Lars-Gunnar Larsson group
The myc oncogenes encode powerful transcription factors that control expression of a vast number of genes involved in, among other functions, cell growth, apoptosis, metabolism and stem cell function. Deregulated MYC contributes to the development of a high proportion of human cancers, in particular at the advanced stages of disease. We are studying the function and regulation of the Myc proteins, with the aim of identifying molecules targeting Myc protein activity.
Function, Regulation and Targeting of the Myc Protein
Recently, we uncovered regulation of cellular senescence, one of the main barriers of tumor development, as a new function of Myc. We found that induction of a Myc antagonist caused cellular senescence in Myc-transformed cells (Wu et al., 2009). Further, Myc represses senescence induced by other oncogenes such as Ras. Repression of senescence by Myc requires phosphorylation of Myc at Ser-62 by the cell cycle kinase Cdk2 (Hydbring et al., 2010). Cdk2 is also required to represses senescence induced by overexpressed Myc itself (Campaner et al., 2010). Importantly, inhibition of Cdk2 by small molecule inhibitors induced senescence in Myc-driven tumor cells (Hydbring et al.,2010; Campaner et al., 2010).
Our future research aims to elucidate exactly how Myc and Cdk2 regulates senescence and to evaluate the in vivo potential of Cdk2 inhibitors in Myc-driven mouse tumor models. Another potential way of combating Myc is to explore and utilize mechanisms of Myc protein destruction via the ubiquitin/ proteasome pathway, and we are continuing our work to identify new E3 ubiquitin ligases targeting Myc. Our recent research also involves identification of low molecular weight inhibitors of interactions between Myc and cofactors crucial for Myc function. Using novel methods for visualizing protein interactions in living or fixed cells, we have screened chemical libraries and identified several molecules selectively targeting Myc. The long-term aim of this work is the development of new drugs for cancer treatment.
Post-translational regulation of Myc through modifications and ubiquitin/proteasome mediated turnover.
Myc is modified at several sites by phosphorylation, acetylation and ubiquitylation, and these modifications are thought to affect Myc function, including its interaction with cofactors and rapid turnover via the ubiquitin/proteasome pathway. We are trying to dissect out the precise role of each of these modifications, to identify new ones and also to identify the players involved.
In 2003, we and Bill Tanseys group at CSHL identified SCFSkp2 as the first E3 ubiquitin ligase degrading c-Myc (von der Lehr et al, 2003). Since then two other E3 ligases, Fbw7/hCdc4 and HectH9 have been shown to target Myc. We have evidence that yet a number of other E3 ligases participate in Myc ubiquitylation and are dissecting the specific roles of each of them play and their relation to each other in this process.
How does Myc regulate transcription?
To understand the function of Myc in transcription regulation it is essential to elucidate its network of interactions with different cofactors and to determine the specific role of each cofactor in Myc-driven transcription. During recent years a picture has emerged of Myc as a 'master mechanic of transcription', with a role in recruitment to promoters all enzymes, adaptors and other factors necessary for this process. These factors may for instance have a role in the regulation of chromatin structure or carry out functions at specific steps in the transcription process such as initiation, elongation or post-transcriptional events. An unexpected finding was that the E3 ligase SCFSkp2, although degrading Myc, acts as a necessary cofactor for Myc-activated transcription and together with the proteasome is recruited by Myc to target promoters. We are therefore particularly interested the function of the ubiquitin/proteasome system in Myc-driven transcription.
To understand Myc function it is also essential to get a complete overview of the target genes under its control, and the contribution of each of these target genes to Myc-induced phenotype, including roles in cell cycle progression, cell growth, cell differentiation, cell survival, cell metabolism etc. Our recent findings suggest that Myc not only regulates transcription of RNA polymerase II (Pol II)- but also of Pol I- and Pol III-driven genes (Arabi et al, 2005), and the question remains whether it carries out similar or different tasks at different promoters.
Visualization of protein-protein interactions between Myc and cofactors in individual cells.
In collaboration with Tom Kerppola (Ann Arbor, Michigan) we have utilized Bimolecular Fluorescence Complementation (BiFC) to study intracellular localization of interactions between Myc and its different cofactors in living cells in real-time (von der Lehr et al, 2003, Arabi et al., 2005). In BiFC, two non-fluorescent fragments of Yellow Fluorescent Protein (YFP) are fused to two interacting proteins of interest, respectively. Upon protein-protein interaction, a functional fluorescent YFP protein is reformed, visualizing the interaction. This technique is now further developed to study multiple transcription factor complexes simultaneously.
In collaboration with Ulf Landegren and Ola Söderberg (Uppsala), we have recently developed Proximity Ligation In Situ Assay (P-LISA). With this method, which is based on DNA-conjugated antibodies and rolling circle DNA amplification, we visualize endogenous interactions between Myc and cofactors at the single molecule level in individual cells (Söderberg et al., 2006). The technique is further developed to study interactions between three or more proteins and also protein/DNA interactions. Applicable for histological tissue sections, it can also be utilized for clinically oriented studies of Myc-driven tumorigenesis (or any interacting proteins of relevance).
