Efficient genome-wide first-generation phenotypic screening system in mice using the piggyBac transposon.

Genome-wide phenotypic screens provide an unbiased way to identify genes involved in particular biological traits, and have been widely used in lower model organisms. However, cost and time have limited the utility of such screens to address biological and disease questions in mammals. Here we report a highly efficient piggyBac (PB) transposon-based first-generation (F1) dominant screening system in mice that enables an individual investigator to conduct a genome-wide phenotypic screen within a year with fewer than 300 cages. The PB screening system uses visually trackable transposons to induce both gain- and loss-of-function mutations and generates genome-wide distributed new insertions in more than 55% of F1 progeny. Using this system, we successfully conducted a pilot F1 screen and identified 5 growth retardation mutations. One of these mutants, a Six1/4 PB/+ mutant, revealed a role in milk intake behavior. The mutant animals exhibit abnormalities in nipple recognition and milk ingestion, as well as developmental defects in cranial nerves V, IX, and X. This PB F1 screening system offers individual laboratories unprecedented opportunities to conduct affordable genome-wide phenotypic screens for deciphering the genetic basis of mammalian biology and disease pathogenesis.

[Basic soil productivity in the double rice cropping system under long-term fertilization regimes in the Poyang Lake region, China]. / 鄱阳湖流域长期施肥下双季稻田的土壤基础地力.

We aimed to explore changes in basic soil productivity (BSP) under different fertilization regimes in the Poyang Lake region, Jiangxi Province, China. Soil samples were collected from a long-term fertilization experiment (since 1981) that included treatments of no fertilization (CK), chemical fertilization (NPK), and combined chemical and organic fertilization (NPKM). Then, a three-year pot experiment (from 2012 to 2014) with double rice cropping was conducted with two different fertilization regimes (no fertilization, F0; fertilization, F1) using CK, NPK and NPKM soils. Grain yield and BSP were analyzed among soils with different fertilization regimes to identify the key factors driving changes in BSP. Results showed that grain yields in NPKM soil were higher than in NPK and CK soils regardless of fertilization in the pot experiment. Under the F0 condition, annual grain yields of NPKM soil were 37.7%-143.9% and 20.8%-66.7% higher than CK and NPK soils, respectively. The BSP values of CK, NPK and NPKM soils in three years were 41.8%-53.1%, 45.2%-62.6% and 59.1%-88.1%, respectively. NPKM soil had significantly higher BSP than NPK and CK soils. Furthermore, there were significant positive correlations between soil organic matter and BSP as well as between organic carbon balance and BSP. These results suggested that long-term application of chemical and organic fertilizers could improve BSP in the double rice cropping system of the Poyang Lake region. In addition, soil organic matter and organic carbon balance are important factors for improving BSP in this region.

EGFR/ARF6 regulation of Hh signalling stimulates oncogenic Ras tumour overgrowth.

Multiple signalling events interact in cancer cells. Oncogenic Ras cooperates with Egfr, which cannot be explained by the canonical signalling paradigm. In turn, Egfr cooperates with Hedgehog signalling. How oncogenic Ras elicits and integrates Egfr and Hedgehog signals to drive overgrowth remains unclear. Using a Drosophila tumour model, we show that Egfr cooperates with oncogenic Ras via Arf6, which functions as a novel regulator of Hh signalling. Oncogenic Ras induces the expression of Egfr ligands. Egfr then signals through Arf6, which regulates Hh transport to promote Hh signalling. Blocking any step of this signalling cascade inhibits Hh signalling and correspondingly suppresses the growth of both, fly and human cancer cells harbouring oncogenic Ras mutations. These findings highlight a non-canonical Egfr signalling mechanism, centered on Arf6 as a novel regulator of Hh signalling. This explains both, the puzzling requirement of Egfr in oncogenic Ras-mediated overgrowth and the cooperation between Egfr and Hedgehog.

[Abundance of Toxic and Non-toxic Microcystis sp. in Lake Hongze and Its Correlation with Environmental Factors].

