Download Figure S2 - Development

bozozok acts as a strong bicoid antimorph in Drosophila
Given the similarity of the homeodomain of Boz to that of the Drosophila gene bicoid (bcd), we argue that Boz
might be able to bind bcd sites in the Drosophila embryo (indeed, we were able to show that Boz binds Bcd-binding
sites, see Fig. 5A). Under the control of the nanos promoter, we expressed an mRNA encoding the full-length boz
ORF, and containing 3 bcd mRNA localization signals, during Drosophila oogenesis. Signals for anterior mRNA
localization remain functional in this hybrid gene (data not shown). Embryos from females carrying one copy of the
bcd-boz transgene express a dominant phenotype resembling that of bcd null mutants (see Fig. S2A-D): head, thorax
and anterior abdomen do not develop. In contrast to the bcd phenotype, however, embryos from bcd-boz transgenic
females do not form a duplicated telson. Embryos from females with one copy of the bcd-boz transgene do not
establish the anterior expression domain of the gap gene hunchback (Fig. S2E,F), which is directly activated by Bcd
in wild-type embryos (Driever et al., 1989). Segmentation is distorted in the thorax and anterior abdomen, as
revealed by even-skipped expression (Fig. S2G,H), which appears similar to or stronger than that observed in bcd
mutant embryos (Driever and Nüsslein-Volhard, 1988).
The differences in phenotypes may be explained by different effects of bcd loss-of-function and bcd antimorphic
activities on development: A Bicoid-repressor fusion would also repress Krüppel, which in bicoid mutants is still
activated by hunchback. This may explain the more pronounced abdominal defects in embryos from bcd-boz
females. Furthermore, given the interactions between the terminal system and boz (increased dosage of boz can
rescue anterior terminal defects in terminal class mutants; bcd and the terminal system have common downstream
targets) (Schaeffer et al., 2000), zygotic anterior terminal genes are also influenced by bcd. A loss-of-function bcd
allele is expected to have different effects on anterior terminal development when compared with the potential
dominant-negative effect of a repressor binding to Bicoid binding sites.
Our data indicate that the bcd-boz gene has an antimorphic activity with respect to bcd as it phenocopies bcd
mutant defects in a dominant fashion. Such a finding is consistent with Boz serving as a transcriptional repressor that
can compete with Bcd for binding to target sites and prevent Bcd-mediated activation of downstream targets.
MATERIALS AND METHODS
For the analysis of boz activity in Drosophila, the boz-coding region was cloned into the P-element-based pCaSpeR vector to be
framed by 1.1 kb of the nanos 5’ upstream region and the bcd 3’ UTR (E. Wimmer, unpublished) (Niessing et al., 1999).
Transgenic lines were generated by P-element-mediated germline transformation (Rivera-Pomar et al., 1995). The boz containing
mRNA was localized correctly at the anterior tip of the embryo (data not shown). Transformed fly strains were maintained
through males only, as the transgene induced lethality in embryos when transmitted through females. Transformants were crossed
to bcdE1 mutants (Frohnhoefer and Nüsslein-Volhard, 1986). Expression of hunchback was visualized using digoxigenin labeled
RNA probes (Roche), and expression of even-skipped by immunohistochemistry using Cy3 (Jackson Laboratories).
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