The effects of low-dose ionizing radiation in the activated rat basophilic leukemia (RBL-2H3) mast cells.

Mast cells play important roles in many biological responses, such as those during allergic diseases and inflammatory disorders. Although laser and UV irradiation have immunosuppressive effects on inflammatory diseases by suppressing mast cells, little is known about the effects of γ-ionizing radiation on mast cells. In this study, we investigated the effects of γ-ionizing radiation on RBL-2H3 cells, a convenient model system for studying regulated secretion by mast cells. Low-dose radiation (<0.1 gray (Gy)) did not induce cell death, but high-dose radiation (>0.5 Gy) induced apoptosis. Low-dose ionizing radiation significantly suppressed the release of mediators (histamine, ß-hexosaminidase, IL-4, and tumor necrosis factor-α) from immunoglobulin E (IgE)-sensitized RBL-2H3 cells. To determine the mechanism of mediator release inhibition by ionizing radiation, we examined the activation of intracellular signaling molecules such as Lyn, Syk, phospholipase Cγ, PKCs, and MAPK, and intracellular free calcium concentrations ([Ca(2+)](i)). The phosphorylation of signaling molecules following stimulation of high-affinity IgE receptor I (FcεRI) was specifically inhibited by low-dose ionizing radiation (0.01 Gy). These results were due to the suppression of FcεRI expression by the low-dose ionizing radiation. Therefore, low-dose ionizing radiation (0.01 Gy) may function as a novel inhibitor of mast cell activation.

TOPORS modulates H2AX discriminating genotoxic stresses.

H2AX plays an important role in chromatin reorganization implicated in DNA repair and apoptosis under various DNA damaging conditions. In this study, the interaction between TOPORS (topoisomerase I-binding protein) and H2AX was verified using mammalian cell extracts exposed to diverse DNA damaging stresses such as ionizing radiation, doxorubicin, camptothecin, and hydrogen peroxide. In vitro assays for ubiquitination revealed that TOPORS functions as a novel E3 ligase for H2AX ubiquitination. TOPORS was found to be dissociated from H2AX proteins when cells were exposed to oxidative stress, but not replication-inducing DNA damaging stress. The protein stability of H2AX was decreased when TOPORS was ectopically expressed in cells, and oxidative stresses such as hydrogen peroxide and ionizing radiation induced recovery of the H2AX protein level. Therefore, these biochemical data suggest that TOPORS plays a key role in the turnover of H2AX protein, discriminating the type of DNA damaging stress.

Differential expression of immune-associated cancer regulatory genes in low- versus high-dose-rate irradiated AKR/J mice.

AKR/J mice carrying leukemia viral inserts develop thymic lymphoma. Recently, we demonstrated that the incidence of thymic lymphoma was decreased when these mice were raised in a low-dose-rate γ-irradiation facility. In contrast, mice irradiated at a high-dose rate developed severe thymic lymphoma and died much earlier. To understand the genetic changes occurred by low- versus high-dose-rate γ-irradiation whole genome microarray was performed. Both groups of mice demonstrated up-regulation of Ifng, Igbp1, and IL7 in their thymuses, however, mice exposed to high-dose-rate γ-irradiation exhibited marked down-regulation of Sp3, Il15, Traf6, IL2ra, Pik3r1, and Hells. In contrast, low-dose-rate irradiated mice demonstrated up-regulation of Il15 and Jag2. These gene expression profiles imply the impaired immune signaling pathways by high-dose-rate γ-irradiation while the facilitation of anti-tumor immune responses by low-dose-rate γ-irradiation. Therefore, our data delineate common and distinct immune-associated pathways downstream of low- versus high-dose-rate irradiation in the process of cancer progression in AKR/J mice.

Phosphorylation of CLK2 at serine 34 and threonine 127 by AKT controls cell survival after ionizing radiation.

