Many animals, including the fruit fly, are sensitive to small differences in ambient temperature. The ability of Drosophila larvae to choose their ideal temperature (18°C) over other comfortable temperatures (19° to 24°C) depends on a thermosensory signaling pathway that includes a heterotrimeric guanine nucleotide-binding protein (G protein), a phospholipase C, and the transient receptor potential TRPA1 channel. We report that mutation of the gene (ninaE) encoding a classical G protein-coupled receptor (GPCR), Drosophila rhodopsin, eliminates thermotactic discrimination in the comfortable temperature range. This role for rhodopsin in thermotaxis toward 18°C was light-independent. Introduction of mouse melanopsin restored normal thermotactic behavior in ninaE mutant larvae. We propose that rhodopsins represent a class of evolutionarily conserved GPCRs that are required for initiating thermosensory signaling cascades.

Female eutherian mammals use X-chromosome inactivation (XCI) to epigenetically regulate gene expression from ~4% of genes. To quantitatively map the topography of XCI for defined cell types at single cell resolution, we have generated female mice that carry X-linked, Cre-activated, and nuclear-localized fluorescent reporters – GFP on one X-chromosome and tdTomato on the other. Using these reporters in combination with different Cre drivers we have defined the topographies of XCI mosaicism for multiple CNS cell types and of retinal vascular dysfunction in a model of Norrie Disease. Depending on cell type, fluctuations in the XCI mosaic are observed over a wide range of spatial scales, from neighboring cells to left vs. right sides of the body. These data imply a major role for XCI in generating female-specific, genetically directed, stochastic diversity in eutherian mammals on spatial scales that would be predicted to affect CNS function within and between individuals.

MicroRNAs (miRNAs) are endogenously expressed small noncoding RNAs. The genomic locations of animal miRNAs are significantly clustered in discrete loci. We found duplication and de novo formation were important mechanisms to create miRNA clusters and the clustered miRNAs tend to be evolutionarily conserved. We proposed a “functional co-adaptation” model to explain how clustering helps newly emerged miRNAs survive and develop functions. We presented evidence that abundance of miRNAs in the same clusters were highly correlated and those miRNAs exerted cooperative repressive effects on target genes in human tissues. By transfecting miRNAs into human and fly cells and extensively profiling the transcriptome alteration with deep-sequencing, we further demonstrated the functional co-adaptation between new and old miRNAs in the miR-17–92 cluster. Our population genomic analysis suggest that positive Darwinian selection might be the driving force underlying the formation and evolution of miRNA clustering. Our model provided novel insights into mechanisms and evolutionary significance of miRNA clustering.

In this work, the interactions between a representative imidazolium-based ionic liquid 1-butyl-3-methylimidazolium tetrafluoroborate ([BMIM][BF4]) and acetonitrile (CH3CN) were investigated in detail using attenuated total reflection infrared spectroscopy (ATR-IR), hydrogen nuclear magnetic resonance ((1)H NMR), and density functional theory calculations. The main conclusions are: (1) a number of species in the [BMIM][BF4]-CH3CN mixtures were identified with the help of excess infrared spectroscopy and quantum chemical calculations. The dilution process of the ionic liquid by acetonitrile was found to be the transformation from ion clusters to ion pairs. (2) The solvent molecules cannot break apart the strong Coulombic interaction between [BMIM](+) and [BF4](-) but can break apart the ion cluster into an ion pair within the concentration range investigated. The strength of hydrogen bonds between the C-Hs of [BMIM](+) and the N of acetonitrile is enhanced during the dilution process. (3) The methyl group of CH3CN locates above/below the imidazolium ring in the solution. These in-depth studies on the properties of the ionic liquid-acetonitrile mixed solvents may shed light on exploring their applications as reaction media in electrochemistry and chemical synthesis.

Transfer RNA-derived small RNAs (tsRNAs) are an emerging class of small RNAs, yet their regulatory roles have not been well understood. Here we studied the molecular mechanisms and consequences of tsRNA-mediated regulation in Drosophila. By analyzing 495 public small RNA libraries, we demonstrate that most tsRNAs are conserved, prevalent and abundant in Drosophila. By carrying out mRNA sequencing and ribosome profiling of S2 cells transfected with single-stranded tsRNA mimics and mocks, we show that tsRNAs recognize target mRNAs through conserved complementary sequence matching and suppress target genes by translational inhibition. The target prediction suggests that tsRNAs preferentially suppress translation of the key components of the general translation machinery, which explains how tsRNAs inhibit the global mRNA translation. Serum starvation experiments confirm tsRNAs participate in cellular starvation responses by preferential targeting the ribosomal proteins and translational initiation or elongation factors. Knock-down of AGO2 in S2 cells under normal and starved conditions reveals a dependence of the tsRNA-mediated regulation on AGO2. We also validated the repressive effects of representative tsRNAs on cellular global translation and specific targets with luciferase reporter assays. Our study suggests the tsRNA-mediated regulation might be crucial for the energy homeostasis and the metabolic adaptation in the cellular systems.

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Avoidance of noxious ambient heat is crucial for survival. A well-known but enigmatic phenomenon is that animals are sensitive to the rate of temperature change. However, the cellular and molecular underpinnings through which animals sense and respond much more vigorously to fast temperature changes are unknown. Using Drosophila larvae, we found that nociceptive rolling behavior was triggered at lower temperatures and at higher frequencies when the temperature increased rapidly. We identified neurons in the brain that were sensitive to the speed of the temperature increase, rather than just the absolute temperature. These cellular and behavioral responses depended on the TRPA1 channel, whose activity was responsive to the rate of temperature increase. We propose that larvae use low threshold sensors in the brain to monitor rapid temperature increases as a protective alert signal to trigger rolling behaviors, allowing fast escape before the temperature of the brain rises to dangerous levels.

SUMMARY Rapsyn, an acetylcholine receptor (AChR)-interacting protein, is essential for synapse formation at the neuromuscular junction (NMJ). Like many synaptic proteins, rapsyn turns over rapidly at synapses. However, little is known about molecular mechanisms that govern rapsyn stability. Using a differential mass-spectrometry approach, we identified heat-shock protein 90β (HSP90β) as a component in surface AChR clusters. The HSP90β-AChR interaction required rapsyn and was stimulated by agrin. Inhibition of HSP90β activity or expression, or disruption of its interaction with rapsyn attenuated agrin-induced formation of AChR clusters in vitro and impaired the development and maintenance of the NMJ in vivo. Finally, we showed that HSP90β was necessary for rapsyn stabilization and regulates its proteasome-dependent degradation. Together, these results indicate a role of HSP90β in NMJ development by regulating rapsyn turnover and subsequent AChR cluster formation and maintenance.