Induction of tumor promotor-inducible genes in murine 3T3 cell lines and tetradecanoyl phorbol acetate-nonproliferative 3T3 variants can occur through protein kinase C-dependent and -independent pathways.

Abstract

We isolated a group of genes that are rapidly and transiently induced in 3T3 cells by tetradecanoyl phorbol acetate (TPA). These genes are called TIS genes (for TPA-inducible sequences). Epidermal growth factor (EGF), fibroblast growth factor (FGF), and TPA activated TIS gene expression with similar induction kinetics. TPA pretreatment to deplete protein kinase C activity did not abolish the subsequent induction of TIS gene expression by epidermal growth factor or fibroblast growth factor; both peptide mitogens can activate TIS genes through a protein kinase C-independent pathway(s). We also analyzed TIS gene expression in three TPA-nonproliferative variants (3T3-TNR2, 3T3-TNR9, and A31T6E12A). The results indicate that (i) modulation of a TPA-responsive sodium-potassium-chloride transport system is not necessary for TIS gene induction either by TPA or by other mitogens and (ii) TIS gene induction is not sufficient to guarantee a proliferative response to mitogenic stimulation.Tetradecanoyl phorbol acetate (TPA) is a potent biological response modulator with a diverse range of effects on cells, in vivo and in cell culture (27). TPA was first identified by using a mouse skin assay, as a powerful tumor promotor. It is also a potent mitogen for a variety of cell types, including murine 3T3 cells (2). TPA stimulates the differentiation of HL60 promyelocytic leukemic cells (20) but exerts an inhibitory effect on the differentiation of Friend erytholeukemic cells (29) and cultured myoblasts (5). TPA is thought to act primarily by binding to and activating the calcium-and phospholipid-dependent protein kinase C (13). To understand more fully the mechanisms by which TPA can modulate such a wide range of biological responses, we isolated, from mouse 3T3 cells stimulated with TPA in the presence of cycloheximide, a set of TPA-inducible primary response genes termed TIS genes (for TPA inducible sequences [10]). The TIS genes exhibit the following characteristics: (i) they are all induced as a primary response (28) to TPA stimulation, and no protein synthesis is necessary for the increase of TIS mRNAs; (ii) the induction is transient; (iii) TIS mRNAs are unstable; and (iv) TIS mRNA accumulation in the presence of TPA can be superinduced by cycloheximide.TIS genes can be induced by other mitogens in 3T3 cells. We anticipated, when we isolated the TPA-inducible TIS genes, that at least some of them might be uniquely responsive to TPA. To determine whether epidermal growth factor (EGF) or fibroblast growth factor (FGF) could induce TIS gene expression in 3T3 cells and to compare induction kinetics of EGF and FGF with that of TPA, we analyzed the time courses of TIS gene induction by EGF and FGF. Like the TPA response (10), induction of the TIS genes (TIS 1, TIS 8, TIS 10, and TIS 11) in response to EGF or FGF generally peaked at about 30 to 60 min and returned to nearly uninduced levels by 3 to 5 h (Fig. 1). Induction of the TIS * Corresponding author.genes in 3T3 cells is thus not unique to TPA but will occur in response to o...

Suppression of Mammary Carcinoma Cell Growth by Retinoic Acid: the Cell Cycle Control Gene Btg2 Is a Direct Target for Retinoic Acid Receptor Signaling

“…Btg2, which was originally identified as an immediate-early gene induced by nerve growth factor in PC12 cells (40,41), is involved in regulation of cell growth and differentiation. It was thus reported that Btg2 is a direct target for the tumor suppressor p53 (42)(43)(44); that it is induced in response to DNA damage in a p53-dependent manner (44,45); that it triggers cell cycle arrest by inhibiting the G 1 -S transition; and that this effect is exerted by decreasing the expression of cyclin D1 (30,46,47). Recent observations showed that, similarly to its activities in PC12 and NIH3T3 cells, overexpression of Btg2 in MCF-7 cells downregulates the expression of cyclin D1 and inhibits cell growth (48).…”

Expression of cyr61, a growth factor-inducible immediate-early gene.

“…Therefore, induction of cyr6l by serum growth factors does not require PKC, although the activation of PKC can lead to induction of cyr6l. Several other genes identified as TPA inducible can also be activated through PKC-independent pathways (30).…”

Granulocyte-macrophage colony-stimulating factor and tetradecanoyl phorbol acetate induce a distinct, restricted subset of primary-response TIS genes in both proliferating and terminally differentiated myeloid cells.

“…We refer to these as TPA-inducible sequences, or TIS cDNAs (17). All of the TIS mRNAs are also induced in 3T3 cells by serum, epidermal growth factor (EGF), and fibroblast growth factor (18). Each TIS mRNA demonstrates a unique developmental profile (25), which suggests that each TIS gene has unique regulatory features.…”