AKT phosphorylates components of the intrinsic cell survival machinery and promotes survival to various stimuli. In the present study, we identified CDC-like kinase 2 (CLK2) as a new substrate of AKT activation and elucidated its role in cell survival to ionizing radiation. AKT directly binds to and phosphorylates CLK2 on serine 34 and threonine 127, in vitro and in vivo. CLK2 phosphorylation was detected in HeLa cells overexpressing active AKT. In addition, we demonstrated that ionizing radiation induces CLK2 phosphorylation via AKT activation. In contrast, the suppression of endogenous AKT expression by siRNA inhibited CLK2 phosphorylation in response to 2 gray of γ-ray or insulin. Furthermore, we examined the effect of CLK2 on the survival of irradiated CCD-18Lu cells overexpressing Myc-CLK2. CLK2 overexpression significantly increased cell growth and inhibited cell death induced by 2 gray. The role of CLK2 in cell survival to ionizing radiation was dependent on the phosphorylation of serine 34 and threonine 127. Our results suggest that AKT activation controls cell survival to ionizing radiation by phosphorylating CLK2, revealing an important regulatory mechanism required for promoting cell survival.

Genome-wide analysis of low-dose irradiated male Drosophila melanogaster with extended longevity.

Ionizing radiation generates oxidative stress, which is thought to be a major cause of aging. Although living organisms are constantly exposed to low levels of radiation, most studies examining the effect of radiation have focused on accelerated aging and diminished life span that result from high-dose radiation. On the other hand, several studies have suggested that low-dose radiation enhances the longevity of Drosophila melanogaster. Therefore, investigation of the biological effects of low-dose radiation could contribute to a more comprehensive understanding of the aging process. In this study, microarray and quantitative real time-PCR were used to measure genome-wide changes in transcript levels in low-dose irradiated fruit flies that showed enhanced longevity. In response to radiation, approximately 13% of the genome exhibited changes in gene expression, and a number of aging-related genes were significantly regulated. These data were compared with quantitative trait loci affecting life-span to identify candidate genes involved in enhanced longevity induced by low-dose radiation. This genome-wide survey revealed novel information about changes in transcript levels in low-dose irradiated flies and identified 39 new candidate genes for molecular markers of extended longevity induced by ionizing radiation. In addition, this study also suggests a mechanism by which low-dose radiation extends longevity.

Alteration of cytokine profiles in mice exposed to chronic low-dose ionizing radiation.

While a high-dose of ionizing radiation is generally harmful and causes damage to living organisms, a low-dose of radiation has been shown to be beneficial in a variety of animal models. To understand the basis for the effect of low-dose radiation in vivo, we examined the cellular and immunological changes evoked in mice exposed to low-dose radiation at very low (0.7mGy/h) and low (3.95mGy/h) dose rate for the total dose of 0.2 and 2Gy, respectively. Mice exposed to low-dose radiation, either at very low- or low-dose rate, demonstrated normal range of body weight and complete blood counts. Likewise, the number and percentage of peripheral lymphocyte populations, CD4(+) T, CD8(+) T, B, or NK cells, stayed unchanged following irradiation. Nonetheless, the sera from these mice exhibited elevated levels of IL-3, IL-4, leptin, MCP-1, MCP-5, MIP-1alpha, thrombopoietin, and VEGF along with slight reduction of IL-12p70, IL-13, IL-17, and IFN-gamma. This pattern of cytokine release suggests the stimulation of innate immunity facilitating myeloid differentiation and activation while suppressing pro-inflammatory responses and promoting differentiation of naïve T cells into T-helper 2, not T-helper 1, types. Collectively, our data highlight the subtle changes of cytokine milieu by chronic low-dose gamma-radiation, which may be associated with the functional benefits observed in various experimental models.

Radiation exposure and cancer incidence in a cohort of nuclear power industry workers in the Republic of Korea, 1992-2005.

