MOLECULAR CHARACTERIZATION OF P26, A CYST-SPECIFIC, SMALL HEAT SHOCK/alfa-CRYSTALLIN PROTEIN FROM ARTEMIA FRANCISCANA
T.H. MacRae, P. Liang
Abstract:
The molecular characteristics of p26, a protein produced in large quantities by encysting Artemia, are reviewed in this paper. Brine shrimp embryos may undergo oviparous development, wherein gastrulae are subject to diapause, cessation of metabolic activity and desiccation. The activated embryos immediately resume growth when rehydrated if conditions are favourable. Mechanisms must therefore exist to preserve the macromolecular components of Artemia exposed to diapause-associated stress. A cyst-specific protein, termed p26, was purified to apparent homogeneity and shown to have molecular chaperone activity in vitro. Chromatography on Sepharose CL-6B columns and centrifugation on sucrose gradients revealed that p26 is a multimer with a molecular mass of approximately 700 kDa. Purified p26 was partially sequenced and degenerative primers suitable for PRC amplification were constructed for use in screening an Artemia cDNA library. One of several cloned cDNAs encoded a protein of 192 animo acid residues which contained an alfa-crystallin domain, the signature sequence of small heat shock/alfa-crystallin proteins. Unusual features of p26 included enrichments of glycine and arginine in its amino terminal variable region, a reduced content of alfa-helix in the alfa-crystallin domain and an extended carboxy terminal tail. Probing of Southern blots indicated a multi-gene family for p26, and two size classes of p26 mRNA occurred on northern blots of poly(A)+ mRNA from encysted gastrulae. Artemia nuclei contained p26, as did the cytoplasm from cysts. In summary, a small heat shock/alfa-crystallin protein of crustacean origin has been described for the first time. Moreover, the data suggest that molecular chaperones protect cells during diapause, thus providing additional insight into the molecular events associated with this process.
(Department of Biology, Dalhousie University, Halifax, N.S., B3H 4J1 Canada)