Regulatory interactions between two actin nucleators, Spire and Cappuccino

Area of Science:

• Cell Biology
• Molecular Biology
• Developmental Biology

Background:

• Spire and Cappuccino are crucial actin nucleation factors for establishing polarity in Drosophila melanogaster oocytes.
• Mutant phenotypes for Spire and Cappuccino are nearly identical, suggesting functional interdependence.
• Biochemical interactions between Spire and Cappuccino have been previously observed.

Purpose of the Study:

• To elucidate the molecular mechanism underlying the interaction between Spire and Cappuccino family proteins.
• To investigate the conservation of this interaction across metazoan phyla.
• To determine the functional consequences of the Spire-Cappuccino interaction on actin nucleation.

Main Methods:

• Biochemical assays to characterize protein-protein interactions.
• In vitro actin nucleation assays.
• Structural analysis of protein domains involved in the interaction.
• Observation of protein localization in Drosophila oocytes.

Main Results:

• The interaction between Spire and Cappuccino is conserved across metazoan phyla.
• The interaction is mediated by the binding of the formin homology 2 (FH2) domain of Cappuccino to the kinase noncatalytic C-lobe domain (KIND) of Spire.
• Two KIND monomers bind to each FH2 dimer with nanomolar affinity, inhibiting actin nucleation.
• The Spire-Cappuccino complex formation enhances actin nucleation by Spire.
• Spire localizes to the oocyte cortex and disappears around stage 10b, coinciding with cytoplasmic streaming onset.

Conclusions:

• The Spire-Cappuccino interaction is a conserved mechanism regulating actin nucleation.
• The KIND-FH2 interaction provides a novel mode of formin regulation.
• This interaction plays a critical role in Drosophila oocyte development and polarity establishment.