Ability of the endangered species Papaver fauriei to produce hybrids with a cultivated poppy (Papaver sp.)

Papaver fauriei is an endemic and endangered species that grows only on the gravelly alpine slopes of Mt. Rishiri, Japan. Cultivated poppy (Papaver sp.), the species name of which is unknown, has been introduced to the natural habitats of P. fauriei through human activities. Because the appearance and internal transcribed spacer (ITS) sequences of these two poppies are highly similar, it is of concern that they could produce hybrids in their natural habitats. Thus, first, the ability of these two poppies to produce hybrids was analyzed by artificial fertilization in this study. A large number of seeds were produced by reciprocal crosses between P. fauriei and the cultivated poppy, comparable with the number of seeds obtained by self‐ or cross‐fertilization of P. fauriei or the cultivated poppy. In addition, high germination was observed for seeds obtained from crosses between the two poppies, and deleterious phenotypes, such as albinism and dwarfism, were not detected in the F₁ generation. These results indicate that after pollination, there is no reproductive isolation between the two poppies. Second, we sequenced the internal transcribed spacer (ITS) region of 240 poppy individuals collected from the gravelly alpine slopes of Mt. Rishiri, and 66 showed the sequence of P. fauriei, whereas 174 showed the sequence of the cultivated poppy. However, the ITS sequence that confirms hybridism between the two poppies was not detected in these individuals, indicating that hybridization of P. fauriei and the cultivated poppy rarely occurs under natural conditions. Unknown mechanism(s) appear to prevent cross‐pollination between the two poppies.     

Hyperthermostable Endoglucanase from Pyrococcus horikoshii

An endoglucanase homolog from the hyperthermophilic archaeon Pyrococcus horikoshii was expressed in Escherichia coli, and its enzymatic characteristics were examined. The expressed protein was a hyperthermostable endoglucanase which hydrolyzes celluloses, including Avicel and carboxymethyl cellulose, as well as β-glucose oligomers. This enzyme is the first endoglucanase belonging to glycosidase family 5 found from Pyrococcus species and is also the first hyperthermostable endoglucanase to which celluloses are the best substrates. This enzyme is expected to be useful for industrial hydrolysis of cellulose at high temperatures, particularly in biopolishing of cotton products.     

Nuclear export signal consensus sequences defined using a localization-based yeast selection system

Proteins bearing nuclear export signals (NESs) are translocated to the cytoplasm from the nucleus mainly through the CRM1-dependent pathway. However, the NES consensus sequence remains poorly defined, and there are currently no high-throughput methods for identifying NESs. In this study, we report the development of an efficient yeast selection system for detecting nuclear export activity as well as several reliable NES consensus sequences identified using this method. Our selection system is based on the nuclear export-dependent rescue of Tys1p, an essential cytoplasmic protein that has been artificially localized to the nucleus in a haploid Delta tys1 knockout strain. A screen of a random peptide library revealed 101 distinct CRM1-dependent NESs, which were classified into six patterns according to the conserved hydrophobic spacing. By combining mutational analyses, we have defined new NES consensus sequences with more specific and redundant residues than the traditional consensus sequence, which are consistent with most experimentally confirmed NESs. These NES consensus sequences should help identify functional NESs, and our selection system can be used to identify other targeting signals or proteins imported to specific subcellular compartments.     

Analysis of changes in DNA copy number in radiation-induced thymic lymphomas of susceptible C57BL/6, resistant C3H and hybrid F1 Mice

Radiation-induced thymic lymphoma in mice is a useful model for studying both the mechanism of radiation carcinogenesis and genetic susceptibility to tumor development. Using array-comparative genomic hybridization, we analyzed genome-wide changes in DNA copy numbers in radiation-induced thymic lymphomas that had developed in susceptible C57BL/6 and resistant C3H mice and their hybrids, C3B6F1 and B6C3F1 mice. Besides aberrations at known relevant genetic loci including Ikaros and Bcl11b and trisomy of chromosome 15, we identified strain-associated genomic imbalances on chromosomes 5, 10 and 16 and strain-unassociated trisomy of chromosome 14 as frequent aberrations. In addition, biallelic rearrangements at Tcrb were detected more frequently in tumors from C57BL/6 mice than in those from C3H mice, suggesting aberrant V(D)J recombination and a possible link with tumor susceptibility. The frequency and spectrum of these copy-number changes in lymphomas from C3B6F1 and B6C3F1 mice were similar to those in C57BL/6 mice. Furthermore, the loss of heterozygosity analyses of tumors in F(1) mice indicated that allelic losses at Ikaros and Bcl11b were caused primarily by multilocus deletions, whereas those at the Cdkn2a/Cdkn2b and Pten loci were due mainly to uniparental disomy. These findings provide important clues to both the mechanisms for accumulation of aberrations during radiation-induced lymphomagenesis and the different susceptibilities of C57BL/6 and C3H mice.     

