Understanding developmental control of cell polarity using single-cell in vivo biochemistry

Project Summary / Abstract Cell polarity is a fundamental feature of eukaryotic cells, and must be coordinated between cells and regulated to allow for normal animal development and tissue homeostasis. Despite genetic identification of proteins involved in cell polarity and a large body of knowledge about their interactions in vitro, it remains unclear how polarity proteins are organized into signaling complexes in cells. This lack of knowledge has prevented the field from understanding mechanisms of developmental control of polarity signaling in vivo. The long-term goal of the proposed research is to resolve the network of protein-protein interactions that supports animal cell polarity and to understand how this network can respond to developmental signals. To enable progress towards this goal, the applicants have developed innovative experimental tools that allow single-molecule measurements of native protein complex abundance in single cells. This project focuses on an evolutionarily conserved protein kinase, called aPKC, that plays a central role in polarity by localizing to one end of a polarized cell and dictating polarized cell behaviors. The applicants will make use of the C. elegans early embryo, in which cells reproducibly polarize in response to multiple spatial and temporal cues, to discover mechanistic links between developmental signals and the polarity machinery. The central hypothesis of this work is that developmental signals control cell polarity by altering the molecular complexes in which aPKC resides. This hypothesis will be explored by elucidating mechanisms that regulate assembly of aPKC into different complexes in the zygote (Aim 1); by determining how polarity is entrained to cell-cell contacts in 8-cell embryos (Aim 2); and by determining how translation of new protein components remodels the polarity system between these two stages (Aim 3). The work proposed in this application is significant because it will reveal fundamental mechanisms controlling cell polarity, and because it places these mechanistic studies in a developmental context. The proposed work is innovative, in the applicant’s opinion, because it uses novel experimental methods to perform biochemical, mechanistic studies in vivo. By studying the biochemical control of aPKC in multiple cellular and developmental contexts in a single experimental system, this work will identify fundamental mechanisms of PAR polarity signaling and to learn how these mechanisms are deployed to achieve different outcomes during development.