MiR-148a Attenuates Paclitaxel Resistance of Hormone-refractory,Drug-resistant Prostate Cancer PC3 Cells by Regulating MSK1 Expression

MicroRNAs are involved in cancer pathogenesis and act as tumor suppressors or oncogenes. It has been recently reported that miR-148a expression is down-regulated in several types of cancer. The functional roles and target genes of miR-148a in prostate cancer, however, remain unknown. In this report, we showed that miR-148a expression levels were lower in PC3 and DU145 hormone-refractory prostate cancer cells in comparison to PrEC normal human prostate epithelial cells and LNCaP hormone-sensitive prostate cancer cells. Transfection with miR-148a precursor inhibited cell growth, and cell migration and invasion, and increased the sensitivity to anti-cancer drug paclitaxel in PC3 cells. Computer-aided algorithms predicted mitogen- and stress-activated protein kinase, MSK1, as a potential target of miR-148a. Indeed, miR-148a overexpression decreased expression of MSK1. Using luciferase reporter assays, we identified MSK1 as a direct target of miR-148a. Suppression of MSK1 expression by siRNA, however, showed little or no effects on malignant phenotypes of PC3 cells. In PC3PR cells, a paclitaxel-resistant cell line established from PC3 cells, miR-148a inhibited cell growth, and cell migration and invasion, and also attenuated the resistance to paclitaxel. MiR-148a reduced MSK1 expression by directly targeting its 3′-UTR in PC3PR cells. Furthermore, MSK1 knockdown reduced paclitaxel-resistance of PC3PR cells, indicating that miR-148a attenuates paclitaxel-resistance of hormone-refractory, drug-resistant PC3PR cells in part by regulating MSK1 expression. Our findings suggest that miR-148a plays multiple roles as a tumor suppressor and can be a promising therapeutic target for hormone-refractory prostate cancer especially for drug-resistant prostate cancer.     

Rapid translocation of nanoparticles from the lung airspaces to the body

Nano-size particles show promise for pulmonary drug delivery, yet their behavior after deposition in the lung remains poorly understood. In this study, a series of near-infrared (NIR) fluorescent nanoparticles were systematically varied in chemical composition, shape, size and surface charge, and their biodistribution and elimination were quantified in rat models after lung instillation. We demonstrate that nanoparticles with hydrodynamic diameter (HD) less than ≈34 nm and a noncationic surface charge translocate rapidly from the lung to mediastinal lymph nodes. Nanoparticles of HD < 6 nm can traffic rapidly from the lungs to lymph nodes and the bloodstream, and then be subsequently cleared by the kidneys. We discuss the importance of these findings for drug delivery, air pollution and carcinogenesis.     

Antimicrobial activity of long-chain (E)-3-alken-2-ones

(E)-3-Tridecen-2-one, a compound identified from the interdigital glands of black-tailed deer (Odocoileus hemionus columbianus), has been shown to inhibit the growth of bacteria and fungi. Homologues of (E)-3-tridecen-2-one were prepared and screened for antimicrobial activity. For the fungus, Trichophyton mentagrophytes, the minimum inhibitory concentration (MIC) of (E)-3-Tetradecen-2-one was 12.5 μg/mL, and for the bacteria, Propionibacterium acnes, the MIC of (E)-3-heptadecen-2-one was 3.13 μg/mL.     

Rapid Accumulation of Virulent Rift Valley Fever Virus in Mice from an Attenuated Virus Carrying a Single Nucleotide Substitution in the M RNA

Background

Rift Valley fever virus (RVFV), a member of the genus Phlebovirus within the family Bunyaviridae, is a negative-stranded RNA virus with a tripartite genome. RVFV is transmitted by mosquitoes and causes fever and severe hemorrhagic illness among humans, while in livestock it causes fever and high abortion rates.

