Krüppel homolog (Kr h) is a dosage-dependent modifier of gene expression in Drosophila

A lethal mutation in the Krüppel homolog (Kr h) was isolated in screens of P-element insertion mutations for modifiers of white gene expression. The mutation occurs in the 5' untranslated region of the Kr h gene and causes a lightening of the eye colour for several alleles of white due to a decrease in white steady-state mRNA levels at pupal stages. Two related genes, scarlet and brown, were significantly affected as well in early pupae. Genetic analysis of different white alleles suggests that enhancer sequences are necessary for interaction with KR H. Thus, the Kr h gene is a member of the dosage-dependent hierarchy effective upon white.     

Krüppel, a gene whose activity is required early in the zygotic genome for normal embryonic segmentation

Embryos homozygous for Krüppel die as late embryos with an altered segmentation pattern. In strong alleles the normal thoracic and anterior abdominal segments are replaced by a partial mirror image duplication of the posterior abdomen. Weak alleles cause smaller pattern deletions in the thorax and abdomen and are not associated with mirror image duplications. The altered segmentation pattern can be traced back to 12 min after the onset of gastrulation, when the shorter germ bands in homozygous Kr embryos provide a first indication of abnormal patterning. The mutant was mapped to position 107.6 at the tip of the right arm of the second chromosome, cytologically to bands 60F2-5. Analysis of homozygous deficiency embryos indicate that the phenotype produced by strong point mutations probably represents the amorphic condition. The requirement for Kr+ gene activity is strictly zygotic. Maternal dosage of Kr+ has no effect on the embryonic phenotype, nor does homozygosity for Kr prevent germ cells from making normal eggs capable of normal embryonic development when fertilized by wild-type sperm. The requirement for Kr+ seems restricted to embryogenesis. Homozygous clones induced in imaginal discs during larval development survive and develop normally and in vivo cultures established from homozygous embryos proliferate normally and metamorphose into adult structures of normal morphology.     

Changes in protein synthetic activity in early Drosophila embryos mutant for the segmentation gene Krüppel

We have identified early embryo proteins related to the segmentation gene Krüppel by [35S]methionine pulse labelling and two-dimensional gel electrophoresis. Protein synthesis differences shared by homozygous embryos of two Krüppel alleles when compared to heterozygous and wild-type embryos are reported. The study was extended to syncytial blastoderm stages by pulse labelling and gel analysis of single embryos, using Krüppel-specific proteins from gastrula stages as molecular markers for identifying homozygous Krüppel embryos. Localized expression of interesting proteins was examined in embryo fragments. The earliest differences detected at nuclear migration stages showed unregulated synthesis in mutant embryos of two proteins that have stage specific synthesis in normal embryos. At the cellular blastoderm stage one protein was not synthesized and two proteins showed apparent shifts in isoelectric point in mutant embryos. Differences observed in older embryos included additional proteins with shifted isoelectric points and a number of qualitative and quantitative changes in protein synthesis. Five of the proteins with altered rates of synthesis in mutant embryos showed localized synthesis in normal embryos. The early effects observed are consistent with the hypothesis that the Krüppel product can be a negative or positive regulator of expression of other loci, while blastoderm and gastrula stage shifts in isoelectric point indicate that a secondary effect of Krüppel function may involve post-translational modification of proteins.     

Correlative changes in homoeotic and segmentation gene expression in Krüppel mutant embryos of Drosophila

Mutations of the segmentation gene Krüppel (Kr) cause deletions of contiguous sets of body segments from the middle region of the Drosophila embryo. We have monitored expression in situ of three other genes implicated in the establishment of the body plan, namely hairy (h), fushi tarazu (ftz) and engrailed (en), in mutant Kr embryos. Our results show that the pattern of expression of all three genes depends upon Kr⁺ activity and are consistent with a hierarchical model of segmentation gene activity. In addition, we find that the initial expression of the homoeotic selector gene Ultrabithorax(Ubx) follows a novel pattern in Kr^- embroys indicating a close integration of the spatial control of homoeotic and segmantation gene expression.     

Regulatory interactions and role in cell type specification of the Malpighian tubules by the cut, Krüppel, and caudal genes of Drosophila.

