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PubMedDate not supplied

Conserved and divergent functions of Drosophila atonal, amphibian, and mammalian Ath5 genes.

Insect and vertebrate eyes differ in their formation, cellular composition, neural connectivity, and visual function. Despite this diversity, Drosophila atona and its vertebrate Ortholog in the eye, Ath5, each regulate determination of the first retinal neuron class-R8 photo-receptors and retinal ganglion cells (RGCs)-in their respective organisms. We have performed a cross-species functional comparison of these genes. In ato mutant Drosophila, ectopic Xenopus Ath5 (Xath5) rescues photoreceptor cell development comparably with atonaI. In contrast, mouse Ath5 (Math5) induces formation of very few ommatidia, and most of these lack R8 cells. In the developing frog eye, ectopic atonal, like Xath5, promotes the differentiation RGCs. Despite strong conservation of atonaI, Xath5, and Math5 structure and shared function, other factors must contribute to the species specificity of retinal neuron determination. These observations suggest that the atonaI family may occupy a position in a gene hierarchy where differences in gene regulation or function can be correlated with evolutionary diversity of eye development.

Amino Acid Sequence↗
PubMed1997

A new nuclear gene for insect phylogenetics: dopa decarboxylase is informative of relationships within Heliothinae (Lepidoptera: Noctuidae).

The lack of a readily accessible roster of nuclear genes informative at various taxonomic levels is a bottleneck for molecular systematics. In this report, we describe the first phylogenetic application of the sequence that encodes the enzyme dopa decarboxylase (DDC). For 14 test species within the noctuid moth subfamily Heliothinae that represent the previously best-supported groupings, a 690-bp fragment of DDC resolved relationships that are largely concordant with prior evidence from elongation factor-1 alpha (EF-1 alpha), morphology, and allozymes. Although both synonymous and nonsynonymous changes occur in DDC substantially more rapidly than they do in EF-1 alpha, DDC divergences within Heliothinae are below saturation at all codon positions. Analysis of DDC and EF-1 alpha in combination resulted in increased bootstrap support for several groupings. As a first estimate of previously unresolved relationships, DDC sequences were analyzed from 16 additional heliothines, for a total of 30 heliothine species plus outgroups. Previous relationships based on DDC were generally stable with increased taxon sampling, although a two- to eightfold downweighting of codon position 3 was required for complete concordance with the 14-species result. The weighted strict consensus trees were largely resolved and were congruent with most although not all previous hypotheses based on either morphology or EF-1 alpha. The proposed phylogeny suggests that the major agricultural pest heliothines belong to a single clade, characterized by polyphagy and associated life history traits, within this largely host-specific moth subfamily. DDC holds much promise for phylogenetic analysis of Tertiary-age animal groups.

Animals↗
PubMed2005

Tight transcriptional regulation of foreign genes in insect cells using an ecdysone receptor-based inducible system.

The use of insect cells has been highly successful for the expression of foreign proteins from baculoviruses or plasmid vectors. Here, we describe a tight transcriptional regulation of foreign genes in insect cells using an ecdysone receptor-based inducible system. The system includes the DEF domains of the spruce budworm (Choristoneura fumiferana) EcR (CfEcR) fused to the Saccharomyces cerevisiae GAL4 DNA-binding domain and the EF domains of mammalian Mus musculus retinoid X receptor (MmRXR) fused to the acidic activation domains (AADs) of the baculovirus transactivators IE1 and IE0. Using a GAL4 response element in reporter constructs, both transient and stable expression in insect lepidopteran cells showed that the chimeric MmRXR and CfEcR only activated the reporter genes in the presence of inducer; no gene expression was detectable in the absence of inducer. Characterization of heterogenous activation domains in insect cells showed that the AADs from Autographa californica multiple nucleopolyhedrovirus (MNPV) IE1 and Orgyia pseudotsugata MNPV IE0 consistently exhibited higher inducible levels than the archetype AAD from herpesvirus VP16 in insect cells. To confirm the tight regulation of this system the highly toxic protein, diphtheria toxin (DT), was used. In the absence of an inducer no cytotoxic effect was observed in insect cells that had been transiently transformed with DT expressing plasmids. This system will therefore be a very useful tool for biotechnology applications expressing highly toxic proteins in insect cells and for studying the functional genomics of insects and microorganisms that infect them.

Animals↗
PubMed2005

Phylogeny of the insect homeobox gene (hox) cluster.

