Deconstructing myc.

Although myc was among the very earliest oncogenes identified and the subject of intense study, it has nonetheless proven to be an enduring enigma. To a large extent the problem derives from the apparent gap between Myc’s biological role and what is surmised to be its molecular function. Myc family proteins (comprising c-, N-, and L-Myc) promote proliferation, growth, and apoptosis; inhibit terminal differentiation; and, when deregulated, are profoundly involved in the genesis of an extraordinarily wide range of cancers (for recent review, see Grandori et al. 2000). Alongside this veritable mountain of biological effects, the molecular characterization of Myc—as a relatively weak transcriptional regulator of uncertain target genes—looks like a molehill. Indeed some have wondered whether the transcriptional activities of Myc might be more apparent than real. The notion that Myc proteins might function in transcription arose over a decade ago, after Myc was shown to be a nuclear protein. An N-terminal fragment of Myc stimulated transcription when fused to a heterologous DNA binding domain, and the C-terminal basic-helixloop-helix-zipper (bHLHZ) of Myc resembled those found in certain families of transcription factors. But full-length Myc did not dimerize or bind DNA and therefore could not be demonstrated to behave as a transcription factor. This picture changed with the discovery of Max, another bHLHZ protein which heterodimerizes with Myc to form a sequence-specific DNA binding complex. Myc–Max heterodimers recognize the E-box sequence CACGTG (as well as related non-canonical sites) and activate transcription from synthetic reporter genes containing multimerized binding sites in mammalian cells as well as yeast (Blackwood and Eisenman 1991; Amati et al. 1992; Kretzner et al. 1992). The effects of Myc on cell proliferation and apoptosis are negated by mutations in the Myc transactivation region, or in the bHLHZ domain required for association with Max and DNA, indicating a tight linkage between the transcriptional and biological activities of Myc. At first glance these findings would be expected to settle the issue of Myc function in favor of the idea that it is a transcription factor which, as a heterodimer with Max, determines cell behavior by activation of a specific set of target genes. However, on second glance a number of complicating issues arise. First, Myc–Max transcriptional activity on either synthetic promoters or putative cellular target genes is disappointingly weak and variable (compared with other activators)—on the order of 2–5 fold. This makes detailed dissection of transcriptional function difficult and, even worse, makes validation of putative target genes both awkward and contentious (for review, see Grandori and Eisenman 1997). Second, although the transcriptional activity of Myc–Max was widely confirmed, there were also reports that Myc could repress transcription of a number of genes dependent on a specific subclass of initiator elements (INRs) located at the transcriptional initiation sites of certain genes (Li et al. 1994). Because the repression experiments did not demonstrate Myc binding to INRs, they left open the possibility that repression might be indirect (e.g. through activation of a repressor by Myc–Max). Third, mutational analyses of Myc Box II, a highly conserved region present in the N-terminal transactivation region of all Myc proteins, produced confounding results. Several reports indicated that mutation of Myc Box II had little effect on transcription activation but abrogated Myc repression activity. Yet other studies indicated that loss of Myc Box II primarily affected activation. Because Myc Box II is crucial for the ability of Myc to transform cells, these results raised the issue of the relative importance of activation and repression (for review, see Grandori et al. 2000). In principle, the importance of Myc transcriptional activity could be settled by identification of specific target genes. The hope was that Myc–Max might modulate expression of a small number of obviously critical genes whose known functions would explain the various biological effects of Myc. However, over the last eight years numerous putative target genes have been identified and, with the advent of global expression arrays, the number that are activated or repressed byMyc continues to grow. Whereas a few of these targets are cell cycle regulators, and therefore might help explain the ability of Myc to drive proliferation, many other candidate target genes have functions relating to metabolism, ribosome biogenesis, or translation, and do not immediately suggest a unitary mechanism for how Myc works. To make matters worse, there has been controversy over the validity of a number of the candidate target genes. For example, in a rat cell line in which c-myc has been deleted, the 1E-MAIL eisenman@fhcrc.org; FAX (206) 667-6522. Article and publication are at http://www.genesdev. org/cgi/doi/10.1101/ gad928101.