A system to efficiently maintain embryonic lethal mutations in the flour beetle Tribolium castaneum

Berghammer, Andreas J.; Bucher, Gregor; Maderspacher, Florian; Klingler, Martin

doi:10.1007/s004270050268

Cited by 37 publications

(27 citation statements)

References 10 publications

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“…As described in the original screening paper [5], the enhancer trap lines are homozygous for the pBac{3xP3-EGFPaf} transposon cassette, which contains the coding sequence for EGFP under the artificial 3xP3 promoter with three binding sites for Pax6 [19], [20]. Hence, all lines have fluorescence in the eyes as a marker for transgenesis, in addition to new expression domains resulting from enhancer trapping.…”

Section: Resultsmentioning

confidence: 99%

Visualizing Late Insect Embryogenesis: Extraembryonic and Mesodermal Enhancer Trap Expression in the Beetle Tribolium castaneum

2014

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The beetle Tribolium castaneum has increasingly become a powerful model for comparative research on insect development. One recent resource is a collection of piggyBac transposon-based enhancer trap lines. Here, we provide a detailed analysis of three selected lines and demonstrate their value for investigations in the second half of embryogenesis, which has thus far lagged behind research on early stages. Two lines, G12424 and KT650, show enhanced green fluorescent protein (EGFP) expression throughout the extraembryonic serosal tissue and in a few discrete embryonic domains. Intriguingly, both lines show for the first time a degree of regionalization within the mature serosa. However, their expression profiles illuminate distinct aspects of serosal biology: G12424 tracks the tissue’s rapid maturation while KT650 expression likely reflects ongoing physiological processes. The third line, G04609, is stably expressed in mesodermal domains, including segmental muscles and the heart. Genomic mapping followed by in situ hybridization for genes near to the G04609 insertion site suggests that the transposon has trapped enhancer information for the Tribolium orthologue of midline (Tc-mid). Altogether, our analyses provide the first live imaging, long-term characterizations of enhancer traps from this collection. We show that EGFP expression is readily detected, including in heterozygote crosses that permit the simultaneous visualization of multiple tissue types. The tissue specificity provides live, endogenous marker gene expression at key developmental stages that are inaccessible for whole mount staining. Furthermore, the nonlocalized EGFP in these lines illuminates both the nucleus and cytoplasm, providing cellular resolution for morphogenesis research on processes such as dorsal closure and heart formation. In future work, identification of regulatory regions driving these enhancer traps will deepen our understanding of late developmental control, including in the extraembryonic domain, which is a hallmark of insect development but which is not yet well understood.

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Section: Resultsmentioning

confidence: 99%

Visualizing Late Insect Embryogenesis: Extraembryonic and Mesodermal Enhancer Trap Expression in the Beetle Tribolium castaneum

2014

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“…A wide variety of genetic and molecular tools have recently become available for Tribolium including screens for zygotic patterning mutants Anderson 1996, 1998;Maderspacher et al 1998;Berghammer et al 1999a), germ line transformation (Berghammer et al 1999b) and parental RNAi (Bucher et al 2002). Beetles and flies diverged approximately 250 Mio years ago and in contrast to other holometabolous insect orders the beetles have retained several primitive features which they share with the more ancestral hemimetabolous insects (Willmann 2003).…”

Section: Introductionmentioning

confidence: 98%

Tribolium castaneum twist: gastrulation and mesoderm formation in a short-germ beetle

et al. 2004

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Mesoderm formation has been extensively analyzed in the long-germ insect Drosophila melanogaster. In Drosophila, both the invagination and specification of the mesoderm is controlled by twist. Here we present a detailed description of mesoderm formation and twist regulation for the short-germ beetle Tribolium castaneum. In contrast to Drosophila, (1) the presumptive mesodermal cells of Tribolium are part of a mitotic domain and divide prior to ventral furrow formation, (2) ventral furrow formation progresses from posterior to anterior, (3) the number of cell layers within the furrow changes from multilayered in caudal to single layered in cephalic regions, and (4) there is a continuous production of mesodermal cells after gastrulation as new segments arise from the posterior growth zone. Tribolium twist (Tc-twist) is initially expressed in all presumptive mesodermal cells; however, after invagination, expression is maintained only in particular locations, which include the anterior compartments of the cephalic segments and a patch of cells at the posterior rim of the growth zone. The growth zone is multilayered with its inner cell layer being continuous with the mesoderm of the newly forming segments where twist expression is re-initiated anterior to the emerging engrailed stripes. A genomic region of Tc-twist was identified which drives ventral expression of a reporter construct in Drosophila. The expression of this Tc-twist construct in the background of Drosophila maternal mutations suggests that the dorsoventral system regulates Tc-twist, but that differences exist in regulation of the Dm-twist and Tc-twist genes by the terminal system.

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“…Second, Tribolium provides many tools for functional studies. It possesses a particularly strong systemic RNAi response -which was the basis for a recent genome-wide RNAi screen -and techniques for forward genetic screens, transgenesis and live imaging are available (Benton et al, 2013;Berghammer et al, 1999Berghammer et al, , 2009Koelzer et al, 2014;Sarrazin et al, 2012). Therefore, we have chosen Tribolium for unbiased genomic approaches like ChIP-seq (D. Stappert, Two novel, complementary next generation sequencing approaches to reveal the dorso-ventral gene regulatory network of Tribolium castaneum.…”

Section: Introductionmentioning

confidence: 99%

Genome wide identification ofTriboliumdorsoventral patterning genes

et al. 2016

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The gene regulatory network controlling dorsoventral axis formation in insects has undergone drastic evolutionary changes. In Drosophila, a stable long-range gradient of Toll signalling specifies ventral cell fates and restricts BMP signalling to the dorsal half of the embryo. In Tribolium, however, Toll signalling is transient and only indirectly controls BMP signalling. In order to gain unbiased insights into the Tribolium network, we performed comparative transcriptome analyses of embryos with various dorsoventral pattering defects produced by parental RNAi for Toll and BMP signalling components. We also included embryos lacking the mesoderm ( produced by Tctwist RNAi) and characterized similarities and differences between Drosophila and Tribolium twist loss-of-function phenotypes. Using stringent conditions, we identified over 750 differentially expressed genes and analysed a subset with altered expression in more than one knockdown condition. We found new genes with localized expression and showed that conserved genes frequently possess earlier and stronger phenotypes than their Drosophila orthologues. For example, the leucine-rich repeat (LRR) protein Tartan, which has only a minor influence on nervous system development in Drosophila, is essential for early neurogenesis in Tribolium and the Tc-zinc-finger homeodomain protein 1 (Tc-zfh1), the orthologue of which plays a minor role in Drosophila muscle development, is essential for maintaining early Tc-twist expression, indicating an important function for mesoderm specification.

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A system to efficiently maintain embryonic lethal mutations in the flour beetle Tribolium castaneum

Cited by 37 publications

References 10 publications

Visualizing Late Insect Embryogenesis: Extraembryonic and Mesodermal Enhancer Trap Expression in the Beetle Tribolium castaneum

Visualizing Late Insect Embryogenesis: Extraembryonic and Mesodermal Enhancer Trap Expression in the Beetle Tribolium castaneum

Tribolium castaneum twist: gastrulation and mesoderm formation in a short-germ beetle

Genome wide identification ofTriboliumdorsoventral patterning genes

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