Development of genetic differentiation and postzygotic isolation in experimental metapopulations of spider mites

We studied the development of genetic differentiation and postzygotic isolation in experimental metapopulations of the two-spotted spider mite, Tetranychus urticae Koch. A genetically diverse starter population was made by allowing six inbred sublines to interbreed. Then three migration patterns were tested: no migration, or one or three immigrants per subpopulation per generation. Variations in four traits were investigated: allozymes, acaricide resistance, diapause, and hatchability. In the allozymes, acaricide resistance, and diapause, genetic variation among subpopulations became high in metapopulations with no migration, but not in the others, which showed that one immigrant is enough to prevent genetic differentiation. Hatchability, which was decreased by interbreeding among the six sublines, gradually recovered in succeeding generations. In metapopulations with no migration, hatchability was reduced again after in-migration at the 15th generation. Different karyotypes or coadapted gene complexes can survive in different subpopulations by genetic drift, and both Wolbachia-infected and -noninfected subpopulations may be selected, which would lead to postzygotic isolation between isolated subpopulations. Our results indicate that sampling effects such as genetic drift or stochastic loss of Wolbachia produce postzygotic isolation in laboratory populations of spider mite.     

Arabidopsis Raf‐like kinases act as positive regulators of subclass III SnRK2 in osmostress signaling

Given their sessile nature, land plants must use various mechanisms to manage dehydration under water‐deficit conditions. Osmostress‐induced activation of the SNF1‐related protein kinase 2 (SnRK2) family elicits physiological responses such as stomatal closure to protect plants during drought conditions. With the plant hormone ABA receptors [PYR (pyrabactin resistance)/PYL (pyrabactin resistance‐like)/RCAR (regulatory component of ABA receptors) proteins] and group A protein phosphatases, subclass III SnRK2 also constitutes a core signaling module for ABA, and osmostress triggers ABA accumulation. How SnRK2 is activated through ABA has been clarified, although its activation through osmostress remains unclear. Here, we show that Arabidopsis ABA and abiotic stress‐responsive Raf‐like kinases (AtARKs) of the B3 clade of the mitogen‐activated kinase kinase kinase (MAPKKK) family are crucial in SnRK2‐mediated osmostress responses. Disruption of AtARKs in Arabidopsis results in increased water loss from detached leaves because of impaired stomatal closure in response to osmostress. Our findings obtained in vitro and in planta have shown that AtARKs interact physically with SRK2E, a core factor for stomatal closure in response to drought. Furthermore, we show that AtARK phosphorylates S171 and S175 in the activation loop of SRK2E in vitro and that Atark mutants have defects in osmostress‐induced subclass III SnRK2 activity. Our findings identify a specific type of B3‐MAPKKKs as upstream kinases of subclass III SnRK2 in Arabidopsis. Taken together with earlier reports that ARK is an upstream kinase of SnRK2 in moss, an existing member of a basal land plant lineage, we propose that ARK/SnRK2 module is evolutionarily conserved across 400 million years of land plant evolution for conferring protection against drought.     

Novel interactions of large P3 moiety and small P4 moiety in the binding of the peptide mimetic factor VIIa inhibitor

Selective factor VIIa-tissue factor complex (FVIIa/TF) inhibition is seen as a promising target for developing new anticoagulant drugs. A novel peptide mimetic factor VIIa inhibitor, ethylsulfonamide-d-biphenylalanine-Gln-p-aminobenzamidine, shows 100-fold selectivity against thrombin in spite of its large P3 moiety, unlike previously reported FVIIa/TF selective inhibitors. X-ray crystal structure analysis reveals that the large P3 moiety, d-biphenylalanine, and the small P4 moiety, ethylsulfonamide, make novel interactions with the 170-loop and Lys192 of FVIIa/TF, respectively, accompanying ligand-induced conformational changes of the 170-loop, Gln217, and Lys192. Structural comparisons of FVIIa with thrombin and amino acid sequence comparisons among coagulation serine proteases suggest that these interactions play an important role in achieving selective inhibition for FVIIa/TF.     

