Functional characterisation of LEA proteins from bdelloid rotifers

Abstract

‘Anhydrobiosis’ or ‘life without water’ is a fascinating phenomenon that was first described by the eminent Dutch microscopist Antony van Leewenhoek in bdelloid rotifers in 1702. Despite being studied for over three hundred years, our understanding of its molecular basis remains largely elusive. Recently, two group 3 LEA (late embryogenesis abundant) proteins, ArLEA1A and ArLEA1B, have been identified in the desiccation-induced gene set in the bdelloid species Adineta ricciae that are hypothesised to protect these animals by preventing protein aggregation and stabilising membranes respectively. In this dissertation, the functional characteristics of bdelloid LEA proteins have been further explored using computational and experimental tools. Analysis of their phylogeny and domain composition reveals that ArLEA1A and ArLEA1B are evolutionarily distinct from other related group 3 LEA proteins. Moreover, unlike some LEA proteins that are unstructured, bdelloid LEA proteins are predicted to form a-helices, with ArLEA1B having an additional propensity to polymerise into tropomyosin like filaments. These proteins are also predicted to localise in the ER (endoplasmic reticulum) and interact with cell signalling molecules. Intracellular localisation analysis of ArLEA1A and ArLEA1B using confocal microscopy confirms that these proteins translocate into the ER as predicted and their distribution within the entire secretory system and the cell exterior is regulated by their N- and C-terminal signals. Preliminary results suggest that over-expressing ArLEA1A fails to provide protection against protein aggregation within the mammalian ER. Lastly, both ArLEA1A and ArLEA1B are found to provide partial protection against desiccation induced damage of fluorescence properties of the red fluorescent protein mCherry. Overall, the results in this dissertation provide important mechanistic insights into the mode of action of bdelloid LEA proteins during anhydrobiosis.