ABSTRACT
3-Aminobenzamide (3AB) is an inhibitor of poly (ADP-ribose) polymerase (PARP), an enzyme implicated in the maintenance of genomic integrity, which is activated in response to radiation-induced DNA strand breaks. cDNA macroarray membranes containing 1536 clones were used to characterize the gene expression profiles displayed by mouse BALB/3T3 fibroblasts (A31 cell line) in response to ionizing irradiation alone or in combination with 3AB. A31 cells in exponential growth were pre-treated with 3AB 4mM 1h before gamma-irradiation (4Gy), remaining in culture during 6h until harvesting time. A31 cells treated with 3AB alone presented a down-regulation in genes involved in protein processing and cell cycle control, while an up-regulation of genes involved in apoptosis and related to DNA/RNA synthesis and repair was verified. A31 cells irradiated with 4Gy displayed 41 genes differentially expressed, being detected a down-regulation of genes involved in protein processing and apoptosis, and genes controlling the cell cycle. Concomitantly, another set of genes for protein processing and related to DNA/RNA synthesis and repair were found to be up-regulated. A positive or negative interaction effect between 3AB and radiation was verified for 29 known genes. While the combined treatment induced a synergistic effect on the expression of LCK proto-oncogene and several genes related to protein synthesis/processing, a negative interaction effect was found for the expression of genes related to cytoskeleton and extracellular matrix assembly (SATB1 and Anexin III), cell cycle control (tyrosine kinase), and genes participating in DNA/RNA synthesis and repair (RNA helicase, FLAP endonuclease-1, DNA-3 glycosylase methyladenine, splicing factor SC35 and Soh1). The present data open the possibility to investigate the direct participation of specific genes, or gene products acting in concert in the mechanism underlying the cell response to radiation-induced DNA damage under the influence of PARP inhibitor.
Subject(s)
3T3 Cells/radiation effects , Benzamides/pharmacology , Enzyme Inhibitors/pharmacology , Gene Expression Regulation/radiation effects , Ribonucleoproteins , 3T3 Cells/drug effects , 3T3 Cells/physiology , Animals , Annexin A3/drug effects , Annexin A3/genetics , Annexin A3/radiation effects , DNA Damage/genetics , DNA Damage/radiation effects , Endodeoxyribonucleases/drug effects , Endodeoxyribonucleases/genetics , Endodeoxyribonucleases/radiation effects , Flap Endonucleases , Gamma Rays , Gene Expression Profiling , Gene Expression Regulation/drug effects , Lymphocyte Specific Protein Tyrosine Kinase p56(lck)/drug effects , Lymphocyte Specific Protein Tyrosine Kinase p56(lck)/genetics , Lymphocyte Specific Protein Tyrosine Kinase p56(lck)/radiation effects , Matrix Attachment Region Binding Proteins/drug effects , Matrix Attachment Region Binding Proteins/genetics , Matrix Attachment Region Binding Proteins/radiation effects , Mice , Nuclear Proteins/drug effects , Nuclear Proteins/genetics , Nuclear Proteins/radiation effects , Oligonucleotide Array Sequence Analysis , Poly(ADP-ribose) Polymerase Inhibitors , Proto-Oncogene Mas , RNA/biosynthesis , RNA/drug effects , RNA/radiation effects , RNA Helicases/drug effects , RNA Helicases/genetics , RNA Helicases/radiation effects , Serine-Arginine Splicing FactorsABSTRACT
The human T cell line Jurkat registers a sinusoidal extremely low frequency (ELF), 0.10 mT magnetic fields (MFs) at the level of the plasma membrane. In this study, the protein tyrosine phosphorylation (PY) of two membrane-associated proteins in Jurkat cells were examined following a short-term MFs exposure, the zeta chains and the Src kinases p56lck. These proteins are interesting to study since the earliest biochemical event upon T cell receptor (TcR) activation is PY of the zeta chains. These signalling chains in the TcR complex was assessed using Western blotting and the activation of the p56lck kinase was analysed by in vitro kinase assay. The MFs exposure of Jurkat for 5 min activated p56lck and resulted in PY of zeta. These findings are in line with earlier reports on how MFs exposure affects signal transduction in Jurkat.
