An XPA gene splicing mutation resulting in trace protein expression in an elderly patient with xeroderma pigmentosum group A without neurological abnormalities.

A certain relationship between XPA gene mutations and the severity of symptoms has been observed in patients with xeroderma pigmentosum group A (XP-A). Patients with mutations within the DNA-binding domain usually exhibit severe symptoms, whereas splicing mutations in the same domain sometimes cause very mild symptoms. This inconsistency can be explained by a small amount of functional XPA protein produced from normally spliced transcripts. We herein report the case of an adult Japanese patient with XP-A with unusually mild symptoms. We identified a homozygous c.529G>A mutation in exon 4 of the XPA gene, which resulted in aberrant splicing with a 29-bp deletion in exon 4 causing a frameshift. Intact mRNA was observable, but a Western blot analysis failed to detect any normal XPA protein. We therefore evaluated the DNA repair capacity in normal cells in which the XPA expression was artificially diminished. The repair capacity was still present in cells with trace levels of the XPA protein. The repair capacity of the cells derived from our patient with mild symptoms was poor by comparison, but still significant compared with that of the cells derived from a patient with XP-A with severe symptoms. These results provide strong evidence that a trace level of XPA protein can still exert a relatively strong repair capacity, resulting in only a mild phenotype.

Expression of human and mouse genes encoding polkappa: testis-specific developmental regulation and AhR-dependent inducible transcription.

BACKGROUND: Human polkappa is a newly identified low-fidelity DNA polymerase. While the enzyme bypasses an abasic site and acetylaminofluorene-adduct in an error-prone manner, it bypasses benzo[a]pyrene-N2-dG lesions in a mostly error-free manner by incorporating predominantly dC opposite the bulky lesions. Benzo[a]pyrene (B[a]P) is activated through intracellular process mediated by the arylhydrocarbon receptor (AhR, also called the dioxin receptor), which is a ligand-activated transcription factor with high affinities for aromatic compounds such as B[a]P and dioxin. RESULTS: We examined promoter structures of the human POLK and mouse Polk genes to study how their expressions are regulated. The mouse Polk gene is developmentally regulated in testis and utilizes two transcription start sites during spermatogenesis, while it utilizes only one site in tissues other than testis. Both of the mouse Polk and the human POLK genes have two AhR-binding sites in the promoter regions and the expression of the mouse Polk gene is indeed enhanced upon AhR-activation. CONCLUSIONS: The AhR activation increases expression of the mouse Polk gene and probably the human POLK gene, the product of which bypasses benzo[a]pyrene-N2-dG lesions in a mostly accurate manner. Thus, polkappa seems to function to reduce mutagenesis at benzo[a]pyrene-adducts, although it may also have a role related to spermatogenesis.

Microalgal cultivation in a solution recovered from the low-temperature catalytic gasification of the microalga.

Microalgal cultivation in a solution recovered from the low-temperature catalytic gasification of the microalga itself was studied. The growth of Chlorella vulgaris in 75-300-fold diluted recovered solution containing phosphate, magnesium ions and micro-elements was comparable to that in the standard culture medium. It was suggested that C. vulgaris could use ammonium in the recovered solution as its nitrogen source and at the same time could provide a source of biomass which was recycled via gasification.

Error-prone bypass of certain DNA lesions by the human DNA polymerase kappa.

The Escherichia coli protein DinB is a newly identified error-prone DNA polymerase. Recently, a human homolog of DinB was identified and named DINB1. We report that the DINB1 gene encodes a DNA polymerase (designated polkappa), which incorporates mismatched bases on a nondamaged template with a high frequency. Moreover, polkappa bypasses an abasic site and N-2-acetylaminofluorene (AAF)-adduct in an error-prone manner but does not bypass a cis-syn or (6-4) thymine-thymine dimer or a cisplatin-adduct. Therefore, our results implicate an important role for polkappa in the mutagenic bypass of certain types of DNA lesions.

Identification of additional genes belonging to the LexA regulon in Escherichia coli.

Exposure of Escherichia coli to a variety of DNA-damaging agents results in the induction of the global 'SOS response'. Expression of many of the genes in the SOS regulon are controlled by the LexA protein. LexA acts as a transcriptional repressor of these unlinked genes by binding to specific sequences (LexA boxes) located within the promoter region of each LexA-regulated gene. Alignment of 20 LexA binding sites found in the E. coli chromosome reveals a consensus of 5'-TACTG(TA)5CAGTA-3'. DNA sequences that exhibit a close match to the consensus are said to have a low heterology index and bind LexA tightly, whereas those that are more diverged have a high heterology index and are not expected to bind LexA. By using this heterology index, together with other search criteria, such as the location of the putative LexA box relative to a gene or to promoter elements, we have performed computational searches of the entire E. coli genome to identify novel LexA-regulated genes. These searches identified a total of 69 potential LexA-regulated genes/operons with a heterology index of <15 and included all previously characterized LexA-regulated genes. Probes were made to the remaining genes, and these were screened by Northern analysis for damage-inducible gene expression in a wild-type lexA+ cell, constitutive expression in a lexA(Def) cell and basal expression in a non-inducible lexA(Ind-) cell. These experiments have allowed us to identify seven new LexA-regulated genes, thus bringing the present number of genes in the E. coli LexA regulon to 31. The potential function of each newly identified LexA-regulated gene is discussed.