Combined effect of multiple carcinogens and synergy index

Tumorigenesis often involves exposure to multiple carcinogens. We here construct a two-stage model, which consists of individuals in three states:normal, intermediate, and malignant. We considered a population exposed to two carcinogens and examined two exposure scenarios: (1) two carcinogens exposed separately; and (2) carcinogens are exposed simultaneously. We calculate the incidence rate of the malignant state, introducing a "synergy index" to quantify the interaction between the two carcinogens. We confirmed the results of previous analyses, but we also calculate more general situation in which each carcinogen affects both steps.     

Global gene expression profiling in early-stage polycystic kidney disease in the Han:SPRD Cy rat identifies a role for RXR signaling

Han:SPRD Cy is a spontaneous rat model of polycystic kidney disease (PKD) caused by a missense mutation in Pkdr1. Cystogenesis in this model is not clearly understood. In the current study, we performed global gene expression profiling in early-stage PKD cyst development in Cy/Cy kidneys and normal (+/+) kidneys at 3 and 7 days of postnatal age. Expression profiles were determined by microarray analysis, followed by validation with real-time RT-PCR. Genes were selected with over 1.5-fold expression changes compared with age-matched +/+ kidneys for canonical pathway analysis. We found nine pathways in common between 3- and 7-day Cy/Cy kidneys. Three significantly changed pathways were designated "Vitamin D Receptor (VDR)/Retinoid X Receptor (RXR) Activation," "LPS/IL-1-Mediated Inhibition of RXR Function," and "Liver X Receptor (LXR)/RXR Activation." These results suggest that RXR-mediated signaling is significantly altered in developing kidneys with mutated Pkdr1. In gene ontology analysis, the functions of these RXR-related genes were found to be involved in regulating cell proliferation and organ morphogenesis. With real-time RT-PCR analysis, the upregulation of Ptx2, Alox15b, OSP, and PCNA, major markers of cell proliferation associated with the RXR pathway, were confirmed in 3- and 7-day Cy/Cy kidneys compared with 3-day +/+ kidneys. The increased RXR protein was observed in both the nucleus and cytoplasm of cystic epithelial cells in early-stage Cy/Cy kidneys, and the RXR-positive cells were strongly positive for PCNA staining. Taken together, cell proliferation and organ morphogenesis signals transduced by RXR-mediated pathways may have important roles for cystogenesis in early-stage PKD in this Pkdr1-mutated Cy rat.     

In vivo kinematics of two-component total ankle arthroplasty during non-weightbearing and weightbearing dorsiflexion/plantarflexion

Relatively high rates of loosening and implant failure have been reported after total ankle arthroplasty, especially in first and second generation implants. Abnormal kinematics and incongruency of the articular surface may cause increased loads applied to the implant with concomitant polyethylene wear, resulting in loosening and implant failure. The purpose of this study was to measure three-dimensional kinematics of two-component total ankle arthroplasty during non-weightbearing and weightbearing activities, and to investigate incongruency of the articular surfaces during these activities. Forty-seven patients with a mean age of 71 years were enrolled. Radiographs were taken at non-weightbearing maximal dorsiflexion and plantarflexion, and weightbearing maximal dorsiflexion, plantarflexion, and neutral position. 3D-2D model-image registration was performed using the radiographs and the three-dimensional implant models, and three-dimensional joint angles were determined. The implanted ankles showed 18.1±8.6° (mean±standard deviation) of plantarflexion, 0.1±0.7° of inversion, 1.2±2.0° of internal rotation, and 0.8±0.6mm of posterior translation of the talar component in the non-weightbearing activity, and 17.8±7.5° of plantarflexion, 0.4±0.5° of inversion, 1.8±2.0° of internal rotation, and 0.7±0.5mm of posterior translation in the weightbearing activity. There were no significant differences between the non-weightbearing and weightbearing kinematics except for the plantarflexion angle. Incongruency of the articular surface occurred in more than 75% of the ankles. Our observations will provide useful data against which kinematics of other implant designs, such as three-component total ankle arthroplasty, can be compared.