DeltaNp73 antisense activates PUMA and induces apoptosis in neuroblastoma cells.
The p73 gene codes for various different protein isoforms. They include proteins expressed under the control of the P1 promoter that contain a transactivation domain and are similar in function to p53 (TAp73 isoforms), as well as proteins regulated by the P2 promoter that lack this domain and function as dominant negative inhibitors of TAp73 and p53 (DeltaNp73 isoforms). Whereas TAp73 functions as a tumor suppressor with pro-apoptotic function, DeltaNp73 is likely to prevent the induction of apoptosis in tumor cells and to participate in oncogenesis. Here we used a loss-of-function strategy to assess the role of DeltaNp73 in SH-SY5Y neuroblastoma cells. An antisense oligonucleotide designed to target DeltaNp73 mRNA, but not TAp73, was used to effectively downregulate this transcript. DeltaNp73 downregulation was accompanied by increased levels of the pro-apoptotic BH3 family member PUMA at the mRNA and protein level, and by conformational activation of BAX which translocated to mitochondria. These DeltaNp73 antisense-mediated alterations led to the induction of apoptosis as detected by decreased cell viability, augmented DNA fragmentation and increased caspase-3 activity in cell lysates. Our results demonstrate the cytoprotective role of DeltaNp73 in neuroblastoma and suggest its use as a target for molecular intervention therapy.
Simões-Wüst
A P
AP
Molecular Oncology Laboratory, University Hospital Zurich, Haeldeliweg 4, CH-8044, Zurich, Switzerland.
Sigrist
B
B
Belyanskaya
L
L
Hopkins Donaldson
S
S
Stahel
R A
RA
Zangemeister-Wittke
U
U
D017209:Apoptosis / Q000235:genetics / Q000502:physiology*; D051017:Apoptosis Regulatory Proteins; D001932:Brain Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D045744:Cell Line, Tumor; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D015536:Down-Regulation; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing / Q000502:physiology; D016147:Genes, Tumor Suppressor / Q000502:physiology; D006801:Humans; D009019:Morphinans / Q000032:analysis; D009447:Neuroblastoma / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D016376:Oligonucleotides, Antisense; D020033:Protein Isoforms; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D000072160:Tumor Protein p73; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D025521:Tumor Suppressor Proteins; D051028:bcl-2-Associated X Protein
D051017:Apoptosis Regulatory Proteins; C494826:BAX protein, human; C438378:BBC3 protein, human; D004268:DNA-Binding Proteins; D009019:Morphinans; D009687:Nuclear Proteins; D016376:Oligonucleotides, Antisense; D020033:Protein Isoforms; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; C000606460:TP73 protein, human; D000072160:Tumor Protein p73; D016159:Tumor Suppressor Protein p53; D025521:Tumor Suppressor Proteins; D051028:bcl-2-Associated X Protein; C000606369:delta Np73 protein, human; C013757:Mr 1257 MS