Identification of c-MYC as a target of the APC pathway

Science

Volumn 281, Issue 5382, 1998, Pages 1509-1512

  He, Tong Chuan ^a   Sparks, Andrew B ^a   Rago, Carlo ^a   Hermeking, Heiko ^a   Zawel, Leigh ^a   Da Costa, Luis T ^a   Morin, Patrice J ^a   Vogelstein, Bert ^a   Kinzler, Kenneth W ^a  

^a NONE

Author keywords

[No Author keywords available]

Indexed keywords

BETA CATENIN;

ARTICLE; CHROMOSOME 5Q; COLON CANCER; COLON CARCINOGENESIS; GENE OVEREXPRESSION; GENE REPRESSION; HUMAN; ONCOGENE C MYC; PRIORITY JOURNAL; TRANSCRIPTION REGULATION; TUMOR SUPPRESSOR GENE;

ADENOMATOUS POLYPOSIS COLI PROTEIN; BETA CATENIN; BINDING SITES; CELL LINE; COLORECTAL NEOPLASMS; CYTOSKELETAL PROTEINS; GENE EXPRESSION REGULATION, NEOPLASTIC; GENES, APC; GENES, MYC; GENES, REPORTER; HT29 CELLS; HUMANS; MUTATION; PROMOTER REGIONS (GENETICS); PROTO-ONCOGENE PROTEINS C-MYC; SIGNAL TRANSDUCTION; TCF TRANSCRIPTION FACTORS; TRANS-ACTIVATORS; TRANSCRIPTION FACTORS; TRANSCRIPTION, GENETIC;

EID: 0032483439     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.281.5382.1509     Document Type: Article

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19. SAGE was performed as in (8) on mRNA from exponentially growing HT29-APC and HT29-β-Gal cells 9 hours after induction. A total of 55,233 and 59,752 tags were obtained from HT29-APC and HT29-β-Gal cells, respectively. Analysis of internal linker controls revealed a sequencing error rate of 0.065 per tag, corresponding to a sequencing error rate of 0.0067 per base. This was in good agreement with instrument specifications and previous estimates of SAGE tag errors based on analysis of the completed yeast genome (8). After correcting for sequencing mistakes, a total of 107,468 tags representing 51,622 and 55,846 from HT29-APC and HT29-β-Gal cells, respectively, were analyzed. These tags represented 14,346 unique transcripts, of which 7811 transcripts appeared at least twice.
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21. A low-basal activity reporter plasmid, pBV-Luc, was first constructed. The pDel-1, pDel-2, pDel-3, pDel-4, pFrag-A, pFrag-B, pFrag-C, pFrag-D, and pFrag-E reporters were constructed by cloning corresponding restriction fragments (illustrated in Fig. 2A) of human c-MYC promoter into pBV-Luc. Details of vector construction are available upon request.
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24. Two TBE-binding elements were identified in the region conferring APC and β-catenin responsiveness. TBE1 (CTTTGAT) was located 1156 bp up-stream of the TATA box at the P1 transcription start site and perfectly matched the consensus for Tcf-binding CTTTG(A/T)(A/T) [M. van de Wetering, M. Oosterwegel, D. Dooijes, H. Clevers, EMBO J. 10, 123 (1991); K. Giese, A. Amsterdam, R. Grosschedl, Genes Dev. 5, 2567 (1991)]. TBE2 was located 589 bp upstream of the TATA box and contained an inverted perfect match (ATCAAAG). A third Tcf-binding site was located 1400 bp up-stream of the TATA box but did not overlap with APC or β-catenin responsiveness.
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54. We thank V. Velculescu, L. Zhang, W. Zhou, and K. Polyak for SAGE advice and C. Geltinger for a genomic clone containing the c-MYC promoter. B.V. is an investigator of the Howard Hughes Medical Institute. Supported by NIH grants GM07309, CA62924, and CA57345. K.W.K. received research funding from Genzyme. Under a licensing agreement between the Johns Hopkins University and Genzyme, SAGE technology is licensed to Genzyme for commercial purposes, and K.W.K. and B.V. are entitled to a share of royalty received by the University from sales of the licensed technology. The SAGE technology is freely available to academia for research purposes. K.W.K. and B.V. are consultants to Genzyme. The University and researchers (K.W.K. and B.V.) own Genzyme stock, which is subject to certain restrictions under University policy. The terms of this arrangement are being managed by the University in accordance with its conflict of interest policies. This work is dedicated to the memory of J.-R. He and J.-X. Yang.