c-Myc A Biological Paradox Ashton Butler

Introduction to c-Myc Normal role in the cell Relation to cancer Table of Contents Introduction to c-Myc Normal role in the cell Relation to cancer Burkitt’s lymphoma

Introduction Oncogene Found on Chromosome 8 Member of Myc protein family Includes N-myc and L-myc Basic helix-loop-helix leucine zipper (bHLH/LZ) transcription factors

bHLH/LZ Transcription Factor Structure -contain the bHLH/LZ domain (basic helix-loop-helix leucine zipper domain) -important in myc’s normal biochemical/biological functions in the cell

Normal Function c-Myc activated through dimerization with Max leads to transcription of target genes Max= another bHLH Leucine zip protein Dimerize at homologous bHLH-LZ domains -leucine zipper domain allows dimerization with Max (another bHLH transcription factor -this allows activation of Myc -leads to transcription of target genes -via histone acetylation -via direct binding of myc-max complex to enhancer box (e-box) of target gene -Ebox element= CACGTG bHLH-LZ domain http://myc-cancer-gene.org/documents/MycReview.pdf E-box

bHLH/LZ Transcription Factor Structure Mad-max dimer represses transcription bHLH-LZ domain Max can also dimerize with Mad (another BHLH-LZ protein), inhibits transcription -recruits corepressors and deacetylases that induce compact chromatin structures and limit access of transcription factors to DNA -Myc must displace Mad to dimerize with Max

Normal Roles Myc has 600+ identified gene targets Main functions: www.myc-cancer-gene.org Main functions: Cell proliferation Apoptosis Hanahan and Weinberg, Cell 100:57-70 (2000) This number underestimated by at least 50% (only targets with E-boxes in their promotors) Myc has many target genes and therefore when activated can have numerous biological effects -Myc has been implicated in all 6 of the hallmarks of cancer

Determining Normal Roles: Mouse Models Homozygous null mutant mice lethal between 9.5 and 10.5 days of gestation indicates role in cell proliferation Over-expressed mutant mice lethal 10 days-8 weeks after birth indicates role in apoptosis http://biology.plosjournals.org/perlserv/?request=getdocument&doi=10.1371%2Fjournal.pbio.0020332 homozygous null mice lethal between 9.5 and 10.5 days of gestation -embryos small, retarded in development/growth -reason for lethality: lacked certain types of cell proliferation -lack hematopoiesis (formation of blood cellular components) and vasculogenesis (formation of vessels) -interesting: important in the vascularization of tumors -myc necessary for development past this time- understandable with wide variety of gene targets and widespread locations throughout the body Overexpressed mutant mice lethal between 10 days and 8 weeks after birth -lethality: stress activates normal apoptosis machinery, massive widespread apoptosis -cells kill themselves before more things go wrong -demonstrates role in apoptosis

Function: Cell Proliferation Stimulates G1-S transition c-Myc low c-Myc high c-Myc is present at very low levels in a resting cell Reception of growth factors results in elevated levels during cell proliferation Elevated levels of c-myc induce cyclins and cdk-activating phosphatases while suppressing cdk-inhibitors if this isn’t tightly regulated, could result in transition occurring too quickly, therefore not allowing time for DNA repair, and other crucial parts of cell checkpoints -overexpression can therefore induce genetic instability Growth Factors act at the Restriction Point

Function: Regulation of Apoptosis Over-expression in normal cells sensitizes the cell to a variety of apoptotic triggers -is not a direct trigger -protective measure -When myc is overexpressed, the cell interprets that as a stress, and it induces apoptosis through various pathways Can act on the Bcl-2 or ARF/p53 pathway to execute apoptosis -1. activate expression of ligand -5. inhibit induction of bcl-2, an anti-apoptotic gene

Function: A Paradox Ability to induce proliferation while at the same time being able to sensitize a cell to apoptosis is a “fail-safe” mechanism a cell that acquires deregulated (overexpressed) myc will also have an enhanced sensitivity to apoptosis so that it can kill itself should anything else go wrong -key point in understanding myc’s vast functions in the cell and understanding myc’s connection to cancer

c-Myc and Cancer: back to the mouse model Double mutant adult mice tumorigenesis Insertion of c-myc near various genetic enhancers tumorigenesis in a variety of tissues Review: homozygous null (cell proliferation) overexpressed (apoptosis) Double mutant adult mice (anti-apoptosis genes, ie BCl-2) **induced overexpression (promotor) -cannot commit apoptosis therefore uncontrolled proliferation and accumulation of DNA errors -go back two slides to point out -because myc can act at all these different points in all of these different pathways, any mutation in this pathway coupled with myc overexpression is going to result in apoptosis evasion and uncontrolled cell proliferation -myc says keep proliferating, and knocked out anti-apoptosis factors are no longer sensitive to increased levels of myc Insertion of c-myc near various genetic enhancers tumorigenesis in a variety of tissues -enhancers cause myc to be overexpressed -whatever tissue it is in, once coupled with another mutation, will cause tumorigenesis -will be seen in Burkitt’s lymphoma shortly Picture: B: lung metastisis histology C: liver tumor histology http://biology.plosjournals.org/perlserv/?request=getdocument&doi=10.1371%2Fjournal.pbio.0020332