Can the acquired knowledge about Myc function and regulation be utilized to combat cancer?
Based on insights into pathways involved in Myc protein interactions, modification, degradation, we are interested in dissecting ways to inhibit Myc function (or to enhance its proapoptotic function) in cancer cells. For instance, using chemical libraries of small molecules, we are screening for specific inhibitors of interactions between Myc and its different cofactors based on some of the assays described above. This could not only give important tools to dissect the specific roles of different interactions in Myc function, but potentially also provide important molecules for cancer therapy.
Giovanna Zinzalla Project
MYC and RAS are unable to cooperate in overcoming cellular senescence and apoptosis in normal human fibroblasts.
Zhang F, Zakaria SM, Högqvist Tabor V, Singh M, Tronnersjö S, Goodwin J, et al
Cell Cycle 2018 ;17(24):2697-2715
MYC Modulation around the CDK2/p27/SKP2 Axis.
Hydbring P, Castell A, Larsson LG
Genes (Basel) 2017 Jun;8(7):
Interferon-γ-induced p27KIP1 binds to and targets MYC for proteasome-mediated degradation.
Bahram F, Hydbring P, Tronnersjö S, Zakaria SM, Frings O, Fahlén S, et al
Oncotarget 2016 Jan;7(3):2837-54
Targeting MYC Translation in Colorectal Cancer.
Castell A, Larsson LG
Cancer Discov 2015 Jul;5(7):701-3
Sin3b interacts with Myc and decreases Myc levels.
Garcia-Sanz P, Quintanilla A, Lafita MC, Moreno-Bueno G, García-Gutierrez L, Tabor V, et al
J. Biol. Chem. 2014 Aug;289(32):22221-36
MYC synergizes with activated BRAFV600E in mouse lung tumor development by suppressing senescence.
Tabor V, Bocci M, Alikhani N, Kuiper R, Larsson LG
Cancer Res. 2014 Aug;74(16):4222-9
The c-MYC oncoprotein, the NAMPT enzyme, the SIRT1-inhibitor DBC1, and the SIRT1 deacetylase form a positive feedback loop.
Menssen A, Hydbring P, Kapelle K, Vervoorts J, Diebold J, Lüscher B, et al
Proc. Natl. Acad. Sci. U.S.A. 2012 Jan;109(4):E187-96
Cellular senescence--a barrier against tumor development?
Semin. Cancer Biol. 2011 Dec;21(6):347-8
Oncogene- and tumor suppressor gene-mediated suppression of cellular senescence.
Semin. Cancer Biol. 2011 Dec;21(6):367-76
Tipping the balance: Cdk2 enables Myc to suppress senescence.
Hydbring P, Larsson LG
Cancer Res. 2010 Sep;70(17):6687-91
The Yin and Yang functions of the Myc oncoprotein in cancer development and as targets for therapy.
Larsson LG, Henriksson MA
Exp. Cell Res. 2010 May;316(8):1429-37
Myc is required for activation of the ATM-dependent checkpoints in response to DNA damage.
Guerra L, Albihn A, Tronnersjö S, Yan Q, Guidi R, Stenerlöw B, et al
PLoS ONE 2010 Jan;5(1):e8924
Cdk2 suppresses cellular senescence induced by the c-myc oncogene.
Campaner S, Doni M, Hydbring P, Verrecchia A, Bianchi L, Sardella D, et al
Nat. Cell Biol. 2010 Jan;12(1):54-9; sup pp 1-14
Phosphorylation by Cdk2 is required for Myc to repress Ras-induced senescence in cotransformation.
Hydbring P, Bahram F, Su Y, Tronnersjö S, Högstrand K, von der Lehr N, et al
Proc. Natl. Acad. Sci. U.S.A. 2010 Jan;107(1):58-63
Direct observation of individual endogenous protein complexes in situ by proximity ligation.
Söderberg O, Gullberg M, Jarvius M, Ridderstråle K, Leuchowius KJ, Jarvius J, et al
Nat. Methods 2006 Dec;3(12):995-1000
c-Myc associates with ribosomal DNA and activates RNA polymerase I transcription.
Arabi A, Wu S, Ridderstråle K, Bierhoff H, Shiue C, Fatyol K, et al
Nat. Cell Biol. 2005 Mar;7(3):303-10
The F-box protein Skp2 participates in c-Myc proteosomal degradation and acts as a cofactor for c-Myc-regulated transcription.
von der Lehr N, Johansson S, Wu S, Bahram F, Castell A, Cetinkaya C, et al
Mol. Cell 2003 May;11(5):1189-200
Myc represses differentiation-induced p21CIP1 expression via Miz-1-dependent interaction with the p21 core promoter.
Wu S, Cetinkaya C, Munoz-Alonso MJ, von der Lehr N, Bahram F, Beuger V, et al
Oncogene 2003 Jan;22(3):351-60
c-Myc hot spot mutations in lymphomas result in inefficient ubiquitination and decreased proteasome-mediated turnover.
Bahram F, von der Lehr N, Cetinkaya C, Larsson LG
Blood 2000 Mar;95(6):2104-10