Hans Schotman , Leena Karhinen , and Catherine Rabouille

1. dGRASP-Mediated NoncanonicalIntegrin SecretionIs Required for Drosophila Epithelial Remodeling Hans Schotman, LeenaKarhinen, and Catherine Rabouille Developmental Cell 14, 171–182, February 2008 Presented by Ian M. Parnham & Steven de Maat, supervised by Dr. Catherine Rabouille

2. Oogenesis • Divided in to early (stages 1–6), mid- (stages 7–10a), and late (stages 10b–14) oogenesis. • During oogenesis, the 15 nurse cells, the oocyte, and a surrounding monolayer of somatically derived follicle cells together comprise the egg chamber

3. Follicle Epithelium (Stage 10) • BC: Border cells • NC: Nurse cells • CC: Centripetal Cells • CFC: Cylindrical Follicular Cells • In stage 10A the Follicular cells migrate, covering the oocyte and changing there shape from cubical to columnar. • In stage 10B follicle cells completely cover oocyte separating nurse cells from the oocyte, elongating there shape

4. What are we going to look at? Immunofluoresence (IF) Immunoelectron microscopy (IEM) Figure 1 ZOC: zone of contact

5. Observations • Architecture of epithelial tissue depends intercellular junctions and adhesion • Adhesion with extracellular matrix components at the basal side is mediated by integrins and integral plasma membrane αβ heterodimers • In the wing imaginal discs of Dropsophila larvae, where αPS1βPS(PS1) and αPS2βPS (PS2) maintain integrity of the developing wing.

6. dGRASP • dGRASP has been considered bona fide Golgi proteins. • Involved in formation and maintenance of the Golgi • GrhA (GRASP65/55 homologue) has been shown to mediate Golgi independent secretion of AcbA in Dictyostelium.

7. Where does dGRASP localize to, during Oocyte development? Figure 1

8. Aim Investigate the role of dGRASP in follicular epithelium surrounding the developing oocyte

9. Do dGRASP binding partners also translocate towards the open ZOC during oocyte development? Figure 1

10. Where does the newly localized dGRASP come from? Figure 2

11. Do dgrasp RNA & dGRASP translocate to the same place? Figure 2

12. Recap • Stage 10A to 10B: formation of open ZOC • Stage 10B dGRASP localizes to the open ZOC, almost no co-localisation with Golgi • Stage 10B dgrasp mRNA also localizes to the open ZOC

13. What does dGRASP do at the open ZOC? Wild Type – Stage 10B dGRASP is important in maintaining follicular epithelium integrity dgrasp100.2 – Stage 10B Figure 3

14. How are integrins deposited during the progression from stage 10A -> 10B Integrin αPS1 Is deposited at the open ZOC in stage 10B Figure 3

15. Is Integrin αPS1 also deposited at the open ZOC of dgrasp100.2in stage 10B? Figure 4

16. Is Integrin αPS1 also deposited at the open ZOC of dgrasp100.2 in stage 10B? Figure 4

17. Is βPS also effected by dgrasp100.2? Figure 4

18. Recap • dGRASP mutant leads to disrupted follicular epithelium • αPS1 is deposited at the open ZOC • αPS1 deposition at the open ZOC is dependent on dGRASP • βPS and other adherens junction proteins were dGRASP independent • How does dGRASP enable αPS1 deposition?

19. Is αPS1 deposition Golgi dependent? Figure 6

20. Is αPS1 deposition dSyntaxin-5 dependent? Figure 6

21. Conclusion • In stage 10B dGRASP localizes to the open ZOC, almost no co-localisation with Golgi • dGRASP mutant leads to disrupted follicular epithelium, by interfering with αPS1 deposition at the open ZOC • dGRASP mediated αPS1 deposition is Golgi independent

22. Discussion Is αPS1 newly synthesized at the open ZOC? Figure 5

23. Discussion Is αPS1 newly synthesized at the open ZOC? Figure 5

24. The End