Identification of a gene, Desiccate, contributing to desiccation resistance in Drosophila melanogaster

Suitable alterations in gene expression are believed to allow animals to survive drastic changes in environmental conditions. Drosophila melanogaster larvae cease eating and exit moist food to search for dry pupation sites after the foraging stage in what is known as the wandering stage. Although the behavioral change from foraging to wandering causes desiccation stress, the mechanism by which Drosophila larvae protect themselves from desiccation remains obscure. Here, we identified a gene, CG14686 (designated as Desiccate (Desi)), whose expression was elevated during the wandering stage. The Desi expression level was reversibly decreased by transferring wandering larvae to wet conditions and increased again by transferring them to dry conditions. Elevation of Desi expression was also observed in foraging larvae when they were placed in dry conditions. Desi encoded a 261-amino acid single-pass transmembrane protein with notable motifs, such as SH2 and PDZ domain-binding motifs and a cAMP-dependent protein kinase phosphorylation motif, in the cytoplasmic region, and its expression was observed mainly in the epidermal cells of the larval integuments. Overexpression of Desi slightly increased the larval resistance to desiccation stress during the second instar. Furthermore, Desi RNAi larvae lost more weight under dry conditions, and subsequently, their mortalities significantly increased compared with control larvae. Under dry conditions, consumption of carbohydrate was much higher in Desi RNAi larvae than control larvae. Based on these results, it is reasonable to conclude that Desi contributes to the resistance of Drosophila larvae to desiccation stress.     

Phase-dependent locomotor activity in first-stadium nymphs of the desert locust, Schistocerca gregaria: effects of parental and progeny rearing density

This study examined the effects of parental and progeny rearing density on locomotor activity of 1st-stadium nymphs of the desert locust, Schistocerca gregaria, using an actograph. Progeny obtained from solitarious (isolated-reared) or gregarious (crowd-reared) locusts were reared in isolation or in a group of 30 nymphs. Crowding after hatching had a slight influence on mean activity shortly after the start of measurements, but no clear effect was detected until day 2, when maximum activity during the 6-24 h of observation was significantly higher than that of the nymphs kept in isolation. On the other hand, the effects of parental rearing density on locomotor activity manifested at all ages examined (0-2 days old). Progeny of gregarious locusts showed consistently higher activity than those of solitarious locusts. In newly hatched nymphs, the effect of parental rearing density was explained by variation in body size at hatching, one of the phase-dependent characteristics. Hatchling body color was also correlated with locomotor activity and body weight. Similar levels of locomotor activity were exhibited when green, solitarious and black, gregarious nymphs were similar in body weight. These results suggested that parental rearing density indirectly influences locomotor activity in the progeny shortly after hatching by affecting their body size as eggs or hatchlings.     

Phosphorus deficiency-induced root elongation and its QTL in rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.)

A significant level of root elongation was induced in rice (Oryza sativa) grown under phosphorus-deficient conditions. The root elongation clearly varied among a total of 62 varieties screened under two different phosphorus levels. Two contrasting varieties, Gimbozu, with a low elongating response and Kasalath, with a high elongating response, were chosen and crossed to produce a hybrid population for QTL analyses. QTLs for the phosphorus deficiency-induced root elongation were detected by two linkage maps, i.e., one with 82 F₃ families constructed by 97 simple sequence repeat (SSR) and sequence-tag site markers and another with 97 F₈ lines by 790 amplified fragment length polymorphism and SSR markers. A single QTL for the elongation response was detected on chromosome 6, with a LOD score of 4.5 in both maps and explained about 20% of total phenotypic variance. In addition, this QTL itself, or a region tightly linked with it, partly explained an ability to reduce accumulation of excess iron in the shoots. The identified QTL will be useful to improve rice varieties against a complex nutritional disorder caused by phosphorus deficiency and iron toxicity.Electronic Supplementary Material Supplementary material is available for this article at     

Aerobic exercise training reduces plasma endothelin-1 concentration in older women.

