Molecular phylogeny and divergence times of drosophilid species

The phylogenetic relationships and divergence times of 39 drosophilid species were studied by using the coding region of the Adh gene. Four genera--Scaptodrosophila, Zaprionus, Drosophila, and Scaptomyza (from Hawaii)--and three Drosophila subgenera--Drosophila, Engiscaptomyza, and Sophophora--were included. After conducting statistical analyses of the nucleotide sequences of the Adh, Adhr (Adh-related gene), and nuclear rRNA genes and a 905-bp segment of mitochondrial DNA, we used Scaptodrosophila as the outgroup. The phylogenetic tree obtained showed that the first major division of drosophilid species occurs between subgenus Sophophora (genus Drosophila) and the group including subgenera Drosophila and Engiscaptomyza plus the genera Zaprionus and Scaptomyza. Subgenus Sophophora is then divided into D. willistoni and the clade of D. obscura and D. melanogaster species groups. In the other major drosophilid group, Zaprionus first separates from the other species, and then D. immigrans leaves the remaining group of species. This remaining group then splits into the D. repleta group and the Hawaiian drosophilid cluster (Hawaiian Drosophila, Engiscaptomyza, and Scaptomyza). Engiscaptomyza and Scaptomyza are tightly clustered. Each of the D. repleta, D. obscura, and D. melanogaster groups is monophyletic. The splitting of subgenera Drosophila and Sophophora apparently occurred about 40 Mya, whereas the D. repleta group and the Hawaiian drosophilid cluster separated about 32 Mya. By contrast, the splitting of Engiscaptomyza and Scaptomyza occurred only about 11 Mya, suggesting that Scaptomyza experienced a rapid morphological evolution. The D. obscura and D. melanogaster groups apparently diverged about 25 Mya. Many of the D. repleta group species studied here have two functional Adh genes (Adh-1 and Adh-2), and these duplicated genes can be explained by two duplication events.      

In Vivo Identification of Nuclear Factors Interacting with the Conserved Elements of Box C/D Small Nucleolar RNAs

