Analysis of the Max-binding protein MNT in human medulloblastomas.
Medulloblastomas (MBs) are the most frequent malignant brain tumors in children. 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 of a tumor-suppressor gene in this region. A candidate for this putative tumor-suppressor is the MNT gene, located at 17p13.3 and encoding a Max-interacting nuclear protein with transcriptional-repressor activity. In this study, 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 expression in all cases analyzed. Endogenous Mnt protein with an apparent molecular 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 electrophoretic mobility-shift assays. These experiments demonstrated that Mnt interacts with Max, and that this heterodimer binds DNA specifically, suggesting a functional bHLHZip domain of MB-derived Mnt. In support, single-strand conformation-polymorphism (SSCP) analyses revealed no mutation in the bHLHZip region. Deletion of the Mnt Sin3 interaction domain was shown to convert Mnt from an inhibitor of myc/ras-co-transformation into a molecule capable of cooperating with Ras in transformation. This region therefore was screened for mutation by SSCP: again, no alterations were found. These findings indicate that the MNT gene located at 17p13.3 is not likely to be involved in the molecular pathogenesis of MBs.
Sommer
A
A
Institut für Molekularbiologie, Medizinische Hochschule Hannover, Hanover, Germany.
Waha
A
A
Tonn
J
J
Sörensen
N
N
Hurlin
P J
PJ
Eisenman
R N
RN
Lüscher
B
B
Pietsch
T
T
D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D050976:Basic-Leucine Zipper Transcription Factors; D002528:Cerebellar Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D002874:Chromosome Mapping; D002886:Chromosomes, Human, Pair 17; D004268:DNA-Binding Proteins / Q000378:metabolism*; D016153:Genes, Suppressor; D016147:Genes, Tumor Suppressor; D018257:Helix-Loop-Helix Motifs; D006801:Humans; D019656:Loss of Heterozygosity; D008527:Medulloblastoma / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism*; D018807:Polymorphism, Single-Stranded Conformational; D012333:RNA, Messenger / Q000235:genetics; D012008:Recurrence; D012097:Repressor Proteins; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D050976:Basic-Leucine Zipper Transcription Factors; D004268:DNA-Binding Proteins; C499330:MAX protein, human; C106465:MNT protein, human; C067850:Myc associated factor X; D009687:Nuclear Proteins; D012333:RNA, Messenger; D012097:Repressor Proteins; D014157:Transcription Factors