Endothelial function deteriorates with aging. On the other hand, exercise training improves the function of vascular endothelial cells. Endothelin-1 (ET-1), which is produced by vascular endothelial cells, has potent constrictor and proliferative activity in vascular smooth muscle cells and, therefore, has been implicated in regulation of vascular tonus and progression of atherosclerosis. We previously reported significantly higher plasma ET-1 concentration in middle-aged than in young humans, and recently we showed that plasma ET-1 concentration was significantly decreased by aerobic exercise training in healthy young humans. We hypothesized that plasma ET-1 concentration increases with age, even in healthy adults, and that lifestyle modification (i.e., exercise) can reduce plasma ET-1 concentration in previously sedentary older adults. We measured plasma ET-1 concentration in healthy young women (21-28 yr old), healthy middle-aged women (31-47 yr old), and healthy older women (61-69 yr old). The plasma level of ET-1 significantly increased with aging (1.02 +/- 0.08, 1.33 +/- 0.11, and 2.90 +/- 0.20 pg/ml in young, middle-aged, and older women, respectively). Thus plasma ET-1 concentration was markedly higher in healthy older women than in healthy young or middle-aged women (by approximately 3- and 2-fold, respectively). In healthy older women, we also measured plasma ET-1 concentration after 3 mo of aerobic exercise (cycling on a leg ergometer at 80% of ventilatory threshold for 30 min, 5 days/wk). Regular exercise significantly decreased plasma ET-1 concentration in the healthy older women (2.22 +/- 0.16 pg/ml, P < 0.01) and also significantly reduced their blood pressure. The present study suggests that regular aerobic-endurance exercise reduces plasma ET-1 concentration in older humans, and this reduction in plasma ET-1 concentration may have beneficial effects on the cardiovascular system (i.e., prevention of progression of hypertension and/or atherosclerosis by endogenous ET-1).     

Structure and Synthesis of Dopastin,an Inhibitor of Dopamine β-Hydroxylase

Dopastin, an inhibitor of dopamine β-hydroxylase of microbial origin, was shown to be N-[2(S)-nitrosohydroxylamino-3-methylbutyl] crotonamide based on chemical, spectroscopic and synthetic studies. The total synthesis of dopastin was completed in 8 steps starting from l-valinol. N-Nitrosohydroxylamino function was introduced through an oxaziran with retention of the absolute configuration in the final product. Thus, the 2S-configuration of dopastin was proved by the total synthesis. Racemic dopastin was also synthesized from isobutyraldehyde in 7 steps.     

The effect of bispecific monoclonal antibody recognizing both hepatoma-specific membrane glycoprotein and anthracycline drugs on the metastatic growth of hepatoma AH66

A monoclonal mouse antibody (MoHG) was produced using in vitro cultured AH66R tumor cells treated with cholesteryl hemisuccinate as an immunogen. The antibody identified a 90 kd membrane glycoprotein (HG-90) which is expressed on in vitro cultured hepatoma cell lines AH66 and AH66R. A monoclonal antibody was prepared to the anthracycline drug daunomycin, and it also reacted with adriamycin. A fusion was made of the hybridoma HG-90 with the hybridoma which recognized daunomycin/adriamycin. This bispecific hybridoma A8C recognized both determinants. We studied the therapeutic effect of the A8C bispecific antibody with adriamycin treatment and compared it to the effect of the bispecific antibody to which adriamycin had been conjugated via an albumin (Alb) bridge. The therapy model used was the tumor AH66R in Donryu rats. Tumor bearing rats had their subcutaneous tumors resected on day 10, a time when distant metastases were present. After the surgical resection of the tumor the rats were injected intravenously for two cycles with the bispecific antibodies, followed by the administration of adriamycin (ADR) or MoHG.Alb.ADR conjugates. A slight therapeutic effect occurred with either MoHG or ADR alone but treatment with the bispecific antibody followed by the administration of ADR or with the MoHG.Alb.ADR conjugates significantly prolonged survival, with 60% of the treated animals being "tumor free" when sacrificed on day 80. Lower serum concentrations of alphafetoprotein were observed with the bispecific antibody and drug treatment. This suggests that the bispecific antibody/drug treatment is potentially more beneficial in the suppression of distant metastases than the MoHG.Alb.ADR conjugate. This may be due to an increase in the local drug concentration of unmodified adriamycin.     

