Graduation Year

2015

Document Type

Thesis

Degree

M.S

Degree Name

Master of Science (M.S.)

Degree Granting Department

Integrative Biology

Major Professor

Kathleen Scott, Ph.D.

Co-Major Professor

Christina Richards, Ph.D.

Committee Member

Christina Richards, Ph.D.

Committee Member

James Garey, Ph.D.

Keywords

chlorophyll a, Elysia chlorotica, endosymbiosis, horizonal gene transfer, kleptoplast, Vaucheria litorea

Abstract

The sacoglossan sea slug, Elysia chlorotica, feeds upon and sequesters plastids from the heterokont alga, Vaucheria litorea, and maintains the metabolically active organelles for up to nine months under starvation conditions while utilizing the photosynthate to survive and reproduce. The photosynthetic pigment, chlorophyll a (Chla), is found in all oxygenic photosynthetic organisms and is responsible for capturing photons of light and converting them into chemical energy. Chlorophyll and its associated proteins involved in the light capturing process are subject to photo oxidative damage and must be continually replaced for ongoing photosynthesis to continue; however, genes encoding these proteins are present in the algal nucleus, presenting a conundrum for sustained plastid photosynthetic activity outside the algal cell. One possibility is that Chla is synthesized by the E. chlorotica-kleptoplast association, due to transfer of algal nuclear genes to the sea slug genome. For this study, molecular and biochemical techniques were employed to determine if Chla is synthesized by the animal. Using algal transcriptome sequences for primer design and amplification of target DNA using polymerase chain reaction (PCR), we have identified and sequenced three algal nuclear-encoded gene fragments that correspond to enzymes in the chlorophyll synthesis pathway and one enzyme in the porphyrin synthesis pathway in adult slug and veliger larvae. Sequences from these genes were nearly identical to those present in the alga. Furthermore, these genes are functional; incubation of slugs with radiolabeled 5-aminolevulinic acid (14C-5-ALA), a precursor of chlorophyll biosynthesis, resulted in production of 14C-labeled chlorophyll, as assayed and identified via HPLC resolution of extracts from slugs. In addition, Chla synthesis in the animal occurs for at least six months under starvation conditions. The discovery of chlorophyll synthesis in E. chlorotica is the first animal known to synthesize Chla; moreover, this finding helps elucidate how ongoing photosynthesis can occur in the sea slug after many months in the absence of its algal food.

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