The homeobox (Hox) genes form an evolutionarily conserved family encoding transcription factors that play major roles in segmental identity and organ specification across species. The canonical grouping of Hox genes present in the HOM-C cluster of Drosophila or related clusters in other organisms includes eight "typical" genes, which are localized in the order labial (lab), proboscipedia (pb), Deformed (Dfd), Sex combs reduced (Scr), Antennapedia (Antp), Ultrabithorax (Ubx), abdominalA (abdA), and AbdominalB (AbdB). The members of Hox cluster are expressed in a distinct anterior to posterior order in the embryo. Analysis of the relatedness of different members of the Hox gene cluster to each other in four evolutionarily diverse insect taxa revealed that the loci pb/Dfd and AbdB, which are farthest apart in linkage, had a high degree of evolutionary relatedness, indicating that pb/Dfd type anterior genes and AbdB are closest to the ancestral anterior and posterior Hox genes, respectively. The greater relatedness of other posterior genes Ubx and abdA to the more anterior genes such as Antp and Scr suggested that they arose by gene duplications in the more anterior members rather than the posterior AbdB.

Animals↗
PubMed1996

Transgenic rice plants harboring an introduced potato proteinase inhibitor II gene are insect resistant.

We introduced the potato proteinase inhibitor II (PINII) gene (pin2) into several Japonica rice varieties, and regenerated a large number of transgenic rice plants. Wound-inducible expression of the pin2 gene driven by its own promoter, together with the first intron of the rice actin 1 gene (act1), resulted in high-level accumulation of the PINII protein in the transgenic plants. The introduced pin2 gene was stably inherited in the second, third, and fourth generations, as shown by molecular analyses. Based on data from the molecular analyses, several homozygous transgenic lines were obtained. Bioassay for insect resistance with the fifth-generation transgenic rice plants showed that transgenic rice plants had increased resistance to a major rice insect pest, pink stem borer (Sesamia inferens). Thus, introduction of an insecticidal proteinase inhibitor gene into cereal plants can be used as a general strategy for control of insect pests.

Animals↗
PubMed2001

[Studies on transgenic tobacco plants expressing two kinds of insect resistant genes].

A synthetic Bt cry1Ac gene fussed with a secretary signal coding sequences at 5' end and a modified gna gene were used to construct a plant expression vector pBSGS1M+ and this vector was transferred into tobacco (Nicotiana tabacum L.) by Agrobacterium-mediated transformation method. Results of PCR, Southern blot and Slot blot analysis indicated that both the chimeric Bt cry1Ac and gna genes were integrated into the genomes of transformed plants. Western blot analysis indicated that at least the cry1Ac protein was produced in transgenic plants. Upon insect bioassay using cotton bollworm (Heliothis armigera Hubner), the mortality of insect larvae on 60% regenerated plants reached 100% in 5 days post infestation and the growth of the survived larvae was seriously inhibited; The results from insect bioassay with peach aphid (Myzus persicae) showed that the transgenic plants were aphid-resistant, evidenced by a 50%-60% reduction in aphid population density, even over 80% for some individual transgenic plants. These results reflect that the modification of the two insect resistant genes and construction of the expression vector are correct and could be valuable for later application in crop breeding for insect resistance.

Animals↗
PubMed2000

Independent evolution of Toll and related genes in insects and mammals.

Toll and Toll-related proteins play an important role in antibacterial innate immunity in insect, plants, and mammals. We present the first comprehensive phylogenetic analyses of Toll-related genes from both insects and mammals. Drosophila melanogaster contains Toll and a highly homologous gene, Tehao. The protein, Dm Tehao, comprises 795 amino acid residues and its cytoplasmic domain shares a striking 61% identity with Dm Toll. Two Toll homologues were found in another dipteran of medical importance, Anopheles gambiae, a vector for human malaria. One Toll-like gene each was identified from Aedes aegypti and Glossina palpalis palpalis, vectors for yellow fever and trypanosomiasis, respectively. Phylogenetic analyses revealed separate clustering of Toll and related proteins from insects and mammals, suggesting independent evolution of the Toll family of proteins and of innate immunity in arthropods and vertebrates. These results also provide new avenues to understanding the function of Toll proteins in insect innate immunity against bacteria, fungi, and protozoans.

Aedes↗
PubMed1995

Transient expression of a promoter-reporter construct in differentiated adult salivary glands and embryos of the mosquito Aedes aegypti.

