Structural elements that regulate pp59c-fyn catalytic activity, transforming potential, and ability to associate with polyomavirus middle-T antigen

J Virol. 1991 Jan;65(1):170-9. doi: 10.1128/JVI.65.1.170-179.1991.

Abstract

Except for its unique amino-terminal region (residues 1 through 83), which possibly dictates substrate recognition, pp59c-fyn bears a high degree of homology with other members of the src family of tyrosine kinases. Here we show that the carboxy terminus of pp59c-fyn is necessary for stable middle-T-antigen association, that pp59c-fyn is normally phosphorylated on both serine and tyrosine residues, and that Tyr-531 and Tyr-420 are phosphorylation sites in vivo and in vitro, respectively. Analysis of a spontaneously generated mutant encoding a truncated form of pp59c-fyn and of variants specifically mutated at the Tyr-531 and Tyr-420 phosphorylation sites indicates that pp59c-fyn has regulatory elements analogous to those that have already been identified for other src-like tyrosine kinases. However, further examination of the pp59c-fyn variants suggests the likelihood of additional means by which its activities might be regulated. Although alteration of Tyr-531 to phenylalanine (531F) in pp59c-fyn results in a protein which is more active enzymatically that the wild type, the enhancement is much less than that for the analogous variant of pp60c-src. Furthermore, contrary to results of similar experiments on other src-like proto-oncogene products, 531F did not induce transformation of NIH 3T3 cells. Studies involving pp59c-fyn-pp60c-src chimeras in which the unique amino-terminal sequences (residues 1 through 83) of the two kinases were precisely interchanged implied that the inability of 531F to induce transformation is probably not caused by the absence of substrates for pp59c-fyn in NIH 3T3 cells but rather by the insufficient enhancement of pp59c-fyn kinase activity. It is therefore probable that the kinase and transforming activities of pp59c-fyn are repressed by additional regulatory elements possibly located in the amino-terminal half of the molecule.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Antigens, Polyomavirus Transforming*
  • Base Sequence
  • Cell Line
  • Chimera
  • Genetic Variation
  • Humans
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Oligonucleotide Probes
  • Peptide Mapping
  • Phosphopeptides / isolation & purification
  • Phosphorylation
  • Plasmids
  • Protein Binding
  • Protein-Tyrosine Kinases / genetics*
  • Proto-Oncogene Proteins / genetics*
  • Proto-Oncogene Proteins / metabolism
  • Proto-Oncogene Proteins c-fyn
  • Proto-Oncogenes*
  • Sequence Homology, Nucleic Acid

Substances

  • Antigens, Polyomavirus Transforming
  • Oligonucleotide Probes
  • Phosphopeptides
  • Proto-Oncogene Proteins
  • Protein-Tyrosine Kinases
  • FYN protein, human
  • Proto-Oncogene Proteins c-fyn