Control of Scribble localization at the cell cortex in migrating astrocytes


Supervising PI

Sandrine Etienne-Manneville (website)

ESR5 Yamini Ravichandran

Project Description

The small G protein Cdc42 plays an evolutionary conserved role in cell polarity. It contributes to the polarization and directed migration of epithelial cells and astrocytes. The goal of this project is to better determine the regulatory mechanisms of Cdc42 localisation at the cell cortex. We expect to provide a clearer view of the fundamental polarity signals and in particular in the molecular mechanisms responsible for Cdc42 recruitment at the cell cortex and the exact role of Scrib in these events.

Summary of Results

Cdc42 is an evolutionary conserved small G protein controlling many cellular processes, such as cell polarity, cytoskeleton dynamics and membrane trafficking, in response to a wide variety of extracellular signals. While inactive Cdc42 localizes in the cytosol, its regulators and effectors are generally membrane- associated. The regulation and functions of Cdc42 at the cell plasma membrane have been extensively studied. However, a significant pool of Cdc42 localizes at the Golgi apparatus where its specific regulation and functions remain elusive. Previous studies have focused on only one isoform of Cdc42, known as the Placental isoform, whereas in vertebrates two isoforms of Cdc42, that are alternative splice variants of one another, are expressed. We have shown that the two isoforms are localized differently in primary rat astrocytes. We use primary rat astrocytes as our working model as they express both the isoforms. While the Placental isoform is mostly found in the cell cytoplasm and at the plasma membrane, the Brain isoform localizes at the Golgi apparatus and on intracellular vesicles. We have also shown that the two isoforms carry out different functions during cell migration, suggesting that the differences between these two isoforms which only differs by the last 10 amino acids are responsible for their distinct localisation and function. Moreover, a patient suffering from a generalized pustular psoriasis disease carries a mutation in the C-ter domain of the Placental isoform which localizes to the Golgi apparatus, suggesting that the amino acid sequence and/or the lipid anchors are key elements in Cdc42 localization and function. The main questions that we ask are: 1) how is the subcellular localization of Cdc42 regulated and 2) how the different subcellular pools of Cdc42 interact. I use Giant Unilamellar Vesicles as model membranes to assess the preferential binding of the Cdc42 isoforms to membranes of different compositions and curvature and I have also developed optogenetic tools to activate and follow the spatio-temporal dynamics of Cdc42 in primary rat astrocytes.

References

  1. Osmani N, Vitale N, Borg J-P, Etienne-Manneville S. Scrib controls Cdc42 localization and activity to promote cell polarization during astrocyte migration. Curr Biol CB. 2006 Dec 19;16(24):2395–405.
  2. Nola S, Sebbagh M, Marchetto S, Osmani N, Nourry C, Audebert S, et al. Scrib regulates PAK activity during the cell migration process. Hum Mol Genet. 2008 Nov 15;17(22):3552–65.
  3. Osmani N, Peglion F, Chavrier P, Etienne-Manneville S. Cdc42 localization and cell polarity depend on membrane traffic. J Cell Biol. 2010 Dec 27;191(7):1261–9.
  4. Boëda B, Etienne-Manneville S. Spectrin binding motifs regulate Scribble cortical dynamics and polarity function. eLife. 2015;4.