NPM1 による新たな遺伝子発現制御機構

筑波大学 (University of Tsukuba)201

Quantifying late-stage host-seeking behaviour of Anopheles gambiae at the insecticidal net interface using a baited-box bioassay

Background Insecticide-treated nets (ITNs) are losing efficacy against pyrethroid-resistant malaria vector populations throughout Africa. Safeguarding bed net efficacy, vital for effective malaria control, requires greater knowledge of mosquito–ITN interactions and how this impacts on the mosquito. Methods A purpose-built benchtop apparatus with a closed 10 cm cubic chamber (the ‘Baited-box’) was used to video record behaviour of individual free-flying female Anopheles gambiae during approach and blood-feeding on a human hand through untreated nets and ITNs at close range. Time and duration of defined behavioural events, and knockdown and mortality at 1- and 24-h post-exposure respectively, were recorded for pyrethroid susceptible and resistant mosquitoes. Results Using three human volunteers differing in relative attractiveness to mosquitoes, 328 mosquitoes were individually tested. There were no significant differences between response rates to ITNs and untreated nets (P > 0.1) or between resistant (Tiassalé) and susceptible (Kisumu) mosquito strains, at untreated nets (P = 0.39) or PermaNet 2.0 (P = 1). The sequence of behavioural events from host-seeking to completion of blood-feeding was consistent in all tests but duration and start time of events involving net contact were reduced or delayed respectively with ITNs. Blood-feeding durations at untreated nets (means from 4.25 to 8.47 min (95% confidence interval (CI) = 3.39–9.89) at 3 human volunteers) were reduced by 37–50% at PermaNet 2.0, in susceptible (mean 2.59–4.72 min, 95% CI = 1.54–5.5, P =  0.2), and neither ITN type showed detectable spatial repellency. After initial contact, blood-feeding commenced later at Olyset (mean 2.76 min, 95% CI = 1.74–3.76, P = 0.0009) and PermaNet (mean 2.4 min, 95% CI = 1.52–3.33, P = 0.0058) than untreated netting (mean 0.68 min, 95% CI = 0.42–0.94). Conclusions The baited box offers a simple method for detailed characterization of mosquito behavioural responses to insecticidal nets, for comparing entomological modes of action between nets and for defining the behavioural responses of particular mosquito strains or populations. The device has potential as a screening assay in the search for novel net treatments and for investigations into behavioural resistance mechanisms

Elimination of Specific miRNAs by Naked 14-nt sgRNAs

tRNase ZL-utilizing efficacious gene silencing (TRUE gene silencing) is a newly developed technology to suppress mammalian gene expression. TRUE gene silencing works on the basis of a unique enzymatic property of mammalian tRNase ZL, which is that it can recognize a pre-tRNA-like or micro-pre-tRNA-like complex formed between target RNA and artificial small guide RNA (sgRNA) and can cleave any target RNA at any desired site. There are four types of sgRNA, 5′-half-tRNA, RNA heptamer, hook RNA, and ∼14-nt linear RNA. Here we show that a 14-nt linear-type sgRNA against human miR-16 can guide tRNase ZL cleavage of miR-16 in vitro and can downregulate the miR-16 level in HEK293 cells. We also demonstrate that the 14-nt sgRNA can be efficiently taken up without any transfection reagents by living cells and can exist stably in there for at least 24 hours. The naked 14-nt sgRNA significantly reduced the miR-16 level in HEK293 and HL60 cells. Three other naked 14-nt sgRNAs against miR-142-3p, miR-206, and miR-19a/b are also shown to downregulate the respective miRNA levels in various mammalian cell lines. Our observations suggest that in general we can eliminate a specific cellular miRNA at least by ∼50% by using a naked 14-nt sgRNA on the basis of TRUE gene silencing

Infrared video tracking of Anopheles gambiae at insecticide-treated bed nets reveals rapid decisive impact after brief localised net contact

