Histone chaperone activity of Fanconi anemia proteins, FANCD2 and FANCI, is required for DNA crosslink repair

Fanconi anaemia (FA) is a rare hereditary disorder characterized by genomic instability and cancer susceptibility. A key FA protein, FANCD2, is targeted to chromatin with its partner, FANCI, and plays a critical role in DNA crosslink repair. However, the molecular function of chromatin-bound FANCD2-FANCI is still poorly understood. In the present study, we found that FANCD2 possesses nucleosome-assembly activity in vitro. The mobility of histone H3 was reduced in FANCD2-knockdown cells following treatment with an interstrand DNA crosslinker, mitomycin C. Furthermore, cells harbouring FANCD2 mutations that were defective in nucleosome assembly displayed impaired survival upon cisplatin treatment. Although FANCI by itself lacked nucleosome-assembly activity, it significantly stimulated FANCD2-mediated nucleosome assembly. These observations suggest that FANCD2-FANCI may regulate chromatin dynamics during DNA repair.     

Efficacy of liraglutide, a glucagon-like peptide-1 (GLP-1) analogue, on body weight, eating behavior, and glycemic control, in Japanese obese type 2 diabetes

ABSTRACT: BACKGROUND: We recently reported that short-term treatment with liraglutide (20.0 +/- 6.4 days) reduced body weight and improved some scales of eating behavior in Japanese type 2 diabetes inpatients. However, it remained uncertain whether such liraglutide-induced improvement is maintained after discharge from the hospital. The aim of the present study was to determine the long-term effects of liraglutide on body weight, glycemic control, and eating behavior in Japanese obese type 2 diabetics. METHODS: Patients with obesity (body mass index (BMI) >25 kg/m2) and type 2 diabetes were hospitalized at Osaka University Hospital between November 2010 and December 2011. BMI and glycated hemoglobin (HbA1c) were examined on admission, at discharge and at 1, 3, and 6 months after discharge. For the liraglutide group (BMI; 31.3 +/- 5.3 kg/m2, n = 29), patients were introduced to liraglutide after correction of hyperglycemic by insulin or oral glucose-lowering drugs and maintained on liraglutide after discharge. Eating behavior was assessed in patients treated with liraglutide using The Guideline For Obesity questionnaire issued by the Japan Society for the Study of Obesity, at admission, discharge, 3 and 6 months after discharge. For the insulin group (BMI; 29.1 +/- 3.0 kg/m2, n = 28), each patient was treated with insulin during hospitalization and glycemic control maintained by insulin after discharge. RESULTS: Liraglutide induced significant and persistent weight loss from admission up to 6 months after discharge, while no change in body weight after discharge was noted in the insulin group. Liraglutide produced significant improvements in all major scores of eating behavior questionnaire items and such effect was maintained at 6 months after discharge. Weight loss correlated significantly with the decrease in scores for recognition of weight and constitution, sense of hunger, and eating style. CONCLUSION: Liraglutide produced meaningful long-term weight loss and significantly improved eating behavior in obese Japanese patients with type 2 diabetes.     

8-hydroxydeoxyguanosine generated in the earthworm Eisenia fetida grown in metal-containing soil

Heavy metal pollution of soil causes biological problems, such as mutagenicity to living organisms, including human beings. However, few methods have been developed to assess metal mutagenicity in soil. To avoid metal mutagenicity, an adequate bio-monitoring method is required. In the present study, to determine if the analysis of oxidative DNA damage generated in the earthworm is a useful bio-monitoring method for soil mutagenicity, the accumulation of 8-hydroxydeoxyguanosine (8-OH-dG), a major form of oxidative DNA damage, in Eisenia fetida (Savigny, 1826) treated with cadmium chloride (CdCl2) or nickel chloride (NiCl2) was analyzed. E. fetida was treated with Cd (10 or 200 microg/g soil) or Ni (10 or 200 microg/g soil) for 1, 2, and 3 weeks or 3 months. After metal exposure, the metal concentration in E. fetida was analyzed by atomic absorption spectrometry and the 8-OH-dG accumulated in E. fetida was analyzed by HPLC analyses and immunohistochemistry. Atomic absorption spectrometry revealed that Cd, but not Ni, accumulated within E. fetida. The 8-OH-dG levels in the DNA of E. fetida treated with Cd for 3 months were significantly higher than those in control E. fetida. Moreover, immunohistochemical analyses revealed that positive signals for 8-OH-dG accumulation in seminal vesicles were detected only in E. fetida treated with 10 microg of Cd for 3 months. Although some points remain unresolved, a bio-monitoring system analyzing the DNA damage generated in the earthworm might be useful for the assessment of the mutagenicity of soil contaminated with various heavy metals, such as Cd.     

