FREQUENT PROVIRUS INSERTIONAL MUTAGENESIS OF NOTCH1 IN THYMOMAS OF MMTV(D) MYC TRANSGENIC MICE SUGGESTS A COLLABORATION OF C-MYC AND NOTCH1FOR ONCOGENESIS/

The MMTV(D)/myc transgenic mice spontaneously develop oligoclonal CD4( +) CD8(+) T-cell tumors. Wt used provirus insertional mutagenesis in t hese mice to identify putative collaborators of c-myc. We found that N otch1 was mutated in a high proportion (52%) of these tumors. Provirus es were inserted upstream of the exon coding for the transmembrane dom ain and in both transcriptional orientations. These mutations led to h igh expression of truncated Notch1 RNAs and proteins (86-110 kD). In a ddition, many Notch1-rearranged tumors showed elevated levels of full- length Notch1 transcripts, whereas nearly all showed increased levels of full-length (330-kD) or close to full-length (280-kD) Notch1 protei ns. The 5' end of the truncated RNAs were determined for some tumors b y use of RT-PCR and 5' RACE techniques. Depending on the orientation o f the proviruses, viral LTR or cryptic promoters appeared to be utiliz ed, and coding potential began in most cases in the transmembrane doma in. Pulse-chase experiments revealed that the 330-kD Notch1 proteins w ere processed into 110- and 280-kD cleavage products. These results su ggest that Notch1 can be a frequent collaborator of c-myc for oncogene sis. furthermore, our data indicate that Notch1 alleles mutated by pro virus insertion can lead to increased expression of truncated and full -length (330/280-kD) Notch1 proteins, both being produced in a cleaved and uncleaved form.