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PubMed2006

A segmentation gene in tribolium produces a polycistronic mRNA that codes for multiple conserved peptides.

Segmentation genes in insects are required for generating the subdivisions of the early embryo. We describe here a new member of the gap family of segmentation genes in the flour beetle Tribolium, mille-pattes (mlpt). mlpt knockdown leads to transformation of the abdominal segments into thoracic segments, providing embryos with up to ten pairs of legs. We show that there are crossregulatory interactions between mlpt and the known gap genes in Tribolium, suggesting that mlpt is itself a gap gene. The mlpt gene reveals an unusual structure, as it encodes a polycistronic mRNA that codes for four peptides. mlpt appears to be the prototype of this previously unknown gene structure in eukaryotes, as we find homologous genes with the same polycistronic arrangement in other insect genomes as well.

Animals↗
PubMedDate not supplied

Expression of a homologue of the fushi tarazu (ftz) gene in a cirripede crustacean.

In Metazoa, Hox genes control the identity of the body parts along the anteroposterior axis. In addition to this homeotic function, these genes are characterized by two conserved features: They are clustered in the genome, and they contain a particular sequence, the homeobox, encoding a DNA-binding domain. Analysis of Hox homeobox sequences suggests that the Hox cluster emerged early in Metazoa and then underwent gene duplication events. In arthropods, the Hox cluster contains eight genes with a homeotic function and two other Hox-like genes, zerknullt (zen)/Hox3 and fushi tarazu (ftz). In insects, these two genes have lost their homeotic function but have acquired new functions in embryogenesis. In contrast, in chelicerates, these genes are expressed in a Hox-like pattern, which suggests that they have conserved their ancestral homeotic function. We describe here the characterization of Diva, the homologue of ftz in the cirripede crustacean Sacculina carcini. Diva is located in the Hox cluster, in the same position as the ftz genes of insects, and is not expressed in a Hox-like pattern. Instead, it is expressed exclusively in the central nervous system. Such a neurogenic expression of ftz has been also described in insects. This study, which provides the first information about the Hoxcluster in Crustacea, reveals that it may not be much smaller than the insect cluster. Study of the Diva expression pattern suggests that the arthropod ftz gene has lost its ancestral homeotic function after the divergence of the Crustacea/Hexapoda clade from other arthropod clades. In contrast, the function of ftz during neurogenesis is well conserved in insects and crustaceans.

Amino Acid Sequence↗
PubMed1971

Insect control by genetic manipulation of natural populations.

The possible use of chromosome rearrangements is considered as a means for introducing genes into insect populations for their own control. The release of laboratory-constructed strains differing from the field population for a number of chromosome interchanges should create an unstable situation leading to the rapid replacement of the field population. This replacement should allow introduction of genes for insecticide susceptibility, cold sensitivity, or the like. The process would produce sterile hybrids while the genetic displacement occurs which itself will contribute to a reduction in pest numbers.

Animals↗
PubMed1992

Variation and the phylogenetic utility of the large ribosomal subunit of mitochondrial DNA from the insect order Hymenoptera.

Nucleotide sequence variation from a 573-bp region of the mitochondrial 16S rRNA gene was determined for representative hymenopteran taxa. An overall bias in the distribution of A and T bases was observed from all taxa; however, the terebrants (parasitoids) displayed significantly lower AT ratios as well as a higher degree of strand asymmetry. Moreover, a strong positive correlation was observed between relative AT richness and sequence divergence, suggesting selection at the nucleotide level for A and T bases as well as functionality. Overall sequence difference ranged from 2.3 to 53.4%, with the maximum divergence between members of the two Hymenopteran suborders. These data were used in a phylogenetic analysis to illustrate the utility and degree of resolution provided by this information at various hierarchical levels within this taxonomically diverse order. Parsimony analysis revealed strong evidence for monophyly of the aculeates and the terebrants. Most noteworthy was a strongly supported clade containing the two terebrant superfamilies Icheumonoidea and Chalcidoidea. Conversely, high sequence divergence values resulted in instability at the base of the tree and limited resolution at the higher taxonomic levels. Nevertheless, these results do identify those taxonomic levels for which 16S rRNA sequences are phylogenetically informative.

Animals↗
PubMed2001

The tc genes of Photorhabdus: a growing family.