Mutation enhancement by DINB1, a mammalian homologue of the Escherichia coli mutagenesis protein dinB.

BACKGROUND: The Escherichia coli dinB gene is an SOS gene known to be required for lambda phage untargeted mutagenesis. When over-expressed, it exhibits a potent mutagenic activity without any exogenous treatment to damage DNA. Frameshift mutations at a run of identical bases are most enhanced. The product DinB is structurally related to the E. coli UmuC protein and the Saccharomyces cerevisiae Rev1 and Rad30 proteins, all of which are shown to be involved in bypass synthesis at a DNA lesion. RESULTS: We have cloned and sequenced human and mouse cDNAs encoding a DinB homologue. Their products are highly similar to DinB and less similar to UmuC, Rev1 or Rad30, and hence the genes were named DINB1 for human and Dinb1 for mouse. Both genes were expressed most abundantly in testis. Transient expression of the mouse cDNA in cultured mouse cells resulted in a nearly 10-fold increase in the incidence of point mutations, among which about 30% were frameshift mutations. CONCLUSIONS: The above results suggest that a mutagenic mechanism, a so-called untargeted type, also operates in mammalian cells. Taken together with recent findings that human cells have multiple DNA polymerases for translesion synthesis which are homologous to the S. cerevisiae Rev3 and Rad30 proteins, our results imply that multiple mutagenic pathways are conserved from bacteria to higher eukaryotes.

Treatment of liquid fraction separated from liquidized food waste in an upflow anaerobic sludge blanket reactor.

Thermochemical liquidization as a pretreatment for anaerobic digestion of food waste was studied using a laboratory-scale upflow anaerobic sludge blanket (UASB) reactor for a period of 82 d. Model food waste (approximately 90 wt% moisture content) was thermochemically liquidized at 175 degrees C for 1 h. The liquidized food waste was separated into a solid phase (6-10 wt%) and a liquid phase (85-89 wt%). The diluted liquid phase was continuously treated by anaerobic digestion using a UASB reactor at 35 degrees C. The volumetric loading rate was increased stepwise to 6.4-7.2 g total organic carbon (TOC)/l-reactor/d. Methane production was found to be approximately 0.35-0.61 l/g-TOC removed. The range of TOC removal efficiencies was 67-69% at an influent TOC concentration of 10.1-11.1 g/l and a volumetric loading rate of 4.8-5.3 g-TOC/l-reactor/d. This treatment process using an UASB reactor could be suitable for resource recovery from food waste.

Effects of glucose addition and light on current outputs in photosynthetic electrochemical cells using Synechocystis sp. PCC6714.

The effects of glucose addition and light on the current outputs in electrochemical cells using a cyanobacterium Synechocystis sp. PCC6714 were investigated under photo- and chemoheterotrophical conditions. The addition of glucose to the anode solutions of the electrochemical cells resulted in a rapid increase in the current outputs under both light and dark conditions. Although the coulombic outputs were almost the same between under light and dark conditions, the rate of glucose consumption was faster under illumination than in the dark. The total sugar content in the cells of strain PCC6714 increased with the addition of glucose and the total sugar accumulated remained intact during the discharge under illumination, while it decreased gradually in the dark. When the light was switched off after the addition of glucose, the current output markedly increased. The coulombic outputs obtained after darkening were 10 to 80 times larger than that obtained by the addition of glucose under the continuous light or dark conditions. Synechocystis sp. completely incorporated 0.14 mM and 0.42 mM glucose for 1 h and 3 h, respectively, under illumination. There was no difference in the coulombic outputs between 1 h and 12 h illumination times in the electrochemical cells with 0.14 mM glucose. When the light was switched off after 1 h illumination in the electrochemical cells with 0.42 mM glucose, the coulombic output obtained from the electrochemical cell was lower than that in the electrochemical cell with 12 h illumination. This indicates that the current output was produced with higher efficiency with glucose incorporated under illumination than that in the case of glucose incorporated after darkening. The highest coulombic yield of 54% in this experiment was obtained by darkening in the electrochemical cell with 0.14 mM glucose.