Poly(ADP-ribose) polymerase 1 (PARP-1) and p53 are two key proteins in the DNA-damage response. Although PARP-1 is known to poly(ADP-ribosyl)ate p53, the role of this modification remains elusive. Here, we identify the major poly(ADP-ribosyl)ated sites of p53 by PARP-1 and find that PARP-1-mediated poly(ADP-ribosyl)ation blocks the interaction between p53 and the nuclear export receptor Crm1, resulting in nuclear accumulation of p53. These findings molecularly link PARP-1 and p53 in the DNA-damage response, providing the mechanism for how p53 accumulates in the nucleus in response to DNA damage. PARP-1 becomes super-activated by binding to damaged DNA, which in turn poly(ADP-ribosyl)ates p53. The nuclear export machinery is unable to target poly(ADP-ribosyl)ated p53, promoting accumulation of p53 in the nucleus where p53 exerts its transactivational function.

Poly(ADP-ribosyl)ation has been suggested to be involved in regulation of DNA repair, transcription, centrosome duplication, and chromosome stability. However, the regulation of degradation of poly(ADP-ribose) and its significance are not well understood. Here we report a loss-of-function mutant Drosophila with regard to poly(ADP-ribose) glycohydrolase, a major hydrolyzing enzyme of poly(ADP-ribose). The mutant lacks the conserved catalytic domain of poly(ADP-ribose) glycohydrolase, and exhibits lethality in the larval stages at the normal development temperature of 25°C. However, one-fourth of the mutants progress to the adult stage at 29°C but showed progressive neurodegeneration with reduced locomotor activity and a short lifespan. In association with this, extensive accumulation of poly(ADP-ribose) could be detected in the central nervous system. These results suggest that poly(ADPribose) metabolism is required for maintenance of the normal function of neuronal cells. The phenotypes observed in the parg mutant might be useful to understand neurodegenerative conditions such as the Alzheimer's and Parkinson's diseases that are caused by abnormal accumulation of substances in nervous tissue. P oly(ADP-ribosyl)ation process involves a posttranslational modification of target proteins catalyzed by the poly(ADPribose) polymerase (PARP) family of enzymes with NADϩ as the substrate, resulting in formation of long-branched polymers of ADP-ribose (1, 2). The covalently attached and negatively charged poly(ADP-ribose) units significantly affect several important biological functions, including DNA repair (3), transcription (4), regulation of telomere length, cell cycle progression, centrosome duplication (5, 6), and chromosome stability (7). One of the major members of the PARP family is PARP-1, which catalyzes poly(ADP-ribosyl)ation in response to DNA strand breaks. Recently, additional members of the PARP family of enzymes have been characterized; PARP-2, -3, tankyrase-1, -2, VPARP, and Ti-PARP (2). In Drosophila, only two PARP family members, corresponding to PARP-1 and tankyrase, have been reported (8, 9).Poly(ADP-ribose) attached to acceptor proteins is hydrolyzed rapidly to produce free ADP-ribose residues by poly(ADPribose) glycohydrolase (PARG) (10-12). In contrast to PARPs, only one gene for PARG has been detected in mammals and insects (13). Thus, it is likely that the regulation of PARG activity is required to complete protein modification cycles initiated by different PARPs. In fact, PARG has been proposed to shuttle between the nucleus and the cytoplasm and becomes localized to the centrosomes during mitosis (14). There is evidence for an alternative form in the cytoplasm (M. K. Jacobson, personal communication), indicating that regulation of poly(ADPribosyl)ation by PARG is very dynamic.Mutation of the parg gene, tej, in plants alters circadian rhythms (15), and increased sensitivity to DNA damage in parg knockout mouse embryonic stem cells has been reported (3). However, little is known about the effects...

