Designed FHIT alleles establish that Fhit-induced apoptosis in cancer cells is limited by substrate binding

Proc Natl Acad Sci U S A. 2003 Feb 18;100(4):1592-7. doi: 10.1073/pnas.0437915100. Epub 2003 Feb 6.


The FHIT gene is inactivated early in the development of many human tumors, and Fhit-deficient mice have increased cancer incidence. Viral reexpression of Fhit kills Fhit-deficient cells by induction of apoptosis. Fhit, a member of branch 2 of the histidine-triad superfamily of nucleoside monophosphate hydrolases and transferases, is a diadenosine polyphosphate hydrolase, the active-site histidine of which is not required for tumor suppression. To provide a rigorous test of the hypothesis that Fhit function depends on forming a complex with substrates, we designed a series of alleles of Fhit intended to reduce substrate-binding andor hydrolytic rates, characterized these mutants biochemically, and then performed quantitative cell-death assays on cancer cells virally infected with each allele. The allele series covered defects as great as 100,000-fold in k(cat) and increases as large as 30-fold in K(M). Nonetheless, when mutant FHIT genes were expressed in two human cancer cell lines containing FHIT deletions, reductions in apoptotic activity correlated exclusively with K(M). Mutants with 2- and 7-fold increases in K(M) significantly reduced apoptotic indices, whereas the mutant with a 30-fold increase in K(M) retained little cellular function. These data indicate that the proapoptotic function of Fhit is limited by substrate binding and is unrelated to substrate hydrolysis.

Publication types

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

MeSH terms

  • Acid Anhydride Hydrolases*
  • Alleles*
  • Apoptosis / physiology*
  • Cell Line
  • Flow Cytometry
  • Humans
  • Kinetics
  • Mutation
  • Neoplasm Proteins / genetics*
  • Neoplasm Proteins / isolation & purification
  • Neoplasm Proteins / metabolism
  • Neoplasm Proteins / physiology
  • Neoplasms / genetics
  • Neoplasms / pathology*
  • Substrate Specificity
  • Tumor Cells, Cultured


  • Neoplasm Proteins
  • fragile histidine triad protein
  • Acid Anhydride Hydrolases