The initiation of pair-rule stripes in the Drosophila blastoderm.

The interactions between the products of gap genes and pair-rule promoters results in the single most dramatic increase in the spatial complexity of gene expression during the segmentation process. We attempt to relate recent findings on the regulation of striped patterns of gene expression in the early Drosophila embryo to general strategies of gene expression and development employed by higher organisms.     

Pair-rule and gap gene mutants in the flour beetle Tribolium castaneum

 Early pattern formation in the Drosophila embryo occurs in a syncytial blastoderm where communication between nuclei is unimpeded by cell walls. During the development of other insects, similar gene expression patterns are generated in a cellular environment. In Tribolium, for instance, pair-rule stripes are transiently expressed near the posterior end of the growing germ band. To elucidate how pattern formation in such a situation deviates from that of Drosophila, functional data about the genes involved are essential. In a genetic screen for Tribolium mutants affecting the larval cuticle pattern, we isolated 4 mutants (from a total of 30) which disrupt segmentation in the thorax and abdomen. Two of these mutants display clear pair-rule phenotypes. This demonstrates that not only the expression, but also the function of pair-rule genes in this short-germ insect is in principle similar to Drosophila. The other two mutants appear to identify gap genes. They provide the first evidence for the involvement of gap genes in abdominal segmentation of short-germ embryos. However, significant differences between the phenotypes of these mutants and those of known Drosophila gap mutants exist which indicates that evolutionary changes occurred in either the regulation or action of these genes. Received: 8 May 1998 / Accepted: 17 June 1998     

Zygotically active genes that affect the spatial expression of the fushi tarazu segmentation gene during early Drosophila embryogenesis

The establishment of the segmental body pattern of Drosophila requires the coordinated functions of three classes of zygotically active genes early in development. We have examined the effects of mutations in these genes on the spatial expression of the fushi tarazu (ftz) pair-rule segmentation gene. Mutations in four gap loci and in three pair-rule loci dramatically affect the initial pattern of transverse stripes of ftz-containing nuclei. Five other pair-rule genes and several other loci that affect the larval cuticular pattern do not detectably affect ftz expression. No simple regulatory relationships can be deduced. Rather, expression of the ftz gene depends upon the interactions among the different segmentation genes active at each position along the anterior-posterior axis of the early embryo.     

Dose-dependent regulation of pair-rule stripes by gap proteins and the initiation of segment polarity

A key step in Drosophila segmentation is the establishment of periodic patterns of pair-rule gene expression in response to gap gene products. From an examination of the distribution of gap and pair-rule proteins in various mutants, we conclude that the on/off periodicity of pair-rule stripes depends on both the exact concentrations and combinations of gap proteins expressed in different embryonic cells. It has been suggested that the distribution of gap gene products depends on cross-regulatory interactions among these genes. Here we provide evidence that autoregulation also plays an important role in this process since there is a reduction in the levels of Kruppel (Kr) RNA and protein in a Kr null mutant. Once initiated by the gap genes each pair-rule stripe is bell shaped and has ill-defined margins. By the end of the fourteenth nuclear division cycle, the stripes of the pair-rule gene even-skipped (eve) sharpen and polarize, a process that is essential for the precisely localized expression of segment polarity genes. This sharpening process appears to depend on a threshold response of the eve promoter to the combinatorial action of eve and a second pair-rule gene hairy. The eve and hairy expression patterns overlap but are out of register and the cells of maximal overlap form the anterior margin of the polarized eve stripe. We propose that the relative placement of the eve and hairy stripes may be an important factor in the initiation of segment polarity.     

The zygotic control of Drosophila pair-rule gene expression. I. A search for new pair-rule regulatory loci

The examination of pair-rule gene expression in wild-type and segmentation mutant embryos has identified many, but not necessarily all, of the elements of the regulatory system that establish their periodic patterns. Here we have conducted a new type of search for previously unknown regulators of these genes by examining pair-rule gene expression in blastoderm embryos lacking parts of or entire chromosomes. This method has the advantage of direct inspection of abnormal pair-rule gene patterns without relying upon mutagenesis or interpretation of larval phenotypes for the identification of segmentation genes. From these experiments we conclude that: (i) most zygotically required regulators of the fushi tarazu (ftz), even-skipped (eve) and hairy (h) pair-rule genes have been identified, except for one or more loci we have uncovered on chromosome arm 2L; (ii) the repression of the ftz and eve genes in the anterior third of the embryo is under maternal, not zygotic control; and (iii) there are no general zygotically required activators of pair-rule gene expression. The results suggest that the molecular basis of pair-rule gene regulation can be pursued with greater confidence now that most key trans-acting factors are already in hand.     

Breakdown of abdominal patterning in the Tribolium Kruppel mutant jaws

During Drosophila segmentation, gap genes function as short-range gradients that determine the boundaries of pair-rule stripes. A classical example is Drosophila Krüppel (Dm'Kr) which is expressed in the middle of the syncytial blastoderm embryo. Patterning defects in Dm'Kr mutants are centred symmetrically around its bell-shaped expression profile. We have analysed the role of Krüppel in the short-germ beetle Tribolium castaneum where the pair-rule stripes corresponding to the 10 abdominal segments arise during growth stages subsequent to the blastoderm. We show that the previously described mutation jaws is an amorphic Tc'Kr allele. Pair-rule gene expression in the blastoderm is affected neither in the amorphic mutant nor in Tc'Kr RNAi embryos. Only during subsequent growth of the germ band does pair-rule patterning become disrupted. However, only segments arising posterior to the Tc'Kr expression domain are affected, i.e. the deletion profile is asymmetric relative to the expression domain. Moreover, stripe formation does not recover in posterior abdominal segments, i.e. the Tc'Kr(jaws) phenotype does not constitute a gap in segment formation but results from a breakdown of segmentation past the 5th eve stripe. Alteration of pair-rule gene expression in Tc'Kr(jaws) mutants does not suggest a direct role of Tc'Kr in defining specific stripe boundaries as in Drosophila. Together, these findings show that the segmentation function of Krüppel in this short-germ insect is fundamentally different from its role in the long-germ embryo of Drosophila. The role of Tc'Kr in Hox gene regulation, however, is in better accordance to the Drosophila paradigm.