Methodology/Principal Findings

Sequence analysis showed that a wild-type RVFV ZH501 preparation consisted of two major viral subpopulations, with a single nucleotide heterogeneity at nucleotide 847 of M segment (M847); one had a G residue at M847 encoding glycine in a major viral envelope Gn protein, while the other carried A residue encoding glutamic acid at the corresponding site. Two ZH501-derived viruses, rZH501-M847-G and rZH501-M847-A, carried identical genomic sequences, except that the former and the latter had G and A, respectively, at M847 were recovered by using a reverse genetics system. Intraperitoneal inoculation of rZH501-M847-A into mice caused a rapid and efficient viral accumulation in the sera, livers, spleens, kidneys and brains, and killed most of the mice within 8 days, whereas rZH501-M847-G caused low viremia titers, did not replicate as efficiently as did rZH501-M847-A in these organs, and had attenuated virulence to mice. Remarkably, as early as 2 days postinfection with rZH501-M847-G, the viruses carrying A at M847 emerged and became the major virus population thereafter, while replicating viruses retained the input A residue at M847 in rZH501-M847-A-infected mice.

Conclusions/Significance

These data demonstrated that the single nucleotide substitution in the Gn protein substantially affected the RVFV mouse virulence and that a virus population carrying the virulent viral genotype quickly emerged and became the major viral population within a few days in mice that were inoculated with the attenuated virus.     

Introgression of Drosophila simulans Nuclear Pore Protein 160 in Drosophila melanogaster Alone Does Not Cause Inviability but Does Cause Female Sterility