PLD activation in Chinese hamster ovary (CHO) cells transfected with PGF2α receptor cDNA

Pertussis toxin-insensitive GTP-binding protein was observed to be involved in prostaglandin F2α(PGF2α)-induced phosphoinositide metabolism in Chinese hamster ovary (CHO) cells transfected with PGF2α receptor cDNA (CHO-PGF2α·R cells) (Ito, S. et al. Biochem. Biophys. Res. Commun. 200: 756, 1994). In the present study, we investigated PGF2α-induced PLD activation in CHO-PGF2α·R cells. PLD activation was examined by measuring the production of [³H]phosphatidylbutanol ([³H]PBut), a specific product of the PLD-catalyzed transphosphatidylation reaction. PGF2α-induced [³H]PBut formation was concentration-dependent with the maximal level obtained at 1 μM PGF2α. The maximal [³H]PBut formation was observed at 2 min after addition of PGF2α. Depletion of extracellular Ca²⁺ with EGTA suppressed PGF2α-induced PLD activation by 50%. PKC inhibitors Ro31–8425 and calphostin C inhibited PGF2α-induced [³H]PBut formation by 50%. PTK inhibitors genistein and herbimycin A failed to inhibit PGF2α-induced PLD activation. A combination of maximal effective concentrations of PGF2α (1 μM) and PMA (100 nM) enhanced PLD activation in an additive manner. Pretreatment of the cells with PMA for 2 h down-regulated PKCα and decreased PGF2α-induced PLD activation. These results suggest that PLD activation by PGF2α is mediated by both PKC-dependent and -independent pathways and that PKCα is involved in the former pathway.     

Genome-wide identification of chromatin-enriched RNA reveals that unspliced dentin matrix protein-1 mRNA regulates cell proliferation in squamous cell carcinoma

Chromatin-enriched noncoding RNAs (ncRNAs) have emerged as key molecules in epigenetic processes by interacting with chromatin-associated proteins. Recently, protein-coding mRNA genes have been reported to be chromatin-tethered, similar with ncRNA. However, very little is known about whether chromatin-enriched mRNA is involved in the chromatin modification process. Here, we comprehensively examined chromatin-enriched RNA in squamous cell carcinoma (SQCC) cells by RNA subcellular localization analysis, which was a combination of RNA fractionation and RNA-seq. We identified 11 mRNAs as highly chromatin-enriched RNAs. Among these, we focused on the dentin matrix protein-1 (DMP-1) gene because its expression in SQCC cells has not been reported. Furthermore, we clarified that DMP-1 mRNA was retained in chromatin in its unspliced form in SQCC in vitro and in vivo. As the inhibition of the unspliced DMP-1 mRNA (unspDMP-1) expression resulted in decreased cellular proliferation in SQCC cells, we performed ChIP-qPCR to identify cell cycle-related genes whose expression was epigenetically modified by unspDMP-1, and found that the CDKN1B promoter became active in SQCC cells by inhibiting unspDMP-1 expression. This result was further validated by the increased CDKN1B gene expression in the cells treated with siRNA for unspDMP-1 and by restoration of the decreased cellular proliferation rate by simultaneously inhibiting CDKN1B expression in SQCC cells. Further, to examine whether unspDMP-1 was able to associate with the CDKN1B promoter region, SQCC cells stably expressing PP7-mCherry fusion protein were transiently transfected with the unspDMP-1 fused to 24 repeats of the PP7 RNA stem loop (unspDMP-1-24xPP7) and we found that unspDMP-1-24xPP7 was efficiently precipitated with the antibody against mCherry and was significantly enriched in the CDKN1B promoter region. Thus, unspDMP-1 is a novel chromatin-enriched RNA that epigenetically regulates cellular proliferation of SQCC.