Functional characterization of artemin, a ferritin homolog synthesized in Artemia embryos during encystment and diapause

Abstract

Oviparously developing embryos of the crustacean Artemia franciscana encyst and enter diapause, exhibiting a level of stress tolerance seldom seen in metazoans. The extraordinary stress resistance of encysted Artemia embryos is thought to depend in part on the regulated synthesis of artemin, a ferritin superfamily member. The objective of this study was to better understand artemin function, and to this end the protein was synthesized in Escherichia coli and purified to apparent homogeneity. Purified artemin consisted of oligomers approximately 700 kDa in molecular mass that dissociated into monomers and a small number of dimers upon SDS/PAGE. Artemin inhibited heat‐induced aggregation of citrate synthase in vitro, an activity characteristic of molecular chaperones and shown here to be shared by apoferritin and ferritin. This is the first report that apoferritin/ferritin may protect cells from stress other than by iron sequestration. Stably transfected mammalian cells synthesizing artemin were more resistant to heat and H2O2 than were cells transfected with vector only, actions also shared by molecular chaperones such as the small heat shock proteins. The data indicate that artemin is a structurally modified ferritin arising either from a common ancestor gene or by duplication of the ferritin gene. Divergence, including acquisition of a C‐terminal peptide extension and ferroxidase center modification, eliminated iron sequestration, but chaperone activity was retained. Therefore, because artemin accumulates abundantly during development, it has the potential to protect embryos from stress during encystment and diapause without adversely affecting iron metabolism.

Stress tolerance during diapause and quiescence of the brine shrimp, Artemia

“…Artemin has a monomeric molecular mass of 27 kDa and, like ferritin, assembles into oligomers of 24 subunits (Chen et al 2007;Hu et al 2011). In contrast to ferritin, the central cavity of oligomerized artemin is occupied by the carboxyl-terminus of constituent monomers and fails to bind iron, a finding augmented by electron microscopy and the paucity in artemin of di-iron ferroxidase center residues required for interaction with iron (De Graaf et al 1990;Chen et al 2003Chen et al , 2007.…”

“…The heat resistance of transformed bacteria synthesizing artemin is enhanced (Rasti et al 2009), and stably transfected mammalian cells containing artemin have increased tolerance to thermal and oxidative stress (Chen et al 2007). That ferritin and artemin are both chaperones suggests that artemin evolved from ferritin to produce an abundant, cyst-specific, molecular chaperone that neither sequesters iron nor disrupts iron metabolism, as would happen if large amounts of intracellular ferritin were synthesized and used as a chaperone.…”

“…Purified artemin prevents heat-induced denaturation of other proteins in the absence of added ATP, an activity shared by ferritin (Chen et al 2007;Hu et al 2011;Shahangian et al 2011). The heat resistance of transformed bacteria synthesizing artemin is enhanced (Rasti et al 2009), and stably transfected mammalian cells containing artemin have increased tolerance to thermal and oxidative stress (Chen et al 2007).…”

The expression pattern of dormancy-associated genes in multiple life-history stages in the rotifer Brachionus plicatilis

“…These generally support previous results (Denekamp et al, 2009), where 55 transcripts constituting *4.5% of the total number of sequenced clones in a resting egg subtractive library, were found for shsp-2, shsp-3, shsp-4 and shsp-5 (but not for shsp-1). Small heat shock proteins are specifically abundant in Artemia cysts and presume to function in maintaining the native folded structure of proteins (Chen et al, 2007).…”

MTH1745, a protein disulfide isomerase-like protein from thermophilic archaea, Methanothermobacter thermoautotrophicum involving in stress response

“…The molarity of MTH1745 in reaction mixtures was based on monomeric molecular masses of MTH1745 variants. Citrate synthase aggregation was monitored by measuring solution turbidity at 360 nm with a SPECTRAmax PLUS spectrophotometer every 4 min for 1 h. Bovine serum albumin (BSA) was used at 150 nM to evaluate nonspecific protection of citrate synthase (Chen et al 2007). …”