Long-lasting insecticidal bed nets (LLINs) protect humans from malaria transmission and are fundamental to malaria control worldwide, but little is known of how mosquitoes interact with nets. Elucidating LLIN mode of action is essential to maintain or improve efficacy, an urgent need as emerging insecticide resistance threatens their future. Tracking multiple free-flying Anopheles gambiae responding to human-occupied bed nets in a novel large-scale system, we characterised key behaviours and events. Four behavioural modes with different levels of net contact were defined: swooping, visiting, bouncing and resting. Approximately 75% of all activity occurred at the bed net roof where multiple brief contacts were focussed above the occupant’s torso. Total flight and net contact times were lower at LLINs than untreated nets but the essential character of the response was unaltered. LLINs did not repel mosquitoes but impacted rapidly: LLIN contact of less than 1 minute per mosquito during the first ten minutes reduced subsequent activity; after thirty minutes, activity at LLINs was negligible. Velocity measurements showed that mosquitoes detected nets, including unbaited untreated nets, prior to contact. This is the most complete characterisation of mosquito-LLIN interactions to date, and reveals many aspects of LLIN mode of action, important for developing the next generation of LLINs

Modulation of Gene Expression by Human Cytosolic tRNase ZL through 5′-Half-tRNA

A long form (tRNase ZL) of tRNA 3′ processing endoribonuclease (tRNase Z, or 3′ tRNase) can cleave any target RNA at any desired site under the direction of artificial small guide RNA (sgRNA) that mimics a 5′-half portion of tRNA. Based on this enzymatic property, a gene silencing technology has been developed, in which a specific mRNA level can be downregulated by introducing into cells a synthetic 5′-half-tRNA that is designed to form a pre-tRNA-like complex with a part of the mRNA. Recently 5′-half-tRNA fragments have been reported to exist stably in various types of cells, although little is know about their physiological roles. We were curious to know if endogenous 5′-half-tRNA works as sgRNA for tRNase ZL in the cells. Here we show that human cytosolic tRNase ZL modulates gene expression through 5′-half-tRNA. We found that 5′-half-tRNAGlu, which co-immunoprecipitates with tRNase ZL, exists predominantly in the cytoplasm, functions as sgRNA in vitro, and downregulates the level of a luciferase mRNA containing its target sequence in human kidney 293 cells. We also demonstrated that the PPM1F mRNA is one of the genuine targets of tRNase ZL guided by 5′-half-tRNAGlu. Furthermore, the DNA microarray data suggested that tRNase ZL is likely to be involved in the p53 signaling pathway and apoptosis

A Supersymmetric Model with an Extra U(1) Gauge Symmetry

In the standard model the proton is protected from decay naturally by gauge symmetries, whereas in the ordinary minimal supersymmetric standard model an ad hoc discrete symmetry is imposed for the proton stability. We present a new supersymmetric model in which the proton decay is forbidden by an extra U(1) gauge symmetry. Particle contents are necessarily increased to be free from anomalies, incorporating right-handed neutrinos. Both Dirac and Majorana masses are generated for neutrinos, yielding non-vanishing but small masses. The superpotential consists only of trilinear couplings and the mass parameter $\mu$ of the minimal model is induced by spontaneous breaking of the U(1) symmetry.Comment: 10 pages, Revte

Supersymmetric Extension of the Standard Model with Naturally Stable Proton

A new supersymmetric standard model based on N=1 supergravity is constructed, aiming at natural explanation for the proton stability without invoking an ad hoc discrete symmetry through R parity. The proton is protected from decay by an extra U(1) gauge symmetry. Particle contents are necessarily increased to be free from anomalies, making it possible to incorporate the superfields for right-handed neutrinos and an SU(2)-singlet Higgs boson. The vacuum expectation value of this Higgs boson, which induces spontaneous breakdown of the U(1) symmetry, yields large Majorana masses for the right-handed neutrinos, leading to small masses for the ordinary neutrinos. The linear coupling of SU(2)-doublet Higgs superfields, which is indispensable to the superpotential of the minimal supersymmetric standard model, is replaced by a trilinear coupling of the Higgs superfields, so that there is no mass parameter in the superpotential. The energy dependencies of the model parameters are studied, showing that gauge symmetry breaking is induced by radiative corrections. Certain ranges of the parameter values compatible with phenomena at the electroweak energy scale can be derived from universal values of masses-squared and trilinear coupling constants for scalar fields at a very high energy scale.Comment: 32 pages, Revtex, 7 figure