A small heat shock protein from Artemia franciscana is phosphorylated at serine 50.
Encysted embryos of Artemia franciscana are exceptionally resistant to stress and an important part of this tolerance involves p26, a small heat shock protein which functions as a molecular chaperone. Cloning and sequencing of randomly selected p26 cDNAs produced by RT-PCR with poly(A)(+) mRNA from encysted embryos as template yielded 10 clones encoding identical polypeptides. The noncoding nucleotide sequences extending from the termination codon to the poly(A) tail of each clone were also identical. These data indicated a single p26 gene is expressed during embryo development. However, two-dimensional gel electrophoresis showed that purified p26 consisted of four isoforms, providing evidence for posttranslational modification of the protein, a possibility supported by mass spectrometry and immunoprobing of Western blots. The major isoform observed in two-dimensional gels, termed a, is the primary gene product, whereas isoform c is phosphorylated at serine 50, a residue located in a protein kinase C reactive site. Isoforms b and d were generated posttranslationally, but by unknown processes. The results represent the first description of posttranslationally modified small heat shock proteins in crustaceans and they expand the phylogenetic range of organisms that possess phosphorylated isoforms of these proteins. At least two small heat shock proteins from other organisms contain serine residues equivalent in position to serine 50 of p26, but neither is phosphorylated.
Qiu
Zhijun
Z
Department of Biology, Dalhousie University, Halifax, NS, Canada B3H 4J1.
Viner
Rosa I
RI
MacRae
Thomas H
TH
Willsie
Julia K
JK
Clegg
James S
JS
D000595:Amino Acid Sequence; D000818:Animals; D001156:Artemia / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D015180:Electrophoresis, Gel, Two-Dimensional; D006360:Heat-Shock Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D010766:Phosphorylation; D010768:Phosphoserine / Q000378:metabolism*; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D012694:Serine / Q000737:chemistry* / Q000378:metabolism*; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D018076:DNA, Complementary; D006360:Heat-Shock Proteins; D020033:Protein Isoforms; D010768:Phosphoserine; D012694:Serine