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Counteracting effects of E1a transformation on cAMP growth inhibition
M Florin-Christensen1, C Missero, J Florin-Christensen
1Department of Internal Medicine, Yale University School of Medicine, New Haven, Connecticut 06510.
Abstract:
Several signaling molecules have been identified which act as inhibitors of epithelial cell growth. The mechanisms for this negative growth regulation are still poorly understood. In the case of TGF-beta, inhibition of keratinocyte cell growth can be totally prevented by transformation with an intact early region 1a (E1a) oncogene. We show here that E1a-transformed keratinocytes become also partially resistant to growth inhibition by elevated 3',5'-cyclic adenosine monophosphate (cAMP) levels, as induced by treatment with forskolin, dibutyryl-cAMP, 8Br-cAMP, or 8Cl-cAMP. Resistance to cAMP is due to interference of E1a with signaling pathways downstream of protein kinase A (PKA) activation, as intracellular cAMP levels and PKA activity were found to be similar in control and E1a-transformed cells. Induction of c-fos expression by 8Br-cAMP occurs at the same time in both cell lines. Interestingly however, this effect is maintained longer in the case of E1a-transformed cells compared to the control. A truncated E1a mutant which is still able to bind to the p105-Rb gene product, p107, and p60/cyclin A, induces cAMP resistance at levels which are only slightly lower than those induced by an intact E1a oncogene. In contrast, an E1a mutant which binds only to a p300 cellular protein and induces a substantial level of TGF-beta resistance fails to induce cAMP resistance. Thus, E1a transformation counteracts the growth-inhibitory effects of cAMP as well as TGF-beta, but to a different degree and through an only partially overlapping mechanism.
Insights
The E1a oncogene confers partial resistance to growth inhibition by cyclic adenosine monophosphate (cAMP) in keratinocytes. This resistance involves E1a interfering with signaling pathways downstream of protein kinase A (PKA).
Area of Science:
- Cell Biology
- Molecular Oncology
- Signal Transduction
Background:
- Epithelial cell growth is regulated by inhibitory signaling molecules, but mechanisms remain unclear.
- Transforming growth factor-beta (TGF-beta) inhibits keratinocyte growth, an effect blocked by the E1a oncogene.
Purpose of the Study:
- To investigate the effect of E1a oncogene transformation on keratinocyte resistance to growth inhibition by cyclic adenosine monophosphate (cAMP).
- To elucidate the molecular mechanisms underlying E1a-mediated resistance to cAMP and TGF-beta.
Main Methods:
- E1a-transformed and control keratinocytes were treated with forskolin, dibutyryl-cAMP, 8Br-cAMP, or 8Cl-cAMP to assess growth inhibition.
- Intracellular cAMP levels and protein kinase A (PKA) activity were measured.
- Expression of c-fos was monitored, and E1a mutants binding to p105-Rb, p107, p60/cyclin A, or p300 were utilized.
Main Results:
- E1a-transformed keratinocytes exhibited partial resistance to cAMP-induced growth inhibition.
- E1a interfered with signaling pathways downstream of PKA activation, as cAMP levels and PKA activity were similar in both cell types.
- E1a mutants binding to p105-Rb, p107, and p60/cyclin A induced significant cAMP resistance, while a mutant binding only to p300 did not.
Conclusions:
- E1a transformation confers partial resistance to cAMP-mediated growth inhibition in keratinocytes.
- E1a's mechanism of cAMP resistance involves interference with downstream signaling, distinct from its TGF-beta resistance pathway.
- Specific E1a protein interactions, particularly with p105-Rb, p107, and p60/cyclin A, are crucial for cAMP resistance.