Vector-borne pathogens develop in close association with specific tissues in their insect hosts. Efforts are being made to characterize insect genes that are expressed in tissues that have important roles in pathogen propagation. Successful transfection and expression of exogenous genes in terminally differentiated tissues of insects has previously proven difficult. Here we report a method that should allow the analysis of genes that are expressed in adult tissues and organs. Transient expression assays have been developed using the salivary glands of the mosquito, Aedes aegypti, which can now be used to analyze salivary gland-specific promoter sequences. A liposome-based transfection reagent was used to transfect cultured adult salivary glands with a DNA construct carrying the luciferase reporter gene under the control of the Drosophila melanogaster heat shock 70 promoter. Luciferase activity was detected in glands 18-20 hr post-transfection. This assay can now be used to determine the regulatory activity of other putative promoter sequences from salivary gland-specific genes. Alternatively, the assay may be used to study the effect of recombinant gene expression on parasite invasion and development. In addition, transient expression of gene constructs in embryos is shown to be a powerful tool for analyzing genes that are expressed at this stage of the mosquito life cycle.

Aedes↗
PubMed2006

Wound and insect herbivory responsive genes in poplar.

Insect herbivory leads to induced resistance to subsequent infestations in plants. This is due in part to feeding-induced expression of genes that can lead to reduced palatability and/or digestibility of the plant material. We identified 57 distinct differentially expressed genes from poplars that were either infested by gypsy moth (Lymantria dispar) or mechanically wounded. Eleven highly insect-inducible genes were also found to be wound-inducible. Time course analysis revealed diverse timing of peak transcript accumulation. Sequence analysis of promoters suggested that the wound responsive elements, W and DRE, and the jasmonic acid responsive H motif, are over-represented in wound-induced poplar promoters and should be investigated further.

Animals↗
PubMed2003

[Study on transgenic elite indica container line D297B containing foreign insect-resistant gene gna].

It was reported in this article that the transgenic elite indica container line D297B containing snowdrop lectin (Galanthus nivalis agglutinin, GNA) gene gna was obtained by biolisties. PCR, Southern blotting and Western blotting indicated that the transgenes were integrated into the genome of D297B and expressed in transgenic plants. Analysis of protein activity showed that product of transgene had activity of agglutinin. Resistance of transgenic seeds to Hygromycin B suggested that the transgenes were integrated in a single locus and inherited according to the pattern of 3:1 in most of transgenic plants. In addition, the PCR analysis revealed that the insect-resistant gene gna and selective marker gene hpt were co-integrated and co-inherited.

Animals↗
PubMed1995

Evolution of segmentation genes in insects.

Systematic genetic analysis of the segmentation process in Drosophila has established a paradigm for the molecular control of the formation of metameric segments. However, it has been suggested that some of the mechanisms involved in this process in Drosophila are uniquely adapted to the syncytial mode of embryogenesis in such higher dipterans. A particularly contentious problem is the role of early segmentation genes in short-germ insects, in which development proceeds by sequential addition of segments in a cellular environment. However, analysis of the expression of presumptive homologs of segmentation genes in holometabolous short-germ insects suggests that they do indeed have a role in segmentation and that the Drosophila paradigm may be more widely applicable than is usually assumed. Most interestingly, these results suggest that the molecular mechanisms of pattern formation in noncellular and in cellular environments may not be as radically different as it is often thought to be.

Animals↗
PubMed1997

Evolution of orthologous intronless and intron-bearing globin genes in two insect species.

While globin genes ctt-2beta and ctt-9.1 in Chironomus thummi thummi each have a single intron, all of the other insect globin genes reported so far are intronless. We analyzed four globin genes linked to the two intron-bearing genes in C. th. thummi. Three have a single intron at the same position as ctt-2beta and ctt-9.1; the fourth is intronless and lies between intron bearing genes. Finally, in addition to its intron, one gene (ctt-13RT) was recently interrupted by retrotransposition. Phylogenetic analyses show that the six genes in C. th. thummi share common ancestry with five globin genes in the distantly related species C. tentans, and that a 5-gene ancestral cluster predates the divergence of the two species. One gene in the ancestral cluster gave rise to ctn-ORFB in C. tentans, and duplicated in C. th. thummi to create ctt-11 and ctt-12. From parsimonious calculations of evolutionary distances since speciation, ctt-11, ctt-12, and ctn-ORFB evolved rapidly, while ctn-ORFE in C. tentans evolved slowly compared to other globin genes in the clusters. While these four globins are under selective pressure, we suggest that most chironomid globin genes were not selected for their unique function. Instead, we propose that high gene copy number itself was selected because conditions favored organisms that could synthesize more hemoglobin. High gene copy number selection to produce more of a useful product may be the basis of forming multigene families, all of whose members initially accumulate neutral substitutions while retaining essential function. Maintenance of a large family of globin genes not only ensured high levels of hemoglobin production, but may have facilitated the extensive divergence of chironomids into as many as 5000 species.

