Differential influence of etoposide on two caspase-2 mRNA isoforms in leukemic cells.
Etoposide (VP-16) is an anticancer agent that induces apoptosis in human leukemic cell lines such as U937 and HL60. We performed RNase protection assays, with two distinct cRNA panels covering most of caspase and BCL-2-related genes, using total RNA from cell lines exposed to various concentrations of the drug. Our results show that VP-16 down-regulates expression of most surveyed genes with the noticeable exception of casp-2S mRNA that is up regulated whereas casp-2L mRNA is decreased. Since these mRNAs are produced by the alternative splicing of exon 9, we devised a reverse transcriptase-polymerase chain reaction method using primers from exons 8 and 10 to demonstrate that VP-16 stimulates the production of exon 9-containing sequences, irrespective of active transcription. However, this effect is specific of the 3'-end of the CASP-2 gene since no difference in the relative amounts of the 5'-end of the mRNA species is detected. Nevertheless, the level of full-length casp-2L mRNA together with that of procaspase-2L protein, which is pro-apoptotic, are decreased under VP-16 treatment, suggesting that an early cell response to treatment by cytotoxic agents is to down-regulate expression of selected pro-apoptotic proteins.
Wotawa
Anne
A
INSERM U517, IFR 100, Faculty of Medicine, 7 boulevard Jeanne d'Arc, 21000, Dijon, France.
Solier
Stéphanie
S
Logette
Emmanuelle
E
Solary
Eric
E
Corcos
Laurent
L
D017398:Alternative Splicing / Q000187:drug effects*; D017209:Apoptosis / Q000187:drug effects; D051017:Apoptosis Regulatory Proteins; D053143:Caspase 2; D020169:Caspases / Q000096:biosynthesis / Q000235:genetics*; D003546:Cysteine Endopeptidases / Q000096:biosynthesis / Q000235:genetics; D003609:Dactinomycin / Q000494:pharmacology; D004790:Enzyme Induction / Q000187:drug effects; D004792:Enzyme Precursors / Q000096:biosynthesis / Q000235:genetics; D005047:Etoposide / Q000494:pharmacology*; D005091:Exons / Q000235:genetics; D015973:Gene Expression Regulation, Leukemic / Q000187:drug effects*; D019254:Genes, bcl-2; D018922:HL-60 Cells / Q000187:drug effects / Q000201:enzymology; D006801:Humans; D007948:Leukemia, Monocytic, Acute / Q000201:enzymology / Q000473:pathology*; D015473:Leukemia, Promyelocytic, Acute / Q000201:enzymology / Q000473:pathology*; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D009363:Neoplasm Proteins / Q000096:biosynthesis / Q000235:genetics*; D014176:Protein Biosynthesis; D011506:Proteins / Q000235:genetics; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis; D011519:Proto-Oncogenes; D012333:RNA, Messenger / Q000096:biosynthesis*; D012334:RNA, Neoplasm / Q000096:biosynthesis*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D020298:U937 Cells / Q000187:drug effects / Q000201:enzymology
D051017:Apoptosis Regulatory Proteins; C491073:BCL2L2 protein, human; D004792:Enzyme Precursors; D064549:Myeloid Cell Leukemia Sequence 1 Protein; D009363:Neoplasm Proteins; D011506:Proteins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D003609:Dactinomycin; D005047:Etoposide; D053143:Caspase 2; D020169:Caspases; D003546:Cysteine Endopeptidases