The mitochondrial protein hTID-1 partners with the caspase-cleaved adenomatous polyposis cell tumor suppressor to facilitate apoptosis.
OBJECTIVE
The adenomatous polyposis cell (APC) tumor suppressor is a multifunctional protein involved in cell migration, proliferation, differentiation, and apoptosis. Cleavage of APC and the subsequent release of an amino-terminal segment are necessary for a transcription-independent mechanism of APC-mediated apoptosis. The aim of the current study is to elucidate the mechanism by which the amino-terminus of APC contributes to the enhancement of apoptosis.


METHODS
Previous yeast 2-hybrid screens, using the armadillo repeat domain of APC as bait, identified hTID-1 as a potential binding partner. Coimmunoprecipitations, coimmunofluorescence, and binding assays confirm a direct interaction between caspase-cleaved APC and hTID-1 in vivo at the mitochondria. Overexpression and small interfering RNA (siRNA) knockdown studies were designed to determine the significance of this interaction.


RESULTS
These experiments have identified hTID-1 as a directly interacting protein partner of caspase-cleaved APC. hTID-1 is an apoptosis modulator: 2 of its known mitochondrial protein isoforms, 43-kilodaltons and 40-kilodaltons, have opposing effects in apoptosis. We demonstrate that the amino-terminal segment of APC interacts with both hTID-1 isoforms directly, although there is a stronger association with the apoptotic suppressor 40-kilodalton isoform in vitro. This interaction localizes to amino acids 202-512 of APC, a region including 2 of the 7 armadillo repeats. Overexpression of the 40-kilodalton hTID-1 isoform partially rescues cells from apoptosis mediated by APC 1-777, whereas siRNA knockdown of this hTID-1 isoform enhances apoptosis.


CONCLUSIONS
These data suggest that the amino-terminal segment of APC promotes cell sensitivity to apoptosis modulated through its binding to 40- and 43-kilodalton hTID-1 isoforms.
Qian
Jiang
J
Department of Molecular Virology, Immunology, and Medical Genetics, The Ohio State University College of Medicine, Columbus, Ohio 43210-2207, USA.
Perchiniak
Erin M
EM
Sun
Kristine
K
Groden
Joanna
J
D025601:Adenomatous Polyposis Coli Protein / Q000737:chemistry / Q000502:physiology*; D017209:Apoptosis; D053148:Caspase 3 / Q000502:physiology; D045744:Cell Line, Tumor; D050956:HSP40 Heat-Shock Proteins / Q000737:chemistry / Q000502:physiology*; D018840:HSP70 Heat-Shock Proteins / Q000502:physiology; D006801:Humans; D047468:Immunoprecipitation; D008928:Mitochondria / Q000378:metabolism; D020033:Protein Isoforms; D034741:RNA, Small Interfering / Q000235:genetics
D025601:Adenomatous Polyposis Coli Protein; C120664:DNAJA3 protein, human; D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; D020033:Protein Isoforms; D034741:RNA, Small Interfering; D053148:Caspase 3