.…”

“…Indeed, previous studies have proposed direct links between Myc activation and cdk activity Daksis et al, 1995;Hanson et al, 1994;Hoang et al, 1994; Jansen-Duerr et al, 1993; Philipp et al, 1994; Rudolph et al, 1996). However, the relationships among Myc's e ects on cdk activity, DNA synthesis and apoptosis remain unclear (Hueber et al, 1997;Pusch et al, 1997;Rudolph et al, 1996;Steiner et al, 1995). We therefore determined whether cells expressing a native human c-myc gene and undergoing apoptosis as a result of serum starvation might also show altered patterns of cdk kinase activity.…”

“…Many regulatory factors that modulate cdk2 activity, including di erential dimerization with cyclins, binding to low molecular weight inhibitors such as p21 WAF1/Cip1 and p27 Kip1 , and reversible posttranslational modification of the cdk2 protein, have been suggested to play a role in the e ect of Myc upon the cell (Galaktionov et al, 1996;Hoang et al, 1994;Rudolph et al, 1996;Steiner et al, 1995). We observed no consistent changes in the abundance of known cdk2 partners, cyclins A, D1 or E, that would correlate with cdk2 activation (data not shown).…”

“…As observed with Myc, elevated cyclin levels can result either in transformation (Motokura et al, 1991;Withers et al, 1991) or in apoptosis (Freeman et al, 1994;Hoang et al, 1994;Lahti et al, 1995;Meikrantz et al, 1994;Meikrantz and Schlegel, 1996;Sherr and Roberts, 1995;Shi et al, 1994), and dominant negative mutants of cdc2, cdk2 and cdk3 can suppress apoptosis (Meikrantz and Schlegel, 1996). Direct links between the activation of Myc and cdks have been proposed (Galaktionov et al, 1996;Hoang et al, 1994;Rudolph et al, 1996;Steiner et al, 1995) and build on the concept that the inappropriate overexpression of c-myc may cause aberrant cell cycle events as a prelude to apoptosis in cells predisposed to this condition. If so, the promotion of apoptosis in cells overexpressing c-myc may be a protective mechanism by the host against the oncogenic potential of Myc protein (Evan and Littlewood, 1993;Harrington et al, 1994), suggesting that cells that secondarily acquire the ability to survive the deleterious consequences of excess Myc (while still bene®ting from its proliferative stimulus) would become fully malignant To explore these complex issues, we have focused our attention on the initial responses of cells to c-myc overexpression in an e ort to emulate the early changes that may arise during oncogenic progression.…”

Mechanisms of apoptosis by c-Myc

“…However, these ideas have been challenged recently because positive associations with death have not been widely generalized. One study that employed a variety of cell cycle inhibitors concluded that Cdks are dispensable for apoptosis (Rudolph et al, 1996). In addition, a more recent study noted that Cdk2 and Cdc25A message levels reacted similarly in Rat1 cells, whether they underwent apoptosis in the presence of deregulated c-Myc or simply exit to G0 in its absence (Helbing et al, 1998).…”