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

“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

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

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

Sacoglossan life cycle

Photosynthetic Machinery Light Dependent Reactions:

Photosynthetic Machinery Light Independent Reactions

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: © The Author(s) Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

Materials and Methods RNA Analysis Transmission Electron Microscopy PAM Measurements

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

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

Wägele H et al. Mol Biol Evol 2011;28:

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

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: © The Author(s) Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

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

“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

Jeopardy Review Sea SlugsBackgroundResultsConclusionRumpho vs. Wagele

Sea Slug are a part of the ________ phylum. Mollusca

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

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.

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

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

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

_____________ 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

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

________ _____ 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

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

, 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

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

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

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

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

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

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

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

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

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!

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

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

Insert Text for Question Category 5 – 30 points

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

Any Questions? What future studies are needed?

Work Cited: