The role of pair-rule genes in Tribolium segmentation

Doctor of PhilosophyDepartment of BiologySusan J. BrownAll arthropods share a segmented body plan. Detailed studies on segmentation mechanisms in the long-germ insect Drosophila melanogaster identified a segmentation hierarchy composed of maternal, gap, pair-rule, and segment polarity genes. In this hierarchy, pair-rule genes play an important role to translate gradients of regional information from maternal and gap genes into segmental expression of segment polarity genes. However, our understanding of the role of pair-rule genes in other short-germ insects and basally branching arthropods is still limited. To gain insights into the role of pair-rule genes in short-germ segmentation, I analyzed genetic interactions as well as expression patterns and functions of homologs of Drosophila pair-rule genes in the short-germ insect Tribolium castaneum. Interestingly, despite the pair-rule like expression patterns of Tribolium homologs of almost all eight canonical Drosophila pair-rule genes, only five have a segmentation function. Knock-down of primary pair-rule genes caused asegmental and truncated phenotypes while knock-down of secondary pair-rule genes caused typical pair-rule phenotypes. Epistatic analysis between the genes revealed that primary pair-rule genes form a gene circuit to prepattern a two-segmental unit, and secondary pair-rule genes are downstream targets of the gene circuit. The typical pair-rule phenotypes observed in secondary pair-rule gene RNAi embryos led to a detailed comparative analysis of the role of paired (prd) and sloppy-paired (slp) between Drosophila and Tribolium. This study revealed that prd is functionally conserved while the functional parasegmental register for Tribolium slp is opposite that of Drosophila slp. The fact that the register of slp function has evolved differently in the lineages leading to Drosophila and Tribolium reveals an unprecedented flexibility in pair-rule patterning. Despite this flexibility in pair-rule patterning between Drosophila and Tribolium, segmental expression of engrailed (en) and wingless (wg) at parasegmental boundaries is conserved in both insects. Analysis of double and triple RNAi for pair-rule genes in Tribolium revealed that the primary pair-rule genes even-skipped and runt are redeployed to directly regulate en and wg with prd or slp at parasegmental boundaries. This redeployment of primary pair-rule genes seem to compensate for the apparently fewer number of functional secondary pair-rule genes in Tribolium segmentation

Wnt-Dependent Epithelial Transitions Drive Pharyngeal Pouch Formation

The pharyngeal pouches, which form by budding of the foregut endoderm, are essential for segmentation of the vertebrate face. To date, the cellular mechanism and segmental nature of such budding have remained elusive. Here, we find that Wnt11r and Wnt4a from the head mesoderm and ectoderm, respectively, play distinct roles in the segmental formation of pouches in zebrafish. Time-lapse microscopy, combined with mutant and tissue-specific transgenic experiments, reveal requirements of Wnt signaling in two phases of endodermal epithelial transitions. Initially, Wnt11r and Rac1 destabilize the endodermal epithelium to promote the lateral movement of pouch-forming cells. Next, Wnt4a and Cdc42 signaling induce the rearrangement of maturing pouch cells into bilayers through junctional localization of the Alcama immunoglobulin-domain protein, which functions to restabilize adherens junctions. We propose that this dynamic control of epithelial morphology by Wnt signaling may be a common theme for the budding of organ anlagen from the endoderm

Genetic regulation of engrailed and wingless in Tribolium segmentation and the evolution of pair-rule segmentation

AbstractIn Drosophila, primary pair-rule genes establish the parasegmental boundaries and indirectly control the periodic expression of the segment polarity genes engrailed (en) and wingless (wg) via regulation of secondary pair-rule genes. Although orthologs of some Drosophila pair-rule genes are not required for proper segmentation in Tribolium, segmental expression of Tc-en and Tc-wg is conserved. To understand how these segment polarity genes are regulated, we examined the results of expressing one or two pair-rule genes in the absence of the other known pair-rule genes. Expression of one or both of the secondary pair-rule genes, Tc-sloppy-paired (Tc-slp) and Tc-paired (Tc-prd), activated Tc-wg in the absence of the primary pair-rule genes, Tc-even-skipped (Tc-eve), Tc-runt (Tc-run) and Tc-odd-skipped (Tc-odd). Tc-eve alone failed to activate Tc-wg or Tc-en, but in combination with Tc-run or Tc-prd activated Tc-en. These results, interpreted within the pair-rule gene expression patterns, suggest separate models for the genetic regulation of the juxtaposed expression of Tc-wg and Tc-en at odd- and even-numbered parasegmental boundaries, respectively. Conserved interactions between eve and prd at the anterior boundary of odd-numbered parasegments may reflect an ancestral segmentation mechanism that functioned in every segment prior to the evolution of pair-rule segmentation