This study examines for the first time cancer incidence between radiation and non-radiation workers in nuclear power facilities in the Republic of Korea. Radiation workers were defined as persons who were issued with a dosimeter at nuclear power facilities, until 2005. All analyses were conducted on male workers only (in total 16,236 individuals) because of the sparseness of females. Statistical analyses were carried out using the standardized incidence ratio (SIR), to compare the cancer risks of radiation and non-radiation workers with those of the general population, and the chi(2) trend test was used to investigate any increase in cancer rates with dose. Poisson regression was also used to estimate the rate ratio (RR) and the excess relative risk (ERR) after considering the confounding effect due to smoking. During 1992-2005, 99 cancer cases in 63,503 person-years were observed among 8,429 radiation workers, while 104 cancer cases were observed in 48,301 person-years among 7,807 non-radiation workers. When compared with the site- and age-specific cancer rates for the male population of Korea, the SIR for all cancers combined was 1.07 [95% confidence interval (CI) 0.87-1.30] for radiation workers, and 0.88 (95% CI 0.72-1.06) for non-radiation workers, respectively. The RR for radiation workers compared with non-radiation workers was 1.18 (95% CI 0.89-1.58) for all cancers combined. The SIRs for thyroid cancer were noticeably high for both radiation and non-radiation workers, possibly due to the screening effect, but analysis of the RR showed that there was no statistically significant difference in thyroid cancer incidence rates between the two groups. For lung cancer, radiation workers showed a higher incidence rate as compared to non-radiation workers, with the RR being 3.48 (95% CI 1.19-11.48). A chi(2) trend test showed that there was no evidence for an increase in cancer rate with increasing cumulative dose for all cancers combined (p = 0.5108). The ERR per Sievert was estimated to be 1.69 (95% CI -2.07 to 8.21) for all cancers combined assuming a 10 years lag time. Consequently, a significant excess of cancer incidence among radiation workers in the nuclear power industry in Korea was not observed. Further follow-up and an expansion of the cohort are needed to overcome the lack of statistical power in the study.

Identification of ionizing radiation-responsive microRNAs in the IM9 human B lymphoblastic cell line.

Ionizing radiation (IR) disrupts cellular homeostasis through multiple mechanisms including changes of the expression profile of genes. Although microRNAs (miRNAs), small single-stranded RNAs, have recently been recognized as important post-transcriptional regulators of gene expression, it is not well investigated if miRNAs function in the cellular response to radiation. Therefore, we determined if IR induces changes in the expression profiles of miRNAs and used this approach to identify IR-responsive miRNAs. To monitor the profiles of miRNAs, microarray analysis was conducted with irradiated IM9 human lymphoblastic cells. The expression levels of specific miRNAs were confirmed by quantitative real-time PCR (qRT-PCR) and statistically analyzed. Finally, the target mRNAs of some IR-responsive miRNAs were predicted with two different prediction programs. IR-exposed human lymphoblastic cells underwent cell cycle arrest and apoptosis. Apoptosis was more significantly increased at a higher radiation dose. There were 73 and 33 human miRNAs in 1 and 10 Gy-irradiated cells, respectively that showed expression level changes of >2-fold. By qRT-PCR analysis, it was revealed that the patterns of miRNA expression were similar to those observed in the microarray data, although the quantitative expression levels were discordant. Prediction of genes targeted by IR-responsive miRNA yielded several genes, many of which are involved in the regulation of apoptosis, the cell cycle, and DNA repair. The expression profiles of miRNAs in the IM9 human B lymphoblastic cells are strongly affected by IR and these changes may be involved in the regulation of cellular response to IR.

Identification of specific microRNAs responding to low and high dose gamma-irradiation in the human lymphoblast line IM9.

MicroRNAs (miRNAs) are short single-stranded RNA molecules that regulate the stability or translational efficiency of target messenger RNAs. Specific miRNAs are required for strict tissue- and developmental stage-specific expression. These miRNAs have roles in many human tumor malignancies and their expression is specifically regulated on each stage of oncogenic process. Therefore, miRNA expression profiling can be used as a new class of biomarker that indicates the development of cancer. Many recent studies indicated that cell exposure to ionizing radiation also induces various physiological responses including DNA repair, cell cycle arrest, cell death and differentiation. In addition, some studies suggest that exposure to low dose radiation induces a favorable effect on cells. However, the functions of miRNAs related to the response of irradiated cells have not been well studied, especially after low dose radiation. In this study, expression profiles of miRNAs isolated from irradiated cells at low and high dose radiation were analyzed with microarrays, and these data were validated using quantitative RT-PCR. Here, we describe specific miRNAs that are expressed in a dose-dependent manner that serve as new markers of irradiated immune cells.

Specific proteolysis of the A-kinase-anchoring protein 149 at the Asp582 residue by caspases during apoptosis.