Change of Morphology and Cytoskeletal Protein Gene Expression during Dibutyryl cAMP-induced Differentiation in C6 Glioma Cells

Elevation of the intracellular cAMP level induces morphological changes of astrocyte-like differentiation in C6 glioma cells. Such changes may be accompanied with expression of cytoskeletal protein genes. We therefore analyzed morphological changes after a treatment with dibutyryl cAMP (dbcAMP) and then assessed the expression of cytoskeletal protein genes by a quantitative real-time polymerase chain reaction. The cell number remained unaltered upon incubation with 1 mM dbcAMP in medium supplemented with 0.1% fetal bovine serum (FBS), whereas the number and lengths of processes increased, when compared with those of cells incubated in medium supplemented with 0.1% or 10% FBS only. The amounts of β-actin, γ-actin, and β-tubulin mRNAs in C6 cells, but not α-tubulin mRNA, increased during the early proliferation in DMEM containing 10% FBS. The expression of cytoskeletal protein genes decreased when incubated with 0.1% FBS or 1 mM dbcAMP in 0.1% FBS, compared with those of cells cultured in 10% FBS. These results indicated that, during the early proliferation in normal culture condition, the expression of cytoskeletal protein genes in C6 cells, except α-tubulin, increased, while in differentiating or differentiated C6 glioma cells, cAMP-induced morphological changes were not accompanied with elevation of gene expression for cytoskeletal proteins, such as actin and tubulin.     

Isolation and characterization of 39 microsatellite loci in the endangered Japanese loach Leptobotia curta

A microsatellite-enriched genomic library was obtained for the endangered Japanese loach Leptobotia curta, and 39 dinucleotide markers were successfully isolated and characterized. These markers had between one and nine alleles, with expected heterozygosity ranging from 0 to 0.839, in a population from the Lake Biwa-Yodo River system of Japan. Linkage equilibrium was observed in most loci, and only one locus showed significant deviation from Hardy-Weinberg equilibrium. These microsatellite markers will be useful for genetic diversity studies of wild and captive L. curta populations.     

Nucleosome formation with the testis-specific histone H3 variant, H3t, by human nucleosome assembly proteins in vitro

Five non-allelic histone H3 variants, H3.1, H3.2, H3.3, H3t and CENP-A, have been identified in mammals. H3t is robustly expressed in the testis, and thus was assigned as the testis-specific H3 variant. However, recent proteomics and tissue-specific RT-PCR experiments revealed a small amount of H3t expression in somatic cells. In the present study, we purified human H3t as a recombinant protein, and showed that H3t/H4 forms nucleosomes with H2A/H2B by the salt-dialysis method, like the conventional H3.1/H4. We found that H3t/H4 is not efficiently incorporated into the nucleosome by human Nap1 (hNap1), due to its defective H3t/H4 deposition on DNA. In contrast, human Nap2 (hNap2), a paralog of hNap1, promotes nucleosome assembly with H3t/H4. Mutational analyses revealed that the Ala111 residue, which is conserved among H3.1, H3.2 and H3.3, but not in H3t, is the essential residue for the hNap1-mediated nucleosome assembly. These results suggest that H3t may be incorporated into chromatin by a specific chaperone-mediated pathway.     

Genetic Population Structure of Chum Salmon in the Pacific Rim Inferred from Mitochondrial DNA Sequence Variation

We examined the genetic population structure of chum salmon, Oncorhynchus keta, in the Pacific Rim using mitochondrial (mt) DNA analysis. Nucleotide sequence analysis of about 500 bp in the variable portion of the 5′ end of the mtDNA control region revealed 20 variable nucleotide sites, which defined 30 haplotypes of three genealogical clades (A, B, and C), in more than 2,100 individuals of 48 populations from Japan (16), Korea (1), Russia (10), and North America (21 from Alaska, British Columbia, and Washington). The observed haplotypes were mostly associated with geographic regions, in that clade A and C haplotypes characterized Asian populations and clade B haplotypes distinguished North American populations. The haplotype diversity was highest in the Japanese populations, suggesting a greater genetic variation in the populations of Japan than those of Russia and North America. The analysis of molecular variance and contingency χ² tests demonstrated strong structuring among the three geographic groups of populations and weak to moderate structuring within Japanese and North American populations. These results suggest that the observed geographic pattern might be influenced primarily by historic expansions or colonizations and secondarily by low or restricted gene flow between local groups within regions. In addition to the analysis of population structure, mtDNA data may be useful for constructing a baseline for stock identification of mixed populations of high seas chum salmon.     

A River Metapopulation Structure of a Japanese Freshwater Goby, Odontobutis Obscura, Deduced from Allozyme Genetic Indices

Gene products of 18 allozyme loci from 1268 individuals of a Japanese freshwater goby called donko, Odontobutis obscura (Odontobutidae; Gobioidei), from 33 localities in the Koya River, Yamaguchi Prefecture, Japan, were investigated to determine the extent of genetic divergence and gene flow within a river metapopulation. Genetic indices including GST(mean FST 0.182), FIT(mean 0.192) and D(mean 0.015) indicated a considerable divergence of local populations in the river. The genetic distance (D) and channel distance between pairs of populations did not show a good correlation, and geographical neighbors were not always genetic neighbors. Therefore, the genetic divergence of populations is attributable to independent genetic drift with restricted gene flow among populations. The agricultural dams and weirs constructed across the river must be responsible for the restricted gene flow. The metapopulation structure of O. obscura in the Koya River may be barely sustained by one-way gene flow only from the upper to the lower populations. An occasional artificial transplantation of some individuals from the lower to the upper populations may be one alternative to maintain a river metapopulation structure safely.