ABSTRACT
As components of the apoptosome, a caspase-activating complex, cytochrome c (Cyt c) and Apaf-1 are thought to play critical roles during apoptosis. Due to the obligate function of Cyt c in electron transport, its requirement for apoptosis in animals has been difficult to establish. We generated "knockin" mice expressing a mutant Cyt c (KA allele), which retains normal electron transfer function but fails to activate Apaf-1. Most KA/KA mice displayed embryonic or perinatal lethality caused by defects in the central nervous system, and surviving mice exhibited impaired lymphocyte homeostasis. Although fibroblasts from the KA/KA mice were resistant to apoptosis, their thymocytes were markedly more sensitive to death stimuli than Apaf-1(-/-) thymocytes. Upon treatment with gamma irradiation, procaspases were efficiently activated in apoptotic KA/KA thymocytes, but Apaf-1 oligomerization was not observed. These studies indicate the existence of a Cyt c- and apoptosome-independent but Apaf-1-dependent mechanism(s) for caspase activation.
Subject(s)
Apoptosis/physiology , Caspases/metabolism , Cytochromes c/genetics , Enzyme Activation/physiology , Proteins/metabolism , Animals , Apoptosis/radiation effects , Apoptotic Protease-Activating Factor 1 , Caspase 1 , Caspases/radiation effects , Electron Transport/physiology , Electron Transport/radiation effects , Enzyme Activation/radiation effects , Enzyme Precursors/metabolism , Enzyme Precursors/radiation effects , Gamma Rays , Lymphocyte Activation/genetics , Mice , Mice, Knockout , Mice, Mutant Strains , Mice, Transgenic , Nervous System Malformations/genetics , Nervous System Malformations/metabolism , T-Lymphocytes/metabolismABSTRACT
The phosphatidylinositol 3' kinase (PI3'K) pathway, which regulates cell survival, is antagonized by the PTEN tumor suppressor. The regulation of PTEN is unclear. A genetic screen of Drosophila gain-of-function mutants identified DJ-1 as a suppressor of PTEN function. In mammalian cells, DJ-1 underexpression results in decreased phosphorylation of PKB/Akt, while DJ-1 overexpression leads to hyperphosphorylation of PKB/Akt and increased cell survival. In primary breast cancer samples, DJ-1 expression correlates negatively with PTEN immunoreactivity and positively with PKB/Akt hyperphosphorylation. In 19/23 primary non-small cell lung carcinoma samples, DJ-1 expression was increased compared to paired nonneoplastic lung tissue, and correlated positively with relapse incidence. DJ-1 is thus a key negative regulator of PTEN that may be a useful prognostic marker for cancer.
Subject(s)
Drosophila Proteins/metabolism , Oncogene Proteins/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Phosphoric Monoester Hydrolases/metabolism , Protein Serine-Threonine Kinases/metabolism , Proto-Oncogene Proteins/metabolism , Animals , Animals, Genetically Modified , Biomarkers, Tumor , Breast Neoplasms/genetics , Breast Neoplasms/metabolism , Breast Neoplasms/pathology , Cell Death , Cell Line , Disease Progression , Drosophila Proteins/genetics , Drosophila melanogaster/physiology , Enzyme Activation , Female , Humans , Intracellular Signaling Peptides and Proteins , Lung Neoplasms/genetics , Lung Neoplasms/metabolism , Lung Neoplasms/pathology , Mice , Mice, Nude , Oncogene Proteins/genetics , PTEN Phosphohydrolase , Phosphoric Monoester Hydrolases/genetics , Phosphorylation , Photoreceptor Cells, Invertebrate/abnormalities , Photoreceptor Cells, Invertebrate/metabolism , Photoreceptor Cells, Invertebrate/ultrastructure , Protein Deglycase DJ-1 , Protein Serine-Threonine Kinases/genetics , Proto-Oncogene Proteins/genetics , Proto-Oncogene Proteins c-akt , Signal Transduction/physiologyABSTRACT
FKHRL1 (FOXO3a) and p53 are two potent stress-response regulators. Here we show that these two transcription factors exhibit "crosstalk" in vivo. In response to DNA damage, p53 activation led to FKHRL1 phosphorylation and subcellular localization change, which resulted in inhibition of FKHRL1 transcription activity. AKT was dispensable for p53-dependent suppression of FKHRL1. By contrast, serum- and glucocorticoid-inducible kinase 1 (SGK1) was significantly induced in a p53-dependent manner after DNA damage, and this induction was through extracellular signal-regulated kinase 1/2-mediated posttranslational regulation. Furthermore, inhibition of SGK1 expression by a small interfering RNA knockdown experiment significantly decreased FKHRL1 phosphorylation in response to DNA damage. Taken together, our observations reveal previously unrecognized crosstalk between p53 and FKHRL1. Moreover, our findings suggest a new pathway for understanding aging and the age dependency of human diseases governed by these two transcription factors.
