Structure of mutant and wild-type MC29 v-myc alleles and biochemical properties of their protein products.
Proviral DNAs of three fibroblast-transforming MC29 deletion mutants (MC29-10A, MC29-10C, MC29-10H) with defects in hemopoietic cell transformation and tumor induction were molecularly cloned and their deletions were defined by nucleotide sequence analysis. The MC29-10C and MC29-10H v-myc alleles have identical internal deletions overlapping with a smaller one in the MC29-10A v-myc allele, and MC29-10H has an additional internal deletion in the partial gag complement. All deletions are in frame, and the deduced sequences of the mutant gag-myc hybrid proteins lack 56 (MC29-10A) or 120 (MC29-10C, MC29-10H) myc-specific and 44 gag-specific (MC29-10H) amino acid residues. The deleted v-myc nucleotide sequences correspond to the 3' end of exon 2 and the 5' end of exon 3 of the cellular c-myc gene including a region that encodes a high number of acidic amino acid residues. Based on these structural analyses, biochemical properties of mutant and wild-type gag-myc hybrid proteins were compared. Tryptic digests of all three mutant proteins lack a large myc-specific peptide that is present in digests of the wild-type protein and is extensively phosphorylated at serine and threonine residues. Concordantly, the sequence analyses predict that such a large tryptic peptide with putative phosphorylation sites at serine and threonine residues is present in the wild-type gag-myc protein but absent in all three mutant proteins due to the v-myc deletions. Chromatography of wild-type and mutant gag-myc proteins on DNA-cellulose revealed that their in vitro DNA affinities are indistinguishable from each other. Correspondingly, the sequence analyses predict that the carboxyl-terminal region rich in basic amino acid residues and with putative DNA affinity is conserved in wild-type and mutant gag-myc proteins. We conclude that the internal v-myc protein sequences defined by the deletions are necessary for hemopoietic cell transformation and complete phosphorylation, but dispensable for fibroblast transformation and in vitro DNA binding.
Otto-Warburg-Laboratorium, Max-Planck-Institut für Molekulare Genetik, Berlin, Dahlem, GFR.
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Bister
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Trachmann
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Jansen
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Schroeer
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Patschinsky
D000595:Amino Acid Sequence; D001483:Base Sequence; D002472:Cell Transformation, Viral; D002872:Chromosome Deletion; D003001:Cloning, Molecular; D004279:DNA, Viral / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D009154:Mutation; D009856:Oncogene Proteins, Viral / Q000235:genetics*; D009857:Oncogenes; D010766:Phosphorylation; D013329:Structure-Activity Relationship; D014760:Viral Fusion Proteins / Q000235:genetics
D004279:DNA, Viral; D004268:DNA-Binding Proteins; D009856:Oncogene Proteins, Viral; D014760:Viral Fusion Proteins