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Domain mapping of tube, a protein essential for dorsoventral patterning of the Drosophila embryo.

ABSTRACT: The tube protein plays an essential role in the signal transduction pathway that establishes dorsoventral polarity in the Drosophila melanogaster embryo. Characterization of each of four tube mutants revealed a substitution or insertion in the amino-terminal half of the protein. This portion of the tube protein is also evolutionarily conserved, as demonstrated by isolation and sequencing of the Drosophila virilis tube gene. Moreover, RNA microinjection assays and germline transformation experiments demonstrated that the amino-terminal domain alone provides substantial levels of gene function: constructs encoding only the amino-terminal domain restore dorsoventral polarity to embryos lacking any maternal tube function. In the carboxyterminal domain, sequence conservation is concentrated in the five octapeptide repeats. Although the repeat-containing domain by itself provides no rescue of the tube maternal effect phenotype, it is necessary for wild-type levels of tube activity. This domain is thus likely to play an ancillary role in axis formation.

SUBMITTER: Letsou A

PROVIDER: S-EPMC413621 | biostudies-other | 1993 Sep

REPOSITORIES: biostudies-other

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Domain mapping of tube, a protein essential for dorsoventral patterning of the Drosophila embryo.

Letsou A A Alexander S S Wasserman S A SA

The EMBO journal 19930901 9

The tube protein plays an essential role in the signal transduction pathway that establishes dorsoventral polarity in the Drosophila melanogaster embryo. Characterization of each of four tube mutants revealed a substitution or insertion in the amino-terminal half of the protein. This portion of the tube protein is also evolutionarily conserved, as demonstrated by isolation and sequencing of the Drosophila virilis tube gene. Moreover, RNA microinjection assays and germline transformation experime ...[more]

PMID: 8253071

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Project description:In early development, genes are expressed in spatial patterns which later define cellular identities and tissue locations. The mechanisms of such pattern formation have been studied extensively in early Drosophila (fruit fly) embryos. The gap gene hunchback (hb) is one of the earliest genes to be expressed in anterior-posterior (AP) body segmentation. As a transcriptional regulator for a number of downstream genes, the spatial precision of hb expression can have significant effects in the development of the body plan. To investigate the factors contributing to hb precision, we used fine spatial and temporal resolution data to develop a quantitative model for the regulation of hb expression in the mid-embryo. In particular, modelling hb pattern refinement in mid nuclear cleavage cycle 14 (NC14) reveals some of the regulatory contributions of simultaneously-expressed gap genes. Matching the model to recent data from wild-type (WT) embryos and mutants of the gap gene Krüppel (Kr) indicates that a mid-embryo Hb concentration peak important in thoracic development (at parasegment 4, PS4) is regulated in a dual manner by Kr, with low Kr concentration activating hb and high Kr concentration repressing hb. The processes of gene expression (transcription, translation, transport) are intrinsically random. We used stochastic simulations to characterize the noise generated in hb expression. We find that Kr regulation can limit the positional variability of the Hb mid-embryo border. This has been recently corroborated in experimental comparisons of WT and Kr- mutant embryos. Further, Kr regulation can decrease uncertainty in mid-embryo hb expression (i.e. contribute to a smooth Hb boundary) and decrease between-copy transcriptional variability within nuclei. Since many tissue boundaries are first established by interactions between neighbouring gene expression domains, these properties of Hb-Kr dynamics to diminish the effects of intrinsic expression noise may represent a general mechanism contributing to robustness in early development.

Project description:BackgroundRobustness to natural temperature fluctuations is critical to proper development in embryos and to cellular functions in adult organisms. However, mechanisms and pathways which govern temperature compensation remain largely unknown beyond circadian rhythms. Pathways which ensure robustness against temperature fluctuations may appear to be nonessential under favorable, uniform environmental conditions used in conventional laboratory experiments where there is little variation for which to compensate. The endo-siRNA pathway, which produces small double-stranded RNAs in Drosophila, appears to be nonessential for robust development of the embryo under ambient uniform temperature and to be necessary only for viral defense. Embryos lacking a functional endo-siRNA pathway develop into phenotypically normal adults. However, we hypothesized that small RNAs may regulate the embryo's response to temperature, as a ribonucleoprotein complex has been previously shown to mediate mammalian cell response to heat shock.Principal findingsHere, we show that the genes DICER-2 and ARGONAUTE2, which code for integral protein components of the endo-siRNA pathway, are essential for robust development and temperature compensation in the Drosophila embryo when exposed to temperature perturbations. The regulatory functions of DICER-2 and ARGONAUTE2 were uncovered by using microfluidics to expose developing Drosophila embryos to a temperature step, in which each half of the embryo develops at a different temperature through developmental cycle 14. Under this temperature perturbation, dicer-2 or argonaute2 embryos displayed abnormal segmentation. The abnormalities in segmentation are presumably due to the inability of the embryo to compensate for temperature-induced differences in rate of development and to coordinate developmental timing in the anterior and posterior halves. A deregulation of the length of nuclear division cycles 10-14 is also observed in dicer-2 embryos at high temperatures.ConclusionsResults presented herein uncover a novel function of the endo-siRNA pathway in temperature compensation and cell cycle regulation, and we hypothesize that the endo-siRNA pathway may regulate the degradation of maternal cell cycle regulators. Endo-siRNAs may have a more general role buffering against environmental perturbations in other organisms.

Dataset Information

Domain mapping of tube, a protein essential for dorsoventral patterning of the Drosophila embryo.

Publications

Domain mapping of tube, a protein essential for dorsoventral patterning of the Drosophila embryo.

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