The toxin complex (tc) genes of Photorhabdus encode insecticidal, high molecular weight Tc toxins. These toxins have been suggested as useful alternatives to those derived from Bacillus thuringiensis for expression in insect-resistant transgenic plants. Although Photorhabdus luminescens is symbiotic with nematodes that kill insects, tc genes have recently been described from other insect-associated bacteria such as Serratia entomophila, an insect pathogen, and Yersinia pestis, the causative agent of bubonic plague, which has a flea vector. Here, recent advances in our understanding of the tc gene family are reviewed in view of their potential development as insect-control agents.

Animals↗
PubMed1996

Production of a polyhydroxyalkanoate biopolymer in insect cells with a modified eucaryotic fatty acid synthase.

A novel pathway for the synthesis of poly-3-hydroxybutyrate has been engineered by simultaneous delivery of two genes into insect cells (Spodoptera frugiperda) by use of individual baculovirus vectors. This system includes expression of a dehydrase-domain mutant rat fatty acid synthase cDNA and the phbC gene encoding polyhydroxyalkanoate synthase from Alcaligenes eutrophus. The dehydrase-deficient fatty acid synthase provides de novo synthesis of R-(-)-3-hydroxybutyryl-coenzyme A as a premature termination product rather than palmityl-coenzyme A, the normal product of wild-type rat fatty acid synthase. High levels of this mutant multifunctional protein provide a suitable precursor pool of R-(-)-3-hydroxybutyryl-coenzyme A for conversion to poly-3-hydroxybutyrate in insect cells coexpressing the phbC gene product. This strategy for redesigning a poly-3-hydroxybutyrate biosynthetic pathway suggests a new method for generating structurally diverse polyhydroxyalkanoates by metabolic engineering.

Animals↗
PubMed2004

Kruppel is a gap gene in the intermediate germband insect Oncopeltus fasciatus and is required for development of both blastoderm and germband-derived segments.

Segmentation in long germband insects such as Drosophila occurs essentially simultaneously across the entire body. A cascade of segmentation genes patterns the embryo along its anterior-posterior axis via subdivision of the blastoderm. This is in contrast to short and intermediate germband modes of segmentation where the anterior segments are formed during the blastoderm stage and the remaining posterior segments arise at later stages from a posterior growth zone. The biphasic character of segment generation in short and intermediate germ insects implies that different formative mechanisms may be operating in blastoderm-derived and germband-derived segments. In Drosophila, the gap gene Krüppel is required for proper formation of the central portion of the embryo. This domain of Krüppel activity in Drosophila corresponds to a region that in short and intermediate germband insects spans both blastoderm and germband-derived segments. We have cloned the Krüppel homolog from the milkweed bug, Oncopeltus fasciatus (Hemiptera, Lygaeidae), an intermediate germband insect. We find that Oncopeltus Krüppel is expressed in a gap-like domain in the thorax during the blastoderm and germband stages of embryogenesis. In order to investigate the function of Krüppel in Oncopeltus segmentation, we generated knockdown phenotypes using RNAi. Loss of Krüppel activity in Oncopeltus results in a large gap phenotype, with loss of the mesothoracic through fourth abdominal segments. Additionally, we find that Krüppel is required to suppress both anterior and posterior Hox gene expression in the central portion of the germband. Our results show that Krüppel is required for both blastoderm-derived and germband-derived segments and indicate that Krüppel function is largely conserved in Oncopeltus and Drosophila despite their divergent embryogenesis.

Amino Acid Sequence↗
PubMed1993

Baculoviruses: high-level expression in insect cells.

Baculoviruses continue to serve as workhorse vectors for the high-level expression of eukaryotic genes in insect cells; however, numerous researchers are also finding novel uses for these vectors by taking advantage of the unique nature of the viruses and their host cells. Concurrently, the technology involved in constructing and utilizing these vectors is being improved so that the time and effort required to construct expression vectors are reduced.

Animals↗
PubMed1997

[Expression of green fluorescent protein with baculovirus vector in insect cells].

