The N-Myc oncoprotein is associated in vivo with the phosphoprotein Max(p20/22) in human neuroblastoma cells.
Proteins encoded by the proto-oncogenes c-myc, L-myc, and N-myc contain at their carboxy-terminus a tripartite segment comprising a basic DNA binding region (BR), a helix-loop-helix (HLH) and a leucine zipper motif (Zip), that are believed to be involved in DNA binding and protein-protein interaction. The N-Myc oncoprotein is overexpressed in certain human tumors that share neuroectodermal features due to amplification of the N-myc gene. Using a monoclonal antibody directed against an N-terminal epitope of the N-Myc protein in immunoprecipitations performed with extracts of neuroblastoma cells, two nuclear phosphoprotein, p20/22, forming a hetero-oligomeric complex with N-Myc are identified. Both proteins are phosphorylated by casein kinase II in vitro. By partial proteolytic maps we show that p20 and p22 are structurally related to each other and that p20 is identical with Max, a recently described in vitro binding partner of myc proteins. Time course experiments show the presence of the complex in cellular extracts immunoprecipitated within a 5 min interval after the preparation of the cell extract. While the expression of N-myc is restricted, expression of both Max(p20/22) and the murine homolog Myn(p20/22) was observed in cells of diverse human and murine embryonal lineages as detected by heterologous complex formation. By introduction of expression vectors containing the wild type N-myc gene or N-myc genes with in frame deletions or point mutations into recipient cells and subsequent immunoprecipitation of the resulting N-Myc proteins we show that the HLH-Zip region is essential to the formation of the N-Myc-p20/22 complex.
Institut für Experimentelle Pathologie, Deutsches Krebsforschungszentrum, Heidelberg, Germany.
A
A
Wenzel
C
C
Cziepluch
U
U
Hamann
J
J
Schürmann
M
M
Schwab
D000818:Animals; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D050976:Basic-Leucine Zipper Transcription Factors; D047388:Casein Kinases; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006801:Humans; D016350:Leucine Zippers; D051379:Mice; D016296:Mutagenesis; D009447:Neuroblastoma / Q000378:metabolism* / Q000473:pathology; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011233:Precipitin Tests; D014176:Protein Biosynthesis; D011494:Protein Kinases / Q000378:metabolism; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism*; D014157:Transcription Factors; 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; C067850:Myc associated factor X; D010750:Phosphoproteins; D016271:Proto-Oncogene Proteins c-myc; D014157:Transcription Factors; C068422:Max protein, mouse; D011494:Protein Kinases; D047388:Casein Kinases