Novel Role Of Translationally Controlled Tumor Protein (TCTP) In The Low Dose Adaptive Response

Exposure to high doses ionizing radiation (X-rays or γ-rays) leads to manyadverse health effects, including cancer, mental retardation and heart disease,which have been fairly well elucidated. In contrast, the health risks of low doseradiation exposure remain ambiguous. As human population is increasinglysubjected to low dose and low dose-rate radiation from activities related todiagnostic radiology，nuclear technology, and the clean-up of sites that arecontaminated with radioactive material, estimating these health risks has been acontinuing endeavor of regulatory agencies for several decades.The “adaptive response” is a phenomenon induced by low dose/low LETradiation that protect cells and whole organisms against endogenous damage ordamage due to a subsequent challenge dose of radiation. Although low doseadaptive response-induced resistance to neoplastic transformation andmicronuclei formation have been suggested to correlate with increased ability torepair DNA damage, the molecular mechanism by which remains largelyunknown. To gain deeper insight into the mechanisms underlying low dose radiationeffects, we used amine-specific isobaric tags for relative and absolutequantitation (iTRAQ)-based approach to identify induced proteolytic events.The latter is a new ’Shotgun proteomics’ method that allows the direct analysisof a mixture of peptides and the generation of a global expression profile ofproteins. The procedure involves sequential two-dimensional liquidchromatography (LC), tandem mass spectrometry (MS/MS) and proteindatabase-searching algorithms. We initiated our studies by analyzing proteinsfrom mitochondria of AG1522normal human diploid fibroblasts exposed to0or10cGy (0.2cGy/h,48h) from cesium-137γ-rays. Cells were harvestedimmediately after the low dose exposure. Among many regulated proteins, theTranslationally Controlled Tumor Protein (TCTP) was found to be mostsensitive, having its level increased by2-fold in low dose-irradiated cells. TCTPis a highly conserved hydrophilic protein that is abundantly expressed in manyeukaryotes. It is regulated in response to a wide range of extracellular stimuliand stresses. It plays essential roles in cell growth, division, and programmedcell death, and has antioxidant properties. However, its role in radiation effectsis poorly understood.We have now confirmed up-regulation of TCTP in several normal humanfibroblast strains exposed to low dose γ-rays or other oxidizing agents (e.g.t-butyl hydroperoxide and hyperthermia). In high dose exposed cells, TCTP wasdown-regulated as a result of protein degradation. Analyses of proteins extractedfrom whole cells in3D cultures showed that TCTP levels are increased by3-foldin human cells exposed to doses as low as1cGy. Cell fraction studies showedthat the greatest increase (9.5-fold) occurred in nuclei of1cGy-irradiated cells.Strikingly, knockdown of TCTP expression by siRNA approach abolished the low dose γ-ray-induced adaptive responses against chromosomal damage, andsuggested a role for TCTP in repair of DNA damage and regulation of theradiation-induced G1checkpoint. In low dose-exposed si-tpt1/TCTP cells,knockdown of TCTP expression was associated with significant decrease in p53.In contrast, in high dose-irradiated cell cultures, down-regulation of TCTP didnot interfere with repair of chromosomal damage or p53expression. Ourongoing studies show that regulation of TCTP by low dose γ-rays is ATM-andDNA-PK-dependent, but p53-independent.Of particular significance, we have also found TCTP regulation to be highlysensitive in tissues of C3H/HeJ mice exposed to10cGy from γ-rays. Similar toour observations in skin fibroblasts maintained in culture, TCTP protein wasincreased (2-fold) in lung and brain tissue of mice exposed to10cGy andsacrificed3h later. No increase occurred in tissue isolated from mice exposed to400cGy.Collectively, our studies reveal the complexity of low dose radiation effects.They indicate that carcinogenic effects at low doses cannot be extrapolated fromeffects observed at high doses as currently adopted in radiation protectionprotocols. TCTP plays different functions in response to low-and high-dose IR,and TCTP may be a new player in DNA repair. In addition, we propose thatTCTP may be a suitable marker for the rapid evaluation of radiation-exposedindividuals.