In the present study, the eutrophic level of 30 water samples collected from Lake hongze in August 2014 were analyzed, and the abundance of toxic and non-toxic Microcystis sp., together with their spatial distribution, was investigated by quantitative real-time PCR techniques. The results showed that the average concentrations of total nitrogen and total phosphorus were 1.63 and 0.11 mg x L(-1), respectively. The trophic state index ( TSI) ranged from 58.1 to 73.6, and the water quality was in the state of eutrophication based on TSI. Toxic Microcystis was widely distributed in Lake Hongze, and its abundance varied sharply, from 1. 13 x 10(4) to 3.51 x 10(6) copies x mL(-1), and the abundance of total Microcystis ranged from 1.06 x 10(5) to 1.10 x 10(7) copies x m(-1), meanwhile, the proportion of toxic Microcystis in the total Microcystis ranged from 8.5% to 38.5%, with the average value of 23.6%. Correlation analysis indicated that there was a significant positive correlation among total Mirocystis, toxic Microcystis and the toxic proportion (P < 0.01). The abundance of total and toxic Microcystis was significantly positively correlated to chlorophyll a ( Chl-a) concentrations and TSI (P < 0.01), but was negatively correlated to transparency (SD) (P < 0.01). The ratio of toxic Microcystis to total Microcystis was significantly positively correlated to Chl-a, TN, TP and TSI (P < 0.01), but significantly negatively correlated to the ratio of TN to TP and SD (P < 0.01). Therefore, reducing total nitrogen and phosphorus concentrations could not only lower the eutrophication level of Lake Hongze, but also inhibit the competition advantage of the toxic Microcystis over non-toxic Microcystis.

γ-Tocotrienol induces paraptosis-like cell death in human colon carcinoma SW620 cells.

Colorectal cancer is one of the most serious illnesses among diagnosed cancer. As a new type of anti-cancer composition from tocotrienol-rich fraction of palm oil, γ-tocotrienol is widely used in anti-cancer research. The objectives of this study were to investigate the effects of γ-tocotrienol on human colon cancer SW620 and HCT-8 cells. We showed that treatment with different concentrations of γ-tocotrienol resulted in a dose dependent inhibition of cell growth. Cell death induced by γ-tocotrienol was mediated by a paraptosis-like cell death in SW620 and HCT-8 cells. Real-time RT-PCR and western blot analyses showed that γ-tocotrienol inhibited the expression level of ß-catenin, cyclin D1 and c-jun. These data suggest that a paraptosis-like cell death induced by γ-tocotrienol in SW620 cells is associated with the suppression of the Wnt signaling pathway, which offers a novel tool for treating apoptosis-resistance colon cancer.

[Regional difference of schistosomiasis knowledge and behavior of population in Mianzhu City].

OBJECTIVE: To understand the status of schistosomiasis knowledge and behavior and analyze the regional difference of population in Mianzhu City. METHODS: Nine towns were divided into 3 groups, and each group had 3 towns. In Group I , there were 5 or more than 5 advanced schistosomiasis patients each town; in Group II , there were 1-4 advanced patients each town; in Group III, there was no advanced patient. A total of 2 743 residents were investigated with questionnaire in all the 9 towns. RESULTS: The overall awareness rate of schistosomiasis control knowledge was 88.12%, and the awareness rates of schistosomiasis control knowledge were 94.55%, 88.21% and 81.10% in Group I , Group II and Group III respectively. The total formation rate of correct behavior was 68.10%, and the formation rates of correct behavior were 73.18%, 67.05% and 63.65% in Group I , Group II and Group III respectively. The awareness rates of schistosome transmission were 95.99%, 89.48% and 79.67%; the awareness rates of Oncomelania snails were 87.67%, 82.54% and 73.92%; the awareness rates of schistosomiasis harm were 95.68%, 93.99% and 80.88%; the rates of residents who thought that schistosomiasis patients did not affect others were 9.97%, 12.83% and 15.58%; the rates of residents who did not know the information of the snails should report to which department were 7.91%, 11.33%, 15.69%; the rates of residents who often had wild bowels were 6.17%, 8.79% and 11.38%; the rates of residents who often washed their hands and feet in ditches and ponds were 58.68%, 58.27%, 61.22%; the rates of residents who would not accept the schistosomiasis checks were 5.86%, 5.66%, 11.49% in Group I , Group II and Group III respectively. CONCLUSION: As a whole, the population of Mianzhu City has positive behaviors to schistosomiasis control. We should still enhance the schistosomiasis control education and interventions according to the characteristics of the different townships.

A paraptosis-like cell death induced by δ-tocotrienol in human colon carcinoma SW620 cells is associated with the suppression of the Wnt signaling pathway.

Tocotrienol is considered a beneficial effect agent on inhibition of tumor development. In this study, we focused on the effects of δ-tocotrienol and its possible mechanism on induction of death in human colon cancer SW620 cells. δ-Tocotrienol inhibited proliferation of SW620 cell in a dose-dependent manner. Our findings showed that δ-tocotrienol effectively induced paraptosis-like death in SW620 cells, correlated with the vacuolation that may be from welling and fusion of mitochondria and/or the endoplasmic reticulum (ER) as well as caspase-3 nonactivated. However, there were no changes in apoptosis based on flow cytometry analysis. Of being noted, δ-tocotrienol reduced the expression of ß-catenin and wnt-1 proteins by about 50% at the highest dose (20µmol/L). δ-Tocotrienol also decreased cyclin D1, c-jun and MMP-7 protein levels in SW620 cells. Altogether, these data indicate that δ-tocotrienol induces paraptosis-like cell death, which is associated with the suppression of the Wnt signaling pathway. Thus, our findings may provide a novel application in treatment of human colon carcinoma.