A-kinase-anchoring protein 149 (AKAP149) is a member of a structurally diverse, though functionally similar anchoring protein family and is localized to the outer membrane of mitochondria and in the endoplasmic reticulum-nuclear envelope network. AKAP149 plays an important role in controlling the subcellular localization and temporal specificity of protein phosphorylation and mRNA metabolism by tethering kinases and phosphatases, such as protein kinase A and type I protein phosphatase, through its N-terminal protein-binding motifs and mRNAs via its C-terminal RNA-binding motifs. It is well recognized that caspases play a central role in transducing and amplifying the intracellular death signal and that apoptosis is executed as a consequence of caspase-mediated cleavage of multiple cellular substrates. The identification of novel death substrates and elucidation of the consequences of their proteolytic cleavages by caspases are therefore crucial for our understanding of cell death and other biological processes. Herein, we demonstrated that AKAP149 is a direct substrate of active caspase-3, -8 -and -10 in vitro and in vivo. 35S-labeled full-length AKAP149 was completely cleaved in vitro by active caspase-3, -8 and -10 into two fragments of approximately 105 and 45 kDa, while caspase-2 cleaved it partially and caspase-1 did not cleave it at all. AKAP149 was also cleaved by caspases during Fas- and staurosporine-induced apoptosis in Jurkat T and HeLa cells, which were blocked by specific inhibitors of caspase-3 and -8. The specific cleavage site for these caspases was mapped in vitro and in vivo to Asp582 at AKAP149, which is located between the protein kinase A regulatory subunit anchoring and KH RNA-binding domains. In addition, HeLa cells transiently overexpressing AKAP149 D582E mutant were resistant to staurosporine-induced HeLa cell apoptosis. Taken together, these data suggest that AKAP149 activity may be deregulated by caspase-dependent proteolysis during apoptotic cell death and may provide useful information for elucidating the apoptosis signaling pathways in detail.

Comprehensive analysis of time- and dose-dependent patterns of gene expression in a human mesenchymal stem cell line exposed to low-dose ionizing radiation.

We focused on the transcriptional responses induced by low and very low doses of ionizing radiation with time effect. Regardless of their importance only a few limited studies have been done. Here we applied a large-scale gene transcript profile to elucidate the genes and biological pathways. Immortalized human mesenchymal stem cells were irradiated with 0.01, 0.05, 0.2 and 1 Gy of gamma radiation and total RNA was extracted from each cell line at 1, 4, 12 and 48 h after exposure. The essential transcriptional responses were identified according to dose and time. A total of 6,016 genes showed altered expression patterns at more than one time point or dose level among the investigated 10,800 genes. Genes that showed dose-dependent expression responses were involved in signal transduction, regulation of transcription, proteolysis, peptidolysis and metabolism. Those that showed time-dependent responses were divided into two distinct groups: the up-and-down group was associated with 'cellular defense mechanisms' such as apoptosis, cell adhesion, stress response and immune response and the down-and-up group with 'fundamental cellular processes' such as DNA replication, mitosis, RNA splicing, DNA repair and translation initiation. Genes showing both dose-and time-dependent responses exhibited a mixture of both features. A highly non-linear relationship between the IR dose and the transcriptional relative response was obtained from the dose-dependent group. The time-dependent group also exhibited a non-linear relationship as the complex effect group did. Some of the early-reactive-phase (1-4 h) genes showed a differential expression response to 0.01, 0.05 and 0.2 Gy but were unresponsive to 1 Gy. Some of the late-recovery-phase (12-48 h) genes showed a differential expression to 1 Gy but were relatively unresponsive to other doses. We further characterized the gene expression patterns that could be implicated in the molecular mechanism of the cellular responses to low and very low-dose irradiation.

Low-dose radiation stimulates the proliferation of normal human lung fibroblasts via a transient activation of Raf and Akt.

