Research objectives

Regulation of secretion during development

Invitation for undergraduate students can be found here

1) Classical and unconventional secretion of junctional and adhesion proteins during epithelium remodeling
The integrity of epithelial tissue depends largely on the formation of physical cell-cell junctions as well as adhesion to the extracellular matrix. Both are mediated by a number of critical transmembrane proteins, such as E-cadherin at the Adherens junctions, claudin/occludin at the tight junctions and connexins at the gap junction as well as the adhesive integrins [1-2]. The Rabouille’s group studies the secretion of these proteins during epithelial development and remodeling in Drosophila and in tissue cultured epithelial mammalian cells. As transmembrane proteins, the adhesion/junctional proteins mentioned above should follow the classical secretory pathway (ER>ER exit sites>Golgi>PM) as described by the Nobel Prize winner George Palade [3]. However, recently, the Rabouille’s group has established that in Drosophila some key proteins in cell adhesion and junction formation bypass the Golgi on their way to the plasma membrane, making them critical and biologically unique [4]. Strikingly, this Golgi bypass requires a peripheral Golgi protein dGRASP [5] to be ectopically localized at the plasma membrane. In Drosophila mutant missing this protein, epithelium is strongly disorganized [6].
We study the relevance of this GRASP mediated Golgi bypass at other stages of Drosophila development, in Tribolium embryogenesis and in mammalian cells, including during tumour formation. We aim at identifying new components and substrates of this pathway as well as understand the developmental regulation eliciting its biogenesis. In this respect, we have established that mechanical stress plays a role [7] and continue to dissect this aspect.

2) Regulation of the functional organisation of the early secretory pathway
A large part of our past activity has been to identify principles underlying the functional organisation of the early secretory pathway (ER exit site-Golgi) in the Drosophila tissue cultured S2 cells [8-9]. In this respect, we have characterised the Drosophila orthologue of the large hydrophilic protein Sec16 that defines ER exit sites [10]. Recently, we have used an RNAi depletion approach of pre-selected putative ER proteins to identify new components involved this organisation [11].
Considering the strong regulation of secretion by signalling [12], we have also depleted 250 kinases and identify a number of involved atypical MAPK that we are now characterising (ms in revision).

3) Asymmetric distribution of mRNAs
Recently, we have developed a method to visualise RNA localisation at the ultrastructural level that is based on RNA in situ hybridisation coupled to immuno-EM on frozen sections [13]. In collaboration with the group of Ilan Davis (Oxford, UK), we have used it to visualise gurken in large cytoplasmic structures in the Drosophila oocyte [14] as well as bicoid RNA [15] (Weil et al, 2010) and we now use it to localise dgrasp mRNA at the basal site of the follicular epithelium, one of the first step in the biogenesis of the GRASP mediated Golgi bypass [6]. We are also interested in identifying cis-acting elements involved in this localisation.

References
[1] Lecuit T (2005) Trend Cell Biol 15:34; [2] Knust E and Bossinger O (2002) Science 298:1955; [3] Mellman I, and Warren W (2000) Cell 100:99. [4] Grieve AG and Rabouille C (2011) The Golgi book. CSH press; [5] Vinke FP, Grieve AG and Rabouille C (2011) Biochem J 433:423; [6] Schotman H, Karhinen L and Rabouille C (2008) Dev Cell 14:171-182; [7] Schotman H, Karhinen L and Rabouille C (2009) J Cell Sci. 122:2662; [8] Kondylis V et al (2007) Dev Cell 12:901; [9] Kondylis V and Rabouille C (2009). FEBS lett. 583:3827; [10] Ivan V et al (2008) Mol. Biol. Cell 19:4352; [11] Kondylis V et al (2011). In press PLOS One; [12] Farhan H and Rabouille C (2011) J Cell Sci124:171; [13] Herpers B, Xanthakis D and Rabouille C (2010) Nat Protoc 5:678; [14] Delanoue R et al (2007) Dev Cell 13:523. [15] Weil T et al (2010) Development 137:169.

About the group leader
Publication list

Undergraduate students
We welcome motivated undergraduate students to participate in one of these projects. Besides learning many cell biology techniques (tissue culture, Westerns, generating DNA constructs, transfections) as well as fly genetics, students can get familiar with imaging techniques depending on their own interests and experience.

If you are interested please contact  Catherine Rabouille  c.rabouille@hubrecht.eu