UV-B light induces an adaptive response to UV-C exposure via photoreactivation activity in Euglena gracilis.

Phytoplankton such as Euglena are constantly exposed to solar light which is used for photosynthesis. Although the solar ultraviolet (UV) induces DNA damage such as cyclobutane-pyrimidine dimers (CPDs), many kinds of living organisms can repair CPDs by photoreactivation (PR) utilizing the near-UV/blue light component in sunlight. Euglena cells are known to possess such PR activity. In the present paper, the formation of CPDs induced by UV-C exposure and the photoreactivation PR repair of these CPDs by UV-A are demonstrated. To clarify the adaptive responses prior UV-B irradiation on PR activity, cells were cultured in the dark or under UV-B light. When the cells were cultured in the dark for 3 d prior to UV-C exposure, PR activity decreased. When the cells were cultured under UV-B light, however, PR activity increased. These results suggest that exposing the cells to UV-B prior to exposure to UV-C induced an adaptive response towards DNA damage caused by UV-C exposure, and this UV-C induced damage was repaired through PR activity.     

Two isoforms of chicken melanopsins show blue light sensitivity

Melanopsin is a vertebrate non-visual opsin and functions as a circadian photoreceptor in mammalian retinas. Here we found the expression of two kinds of melanopsin genes in the chicken pineal gland and identified the presence of five isoforms derived from these two genes. Reconstitution of the recombinant proteins with 11-cis-retinal revealed that at least two of these melanopsin protein isoforms can function as blue-sensitive photopigments with absorption maxima at 476-484nm. These values are consistent with maximal sensitivities of action spectra determined from the physiological and behavioral studies on mammalian melanopsins. The melanopsin isoforms found in this study may function as pineal circadian photoreceptors.     

Abiotic stress-inducible receptor-like kinases negatively control ABA signaling in Arabidopsis

Membrane-anchored receptor-like protein kinases (RLKs) recognize extracellular signals at the cell surface and activate the downstream signaling pathway by phosphorylating specific target proteins. We analyzed a receptor-like cytosolic kinase (RLCK) gene, ARCK1, whose expression was induced by abiotic stress. ARCK1 belongs to the cysteine-rich repeat (CRR) RLK sub-family and encodes a cytosolic protein kinase. The arck1 mutant showed higher sensitivity than the wild-type to ABA and osmotic stress during the post-germinative growth phase. CRK36, an abiotic stress-inducible RLK belonging to the CRR RLK sub-family, was screened as a potential interacting factor with ARCK1 by co-expression analyses and a yeast two-hybrid system. CRK36 physically interacted with ARCK1 in plant cells, and the kinase domain of CRK36 phosphorylated ARCK1 in vitro. We generated CRK36 RNAi transgenic plants, and found that transgenic plants with suppressed CRK36 expression showed higher sensitivity than arck1-2 to ABA and osmotic stress during the post-germinative growth phase. Microarray analysis using CRK36 RNAi plants revealed that suppression of CRK36 up-regulates several ABA-responsive genes, such as LEA genes, oleosin, ABI4 and ABI5. These results suggest that CRK36 and ARCK1 form a complex and negatively control ABA and osmotic stress signal transduction.     

Characterization of the tissue-specific expression of phenylalanine ammonia-lyase gene promoter from loblolly pine (<Emphasis Type="Italic">Pinus taeda</Emphasis>) in <Emphasis Type="Italic">Nicotiana tabacum</Emphasis>

We isolated the 5′ flanking region of a gene for phenylalanine ammonia-lyase (PAL; EC 4.3.1.5) from Pinus taeda, PtaPAL. To investigate the tissue-specific expression of the PtaPAL promoter, histochemical assay of GUS activity was performed using the transgenic tobacco expressing the PtaPAL promoter-GUS. The region of −897 to −420 in PtaPAL promoter showed high activities in the secondary xylem and response to bending stress. To characterize the cis-regulatory functions of the promoters for enzymes in phenylpropanoid biosynthesis, we examined the activity of chimeric promoters of PtaPAL and a 4-coumarate CoA ligase, Pta4CLα. The chimeric promoter showed similar activity as the Pta4CLα promoter. Electrophoretic mobility shift assays implicated −897 to –674 of PtaPAL promoter containing cis-elements of the expression in xylem of Pinus taeda. The results suggested that AC elements of PtaPAL have multiple functions in the expression under the various developmental stages and stress conditions in the transgenic tobacco. The nucleotide sequence data reported will appear in the EMBL, GenBank, and DDBJ Nucleotide Sequence Databases under the accession number AB449103 (PtaPAL promoter sequence).     

