A novel group 6 LEA protein from diapause embryos of Artemia franciscana is cytoplasmically localized

Abstract

The expression of late embryogenesis abundant (LEA) proteins is one mechanism by which anhydrobiotic organisms survive periods of severe water loss. Artemia franciscana is an animal extremophile that uniquely expresses LEA proteins concurrently from Groups 1, 3, and 6. In this study we examine the subcellular localization of AfrLEA6, a Group 6 LEA protein from embryos of A. franciscana. Immunohistochemistry reveals that AfrLEA6 is located in the cytoplasm of diapause embryos and does not co-localize with nuclei or mitochondria. Due to a trace contaminant arising from chitin-based affinity chromatography during AfrLEA6 purification, the primary antiserum displayed affinities for both AfrLEA6 as well as Artemia chitin-binding proteins. This contaminant (fusion protein of intein plus chitin binding domain) co-migrates with AfrLEA6 during SDS-PAGE. Pre-adsorption of the antiserum with dechorionated embryos was required to remove the non-specific fluorescence in the embryonic cuticular membrane. Results of this study are consistent with the apparent importance of distributing multiple types of LEA proteins across many subcellular locations in anhydrobiotic organisms.