1. Presented by: Madeleine Cook & Tiffany Irizarry Transcriptomic Evidence That Longevity of Acquired Plastids in The Photosynthetic Slugs Elysia timida and Plakobranchus ocellatus Does Not Entail Lateral Transfer of Algal Nuclear Genes

2. “Solar-powered Sea slugs” Mollusca > Gastopoda > Opisthobranch > Sacoglossa Distribution: Mediterranean, Canary Islands, Caribbean, Eastern Atlantic Kleptoplasty (chloroplast symbiosis) –Elysia chlorotica –Elysia crispate –Elysia timida –Plakobranchus ocellatus * Only plastids of these species show a high photosynthetic rate over long periods of time. E. chlorotica E. crispate

3. Elysia timida Possess large red/orange spots on parapodia, head and rhinophores. Depth: 1 to 2 meters Feed on green algae (chlorophytes) –juveniles preferring Cladophora –adults preferring Acetabularia. E. timida Acetabularia Cladophora

4. Plakobranchus ocellatus Depth: < 1 to 18 meters Feed on wide variety of at least 5 marine species of ulvophyceaen chlorophytes P. ocellatus Ulvophyceaen chlorophyte

5. Sacoglossan life cycle

6. Photosynthetic Machinery Light Dependent Reactions:

7. Photosynthetic Machinery Light Independent Reactions

8. Hypothesis and Purpose “ If algal nuclear genes are transferred to the animal either during feeding or in the germ line, and it they are expressed, then they should be readily detectable with deep- sequencing methods” Wägele H et al. Mol Biol Evol 2011;28:699-706 © The Author(s) 2010. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

9. Materials and Methods RNA Analysis Transmission Electron Microscopy PAM Measurements

10. Photosynthetically active sacoglossans and transmission electron micrographs (TEMs) of their plastids. Wägele H et al. Mol Biol Evol 2011;28:699-706 © The Author(s) 2010. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

11. Photosynthetic activity of species investigated in this study. Wägele H et al. Mol Biol Evol 2011;28:699-706 © The Author(s) 2010. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

12. Wägele H et al. Mol Biol Evol 2011;28:699-706

13. Overview of phylogenetic relationships of photosynthetically active slugs within the Plakobranchidae (Sacoglossa). Wägele H et al. Mol Biol Evol 2011;28:699-706 © The Author(s) 2010. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

14. Expressed genes in Plakobranchus ocellatus and Elysia timida having nuclear-encoded homologues for chloroplast proteins in Arabidopsis. Wägele H et al. Mol Biol Evol 2011;28:699-706 © The Author(s) 2010. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

15. Conclusion: Essential plastid proteins are not provided by the slugs. The slugs maintain their long-lived plastids without the help of algal nuclear genes. –This does not exclude the possibility that some genes might have been ‘‘transferred’’ from the algae to the slugs. –But, it does exclude the possibility that plastid longevity in these two sacoglossans depend upon the expression of… Acquired algal photosynthesis genes (germ line or otherwise) Sequestered algal photosynthesis genes Sequestered algal photosynthesis mRNAs

16. “Bones to pick” with Rumpho et al… Rumpho Nuclear-encoded psbO gene of V. litorea had been transferred from the algal to E. chlorotica genome and is functionally expressed. Wa ̈ gele But targeting this gene product into the plastid would entail some considerable difficulties, because of the different number of membranes surrounding the V. litorea plastids in the algal and the slug! If the psbO has been transferred in the E. chlorotica, how does the protein get inside the kleptoplast? Bone

17. Jeopardy Review Sea SlugsBackgroundResultsConclusionRumpho vs. Wagele 10 20 30 40 50

18. Sea Slug are a part of the ________ phylum. Mollusca

19. The process of sequestering & utilizing living chloroplast from the algae they eat is known as Kleptoplasty

20. Sacoglossan sea slugs are unique in the animal kingdom in that they sequester and maintain active plastids that they acquire from the _____________ upon which they feed, making the animals photosynthetic. siphonaceous algae.