c-Myc and Cancer Model: Unregulated c-Myc creates genetic instability a link between c-myc and cancer is well established, but the molecular cellular mechanisms of c-myc-mediated transformation is not fully known Model: unregulated c-Myc creates genetic instability -Acceleration through G1 and S phase results in bypassing of cell cycle checkpoints -creates genetic instability because DNA errors go unchecked, promotes cell proliferation with this instability -Normally results in apoptosis in a normal cell - the cell would respond to the stress and protect itself by committing suicide -Pre-cancer cell: paired with additional mutations that activate and overexpress anti-apoptotic signals (BCL- 2 or p53), sets the stage for a full-blown cancerous cell -cell won’t respond to increased levels of myc expression and therefore won’t kill itself if it needs to (because of genetic instability or other stresses)- safeguard against cancer is gone G1-S checkpoint

c-Myc and Cancer Over-expressed in 70% of all human cancers Translocated in 90% of all Burkitt’s lymphoma cases 90% of gynecological cancers 80% of breast cancers 70% of colon cancers Contributes to more than 70,000 cancer deaths annually in the U.S. Due to the extensiveness of myc’s target genes (600+), myc is a factor in a wide variety of cancers (goes back to being associated with the 6 hallmarks of cancer)- lung, prostate, skin, leukemias, lymphomas -linked to activation of a key regulator of metastasis, MTA1 (discovered in 2005) -first generation of drugs being developed to block Myc’s activation of MTA1 Myc-cancer-gene.org

c-Myc and Cancer: Burkitt’s Lymphoma c-Myc was first discovered in lymphoma patients Reciprocal translocation from chromosome 8 to chromosome 14 Translocation moved to a location close to immunoglobulin enhancers results in uncontrolled cell proliferation Still not sure how or why translocation occurs, but using mouse models Enhancers: -now myc is vigorously transcribed, and c-myc is overproduced in B-lymphocytes, -coupled with second mutation, get solid tumors http://www.ncbi.nlm.nih.gov/books/bv.fcgi?call=bv.View..ShowSection&rid=gnd.section.92 http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/B/BurkittLymphoma.html

c-Myc and Cancer: Burkitt’s Lymphoma Rare but extremely aggressive cancer Predominantly affects children in Southern Africa Solid tumor of B lymphocytes High tendency to spread to CNS, bone marrow, other blood elements type of a group of malignant diseases know as the Non-Hodgkin's Lymphomas (NHL). -very similar to leukemias Rare cancer -only about 300 new cases in the US each year -usually affects between the ages of 12-30 -one of the most aggressive forms of human cancer (can go from healthy to critcally ill in a matter of weeks) Predominantly affects children in Southern Africa Associated with infection by the Epstein-Barr virus -Same virus that cause mononucleosis B lymphocytes (make antibodies) Burkitt lymphoma is characterized by a starry-sky appearance. due to the presence of macrophages with relatively clear cytoplasm as seen in the inset light-colored histiocytes against the dark lymphoma cells. -lymphoma cells may accumulate in lymph nodes, resulting in swelling, often in neck and jaw -secondary tumors may form in jaw and neck most frequent, visible and characteristic clinical feature High rate of cure (70-80%) using chemotherapy IF caught early, and preventative measures against CNS invasion are esstential http://www.brown.edu/Courses/Digital_Path/systemic_path/female/burkitt.html http://tmcr.usuhs.mil/tmcr/chapter41/clinical.htm

Summary of c-Myc Transcription factor Paradox: Functions in cell proliferation and apoptosis Mouse model Knockout: lethal (lack of cell proliferation) Over-expressed: lethal (apoptosis) Oncogene Cancer: over-expression coupled with other mutations (i.e. tumor suppressors) leads to tumorigenesis in a variety of tissues

References Cited Beer S, Zetterberg A, Ihrie RA, et al:Developmental Context Determines Latency of MYC-Induced Tumorigenesis. PLoS Biology 2004, 2(11):e332. Burkitt lymphoma. Genes and Disease. http://www.ncbi.nlm.nih.gov/books/bv .fcgi?call=bv.View..ShowSection&rid=gnd.section.92 Burkitt’s lymphoma. 27 Aug. 2004. http://users.rcn.com/jkimball.ma.ultranet/ Biology Pages/B/BurkittLymphoma.html Cosel H: Scientists discover role for c-myc gene in tumor angiogenesis. Innovations Report 1 Jan. 2002. http://www.innovations-report.com/ specials/ printa.php?id=13299. Gardner L, Lee L, Dang C: The c-Myc Oncogenic Transcription factor. http://myc-cancer-gene.org/documents/ MycReview.pdf Pelengaris S, Rudolph B, Littlewood T: Action of Myc in vivo- proliferation and apoptosis. Current Opinion in Genetics and Development 2000, 10:100-105. Nilsson JA, Cleveland JL: Myc pathways provoking cell suicide and cancer. Oncogene 2003, 22:9007-9021. Weinberg, R: The Biology of Cancer. 2007. Werner L, Leon J, Eilers M: Contributions of Myc to tumorigenesis. Biochimica et Biophysica Acta 2001, 1602:61-71. Wyce M: Myc activates metastasis gene. Medical News Today 14 Sept. 2005. http://www.medicalnewstoday.com/medicalnews. php?newsid=30583.