Krüppel and caudal genes are both required for normal segmentation of the embryo, and the developmental regulatory gene cut is necessary for the normal specification of external sensory organs. These three genes are also expressed in the Malpighian tubules before and during differentiation. Two of the genes, Krüppel and cut, are known to be required for development of the tubules. We report that the absence of maternal and zygotic caudal function reduces their normal growth and elongation. Normal Krüppel function, which is known to be required for caudal expression, is also required for cut expression, while cut and caudal are expressed independently of each other. Cell type transformations of Malpighian tubules were studied by examining the effects of mutations on the expression of markers specific to Malpighian tubules, hindgut, or midgut of normal embryos. Loss of Krüppel activity confers hindgut characteristics on those cells that normally form the Malpighian tubules with all markers tested. Loss of cut function alters the expression of some markers but not others. The pathway of tissue specific gene regulation, apparently, branches beyond Krüppel to form at least a cut and a caudal branch.     

Gradients of Krüppel and knirps gene products direct pair-rule gene stripe patterning in the posterior region of the Drosophila embryo

Abdominal segmentation of the Drosophila embryo requires the activities of the gap genes Krüppel (Kr), knirps (kni), and tailless (tll). They control the expression of the pair-rule gene hairy (h) by activating or repressing independent cis-acting units that generate individual stripes. Kr activates stripe 5 and represses stripe 6, kni activates stripe 6 and represses stripe 7, and tll activates stripe 7. Kr and kni proteins bind strongly to h control units that generate stripes in areas of low concentration of the respective gap gene products and weakly to those that generate stripes in areas of high gap gene expression. These results indicate that Kr and kni proteins form overlapping concentration gradients that generate the periodic pair-rule expression pattern.     

Analysis of maternal effect mutant combinations elucidates regulation and function of the overlap of hunchback and Krüppel gene expression in the Drosophila blastoderm embryo

The metameric organisation of the Drosophila embryo is generated early during development, due to the action of maternal effect and zygotic segmentation and homeotic genes. The gap genes participate in the complex process of pattern formation by providing a link between the maternal and the zygotic gene activities. Under the influence of maternal gene products they become expressed in distinct domains along the anteroposterior axis of the embryo; negative interactions between neighboring gap genes are thought to be involved in establishing the expression domains. The gap gene activities in turn are required for the correct patterning of the pair-rule genes; little is known, however, about the underlying mechanisms. We have monitored the distribution of gap and pair-rule genes in wild-type embryos and in embryos in which the anteroposterior body pattern is greatly simplified due to combinations of maternal effect mutations (staufen exuperantia, vasa exuperantia, vasa exuperantia, bicoid oskar, bicoid oskar torsolike, vasa torso exuperantia). We show that the domains of protein distribution of the gap genes hunchback and Krüppel overlap in wild-type embryos. Based on the analysis of the maternal mutant combinations, we suggest an explanation of how this overlap is generated. Furthermore, our data show that different constellations of gap gene activities provide different input for the pair-rule genes, and thus strongly suggest that the overlap of hunchback and Krüppel in wild-type is functional in the formation of the patterns of pair-rule genes.     

An RNAi screen identifies additional members of the Drosophila Integrator complex and a requirement for cyclin C/Cdk8 in snRNA 3′-end formation

Formation of the 3′ end of RNA polymerase II–transcribed snRNAs requires a poorly understood group of proteins called the Integrator complex. Here we used a fluorescence-based read-through reporter that expresses GFP in response to snRNA misprocessing and performed a genome-wide RNAi screen in Drosophila S2 cells to identify novel factors required for snRNA 3′-end formation. In addition to the known Integrator complex members, we identified Asunder and CG4785 as additional Integrator subunits. Functional and biochemical experiments revealed that Asunder and CG4785 are additional core members of the Integrator complex. We also identified a conserved requirement in both fly and human snRNA 3′-end processing for cyclin C and Cdk8 that is distinct from their function in the Mediator Cdk8 module. Moreover, we observed biochemical association between Integrator proteins and cyclin C/Cdk8, and that overexpression of a kinase-dead Cdk8 causes snRNA misprocessing. These data functionally define the Drosophila Integrator complex and demonstrate an additional function for cyclin C/Cdk8 unrelated to its function in Mediator.     

A genome-wide screen identifies a single β-defensin gene cluster in the chicken: implications for the origin and evolution of mammalian defensins

Background  

Defensins comprise a large family of cationic antimicrobial peptides that are characterized by the presence of a conserved cysteine-rich defensin motif. Based on the spacing pattern of cysteines, these defensins are broadly divided into five groups, namely plant, invertebrate, α-, β-, and θ-defensins, with the last three groups being mostly found in mammalian species. However, the evolutionary relationships among these five groups of defensins remain controversial.