The CCNG2 gene that encodes the unconventional
cyclin G2 was one of the few genes up-regulated on anti-
human epidermal growth factor receptor 2 (HER2) antibody-mediated inhibition of HER2 signaling. The purpose of this study was to explore how HER2 signaling modulates
cyclin G2 expression and the effect of elevated
cyclin G2 on
breast cancer cell growth. Treatment of
breast cancer cells that overexpress HER2 (BT474, SKBr3, and MDAMB453) with the anti-HER2 antibody
trastuzumab or its precursor 4D5 markedly up-regulated
cyclin G2 mRNA in vitro and in vivo, as shown by real-time PCR. Immunoblot and immunofluorescence analysis with specific
antibodies against
cyclin G2 showed that anti-HER2 antibody significantly increased
cyclin G2 protein expression and translocated the
protein to the nucleus.
Trastuzumab was not able to induce
cyclin G2 expression in cells weakly expressing HER2 (MCF7) or in cells that had developed resistance to
trastuzumab. Enforced expression of HER2 in T47D and MDAMB435
breast cancer cells reduced
cyclin G2 levels. Collectively, these data suggest that HER2-mediated signaling negatively regulates
cyclin G2 expression. Inhibition of
phosphoinositide 3-kinase (
LY294002), c-jun NH(2)-terminal
kinase (
SP600125), and
mammalian target of rapamycin (mTOR)/
p70 S6 kinase (
p70S6K;
rapamycin) increased
cyclin G2 expression. In contrast, treatment with inhibitors of
p38 mitogen-activated protein kinase (
SB203580),
mitogen-activated protein kinase/
extracellular signal-regulated kinase kinase 1/2 (
U0126), or
phospholipase Cgamma (
U73122) did not affect
cyclin G2 expression. Anti-HER2 antibody in combination with
LY294002,
rapamycin, or
SP600125 induced greater
cyclin G2 expression than either agent alone. Ectopic expression of
cyclin G2 inhibited
cyclin-dependent kinase 2 activity, Rb phosphorylation, cell cycle progression, and cellular proliferation without affecting p27(Kip1) expression. Thus,
cyclin G2 expression is modulated by HER2 signaling through multiple pathways including
phosphoinositide 3-kinase, c-jun NH(2)-terminal
kinase, and mTOR signaling. The negative effects of
cyclin G2 on cell cycle and cell proliferation, which occur without altering p27(Kip1) levels, may contribute to the ability of
trastuzumab to inhibit
breast cancer cell growth.