Interaction between growth arrest-DNA damage protein 34 and Src kinase Lyn negatively regulates genotoxic apoptosis

Proc Natl Acad Sci U S A. 2001 Aug 28;98(18):10172-7. doi: 10.1073/pnas.191130798. Epub 2001 Aug 21.

Abstract

Genotoxic stresses activate intracellular signaling molecules, which lead to growth arrest, DNA repair, and/or apoptosis. Among these molecules are the growth arrest and DNA damage protein 34 (GADD34) and the Src-related protein tyrosine kinase Lyn. Here, we report that these two proteins physically and functionally interact to regulate DNA damage-induced apoptosis. Multiple isolates of GADD34 and the related murine protein MyD116 were identified as binding partners of Lyn in a yeast two-hybrid screen. The specific interaction was confirmed by in vitro association of GADD34 with glutathione S-transferase fusion proteins containing the Src Homology 3 (SH3) domain of Lyn, as well as coimmunoprecipitation of GADD34 and Lyn from mammalian cells. GADD34 was tyrosine-phosphorylated in vivo in a Lyn-dependent manner. Lyn efficiently phosphorylated affinity-purified GADD34 in vitro. Lyn negatively regulated the proapoptotic function of GADD34 in a kinase-dependent manner. Expression of wild-type, but not kinase-inactive, Lyn weakened promotion of apoptosis by GADD34 following treatment with methyl-methanesulfonate or ionizing radiation in HEK293 and HeLa cells. In contrast, pretreatment of cells with the Src-specific tyrosine kinase inhibitor PP1 strengthened promotion of apoptosis by GADD34. We propose that Lyn regulates the proapoptotic function of GADD34 by binding and phosphorylating it.

Publication types

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

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Antigens, Differentiation
  • Apoptosis / drug effects
  • Apoptosis / physiology*
  • Base Sequence
  • Cell Cycle Proteins
  • Cell Line
  • Chickens
  • DNA Damage
  • DNA Primers / genetics
  • HeLa Cells
  • Humans
  • Mice
  • Molecular Sequence Data
  • Mutagens / toxicity
  • Phosphorylation
  • Protein Phosphatase 1
  • Proteins / genetics
  • Proteins / metabolism*
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Sequence Homology, Amino Acid
  • Transfection
  • Two-Hybrid System Techniques
  • src-Family Kinases / genetics
  • src-Family Kinases / metabolism*

Substances

  • Antigens, Differentiation
  • Cell Cycle Proteins
  • DNA Primers
  • Mutagens
  • Proteins
  • Recombinant Proteins
  • lyn protein-tyrosine kinase
  • src-Family Kinases
  • PPP1R15A protein, human
  • Ppp1r15a protein, mouse
  • Protein Phosphatase 1