<em>twist</em> is a potential oncogene that inhibits apoptosis
Abstract
Oncogene activation increases susceptibility to apoptosis. Thus, tumorigenesis must depend, in part, on compensating mutations that protect from programmed cell death. A functional screen for cDNAs that could counteract the proapoptotic effects of the myc oncogene identified two related bHLH family members, Twist and Dermo1. Both of these proteins inhibited oncogene- and p53-dependent cell death. Twist expression bypassed p53-induced growth arrest. These effects correlated with an ability of Twist to interfere with activation of a p53-dependent reporter and to impair induction of p53 target genes in response to DNA damage. An underlying explanation for this observation may be provided by the ability of Twist to reduce expression of the ARF tumor suppressor. Thus, Twist may affect p53 indirectly through modulation of the ARF/MDM2/p53 pathway. Consistent with a role as a potential oncoprotein, Twist expression promoted colony formation of E1A/ras-transformed mouse embryo fibroblasts (MEFs) in soft agar. Furthermore, Twist was inappropriately expressed in 50% of rhabdomyosarcomas, a tumor that arises from skeletal muscle precursors that fail to differentiate. Twist is known to block myogenic differentiation. Thus, Twist may play multiple roles in the formation of rhabdomyosarcomas, halting terminal differentiation, inhibiting apoptosis, and interfering with the p53 tumor-suppressor pathway.
A defining characteristic of tumor cells is the escape from regulatory mechanisms that normally restrain cell proliferation. This is accomplished through the accumulation of multiple genetic alterations. Among these are the inactivation of key tumor suppression pathways and the activation of oncogenes (for review, see Vogelstein and Kinzler 1998).
The products of cellular oncogenes such as ras and myc are components of normal growth control pathways. These form part of the program that promotes entry into the division cycle in response to appropriate environmental cues. However, in tumor cells, the normal function of these genes is subverted to provide hyperactive proliferative signals. It is becoming increasingly clear that normal cells respond to inappropriate growth signals by activating homeostatic growth control pathways that protect multicellular organisms from tumor formation.
Constitutive activation of Ras promotes transformation of some immortalized cells. However, Ras activation in normal cells provokes cellular senescence. This irreversible growth arrest probably negates the ability of these cells to contribute to tumor formation (Serrano et al. 1997; Lin et al. 1998). In contrast, other cellular and viral oncogenes sensitize cells to undergo programmed cell death on exposure to stimuli that might normally cause a reversible growth arrest. For example, Myc induces apoptosis on removal of serum survival factors from primary cells or Rat1 fibroblasts (Evan et al. 1992; Hermeking and Eick 1994; Wagner et al. 1994). Similarly, primary mouse embryo fibroblasts (MEFs) that express E1A are sensitized to programmed cell death in response to contact inhibition, growth factor withdrawal, and DNA damage (Debbas and White 1993; Lowe and Ruley 1993; Lowe et al. 1993, 1994).
The growth inhibitory properties of many oncogenes dictate that tumorigenesis requires the cooperation of different classes of genes. For example, transformation by Ras can proceed only in the presence of additional mutations that prevent Ras-induced senescence (Serrano et al. 1997). Furthermore, the ability of Myc or E1A to promote tumorigenesis requires that transformed cells be protected from the proapoptotic effects of these oncoproteins. An understanding of oncogene cooperation requires both a knowledge of the mechanisms by which oncogene activation provokes homeostatic responses and a clarification of the routes through which cooperating oncogenic events defeat these protective controls.
It is clear that oncogene-transformed cells require additional genetic alterations that render them resistant to apoptotic stimuli. Such genetic changes are likely to be obligate for oncogenesis. However, the full spectrum of these events has proven difficult to elucidate, because protective genes are not likely to be discovered via the cellular transformation assays that have proven a rich source of new oncogenes. For example, bcl-2 synergizes with myc in the generation of Burkitt lymphoma but does not cooperate with myc in transformation of mouse fibroblasts in vitro (Vaux et al. 1988). As an approach to the discovery of potential oncogenes that might elude conventional methods, we have undertaken a search for cellular genes that can counter the proapoptotic effects of myc activation.
Acknowledgments
R.M. was supported by a grant from the Italian Association for Cancer Research (AIRC). L.K. is supported in part by a grant from the National Institutes of Health (NIH). Y.H. is supported by an Initial Investigatorship (1104-FI1) and V.S. by a grant-in-aid (1060-G1) from the American Heart Association of Greater Los Angeles. D.B. is the Hugh and Catherine Stevenson Chair in Cancer Biology. G.J.H. is supported by grants from the US Army (DAMD 17-96-1-6053), the NIH, and the Stewart Trust and is a Pew Scholar in the Biomedical Sciences. We thank Chris McCollough, Sara Piccinin, and Martina Fabris for their help and support, Bert Vogelstein for providing the PG-13 reporter construct, and Scott Lowe and Linda Penn for C8 MEF and Rat-1/MycER cells, respectively.
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Footnotes
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