| dc.description | During diapause and quiescence, encysted Artemia gastrulae bring their metabolism to a reversible standstill and demonstrate a remarkable resistance to environmental stress. Previous results suggested that p26, an abundant, low molecular weight protein in Artemia cysts, is a small heat shock/alpha-crystallin protein and that it has a protective function in these embryos. In this study, p26 was purified to apparently homogeneity from cell-free extracts of Artemia cysts. p26 existed as multimers up to 700 kD in mass and multiple isoforms of the protein were detected on 2D-gels. A complete cDNA for p26 was cloned and sequenced, revealing an encoded polypeptide of 192 amino acids with a conserved "alpha-crystallin domain". Unusual characteristics of p26 were the presence of glycine- and arginine-rich regions in the N-terminal domain, the longest C-terminal extension yet observed in a small heat shock/alpha-crystallin protein, a lack of predicted alpha-helix structure in alpha-crystallin domain and the absence of the "Arg-X-X-Ser" phosphorylation motif. Artemia had multiple p26 genes and they were exclusively expressed in embryos undergoing encystment. During encystment, p26 mRNA first appeared in two day embryos, while p26 protein appeared in three day embryos. Both p26 mRNA and protein reached their highest amounts before the cysts were released and they remained at this level in activated cysts. p26 mRNA decreased rapidly during post-gastrula development and disappeared completely after emergence. The protein remained at a high level until emergence, after which it decreased quickly and had disappeared from the western blots of instar II larvae cell-free extracts. p26 occurred in the cytoplasm as well as in nuclei, wherein it was sometime found in discrete compartments. p26 had chaperone activity in vivo, as demonstrated by the enhanced thermoresistance of bacteria expressing p26. In conclusion, p26 represents the only known small heat shock/alpha-crystallin protein from a crustacean and it is the only protein in this family thought to function in diapause. p26 may bind to proteins and other macromolecules in encysted Artemia embryos under stress and prevent their irreversible aggregation. This allows spontaneous and/or assisted folding of proteins, permitting rapid resumption of cyst metabolism under favourable growth conditions. | en_US |