We have been analyzing genes for reproductive isolation by replacing Drosophila melanogaster genes with homologs from Drosophila simulans by interspecific backcrossing. Among the introgressions established, we found that a segment of the left arm of chromosome 2, Int(2L)S, carried recessive genes for hybrid sterility and inviability. That nuclear pore protein 160 (Nup160) in the introgression region is involved in hybrid inviability, as suggested by others, was confirmed by the present analysis. Male hybrids carrying an X chromosome of D. melanogaster were not rescued by the Lethal hybrid rescue (Lhr) mutation when the D. simulans Nup160 allele was made homozygous or hemizygous. Furthermore, we uniquely found that Nup160 is also responsible for hybrid sterility. Females were sterile when D. simulans Nup160 was made homozygous or hemizygous in the D. melanogaster genetic background. Genetic analyses indicated that the D. simulans Nup160 introgression into D. melanogaster was sufficient to cause female sterility but that other autosomal genes of D. simulans were also necessary to cause lethality. The involvement of Nup160 in hybrid inviability and female sterility was confirmed by transgene experiment.INVESTIGATING the genetic bases of reproductive isolation is important for understanding speciation (Sawamura and Tomaru 2002; Coyne and Orr 2004; Wu and Ting 2004; Noor and Feder 2006; Presgraves 2010). In fact, continued interest in this issue has led to the isolation of several genes that are responsible for hybrid sterility and inviability in Drosophila (Ting et al. 1998; Barbash et al. 2003; Presgraves et al. 2003; Brideau et al. 2006; Masly et al. 2006; Phadnis and Orr 2009; Prigent et al. 2009; Tang and Presgraves 2009). Drosophila melanogaster and Drosophila simulans are the best pair for such genetic analyses (Sturtevant 1920). Hybrid male lethality in the cross between D. melanogaster females and D. simulans males is caused by incompatibility involving chromatin-binding proteins (Barbash et al. 2003; Brideau et al. 2006), and hybrid female lethality in the reciprocal cross is caused by incompatibility between a maternally supplied factor and a repetitive satellite DNA (Sawamura et al. 1993a; Sawamura and Yamamoto 1997; Ferree and Barbash 2009). Furthermore, individuals with the genotype equivalent to the backcrossed generation exhibit different incompatibilities (Pontecorvo 1943; Presgraves 2003), two components of which have been identified (Presgraves et al. 2003; Tang and Presgraves 2009). Because of the discovery of rescuing mutations that prevent hybrid inviability and sterility (Watanabe 1979; Hutter and Ashburner 1987; Sawamura et al. 1993a,b; Davis et al. 1996; Barbash and Ashburner 2003), chromosome segments from D. simulans can be introgressed into the D. melanogaster genome (Sawamura et al. 2000; Masly et al. 2006). For example, introgression of the D. simulans chromosome 4 or Y into D. melanogaster results in male sterility (Muller and Pontecorvo 1940; Orr 1992), and the recessive sterility by the chromosome 4 introgression is attributed to an interspecific gene transposition between chromosomes (Masly et al. 2006).The other successful introgressions of this type are the tip and the middle regions of the left arm of chromosome 2, Int(2L)D and Int(2L)S, respectively (Sawamura et al. 2000). Both female and male Int(2L)S homozygotes are sterile (Figure 1A), and the recessive sterility genes have been mapped with recombination and complementation assays against deficiencies. The male sterility genes are polygenic and interact epistatically with each other (Sawamura and Yamamoto 2004; Sawamura et al. 2004b), but the female sterility gene has been mapped to a 170-kb region containing only 20 open reading frames (ORFs) (Sawamura et al. 2004a). Interestingly, Int(2L)S also carries a recessive lethal gene whose effect is detected only in a specific genotype (Figure 1B). Lethality in hybrid males from the cross between D. melanogaster females and D. simulans males is rescued by the Lethal hybrid rescue (Lhr) mutation in D. simulans (Watanabe 1979), but the hybrid males cannot be rescued if they carry the introgression, presumably because of incompatibility between an X-linked gene(s) of D. melanogaster and a homozygous D. simulans gene in the Int(2L)S region (Sawamura 2000). Because the female sterility gene and the lethal gene were not separated by recombination, Sawamura et al. (2004a) suggested that female sterility and lethality may be a consequence of the pleiotropic effects of a single gene.Open in a separate windowFigure 1.—Viability and fertility of flies with various genotypes. (A) Females and males that are heterozygous or homozygous for the D. simulans introgression Int(2L)S (Int) in the D. melanogaster genetic background. (B) Four genotypic classes from the cross between introgression heterozygote [Int(2L)S/CyO] females and D. simulans Lethal hybrid rescue (Lhr) males. (C) Four genotypic classes from the cross between D. melanogaster females with a deficiency (Df) [Df(2L)/CyO] and D. simulans Lhr males. Open chromosome regions are from D. melanogaster, and shaded ones are from D. simulans.Because the hybrid lethal gene on Int(2L)S is recessive, the gene can be mapped by deficiencies instead of using introgression (Figure 1C) (Sawamura 2000; Sawamura et al. 2004a). In fact, hybrid males carrying a deficiency encompassing this region (hemizygous for the D. simulans genes) and the D. melanogaster X chromosome are lethal even if they carry the hybrid rescue mutation (see also Presgraves 2003). Tang and Presgraves (2009) subsequently narrowed down this region with multiple deficiencies and identified the hybrid lethal gene with a complementation test and transformation. We confirmed their conclusion and report our data here. In the transformation experiment, we used the natural promoter of the gene, instead of overexpressing the gene (Tang and Presgraves 2009), and we directly indicated, for the first time, that the hybrid lethal gene is also responsible for the female sterility of introgression homozygotes. The D. simulans allele of the gene seems to be nonfunctional on the genetic background of D. melanogaster. Moreover, our results indicated that this gene and chromosome X of D. melanogaster are not sufficient to explain the inviability and that another autosomal gene(s) in D. simulans is required.     

