FUNCTIONAL-ANALYSIS OF THE AUG-INITIATED AND CUG-INITIATED FORMS OF THE C-MYC PROTEIN

Activation of the c-myc proto-oncogene by chromosomal translocation or proviral insertion frequently results in the separation of the c-myc coding region from its normal regulatory elements. Such rearrangements are often accompanied by loss or mutation of c-myc exon 1 sequences. These genetic alterations do not affect synthesis of the major c-myc p rotein, p64, which is initiated from the first AUG codon in exon 2. Ho wever they can result in mutation or loss of the CUG codon located in exon 1 that normally serves as an alternative translational initiation codon for synthesis of an N-terminally extended form of c-Myc (p67). It has been hypothesized that p67 is a functionally distinct form of c -Myc whose specific loss during c-myc rearrangements confers a selecti ve growth advantage. Here we describe experiments designed to test the functional properties of the two c-Myc protein forms. We introduced m utations within the translational initiation codons of a normal human c-myc cDNA that alter the pattern of Myc protein synthesis (p64 vs, p6 7). The functions of each of these proteins were experimentally addres sed using co-transformation and transcriptional activation assays. Bot h the p64 and p67 c-Myc proteins were independently able to collaborat e with bcr-abl in the transformation of Rat-1 fibroblasts. In addition , both the exon 1- and exon 2-initiated forms of the c-Myc protein sti mulated transcription of a Myc/Max-responsive reporter construct to a similar level. Given the apparent absence of functional differences be tween p64 and p67, we conclude that the basis for c-Myc oncogenic acti vation lies primarily in the overall deregulation of its expression an d not in alterations in the protein. The existence of the CUG translat ional initiator may reflect a mechanism for the continued synthesis of c-Myc protein under conditions where AUG initiation is inhibited.