DNA damage-induced Bcl-xL deamidation is mediated by NHE-1 antiport regulated intracellular pH.
The pro-survival protein Bcl-xL is critical for the resistance of tumour cells to DNA damage. We have previously demonstrated, using a mouse cancer model, that oncogenic tyrosine kinase inhibition of DNA damage-induced Bcl-xL deamidation tightly correlates with T cell transformation in vivo, although the pathway to Bcl-xL deamidation remains unknown and its functional consequences unclear. We show here that rBcl-xL deamidation generates an iso-Asp(52)/iso-Asp(66) species that is unable to sequester pro-apoptotic BH3-only proteins such as Bim and Puma. DNA damage in thymocytes results in increased expression of the NHE-1 Na/H antiport, an event both necessary and sufficient for subsequent intracellular alkalinisation, Bcl-xL deamidation, and apoptosis. In murine thymocytes and tumour cells expressing an oncogenic tyrosine kinase, this DNA damage-induced cascade is blocked. Enforced intracellular alkalinisation mimics the effects of DNA damage in murine tumour cells and human B-lineage chronic lymphocytic leukaemia cells, thereby causing Bcl-xL deamidation and increased apoptosis. Our results define a signalling pathway leading from DNA damage to up-regulation of the NHE-1 antiport, to intracellular alkalanisation to Bcl-xL deamidation, to apoptosis, representing the first example, to our knowledge, of how deamidation of internal asparagine residues can be regulated in a protein in vivo. Our findings also suggest novel approaches to cancer therapy.
Zhao
Rui
R
Laboratory of Lymphocyte Signalling and Development, The Babraham Institute, Babraham, Cambridge, United Kingdom.
Oxley
David
D
Smith
Trevor S
TS
Follows
George A
GA
Green
Anthony R
AR
Alexander
Denis R
DR
D000577:Amides / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D001483:Base Sequence; D027682:Cation Transport Proteins / Q000502:physiology*; D002460:Cell Line; D004249:DNA Damage; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008565:Membrane Proteins / Q000502:physiology*; D051379:Mice; D008969:Molecular Sequence Data; D015398:Signal Transduction; D000074082:Sodium-Hydrogen Exchanger 1; D017923:Sodium-Hydrogen Exchangers / Q000502:physiology*; D051020:bcl-X Protein / Q000235:genetics*
D000577:Amides; D027682:Cation Transport Proteins; D017931:DNA Primers; D008565:Membrane Proteins; C490997:Slc9a1 protein, mouse; D000074082:Sodium-Hydrogen Exchanger 1; D017923:Sodium-Hydrogen Exchangers; D051020:bcl-X Protein