Use of digitally modified videos to examine female mate preference for orange spot coloration of males in the guppy, Poecilia reticulata

Digitally modified video techniques may provide some advantages for the behavioral study of animals, including fishes. However, the validity of the experiments that are conducted using digitally modified videos should be verified. In the present study, we examined female mate preference in the guppy, Poecilia reticulata in a Japanese population by exploring an appropriate method involving the use of digitally modified video playbacks. For the male stimulus, two video images were produced by digital modification of a video recording of a male; the brightness of the orange spot coloration of the male was enhanced (bright male) in one video and decreased (dull male) in the other video. In experiment 1, female choice was examined according to the method described in previous studies, i.e., two monitors were set on opposite sides of the test aquarium to simultaneously present two stimulus male images to the females. However, in this experiment, the stimulus male images did not attract the females to a greater extent than the blank aquarium images. In addition, although the first trial showed a female preference for bright males, in the second trial, in which each stimulus male image was shown from the monitor opposite to that used in the first trial, a significant preference for bright males could not be detected. In experiment 2, a monitor was placed on one side of the aquarium and the left and right sides of the monitor showed the two stimulus male images simultaneously. In addition, a 24-h time interval was set between the first and second trials. Females spent more time in the proximity of the male images than that of the blank images, and a female preference for bright males was observed in both trials. The results of this study clearly indicate the female preference for males possessing bright coloration of orange spots. These results also suggest that the one-sided monitor system adopted in experiment 2 is a more appropriate method than the two-sided monitor system to examine female mate preferences by using digitally modified video playbacks.     

A novel branching enzyme of the GH-57 family in the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1

Branching enzyme (BE) catalyzes formation of the branch points in glycogen and amylopectin by cleavage of the alpha-1,4 linkage and its subsequent transfer to the alpha-1,6 position. We have identified a novel BE encoded by an uncharacterized open reading frame (TK1436) of the hyperthermophilic archaeon Thermococcus kodakaraensis KOD1. TK1436 encodes a conserved protein showing similarity to members of glycoside hydrolase family 57 (GH-57 family). At the C terminus of the TK1436 protein, two copies of a helix-hairpin-helix (HhH) motif were found. TK1436 orthologs are distributed in archaea of the order Thermococcales, cyanobacteria, some actinobacteria, and a few other bacterial species. When recombinant TK1436 protein was incubated with amylose used as the substrate, a product peak was detected by high-performance anion-exchange chromatography, eluting more slowly than the substrate. Isoamylase treatment of the reaction mixture significantly increased the level of short-chain alpha-glucans, indicating that the reaction product contained many alpha-1,6 branching points. The TK1436 protein showed an optimal pH of 7.0, an optimal temperature of 70 degrees C, and thermostability up to 90 degrees C, as determined by the iodine-staining assay. These properties were the same when a protein devoid of HhH motifs (the TK1436DeltaH protein) was used. The average molecular weight of branched glucan after reaction with the TK1436DeltaH protein was over 100 times larger than that of the starting substrate. These results clearly indicate that TK1436 encodes a structurally novel BE belonging to the GH-57 family. Identification of an overlooked BE species provides new insights into glycogen biosynthesis in microorganisms.     

Variable dependence on detrital and grazing food webs by generalist predators: aerial insects and web spiders

Recent studies have shown that organisms from the detritus food web subsidize generalist predators in aboveground food webs, but its significance in space and time is largely unknown. Here we report seasonal dynamics of aerial insects from grazing and detritus food webs in both forest and grassland habitats, and show how these patterns influence the dependence of web spiders on the detritus food web. Detrital insects were more abundant in spring, decreased in summer, and then increased slightly in autumn. This pattern was most conspicuous in Nematocera. Due to different seasonal activity patterns of grazing and detrital insects, the proportion of detrital insects was greater in spring and autumn. Detrital insects were relatively more abundant in the forest than in the grassland. Prey captured by web spiders generally reflected seasonal and spatial patterns of aerial insect abundance. In particular, Leucauge spiders reversed their dependence on the two food webs seasonally. Body size of spiders was negatively correlated with the proportion of detrital prey, suggesting that the detrital subsidy is essential for relatively small predators. This size effect probably resulted from interaction of the following two factors: 1) the maximum body size of prey that can be caught increased with spider body size, 2) larger body size classes of aerial insects included a higher proportion of insects from the grazing food web.