Abstract IA18: “Amplification versus Gene-Specific Models”: What is this session about?
Authors: Amati Bruno 
Published: October 2015

Transcription factors (TFs) bind specific DNA-sequence motifs in genomic regulatory elements (such as promoters and enhancers) to modulate the transcriptional activity of their target genes. DNA sequence alone is not sufficient to define target elements in the genome in vivo, with chromatin-associated features (e.g. general accessibility, histone marks, presence of the transcriptional machinery, other factors, etc...) acting as primary determinants in helping TFs to discriminate between functional target sites and spurious consensus motifs [1]. This is particularly true for Myc, which requires a pre-existing active/poised chromatin environment to bind DNA in vivo [2] and recognizes a relatively simple consensus motif. When over-expressed (such as in tumors) or hyper-activated by acute mitogenic stimuli (such as LPS treatment in B cells) Myc becomes detectable by ChIP-seq on a majority, if not all active regulatory elements in the genome, a phenomenon termed “invasion” [3-6]. Two very different interpretations of this phenomenon have been provided: the first calls for a functionally productive engagement of Myc at all invaded loci, and hence the direct up-regulation - also termed “amplification” - of their transcriptional output [3, 4]; the second posits that, upon over-expression, the low-affinity interactions that Myc uses to scan active chromatin become detectable above background, and should not automatically be equated with productive gene regulation [6, 7]. As a corollary, over-expression or hyper-activation of any TF is likely to give rise to apparent chromatin invasion when profiled by ChIP-seq, as observed for example for p53 (see poster by Tonelli et al.).

Most importantly in this setting, Myc activity (whether normal or pathological) endows cells with a series of physiological changes (e.g. in cell size, energy metabolism, translation, nucleotide biosynthesis) that have the potential to feedback on global RNA production [8, 9]. These processes are controlled by a variety of Myc-regulated genes - yet to be fully identified - thereby indirectly connecting Myc activity to general RNA amplification. Indeed, amplification has been observed when comparing cells in different physiologic states, such as quiescent vs. activated, normal vs. tumor, or different tumors with variable Myc levels [3, 4, 6]. It is noteworthy here that total RNA content has been used for years to discriminate proliferating from quiescent cells [10].

Conceptually, we are thus facing two opposite mechanisms with similar phenotypic outcomes. Discriminating between the direct and indirect amplification models will be relevant to the field not only as an academic exercise, but mainly as a basis for the full understanding of Myc' s mechanisms of action in oncogenesis and their potential exploitation in therapeutic development. In this situation, it is thus essential to consider and formally interpret all existing data. Upon careful scrutiny, various datasets, including those based on which direct amplification was originally proposed [3, 4], part of our own [6] and others (e.g. [11, 12]), simply do not allow a formal discrimination between direct and indirect RNA amplification phenomena. However, as will be discussed in this presentation, situations exist for which the sole plausible explanation appears to be differential up- and down-regulation of transcription by Myc, coupled - but not always - to indirect RNA amplification [5, 6] (see also posters by Sabò et al., Tesi et al., Kress et al.).

Altogether, we conclude that (i.) at this time the only model that appears consistent with all existing data is differential gene regulation by Myc; (ii.) the existence of direct transcriptional amplification remains to be formally proven, and is not strictly required to explain existing experimental observations.

References:

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2. Guccione, E., et al., Myc-binding-site recognition in the human genome is determined by chromatin context. Nat Cell Biol, 2006. 8(7): p. 764-70.

3. Lin, C.Y., et al., Transcriptional amplification in tumor cells with elevated c-Myc. Cell, 2012. 151(1): p. 56-67.

4. Nie, Z., et al., c-Myc is a universal amplifier of expressed genes in lymphocytes and embryonic stem cells. Cell, 2012. 151(1): p. 68-79.

5. Walz, S., et al., Activation and repression by oncogenic MYC shape tumour-specific gene expression profiles. Nature, 2014. 511(7510): p. 483-7.

6. Sabò, A., et al., Selective transcriptional regulation by Myc in cellular growth control and lymphomagenesis. Nature, 2014. 511(7510): p. 488-92.

7. Sabo, A. and B. Amati, Genome recognition by MYC. Cold Spring Harbor perspectives in medicine, 2014. 4(2).

8. Marguerat, S. and J. Bahler, Coordinating genome expression with cell size. Trends in genetics : TIG, 2012. 28(11): p. 560-5.

9. Cunningham, J.T., et al., Protein and nucleotide biosynthesis are coupled by a single rate-limiting enzyme, PRPS2, to drive cancer. Cell, 2014. 157(5): p. 1088-103.

10. Darzynkiewicz, Z., F. Traganos, and M.R. Melamed, New cell cycle compartments identified by multiparameter flow cytometry. Cytometry, 1980. 1(2): p. 98-108.

11. Chipumuro, E., et al., CDK7 Inhibition Suppresses Super-Enhancer-Linked Oncogenic Transcription in MYCN-Driven Cancer. Cell, 2014. 159(5): p. 1126-39.

12. Rahl, P.B., et al., c-Myc regulates transcriptional pause release. Cell, 2010. 141(3): p. 432-45.

Citation Format: Bruno Amati. “Amplification versus Gene-Specific Models”: What is this session about? [abstract]. In: Proceedings of the AACR Special Conference on Myc: From Biology to Therapy; Jan 7-10, 2015; La Jolla, CA. Philadelphia (PA): AACR; Mol Cancer Res 2015;13(10 Suppl):Abstract nr IA18.