Amino Acid Sequence↗
PubMed1991

A baculovirus dual expression vector derived from the Autographa californica nuclear polyhedrosis virus polyhedrin and p10 promoters: co-expression of two influenza virus genes in insect cells.

A baculovirus transfer vector, pAcUW3, was developed to facilitate the insertion of two influenza virus genes, those encoding the haemagglutinin (HA) and neuraminidase (NA) membrane glycoproteins, into the Autographa californica nuclear polyhedrosis virus genome in a single cotransfection experiment. The NA gene was inserted in place of the polyhedrin coding sequences under the control of the polyhedrin promoter, whereas the HA gene was placed under the control of a copy of the p10 promoter at a site upstream of and in opposite orientation to the polyhedrin promoter. After infection of Spodoptera frugiperda cells with the recombinant virus, AcUW3HANA, both HA and NA were expressed in the very late phase of infection and were shown to be functional in appropriate assays. Immunofluorescence assays demonstrated their localization at the surface of infected insect cells. The expression of both foreign genes in the recombinant virus was found to be stable for at least 12 passages in cell culture.

Animals↗
PubMed1998

The expression of two engrailed-related genes in an apterygote insect and a phylogenetic analysis of insect engrailed-related genes.

Homologues of the Drosophila segment polarity gene engrailed have been cloned from many insect species, as well as other arthropods and non-arthropods. We have cloned partial cDNAs of two engrailed homologues, which we call engrailed-related genes, from the phylogenetically basal insect, Thermobia domestica (Order Thysanura) and possibly as many as four engrailed-related genes from the phylogenetically intermediate insect, Oncopeltus fasciatus (Order Hemiptera). Previous to our findings, only single engrailed-related homologues had been found in phylogenetically intermediate insect species (Tribolium and Schistocerca) and in the crustacean Artemia, while two engrailed-related homologues have been found in more derived orders (Hymenoptera and the engrailed and invected genes of lepidopterans and dipterans). Consequently, we performed a phylogenetic analysis of insect engrailed-related genes to determine whether insects ancestrally had one or two engrailed-related genes. We have found evidence of concerted evolution among engrailed-related paralogues, however, that masks the true phylogenetic history of these genes; the phylogeny may only be decipherable, therefore, by examining the presence or absence of engrailed-specific and invected-specific motifs, which will require cloning the full length cDNAs from more species. In addition, we examined the embryonic expression pattern of the two Thermobia engrailed-related genes; like Drosophila engrailed and invected, they are expressed in very similar patterns, but show one temporal difference in pregnathal segments that correlates with the tentative phylogenetic placement of the genes. Thermobia engrailed-related expression also confirms that the dorsal ridge is an ancient structure in insects.

Amino Acid Sequence↗
PubMed2005

Concurrent use of transgenic plants expressing a single and two Bacillus thuringiensis genes speeds insect adaptation to pyramided plants.

Transgenic plants expressing insecticidal proteins from the bacterium Bacillus thuringiensis (Bt) were grown on over 13 million ha in the United States and 22.4 million ha worldwide in 2004. Preventing or slowing the evolution of resistance by insects ("resistance management") is critical for the sustainable use of Bt crops. Plants containing two dissimilar Bt toxin genes in the same plant ("pyramided") have the potential to delay insect resistance. However, the advantage of pyramided Bt plants for resistance management may be compromised if they share similar toxins with single-gene plants that are deployed simultaneously. We tested this hypothesis using a unique model system composed of broccoli plants transformed to express different Cry toxins (Cry1Ac, Cry1C, or both) and a synthetic population of the diamondback moth (Plutella xylostella) carrying genes for resistance to Cry1Ac and Cry1C at frequencies of approximately 0.10 and 0.34, respectively. After 24-26 generations of selection in the greenhouse, the concurrent use of one- and two-gene plants resulted in control failure of both types of Bt plants. When only two-gene plants were used in the selection, no or few insects survived on one- or two-gene Bt plants, indicating that concurrent use of transgenic plants expressing a single and two Bt genes will select for resistance to two-gene plants more rapidly than the use of two-gene plants alone. The results of this experiment agree with the predictions of a Mendelian deterministic simulation model and have important implications for the regulation and deployment of pyramided Bt plants.

Adaptation, Biological↗