The U16 small nucleolar RNA (snoRNA) is encoded by the third intron of the L1 (L4, according to the novel nomenclature) ribosomal protein gene of Xenopus laevis and originates from processing of the pre-mRNA in which it resides. The U16 snoRNA belongs to the box C/D snoRNA family, whose members are known to assemble in ribonucleoprotein particles (snoRNPs) containing the protein fibrillarin. We have utilized U16 snoRNA in order to characterize the factors that interact with the conserved elements common to the other members of the box C/D class. In this study, we have analyzed the in vivo assembly of U16 snoRNP particles in X. laevis oocytes and identified the proteins which interact with the RNA by label transfer after UV cross-linking. This analysis revealed two proteins, of 40- and 68-kDa apparent molecular size, which require intact boxes C and D together with the conserved 5′,3′-terminal stem for binding. Immunoprecipitation experiments showed that the p40 protein corresponds to fibrillarin, indicating that this protein is intimately associated with the RNA. We propose that fibrillarin and p68 represent the RNA-binding factors common to box C/D snoRNPs and that both proteins are essential for the assembly of snoRNP particles and the stabilization of the snoRNA.One of the most interesting recent findings related to ribosome biogenesis has been the identification of a large number of small RNAs localized in the nucleolus (snoRNAs). So far, more than 60 snoRNAs have been identified in vertebrates (17), and more than 30 have been identified in yeast (2). The total number of snoRNAs is not known, but it is likely to be close to 200 (33, 38). These snoRNAs, with the exception of the mitochondrial RNA processing (MRP) species (38), can be grouped into two major families on the basis of conserved structural and sequence elements. The first group includes molecules referred to as box C/D snoRNAs, whereas the second one comprises the species belonging to the box H/ACA family (2, 15).The two families differ in many aspects. The box C/D snoRNAs are functionally heterogeneous. Most of them function as antisense RNAs in site-specific ribose methylation of the pre-rRNA (1, 10, 17, 26); a minority have been shown to play a direct role in pre-rRNA processing in both yeast and metazoan cells (11, 21). The box C/D snoRNAs play their role by means of unusually long (up to 21 contiguous nucleotides) regions of complementarity to highly conserved sequences of 28S and 18S rRNAs (1). In contrast, several members of the H/ACA RNA family have been shown to direct site-specific isomerization of uridines into pseudouridines and to display shorter regions of complementarity to rRNA (14, 24). Mutational analysis suggests that H/ACA snoRNAs can also play a role as antisense RNAs by base pairing with complementary regions on rRNA (15, 24).Another difference between the two families can be seen by comparison of secondary structures. A Y-shaped motif, where a 5′,3′-terminal stem adjoins the C and D conserved elements, has been proposed for many box C/D snoRNAs (16, 26, 40, 42), whereas box H/ACA snoRNAs have been proposed to fold into two conserved hairpin structures connected by a single-stranded hinge region, followed by a short 3′ tail (15).Despite these differences, analogies have been found in the roles played by the conserved box elements. Mutational analysis and competition experiments indicated that C/D and H/ACA boxes are required both for processing and stable accumulation of the mature snoRNA, suggesting that they represent binding sites for specific trans-acting factors (2, 3, 8, 15, 16, 28, 36, 41).All snoRNAs are associated with proteins to form specific ribonucleoparticles (snoRNPs). The study of these particles began only recently, and so far, very few aspects of their structure and biosynthesis have been clarified. The only detailed analysis performed was on the mammalian U3 (19) and the yeast snR30 (20) snoRNPs. Of the identified components, a few appear to be more general factors: fibrillarin, which was shown to be associated with C/D snoRNPs (3, 4, 8, 13, 28, 31, 39), and the nucleolar protein GAR1, which was found associated with H/ACA snoRNAs in yeast (20). Just as the study of small nuclear RNP (snRNP) particles was crucial to the understanding of the splicing process, a detailed structural and functional analysis of snoRNP particles will be essential to elucidate the complex process of ribosome biosynthesis.In this study, we have analyzed the snoRNP assembly of wild-type and mutant U16 snoRNAs by following the kinetics of complex formation in the in vivo system of the Xenopus laevis oocyte. By a UV cross-linking technique, we have identified two proteins, of 40- and 68-kDa apparent molecular mass, which require intact boxes C and D together with the terminal stem for their binding. The 40-kDa species is specifically recognized by fibrillarin antibodies, indicating that this protein is intimately associated with the RNA.     

东亚飞蝗hunchback基因在卵子形成和胚胎发育过程中的原位表达

hb（hunchback）基因是昆虫胚胎前后轴模式形成的关键基因．对东亚飞蝗（Locusta migratoria manilensis,Meyen）hb基因的功能已有报道,但其表达模式还不清楚．为了研究胁基因在东亚飞蝗卵子形成和胚胎发育过程中的时空表达情况,本研究采用免疫组化方法在蛋白质水平上检测了hb基因的时空表达模式．在卵子形成过程中,hb基因局限在卵细胞核区中表达,随着卵子的发育逐渐移至卵细胞的后端;卵受精后,核区里的Hb蛋白向外扩散,在卵后端形成浓度梯度;胚盘期,hb基因在胚盘中央呈带状表达;胚盘分化为原头和原躯干后,表达条带变宽,并呈现出梯度表达,该表达区域将形成颌、胸部的部分体节;随着腹节开始形成,hb基因在颌胸部的表达逐渐减弱,而在腹部后端的“生长区”表达,并呈现出不连续性．经比较,hb易基因在昆虫颌胸部的表达较为保守,而在卵子形成过程中和腹部的表达具有较大的变异性．与黑腹果蝇等长胚带昆虫相比,东亚飞蝗hb基因在体节形成的基因级联调控中具有更重要、更直接的调控作用．     

Global transcriptional response of pig brain and lung to natural infection by Pseudorabies virus

Background  

Pseudorabies virus (PRV) is an alphaherpesviruses whose native host is pig. PRV infection mainly causes signs of central nervous system disorder in young pigs, and respiratory system diseases in the adult.     