Infection with Opisthorchis viverrini (OV) is associated with cholangiocarcinoma. OV is common in northeast Thailand, but less than 10% of the inhabitants develop cholangiocarcinoma. Animal experiments suggest that OV infection alone does not cause cholangiocarcinoma, and thus other environmental and genetic factors may play a role in causation. We conducted a population-based case-control study in which sex, age and place of residence were matched individually. Polymorphisms of GSTM1 and GSTT1 alone were not associated with risk for cholangiocarcinoma, while an elevated level of antibodies against OV (ELISA) 0.200 was the strongest risk indicator (odds ratio as compared to that <0. Key words: population-based case-control study; inflammation; polymorphism; alcohol drinking; nitrosamine Cholangiocarcinoma is uncommon, and the aetiology has been elucidated only for a limited portion of cases.1 Northeast Thailand has an exceptionally high incidence of liver cancer of up to 87.5 (male) and 37.2 (female) per 100,000 (world population-adjusted incidence rate). Most of these cases have been attributed to cholangiocarcinoma.2 Infestation of the liver fluke, Opisthorchis viverrini (OV), has been causally related to cholangiocarcinoma in this region. 3 The prevalence of OV infection in Nakhon Phanom province was between 30 and 70% with substantial variation between villages; 4 the technique for estimation of prevalence was not explicitly mentioned but supposed OV egg counting in the faeces. However, it is unknown why only a limited proportion of the inhabitants (<10%) develop cholangiocarcinoma.In our study, we hypothesised that genetic background may be a contributing factor in the occurrence of the cholangiocarcinoma. Hence, we examined the association between polymorphisms of genes for glutathione-S-transferase enzymes, GSTM1 and GSTT1, and the risk for cholangiocarcinoma. Because both enzymes function in carcinogen detoxification systems, 5 it seems reasonable to speculate that null variants of these genes may be associated with increased risk of cancer. 6The association between OV infection and cancer risk has been a common assumption, but adequate analytic epidemiological studies are sparse. Parkin and colleagues 7 reported in their casecontrol study that past or present infection with OV, as indicated by serum antibodies against OV, is associated with a 5-fold increase in cancer risk. Although the anti-OV antigen titre was elevated and reached a plateau at an approximately 2 months after OV inoculation in hamsters, 8 the transition of the antibody level over the lifespan remains uncertain in both humans and animals. We thus examined the risk of foods possibly containing OV metacercariae as well as that of OV infection as indicated by serum antibody response.In animal experiments, OV infection causes cholangiocarcinoma with coadministration of nitrosamine 9 in a dose-dependent manner.10 Diet and cigarette smoking are major sources of nitrosamine exposure, 11 and several foods and foodstuffs consumed in the study ar...

Cholangiocarcinoma (CCA) induced by liver fluke (Opisthorchis viverrini, Ov) infection is one of the most common and serious disease in northeast Thailand. To elucidate the molecular mechanism of cholangiocarcinogenesis induced by Ov infection, we employed a hamster model of CCA induced by Ov and N-nitrosodimethylamine and analyzed candidate genes involved in CCA using fluorescence differential display-PCR. Of 149 differentially amplified bands we identified, the upregulation of 23 transcripts and downregulation of 1 transcript related to CCA hamsters were confirmed by a reverse northern macroarray blot. The upregulated genes include signal transduction protein kinase A regulatory subunit Ialpha (Prkar1a), myristoylated alanine-rich protein kinase C substrate, transcriptional factor LIM-4-only domain, oxysterol-binding protein involved in lipid metabolism, splicing regulatory protein 9, ubiquitin conjugating enzyme involved in protein degradation, beta tubulin, beta actin, and collagen type VI. Quantitative real-time PCR confirmed that the expression of Prkar1a was significantly higher in CCA and its precursor lesion when compared with normal liver and normal gall bladder epithelia (P<0.05). Prkar1a expression tended to increase along with the progression of biliary transformation from hyperplasia and precancerous lesions to carcinoma. These findings contribute to our understanding of the processes involved in the molecular carcinogenesis of CCA in order to provide a unique perspective on the development of new chemotherapeutics in future.

No abstract

Although feeding time is a dominant cue for circadian rhythms in mammalian peripheral tissue, the effect of feeding and fasting on circadian gene expression and behaviour is unknown. Here we report that fasting does not affect the phase of rhythmic mRNA expression levels of the clock genes, mPer1, mPer2 and of the clock controlled gene, mDBP. However, the levels of each of these genes were significantly altered in different ways and recovered by feeding. We also found that feeding enhances phase-shifting to a new light-dark cycle of rhythmic mPer2 mRNA expression in the heart. Furthermore, feeding enhances the phase-shifting to new light-dark cycle of behaviour more than fasting. Our data indicate that feeding is an important cue for circadian behaviour rhythms as well as for the photo-entrainment of peripheral clock gene expression.

No abstract

Gaucher disease (GD) is the most common of the lysosomal storage disorders and is caused by defects in the GBA gene encoding glucocerebrosidase (GlcCerase). The accumulation of its substrate, glucocylceramide (GlcCer) is considered the main cause of GD. We found here that the expression of human mutated GlcCerase gene (hGBA) that is associated with neuronopathy in GD patients causes neurodevelopmental defects in Drosophila eyes. The data indicate that endoplasmic reticulum (ER) stress was elevated in Drosophila eye carrying mutated hGBAs by using of the ER stress markers dXBP1 and dBiP. We also found that Ambroxol, a potential pharmacological chaperone for mutated hGBAs, can alleviate the neuronopathic phenotype through reducing ER stress. We demonstrate a novel mechanism of neurodevelopmental defects mediated by ER stress through expression of mutants of human GBA gene in the eye of Drosophila.