Identification of the S6 kinase activity stimulated in quiescent brine shrimp embryos upon entry to preemergence development as p70 ribosomal protein S6 kinase: Isolation of Artemia franciscana p70(S6k) cDNA

J. Santiago, T.W. Sturgill-2001

Biochemistry and Cell Biology, 79(2): 141-152  (from Current Contents)

Abstract:

We previously demonstrated that a protein kinase responsible for phosphorylating 40S ribosomal subunits is activated in quiescent Artemia franciscana embryos within 15 min of restoration of normal tonicity and incubation at 30 °C. Here, we identify the activated S6 kinase as A. franciscana p70 ribosomal S6 kinase (p70(S6k)) subsequent to the isolation of an Artemia p70(S6k) cDNA. The protein conceptually translated from cDNA has 70% similarity and 64% identity to both Drosophila melanogaster and human p70(S6k). Southern blot analysis is consistent with presence of a single p70(S6k) gene. Two transcripts of 5.4 and 2.7 kb were found. Abundance of both mRNAs increased dramatically around 4 h of preemergence development, and exhibited different steady-state level variation thereafter. Stimulated S6 kinase activity, partially purified by Superose 6 chromatography, correlated best with the slowest migrating, similar to 65 kDa, form detected by Western analysis using a specific polyclonal antibody made to a peptide from the predicted p70(S6k) NH2-terminus. Furthermore, the A. franciscana p70(S6k) was immunoprecipitated with the same antibody, showing in parallel an S6 kinase activity similar to peak profiles. We conclude that the stimulated S6 kinase activity is that of an ortholog of human p70(S6k) that may be involved in the regulation of protein synthesis during preemergence development in A. franciscana species.

(Univ Virginia, Hlth Sci Ctr, Box 577, Charlottesville, VA 22908, USA, e-mail of T.W. Sturgill: tws7w@Virginia.edu)

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