The green fluorescent protein (GFP) gene was subcloned into the transfer vector pVLneo downstream of the polyhedrin gene (ocu) promoter. Insect cells were cotransfected with recombinant plasmid and Autographa californica Nuclear Polyhedrosis Virus (AcNPV) DNA. In the presence of G418, the recombinant virus containing GFP gene was purified. The GFP expressed in insect cells with a Mw of 30 kDa is observable by strong green light under a fluorescent microscope. Excitation and emission spectra of the GFP were 395 nm and 509 nm respectively. Integration of GFP gene on AcNPV genome was identified directly by Southern blot which gave strong hybridization signal between GFP cDNA probe and 1 kb EcoRI fragment of recombinant virus.

Animals↗
PubMed1994

VSG gene expression site control in insect form Trypanosoma brucei.

When the African trypanosome Trypanosoma brucei is taken up from mammals by a tse-tse fly, it replaces its variant surface glycoprotein (VSG) coat by a procyclin coat. Transcription of VSG genes stops in the fly, but transcription of sequences derived from the promoter area of the VSG expression site(s) remains high. Whether this is due to continuing high activity of one promoter or to low activity of many promoters was unclear. We have used the small differences between the sequences of different expression sites to show that multiple expression site promoters are active in insect form trypanosomes. This is confirmed by the low expression of single copy marker genes introduced into the transcribed area. However, if the expression site promoter is removed from the genomic location of the expression site and inserted in the non-transcribed spacer of the ribosomal DNA (rDNA), it is derepressed. Derepression of transcription can also be accomplished by replacing the promoter of an expression site by an rDNA promoter. We conclude that the down-regulation of VSG gene expression site promoters in insect form trypanosomes is affected by both the DNA sequence of the promoter and the genomic context in which it resides.

Animals↗
PubMed1991

Murine genes related to the Drosophila AbdB homeotic genes are sequentially expressed during development of the posterior part of the body.

The cloning, characterization and developmental expression patterns of two novel murine Hox genes, Hox-4.6 and Hox-4.7, are reported. Structural data allow us to classify the four Hox-4 genes located in the most upstream (5') position in the HOX-4 complex as members of a large family of homeogenes related to the Drosophila homeotic gene Abdominal B (AbdB). It therefore appears that these vertebrate genes are derived from a selective amplification of an ancestral gene which gave rise, during evolution, to the most posterior of the insect homeotic genes so far described. In agreement with the structural colinearity, these genes have very posteriorly restricted expression profiles. In addition, their developmental expression is temporally regulated according to a cranio-caudal sequence which parallels the physical ordering of these genes along the chromosome. We discuss the phylogenetic alternative in the evolution of genetic complexity by amplifying either genes or regulatory sequences, as exemplified by this system in the mouse and Drosophila. Furthermore, the possible role of 'temporal colinearity' in the ontogeny of all coelomic (metamerized) metazoans showing a temporal anteroposterior morphogenetic progression is addressed.

Amino Acid Sequence↗
PubMed2003

Length analyses of Drosophila odorant receptors.

Odorant receptors comprise a unique family of G-protein-coupled seven-transmembrane receptors both in mammals and insects. In the fruit fly Drosophila melanogaster, all 61 candidate odorant receptor genes have been identified based on the complete genome sequence, and their expression patterns have been examined. A given odorant receptor is expressed in the antenna or maxillary palp, or not expressed at all. Here we have applied a set of statistical analyses to the length of the extra- and intracellular loops and terminals (LTs) of Drosophila odorant receptors to examine possible inter- and intramolecular relations at the population level. We have first provided some useful statistical information such as mean length values and length histograms to depict a general nature of Drosophila odorant receptors at the population level, after focusing on discrepancy on assigning transmembrane domains between researchers. In a preferable transmembrane assignment, all extracellular LTs, especially the second extracellular loops, were relatively large in length, suggesting their functional significance. Somewhat surprisingly, principle component analysis (PCA) indicated that the maxillary palp receptors were almost as diverse as the antenna receptors despite their much smaller population size. PCA together with histograms also revealed that receptors with an abnormal length configuration tended not to be expressed, suggesting that LT length deviations are related to transcriptional silencing of odorant receptor genes. Rank transformation tests pointed out possible LTs that could have different length between differently expressed receptors at the population level. Taken together, length analyses provide us with a general picture, i.e. "length configuration," of Drosophila odorant receptors at the population level that could point out putatively important functional sites for experimental studies.

Amino Acid Sequence↗
PubMed1997

Isolation of a functional ecdysteroid receptor homologue from the ixodid tick Amblyomma americanum (L.).