[Surveillance of schistosomiasis of reconstruction personnel after earthquake in Mianzhu City from 2009 to 2011].

After "5.12" earthquake, 5 128 reconstruction people from schistosomiasis endemic areas were surveyed for schistosome infection from 2009 to 2001. There were 261 seropositive persons with the positive rate of 5.09%, but there were no persons with positive stool examination. The seropositive rate was higher in reconstruction persons from schistosomiasis endemic areas than that in local residents, therefore, we still should strengthen the active schistosomiasis surveillance of reconstruction people from schistosomiasis endemic areas in order to prevent input of infection source so as to consolidate the achievements of schistosomiasis control.

LATS1 tumour suppressor affects cytokinesis by inhibiting LIMK1.

LATS (large tumour suppressor) is a family of conserved tumour suppressors identified in Drosophila and mammals. Here we show that human LATS1 binds to LIMK1 in vitro and in vivo and colocalizes with LIMK1 at the actomyosin contractile ring during cytokinesis. LATS1 inhibits both the phosphorylation of cofilin by LIMK1 and LIMK1-induced cytokinesis defects. Inactivation of LATS1 by antibody microinjection or RNA-mediated interference in cells, or gene knockout in mice, abrogates cytokinesis and increases the percentage of multinucleate cells. Our findings indicate that LATS1 is a novel cytoskeleton regulator that affects cytokinesis by regulating actin polymerization through negative modulation of LIMK1.

Drosophila Src-family kinases function with Csk to regulate cell proliferation and apoptosis.

Elevated Src protein levels and activity are associated with the development and progression of a variety of cancers. The consequences of deregulated Src activity have been studied extensively in cell culture; however, the effects of this deregulation in vivo, as well as the mechanisms of Src-induced tumorigenesis, remain poorly understood. In this study, the effect of expressing wild-type and constitutively active Drosophila Src-family kinases (SFKs) in the developing eye was examined. Overexpression of either wild-type Drosophila SFK (Src64 and Src42) is sufficient to induce ectopic proliferation in G1/G0-arrested, uncommitted cells in eye imaginal discs. In addition, both kinases trigger apoptosis in vivo, in a dosage-dependent manner. Constitutively active mutants are hypermorphic as they trigger proliferation and death more potently than their wild-type counterparts. Moreover, SFK-induced proliferation and apoptosis are largely independent events, as blocking ectopic proliferation does not block cell death. Further, DCsk (the Drosophila homolog of the C-terminal Src kinase) phosphorylates and interacts genetically with the wild-type SFKs, but not with the constitutively active mutants in which a conserved C-terminal tyrosine was mutated to phenylalanine, providing the first in vivo evidence that Csk regulates SFKs during development through phosphorylation of their C-terminal tyrosine.

A genetic screen for modifiers of the lats tumor suppressor gene identifies C-terminal Src kinase as a regulator of cell proliferation in Drosophila.

Disrupting mechanisms that control cell proliferation, cell size and apoptosis can cause changes in animal and tissue size and contribute to diseases such as cancer. The LATS family of serine/threonine kinases control tissue size by regulating cell proliferation and function as tumor suppressor genes in both Drosophila and mammals. In order to understand the role of lats in size regulation, we performed a genetic modifier screen in Drosophila to identify components of the lats signaling pathway. Mutations in the Drosophila homolog of C-terminal Src kinase (dcsk) were identified as dominant modifiers of both lats gain-of-function and loss-of-function phenotypes. Homozygous dcsk mutants have enlarged tissue phenotypes similar to lats and FACS and immunohistochemistry analysis of these tissues revealed that dcsk also regulates cell proliferation during development. Animals having mutations in both dcsk and lats display cell overproliferation phenotypes more severe than either mutant alone, demonstrating these genes function together in vivo to regulate cell numbers. Furthermore, homozygous dcsk phenotypes can be partially suppressed by overexpression of lats, indicating that lats is a downstream mediator of dcsk function in vivo. Finally, we show that dCSK phosphorylates LATS in vitro at a conserved C-terminal tyrosine residue, which is critical for normal LATS function in vivo. Taken together, these results demonstrate a role for dCSK in regulating cell numbers during development by inhibiting cell proliferation and suggest that lats is one of the mediators of the dcsk phenotype.