Origins of sequence diversity in the malaria vaccine candidate merozoite surface protein-2 (MSP-2) in Amazonian isolates of Plasmodium falciparum

The recent evolution of Plasmodium falciparum is at odds with the extensive polymorphism found in most genes coding for antigens. Here, we examined the patterns and putative mechanisms of sequence diversification in the merozoite surface protein-2 (MSP-2), a major malarial repetitive surface antigen. We compared the msp-2 gene sequences from closely related clones derived from sympatric parasite isolates from Brazilian Amazonia and used microsatellite typing to examine, in these same clones, the haplotype background of chromosome 2, where msp-2 is located. We found examples of msp-2 sequence rearrangements putatively created by nonreciprocal recombinational events, such as replication slippage and gene conversion, while maintaining the chromosome haplotype. We conclude that these nonreciprocal recombination events may represent a major source of antigenic diversity in MSP-2 in P. falciparum populations with low rates of classical meiotic recombination.     

Cyclin A2 confers cisplatin resistance to endometrial carcinoma cells via up‐regulation of an Akt‐binding protein,periplakin

Although overexpression of cyclin A2 is reportedly an indicator of a poor prognosis of various malignancies including endometrial carcinoma, its molecular mechanism remains undetermined. To address this issue, we examined the effect of cyclin A2 on the development of resistance to chemotherapeutic drugs. The expression of cyclin A2 protein was increased in advanced‐stage and chemotherapy‐refractory stage endometrial carcinomas compared with that in early‐stage tumours. The expression levels of cyclin A2 in endometrial carcinoma cell lines correlated positively with the IC₅₀ for cisplatin. Endometrial carcinoma HHUA cells that overexpressed cyclin A2 showed increased resistance to cisplatin in vitro and in vivo, via the activation of a survival pathway, the inositol‐3 phosphate kinase (PI3K) cascade. The use of a cDNA microarray identified an Akt‐binding protein, periplakin, as a novel target of cyclin A2. The cyclin A2‐induced up‐regulation of periplakin was mediated via direct binding of Sp1 to the promoter that was activated by cyclin A2 along with chromatin remodelling involving CBP/p300, and the siRNA‐mediated silencing of periplakin suppressed the PI3K pathway. These results indicate cyclin A2 to be involved in the acquisition of aggressive behaviour of tumour cells through the activation of PI3K by cyclin A2‐induced periplakin, and to be a promising therapeutic target.     

Streptococcus panodentis sp. nov. from the oral cavities of chimpanzees

Three strains TKU9, TKU49 and TKU50^T, were isolated from the oral cavities of chimpanzees (Pan troglodytes). The isolates were all gram‐positive, facultative anaerobic cocci that lacked catalase activity. Analysis of partial 16S rRNA gene sequences showed that the most closely related species was Streptococcus infantis (96.7%). The next most closely related species to the isolates were S. rubneri, S. mitis, S. peroris and S. australis (96.6 to 96.4%). Based on the rpoB and gyrB gene sequences, TKU50^T was clustered with other member of the mitis group. Enzyme activity and sugar fermentation patterns differentiated this novel bacterium from other members of the mitis group streptococci. The DNA G + C content of strain TKU50^T was 46.7 mol%, which is the highest reported value for members of the mitis group (40–46 mol%). On the basis of the phenotypic characterization, partial 16S rRNA gene and sequences data for two housekeeping gene (gyrB and rpoB), we propose a novel taxa, S. panodentis for TKU 50^T (type strain = CM 30579^T = DSM 29921^T), for these newly described isolates.