The biological effects of low-dose radiation have been investigated and debated for more than a century, but its cellular effects and regulatory mechanisms remain poorly understood. This study shows the human cellular responses to low-dose radiation in CCD-18 Lu cells, which are derived from normal human lung fibroblasts. We examined a colony-forming assay for cell survival by ionizing radiation. Live cell counting and cell cycle analysis were measured for cell proliferation and cell cycle progression following low-dose irradiation. We examined Raf and Akt phosphorylation to determine the proliferation mechanism resulting from low-dose radiation. We also observed that p53 and p21 were related to cell cycle response. We found that 0.05 Gy of ionizing radiation enhanced cell proliferation and did not change the progression of the cell cycle. In addition, 0.05 Gy of ionizing radiation transiently activated Raf and Akt, but did not change phospho-p53, p53 and p21 in CCD-18 Lu cells. However, 2 Gy of ionizing radiation induced cell cycle arrest, phosphorylation of p53, and expression of p53 and p21. The phosphorylation of Raf and Akt proteins induced by 0.05 Gy of ionizing radiation was abolished by pre-treatment with an EGFR inhibitor, AG1478, or a PI3k inhibitor, LY294002. Cell proliferation stimulated by 0.05 Gy of ionizing radiation was blocked by the suppression of Raf and Akt phosphorylation with these inhibitors. These results suggest that 0.05 Gy of ionizing radiation stimulates cell proliferation through the transient activation of Raf and Akt in CCD-18 Lu cells.

Low-dose of ionizing radiation enhances cell proliferation via transient ERK1/2 and p38 activation in normal human lung fibroblasts.

This study shows the human cellular responses and the mechanism of low-dose ionizing radiation in CCD 18 Lu cells, which are derived from normal human lung fibroblasts. Cell proliferation and viability assay were measured for the cells following gamma-irradiation using trypan blue, BrdU incorporation, and Wst-1 assay. We also examined genotoxicity using a micronuclei formation assay. The activation of the MAPKs pathway was determined by Western blot analysis, and the siRNA system was used to inhibit the expression of ERK1/2 and p38. We found that 0.05 Gy of ionizing radiation stimulated cell proliferation and did not change Micronuclei frequencies. In addition, 0.05 Gy of ionizing radiation activated ERK1/2 and p38, but did not activate JNK1/2 in cells. A specific ERK1/2 inhibitor, U0126, decreased the phosphorylation of ERK1/2 proteins induced by 0.05 Gy of ionizing radiation, and a similar suppressive effect was observed with a p38 inhibitor, PD169316. Suppression of ERK1/2 and p38 phosphorylation with these inhibitors decreased cell proliferation, which was stimulated by 0.05 Gy of ionizing radiation. Furthermore, downregulation of ERK1/2 and p38 expression using siRNA blocked the cell proliferation that had been increased by 0.05 Gy of ionizing radiation. These results suggest that 0.05 Gy of ionizing radiation enhances cell proliferation through the activation of ERK1/2 and p38 in normal human lung fibroblasts.

Site-Specific Phosphorylation of Ikaros Induced by Low-Dose Ionizing Radiation Regulates Cell Cycle Progression of B Lymphoblast Through CK2 and AKT Activation.

PURPOSE: To determine how low-dose ionizing radiation (LDIR) regulates B lympho-proliferation and its molecular mechanism related with Ikaros, transcription factor. METHODS AND MATERIALS: Splenocytes and IM-9 cells were uniformly irradiated with various doses of a (137)Cs γ-source, and cell proliferation was analyzed. To determine the LDIR-specific phosphorylation of Ikaros, immunoprecipitation and Western blot analysis were performed. To investigate the physiologic function of LDIR-mediatied Ikaros phosphorylation, Ikaros mutants at phosphorylation sites were generated, and cell cycle analysis was performed. RESULTS: First, we found that LDIR enhances B lymphoblast proliferation in an Ikaros-dependent manner. Moreover, we found that LDIR elevates the phosphorylation level of Ikaros protein. Interestingly, we showed that CK2 and AKT are involved in LDIR-induced Ikaros phosphorylation and capable of regulating DNA binding activity of Ikaros via specific phosphorylation. Finally, we identified LDIR-specific Ikaros phosphorylation sites at S391/S393 and showed that the Ikaros phosphorylations at these sites control Ikaros's ability to regulate G1/S cell cycle progression. CONCLUSION: Low-dose ionizing radiation specifically phosphorylates Ikaros protein at Ser 391/393 residues to regulate cell cycle progression in B lymphoblast.

Immunomodulatory function of murine NK cell activity by alginate.

The in vivo immunomodulatory function of the activity of murine natural killer (NK) cells induced by high mannuronic acid-containing alginate (HMA) was examined. HMA was injected i.p at doses of 25 and 100 mg/kg. The NK activity was 3 times higher with 100 mg/kg HMA than the baseline. In addition, in vitro studies of splenocytes cultured with HMA for 20 h showed a significant increase in NK activity at E:T ratio of 100:1; a 160% and 210% increase at 10 and 100 microg/mL, respectively. There was a six fold increase in interferon-gamma production in a postculture of splenocytes with 100 microg/mL HMA. HMA had no suppressive effects on the lymphocyte function in the presence or absence of mitogens. This suggests that HMA is useful in cancer immunotherapy.

Role of AKT and ERK pathways in controlling sensitivity to ionizing radiation and adaptive response induced by low-dose radiation in human immune cells.

Despite many studies of the effect of ionizing radiation, biological mechanisms of action might differ greatly depend on dose, dose rate, and cell type. This study was performed to explore the effects of low- and high-dose radiation in human immune cell lines. We examined cell sensitivity after irradiation with 0.05, 0.1, or 2Gy in two normal cell lines and three tumor cell lines. Low-dose radiation of 0.05 and 0.1Gy had no effect on cell survival in any tested cell line, with the exception of IM-9 cells, whose viability was transiently increased. However, IM-9 and C1R-sB7 cells were very sensitive to high-dose radiation-induced cell death, whereas Jurkat and JM1 cells showed moderate sensitivity, and THP-1 cells were completely resistant. This radiosensitivity was correlated with basal AKT activation, which is induced by phosphorylation. In radiosensitive IM-9 cells, priming with chronic low-dose irradiation blocked cell death induced by high-dose radiation challenge via inhibition of caspase activation and PARP cleavage. AKT phosphorylation was not altered in IM-9 cells, but ERK phosphorylation was greatly elevated immediately after chronic low-dose irradiation. Taken together, our results suggest that the different responses of normal and tumor cells to low-dose and high-dose radiation depend on AKT activation, which is regulated by protein phosphatase 2 (PP2A). In radiosensitive normal cells lacking basal AKT activity, chronic low-dose radiation increases activation of the ERK pathway, which plays an important role in the adaptive response to radiation, providing a very important insight into understanding the effects of ionizing radiation on health.

The Inhibitory Effects of Low-Dose Ionizing Radiation in IgE-Mediated Allergic Responses.

Ionizing radiation has different biological effects according to dose and dose rate. In particular, the biological effect of low-dose radiation is unclear. Low-dose whole-body gamma irradiation activates immune responses in several ways. However, the effects and mechanism of low-dose radiation on allergic responses remain poorly understood. Previously, we reported that low-dose ionizing radiation inhibits mediator release in IgE-mediated RBL-2H3 mast cell activation. In this study, to have any physiological relevance, we investigated whether low-dose radiation inhibits allergic responses in activated human mast cells (HMC-1(5C6) and LAD2 cells), mouse models of passive cutaneous anaphylaxis and the late-phase cutaneous response. High-dose radiation induced cell death, but low-dose ionizing radiation of <0.5 Gy did not induce mast cell death. Low-dose ionizing radiation that did not induce cell death significantly suppressed mediator release from human mast cells (HMC-1(5C6) and LAD2 cells) that were activated by antigen-antibody reaction. To determine the inhibitory mechanism of mediator released by low-dose ionizing radiation, we examined the phosphorylation of intracellular signaling molecules such as Lyn, Syk, phospholipase Cγ, and protein kinase C, as well as the intracellular free Ca2+ concentration ([Ca2+]i). The phosphorylation of signaling molecules and [Ca2+]i following stimulation of FcεRI receptors was inhibited by low dose ionizing radiation. In agreement with its in vitro effect, ionizing radiation also significantly inhibited inflammatory cells infiltration, cytokine mRNA expression (TNF-α, IL-4, IL-13), and symptoms of passive cutaneous anaphylaxis reaction and the late-phase cutaneous response in anti-dinitrophenyl IgE-sensitized mice. These results indicate that ionizing radiation inhibits both mast cell-mediated immediate- and delayed-type allergic reactions in vivo and in vitro.