21. List one of the four Kleptoplasty (Chloroplast) Symbiosis : Elysia chlorotica Elysia crispate Elysia timida Plakobranchus ocellatus

22. Name the preference of the green Agal for the E. timida juvenilies Cladophora

23. ___________ is a division of green algae, which are associated with mostly aquatic photosynthetic eukaryotic organism. Chlorophyta

24. _____________ is the most commonly seen plakobranchid in sandy and mixed habitats from < 1 to 18 m (3-60 ft). It occurs in highly protected to moderately protected back reef areas Plakobranchus ocellatus

25. Animal on left with its parapodial flaps pulled open to show the longitudinal ridges which line the inside of the ____________ "parapodial cavity” P. ocellatus

26. ________ _____ are surrounded by two membranes and require a nuclear-encoded RuBisCO small subunit (RbcS) and intrinsic antenna proteins (light harvesting complex protein [LHCP]) for function. (E.timida) Chlorophyte plastids

27. ONLY ______ ________ (NOT THE ALGAL NUCLEI) ARE SEQUESTERED BY THE ANIMALS DURING FEEDING! ALGAL PLASTIDS

28. , A sacoglossan with short-term plastid retention (Thuridilla hopei) shows degrading chloroplasts in the _______ _______ cells when collected and mainly plastid remnants after 15 days of starvation Digestive glands

29. Used a deep-sequencing approach to focus on expressed genes from photosynthesizing animals, in order to identify expressed genes that might have been acquired from __________. Algae

30. The Acetabularia data were obtained from another database and show that ulvophyceaen nuclear sequences specific to photosynthesis, had they been present in the __________ data, would have readily been detected with the method use Sacoglossan

31. Expressed genes in Plakobranchus ocellatus and Elysia timida having nuclear-encoded homologues for chloroplast proteins in ____________. Arabidopsis

32. Major ________ ________ _____ required by typical chlorophyte plastids (RbcS, LHCP, photosystem I and II components, and Calvinvcycle enzymes) are not expressed by the slugs slugs. Nuclear Encoded Portion

33. The slugs maintain their long-lived plastids without the help of _____ ______ ______ Algal Nuclear Genes

34. Genes are often transferred from symbiont to host chromosomes in the wake of organelle origins during ____________ (You-are-what-you-eat Theory) Endosymbiosis

35. These results stand in marked contrast to recent reports of gene transfer in a different sacoglossan species with long- term plastid maintenance to who? Rumpho

36. This paper exclude the possibility that plastid longevity in these two sacoglossans depend upon the expression of… * Acquired algal photosynthesis genes (germ line or otherwise) *Sequestered algal photosynthesis genes * Sequestered algal photosynthesis mRNAs

37. WHY CAN WE ASSUME THAT THIS IS THE CASE?!?!?. These results stand in marked contrast to recent reports of gene transfer in a different sacoglossan species with long- term plastid maintenance!

38. True or False: Both Rumpho and Wagele suggested that there is lateral transfer of algal Nuclear Genes False

39. How does the case of sacoglossan puts that theory of “You are what you eat” to the test because ….. The physical interactions between food plastids and their feeding host could hardly be more intimate

40. Insert Text for Question Category 5 – 30 points

41. According to Rumpho, The _____ and _____ peptide are responsible for the targeting across a series of four translocons. Signal and Transit

42. Any Questions? What future studies are needed?

43. Work Cited: http://www.newscientist.com/data/images/ns/cms/dn16124/dn16124-1_300.jpg http://www.seaslugforum.net/images/jefftd2a.jpg http://www.emop.org/emop%20images/e.timida01.jpg http://www.seaslugforum.net/find/elystimi http://www.dr-ralf-wagner.de/Bilder/Cladophora_glomerata-DF.jpg http://web.ku.edu/~ifaa/jpg/Inouye/D-Acetabularia_ryukyuensis_1.jpg http://www.seaslugforum.net/find/placocel