Ecdysteroids are assumed to be the major steroid hormones in arthropods. However, with the exception of insects and crustaceans, very little is known about ecdysteroid action in other arthropods. To determine if ecdysteriods play a functional role in the ixodid tick, Amblyomma americanum (L.), we isolated cDNAs encoding three presumed ecdysteroid receptor isoforms (AamEcRA1, AamEcRA2, and AamEcRA3) that have common DNA and ligand binding domains linked to distinct amino termini. The DNA and ligand binding domains share an average of 86 and 64% identity, respectively with DNA and ligand binding domains from insect EcR proteins. The amino termini are highly divergent and the AamEcRs lack the 'F' domain found in the insect EcRs. Analysis of AamEcR cDNAs show that processing of the AamEcR gene is complex, producing multiple transcripts with unique 5' and 3' termini as well as splicing variants with incomplete open reading frames. AamEcR mRNA profiles in whole animals and isolated tissues are consistent with complex regulation of AamEcR expression. We also examined the ability of AamEcRA1, when paired with an AamRXR, to activate transcription of an ecdysone response element containing reporter, and demonstrate that the AamEcR gene encodes a functional ecdysteroid receptor.

Amino Acid Sequence↗
PubMed1999

A peritrophin-like protein expressed in the embryonic tracheae of Drosophila melanogaster.

We have cloned and sequenced a cDNA from Drosophila melanogaster that encodes a protein homologous to the peritrophins, a family of chitin-binding proteins from the peritrophic matrix of insects. Unexpectedly, the gene, Gasp, is expressed in the embryonic tracheae. We suggest that this family of proteins may be present in other tissues than the peritrophic matrix, particularly where nutrient or gas exchange are important, and/or where invasion by parasites or viruses is possible. We have also mapped two similar genes that had been sequenced by the Berkeley Drosophila Genome Project, and find that these three very similar genes are not clustered, but are located on three different chromosomes.

Amino Acid Sequence↗
PubMed2004

Transcription regulation within the circadian clock: the E-box and beyond.

The circadian oscillator is composed of transcriptional feedback loops in organisms ranging from cyanobacteria to humans. These transcriptional feedback loops are so named because transcriptional regulators accumulate to high levels and then feed back to control their own genes' transcription, thus generating a self-sustaining rhythm in gene expression. In insects and vertebrates, the genes that encode these feedback regulators are remarkably well conserved and function to control not 1 but 2 feedback loops. These feedback loops control rhythmic transcription in opposite phases of the circadian cycle, yet they are interlocked because they share a number of components. In this review, the author will compare transcriptional regulatory mechanisms within the Drosophila and mammalian feedback loops and outline remaining questions concerning transcriptional regulation within and downstream of these feedback loops.

Animals↗
PubMed1999

An extraretinally expressed insect cryptochrome with similarity to the blue light photoreceptors of mammals and plants.

Photic entrainment of insect circadian rhythms can occur through either extraretinal (brain) or retinal photoreceptors, which mediate sensitivity to blue light or longer wavelengths, respectively. Although visual transduction processes are well understood in the insect retina, almost nothing is known about the extraretinal blue light photoreceptor of insects. We now have identified and characterized a candidate blue light photoreceptor gene in Drosophila (DCry) that is homologous to the cryptochrome (Cry) genes of mammals and plants. The DCry gene is located in region 91F of the third chromosome, an interval that does not contain other genes required for circadian rhythmicity. The protein encoded by DCry is approximately 50% identical to the CRY1 and CRY2 proteins recently discovered in mammalian species. As expected for an extraretinal photoreceptor mediating circadian entrainment, DCry mRNA is expressed within the adult brain and can be detected within body tissues. Indeed, tissue in situ hybridization demonstrates prominent expression in cells of the lateral brain, which are close to or coincident with the Drosophila clock neurons. Interestingly, DCry mRNA abundance oscillates in a circadian manner in Drosophila head RNA extracts, and the temporal phasing of the rhythm is similar to that documented for the mouse Cry1 mRNA, which is expressed in clock tissues. Finally, we show that changes in DCry gene dosage are associated predictably with alterations of the blue light resetting response for the circadian rhythm of adult locomotor activity.

Amino Acid Sequence↗