Sam68 is cleaved by caspases under apoptotic cell death induced by ionizing radiation.

The RNA-binding protein Sam68, a mitotic substrate of tyrosine kinases, has been reported to participate in the cell cycle, apoptosis, and signaling. In particular, overexpression of Sam68 protein is known to suppress cell growth and cell cycle progression in NIH3T3 cells. Although Sam68 is involved in many cellular activities, the function of Sam68, especially in response to apoptotic stimulation, is not well understood. In this study, we found that Sam68 protein is cleaved in immune cells undergoing apoptosis induced by γ-radiation. Moreover, we found that Sam68 cleavage was induced by apoptotic stimuli containing γ-radiation in a caspase-dependent manner. In particular, we showed that activated casepase-3, 7, 8 and 9 can directly cleave Sam68 protein through in vitro protease cleavage assay. Finally, we found that the knockdown of Sam68 attenuated γ-radiation-induced cell death and growth suppression. Conclusively, the cleavage of Sam68 is a new indicator for the cell damaging effects of ionizing radiation.

Chronic low-dose γ-irradiation of Drosophila melanogaster larvae induces gene expression changes and enhances locomotive behavior.

Although radiation effects have been extensively studied, the biological effects of low-dose radiation (LDR) are controversial. This study investigates LDR-induced alterations in locomotive behavior and gene expression profiles of Drosophila melanogaster. We measured locomotive behavior using larval pupation height and the rapid iterative negative geotaxis (RING) assay after exposure to 0.1 Gy γ-radiation (dose rate of 16.7 mGy/h). We also observed chronic LDR effects on development (pupation and eclosion rates) and longevity (life span). To identify chronic LDR effects on gene expression, we performed whole-genome expression analysis using gene-expression microarrays, and confirmed the results using quantitative real-time PCR. The pupation height of the LDR-treated group at the first larval instar was significantly higher (∼2-fold increase in PHI value, P < 0.05). The locomotive behavior of LDR-treated male flies (∼3 - 5 weeks of age) was significantly increased by 7.7%, 29% and 138%, respectively (P < 0.01), but pupation and eclosion rates and life spans were not significantly altered. Genome-wide expression analysis identified 344 genes that were differentially expressed in irradiated larvae compared with in control larvae. We identified several genes belonging to larval behavior functional groups such as locomotion (1.1%), oxidation reduction (8.0%), and genes involved in conventional functional groups modulated by irradiation such as defense response (4.9%), and sensory and perception (2.5%). Four candidate genes were confirmed as differentially expressed genes in irradiated larvae using qRT-PCR (>2-fold change). These data suggest that LDR stimulates locomotion-related genes, and these genes can be used as potential markers for LDR.

Altered microRNA expression profiles are involved in resistance to low-dose ionizing radiation in the absence of BMI1 in human dermal fibroblasts.

The polycomb group RING finger protein, B-cell­specific moloney murine leukemia virus integration site 1 (BMI1), has emerged as a key regulator of cell proliferation, cell cycle, cell immortalization, chemoresistance and radioresistance. Although the radioresistant effect of BMI1 has been thoroughly investigated, the effectiveness of this factor on low-dose radiation (LDR) resistance has not been explored. Here, we demonstrate that BMI1 is not critical for altering cell viability or cell growth in response to LDR, but BMI1 changes cellular gene expression profiles in response to LDR. Normal human dermal fibroblasts (NHDFs) stably expressing BMI1 short hairpin RNA (shRNA) did not exhibit changes in cell viability or cell cycle distribution assays following exposure to 0.1 Gy of γ-radiation. However, microRNA (miRNA) microarrays revealed that a lack of BMI1 leads to changes in miRNA expression in response to LDR. Bioinformatics analyses demonstrated that predicted target genes of the altered miRNAs are functionally involved in both negative and positive regulation of cell growth, cell proliferation, cell cycle and apoptosis. Therefore, these results indicate that low radiosensitivity even in the absence of the radioresistant factor BMI1 is related with the altered miRNA expression profiles in NHDF.