Participation of cyclin A in Myc-induced apoptosis.

Abstract

The involvement of c-Myc in cellular proliferation or apoptosis has been linked to differential cyclin gene expression. We observed that in both proliferating cells and cells undergoing apoptosis, cyclin A (but not B. C, D1, and E) mRNA level was elevated in unsynchronized Myc-overexpressing cells when compared with parental Ratla fibroblasts. We further demonstrated that Zn2+-inducible cyclin A expression was sufficient to cause apoptosis When Myc-induced apoptosis was blocked by coexpression ofBc1-2, the levels ofcyclin C, D1, and E mRNAs were also elevated. Thus, while apoptosis induced by c-Myc is associated with an elevated cyclin A mRNA level, protection from apoptouis by coexpressed Bc1-2 is associated with a complementary increase in cycin C, D1, and E mRNAs.c-Myc is a helix-loop-helix leucine-zipper transcriptional factor that participates in opposite cellular fates of proliferation or programmed cell death (apoptosis) (1-4). Under normal cell culture conditions (i.e., high serum), overexpression of c-Myc causes cells to proliferate. Upon serum starvation or growth factor withdrawal, however, certain cells that overexpress c-Myc undergo apoptosis instead of growth arrest (1, 2). Although the domains of c-Myc protein required for neoplastic transformation and apoptosis are the same, the molecular events underlying these divergent cellular fates remain unknown (2-4).Recent studies on the interleukin 2-dependent BAF-B03 pre-B cell line indicate that c-Myc alters cyclin gene expression and affects the transition of cells into the G2 phase (5). Inducible c-Myc activity in a Myc-estrogen receptor chimeric system augments cyclin A and E gene expression and stimulates mitogenesis in rat fibroblasts (6). We sought to define further the possible connections between c-Myc, cyclin gene expression and cell fates. Specifically, we hypothesized that cell death might result from constitutive imbalanced cyclin gene expression induced by c-Myc in the setting of growth factor withdrawal (1, 2, 7). In addition, because Bcl-2 could counteract c-Myc-induced apoptosis (8, 9), the Bcl-2 effect may also be manifested through altered cycin gene expression. (10). Polypeptides from one-tenth of total cell lysates from each 100 x 20 mm plate were resolved by SDS/10%o PAGE and subjected to immunoblot analysis with either anti-Myc 9E10 monoclonal antibody (1:100 dilution) (2) or anti-Bcl-2 monoclonal antibody (1:100 dilution) (Dako). After incubation of blots with a secondary goat anti-mouse horseradish peroxidase-conjugated antibody (Bio-Rad) (1:10,000 dilution), reactive polypeptides were detected by the enhanced chemiluminescence (ECL) system (Amersham). Luminograms from the same blot developed with either antiMyc or anti-Bcl-2 primary antibodies were superimposed and shown with molecular weight standards for clarity of presentation. MATERIALS AND METHODSExpression of Zn2+-inducible human cyclin A in cell lines was detected 24 hr after induction with various final concentrations of ZnSO4. Total cell lysates were prepared,...

Myc down-regulation induces apoptosis in M14 melanoma cells by increasing p27kip1 levels

“…These observations are in agreement with results from other authors, who demonstrated that a deregulation of c-myc is associated to an increase of cyclin E expression (Jansen-Durr et al, 1993;Daksis et al, 1994;Hanson et al, 1994;Hoang et al, 1994). However, even though evidences have been provided that c-myc is able to directly transactivate the expression of cyclin E, the mechanism involved remains unclear (Leone et al, 1997;PerezRoger et al, 1997).…”

Mechanisms of apoptosis by c-Myc

“…Several genes that promote cell cycle transit have been suggested as c-Myc targets (Dang, 1999) and two of these, cyclin A and Cdc25A, have been suggested to participate in apoptosis by c-Myc (Hoang et al, 1994;Galaktionov et al, 1996). Integrins regulate cyclin A, which associates with CDK2 and promotes S phase progression.…”

“…Integrins regulate cyclin A, which associates with CDK2 and promotes S phase progression. Enforced expression of cyclin A in Rat1 cells is su cient to confer anchorage-independent growth capacity and susceptibility to apoptosis by serum deprival (Hoang et al, 1994). Cdc25A is a phosphatase responsible for activating cdc2.…”

Apoptosis and Proliferation of Fibroblasts During Postnatal Skin Development and Scleroderma in the Tight-skin Mouse

“…In general, cultured young or c-myc -expressing fibroblasts, with greater proliferation rates, exhibit lesser resistance to apoptosis than senescent fibroblasts (Wang 1995;Evan et al 1992). This may be related to the need of cell cycle progres-sion for both proliferation and apoptosis (Hoang et al 1994;Wu and Levine 1994). In addition, the Bcl-2 protein, an inhibitor of apoptosis, is differentially expressed in young and senescent fibroblasts and is involved in their different susceptibility to apoptosis (Wang 1995).…”