Molecular characterization of artemin and ferritin from Artemia franciscana

Abstract

Embryos of the brine shrimp, Artemia franciscana, exhibit remarkable resistance to physiological stress, which is temporally correlated with the presence of two proteins, one a small heat shock/a-crystallin protein termed p26 and the other called artemin, of unknown function. Artemin was sequenced previously by Edman degradation, and its relationship to ferritin, an iron storage protein, established. The isolation from an Artemia expressed sequence tag library of artemin and ferritin cDNAs extends this work. Artemin cDNA was found to contain an ORF of 693 nucleotides, and its deduced amino-acid sequence, except for the initiator methionine, was identical with that determined previously. Ferritin cDNA is 725 bp in length with an ORF of 516 nucleotides. Artemin amino-acid residues 32-185 are most similar to ferritin, but artemin is enriched in cysteines. The abundance of cysteines and their intramolecular spatial distribution suggest that artemin protects embryos against oxidative damage and/or that its function is redox regulated.The conserved regions in artemin and ferritin monomers are structurally similar to one another and both proteins assemble into oligomers. However, modeling of the quaternary structure indicated that artemin multimers lack the central space used for metal storage that characterizes ferritin oligomers, implying different roles for this protein.Probing of Northern blots revealed two artemin transcripts, one of 3.5 kb and another of 2.2 kb. These transcripts decreased in parallel and had almost disappeared by 16 h of development. The ferritin transcript of 0.8 kb increased slightly during reinitiation of development, then declined, and was almost completely gone by 16 h. Clearly, the loss of artemin and ferritin during embryo development is due to transcriptional regulation and proteolytic degradation of the proteins.

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

“…The second abundant cyst protein termed artemin (Slobin 1980) or the 19S complex (De Herdt et al 1979 is similar in amino acid sequence to ferritin, but artemin is enriched in cysteine and possesses an extended carboxylterminal tail not found in ferritin (De Graaf et al 1990;Chen et al 2003;Rasti et al 2009). 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).…”

“…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.…”

“…Most artemin mRNA is gone from early cyst-derived nauplii (Chen et al 2003) but when the protein disappears is unknown. Artemin is extremely stable, enduring extended cyst storage, years of anoxia, and heating in vitro at 70°C which causes binding of non-polyadenylated RNA (Warner et al 2004;Shahangian et al 2011).…”

Artemin, a Diapause-Specific Chaperone, Contributes to the Stress Tolerance of Artemia Cysts and Influences Their Release from Females

“…Artemin, an abundant, ATP-independent, diapause-specific molecular chaperone shown to protect transfected mammalian cells from stress and prevent protein denaturation in vitro, is a ferritin homolog (Chen et al, 2003;Warner et al, 2004;Chen et al, 2007;Hu et al, 2011;Shirzad et al, 2011;Shahangian et al, 2011). Artemin assembles oligomers of ~600-700 kDa formed from 24 monomers, each ~26 kDa (De Graaf et al, 1990;Chen et al, 2007;Clegg, 2011;Hu et al, 2011).…”

“…Artemin fails to bind iron because the carboxyl terminal extension of protein monomers thought to have a role in chaperoning (Shirzad et al, 2011) fills the oligomer cavity which is equivalent to the space where iron resides in ferritin. Additionally, artemin lacks all but a single residue of the di-iron ferroxidase center employed by ferritin in iron binding (Harrison and Arosio, 1996;Chen et al, 2003;Chen et al, 2007;Crichton and Declercq, 2010). Artemin is very thermotolerant, surviving at 90°C for 30 min, and heated artemin binds non-polyadenylated RNA (Warner et al, 2004).…”