Analysis of the Max-binding protein MNT in human medulloblastomas

Medulloblastomas (MBs) are the most frequent malignant brain tumors in chil dren. The molecular pathogenesis of these tumors is still poorly understood . Microsatellite and restriction-fragment-length polymorphism studies have revealed allelic loss of genetic material on the short arm of chromosome 17 in the region 17p13 in approximately 50% of MBs, suggesting the presence o f a tumor-suppressor gene in this region. A candidate for this putative tum or-suppressor is the MNT gene, located at 17p13.3 and encoding a Max-intera cting nuclear protein with transcriptional-repressor activity. In this stud y, we analyzed MNT mRNA and protein expression in 44 MB samples, including 32 primary tumors, 3 recurrent tumors and 9 MB cell lines. Allelic loss at 17p13.3 was found in 49% of informative cases. RT-PCR showed MNT mRNA expre ssion in all cases analyzed. Endogenous Mnt protein with an apparent molecu lar weight of 72 to 74 kDa was detected in lysates from MB cell lines. The presence and functional integrity of Mnt in MBs were tested in electrophore tic mobility-shift assays. These experiments demonstrated that Mnt interact s with Max, and that this heterodimer binds DNA specifically, suggesting a functional bHLHZip domain of MB-derived Mnt. In support, single-strand conf ormation-polymorphism (SSCP) analyses revealed no mutation in the bHLHZip r egion. Deletion of the Mnt Sin3 interaction domain was shown to convert Mnt from an inhibitor of myc/rasco-transformation into a molecule capable of c ooperating with Ras in transformation. This region therefore was screened f or mutation by SSCP: again, no alterations were found. These findings indic ate that the MNT gene located at 17p13.3 is not likely to be involved in th e molecular pathogenesis of MBs. (C) 1999 Wiley-Liss, Inc.