The c-myc proto-oncogene is a transcription-activating protein which has been linked to malignancies including B cell leukemia and lymphomas, breast tumors, small cell lung carcinomas and brain tumors. We recently identified two genes whose expression increases in response to growth induction by c-myc; they include the mRNA cap binding protein (translation initiation factor elF-2alpha). In preliminary studies, elF-4E and elF-2alpha mRNA increased in parallel with c-myc in serum-stimulated cells. In addition, myc-transfected fibroblasts overexpressed both elF-4E and elF-2alpha. Finally, both elF- 4E and elF-2alpha were transcriptionally regulated in response to estradiol activation of an estrogen-regulated c-myc allele. This induction of elF-4E and elF-2alpha expression may be a uniquely important function for c-myc because both elF-4E and elF-2alpha can act individually as transforming genes. Although we found a correlation between c-myc and expression of elF-4E and elF-2alpha, the mechanism by which c-myc regulates these factors remains unclear. To examine myc regulation of elF-4E, we will clone and characterize promoter sequences of elF-4E. Our preliminary studies identified a DNA-binding site in the elF-2alpha promoter [TCCGCATGCG-NRF- 1] as a potential site of interaction with c-myc. Consequently, we will further characterize the response of this or other conserved sequences in the elF-4E and elF-2alpha promoters to c-myc using hypersensitivity site mapping, methylation interference, gel-shift assay and reporter gene-co-transfection.
The aim of these studies is to identify sequences that may directly interact with c-myc. In addition, the potential significance of myc interactions with elF-4E and elF-2alpha will be examined in correlative and functional studies. Breast cancers often carry amplified c-myc sequences; thus, we will examine breast cancers for correlations between increased elF-4E and elF- 2alpha mRNA and increased c-myc. Furthermore, one mechanism by which interferon inhibits cell growth is to increase the elF-2alpha kinase protein which inactivates elF-2alpha. Consequently, we will examine the potential for the elF-2alpha kinase to suppress cellular transformation by myc. These correlative and functional studies should assess the significance of the upregulation of elF-4E and elF-2alpha by c-myc as a potential target for therapeutic manipulations.
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