Molecular Regulation of Developmental Pathways

Life-cycle of Drosophila
Embryo hatches as larva 1 day after fertilization
Becomes pupa (hard casing)
Hatches as adult fly
Total process is about 10 days (rapid development)
Embryo has dorsal and ventral side
Jani Nusslein-Volhard and Eric Wieschaus
Performed the Heidelberg Screen to identify genes in early embryonic development
(saturation screen)
Saturation Screen: produced many mutations and screened them to identify allele
Heidelberg Screen
Screened ~27k mutations
548 embryonic lethal mutations showed abnormalities in body plan of the embryo
Genes that influence the spatial patterning
Located in 139 genes
Found mutants that failed to form ventral structure (became dorsalized)
Found mutants that failed to form dorsal structure (became ventralized)
Both maternal-effect
Drosophila Development
Nuclear division is separate rom cell division in early embryo
Syncytial Stage: thousands of nuclei in cytoplasm
Nuclei migrate to outer edge of egg and cell boundaries form
Pole cells produce germ line, rest are somatic
Embryonic Regions
Different tissues develop from cells on axis
Mesoderm (ventral side): makes muscle and connective tissue
Neurogenic Ectoderm: makes central nervous system and skin
Dorsal Skin: makes dorsal skin
Amnion (dorsal side): thrown away after fly embryo is developed
Concentration Gradient in Embryo
Toll: receptor that covers egg cell membrane
Spatzle: high in concentration on ventral side (almost none on dorsal side)
Spatzle binds to Toll
Pelle (and Tube): activated by Toll receptor and becomes kinase
Cactus: phosphorylated by Pelle and broken down
Dorsal: TF bound to Cactus and released when Cactus is phosphorylated
Nucleus: Dorsal enters nucleus and activates transcription

Ventral-Dorsal Gradients
Extracellular gradient of Spatzle concentration
Gradient of Toll activation in cell membrane
Gradient of Dorsal concentration in nucleus
Spatzle is a morphogen: extracellular concentration specifies states of transcription
in nucleus
Dorsal is the TF responsible for making the ventralized side of the embryo
If Cactus is not functional, Dorsal will always enter and entire embryo is ventralized
If Dorsal never enters nucleus, embryo is dorsalized
All of these proteins are synthesized from mothers DNA (maternal effect)
Zygotic Effect Expression: Embryos own genotype determines its phenotype
Genes in Each Region
Twist and Snail: expressed in mesoderm
Sog and Rhomboid: expressed in neurogenic ectoderm
Dpp: expressed in dorsal skin
Dpp: Repressed by presence of Dorsal (Dorsal is acting as a repressor)
Sog and Rhomboid: activated in lower concentrations of Dorsal
Snail and Twist: activated in higher concentrations of Dorsal
Binding Site Affinity
Sog: expressed at all concentrations of Dorsal (4 high affinity binding sites)
Rhomboid: expressed only at medium and high levels (1 high affinity binding site)
Twist: only expressed at high level (low affinity binding sites)
If rhomboid lost its high affinity binding site, what would happen to rhomboid
expression? It would still have low affinity sites, but require high concentrations of
Dorsal
Repression of Snail
Expression of Rho and Sog, repression of Snail
Expression of Snail, repression of Rho and Sog
If Snail is inactivated, Rho expands past ventral midline instead of staying at outer
regions of embryo
Gene Regulatory Network (GRN): flowcharts that show set of genes that control
another set of genes