The draft genome of the pest tephritid fruit fly Bactrocera tryoni: resources for the genomic analysis of hybridising species

Background

The tephritid fruit flies include a number of economically important pests of horticulture, with a large accumulated body of research on their biology and control. Amongst the Tephritidae, the genus Bactrocera, containing over 400 species, presents various species groups of potential utility for genetic studies of speciation, behaviour or pest control. In Australia, there exists a triad of closely-related, sympatric Bactrocera species which do not mate in the wild but which, despite distinct morphologies and behaviours, can be force-mated in the laboratory to produce fertile hybrid offspring. To exploit the opportunities offered by genomics, such as the efficient identification of genetic loci central to pest behaviour and to the earliest stages of speciation, investigators require genomic resources for future investigations.

Results

We produced a draft de novo genome assembly of Australia’s major tephritid pest species, Bactrocera tryoni. The male genome (650 -700 Mbp) includes approximately 150Mb of interspersed repetitive DNA sequences and 60Mb of satellite DNA. Assessment using conserved core eukaryotic sequences indicated 98% completeness. Over 16,000 MAKER-derived gene models showed a large degree of overlap with other Dipteran reference genomes. The sequence of the ribosomal RNA transcribed unit was also determined. Unscaffolded assemblies of B. neohumeralis and B. jarvisi were then produced; comparison with B. tryoni showed that the species are more closely related than any Drosophila species pair. The similarity of the genomes was exploited to identify 4924 potentially diagnostic indels between the species, all of which occur in non-coding regions.

Conclusions

This first draft B. tryoni genome resembles other dipteran genomes in terms of size and putative coding sequences. For all three species included in this study, we have identified a comprehensive set of non-redundant repetitive sequences, including the ribosomal RNA unit, and have quantified the major satellite DNA families. These genetic resources will facilitate the further investigations of genetic mechanisms responsible for the behavioural and morphological differences between these three species and other tephritids. We have also shown how whole genome sequence data can be used to generate simple diagnostic tests between very closely-related species where only one of the species is scaffolded.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-1153) contains supplementary material, which is available to authorized users.     

Sulfate reduction and S-oxidation in a moorland pool sediment

In an oligotrophic moorland pool in The Netherlands, S cycling near the sediment/water boundary was investigated by measuring (1) SO₄ ^2– reduction rates in the sediment, (2) depletion of SO₄ ^2– in the overlying water column and (3) release of³⁵S from the sediment into the water column. Two locations differing in sediment type (highly organic and sandy) were compared, with respect to reduction rates and depletion of SO₄ ^2– in the overlying water.Sulfate reduction rates in sediments of an oligotrophic moorland pool were estimated by diagenetic modelling and whole core³⁵SO₄ ^2– injection. Rates of SO₄ ^2– consumption in the overlying water were estimated by changes in SO₄ ^2– concentration over time in in situ enclosures. Reduction rates ranged from 0.27–11.2 mmol m^–2 d^–1. Rates of SO₄ ^2– uptake from the enclosed water column varied from –0.5, –0.3 mmol m^–2 d^–1 (November) to 0.43–1.81 mmol m^–2 d^–1 (July, August and April). Maximum rates of oxidation to SO₄ ^2– in July 1990 estimated by combination of SO₄ ^2– reduction rates and rates of in situ SO₄ ^2– uptake in the enclosed water column were 10.3 and 10.5 mmol m^–2 d^–1 at an organic rich and at a sandy site respectively.Experiments with³⁵S^2– and³⁵SO₄ ^2– tracer suggested (1) a rapid formation of organically bound S from dissimilatory reduced SO₄ ^2– and (2) the presence of mainly non SO₄ ^2–-S derived from reduced S transported from the sediment into the overlying water. A³⁵S^2– tracer experiment showed that about 7% of³⁵S^2– injected at 1 cm depth in a sediment core was recovered in the overlying water column.Sulfate reduction rates in sediments with higher volumetric mass fraction of organic matter did not significantly differ from those in sediments with a lower mass fraction of organic matter.Corresponding author