Bonini, Nancy M. and Nelson, David L. (1990) Phosphoproteins associated with cyclic nucleotide stimulation of ciliary motility in Paramecium. Journal of Cell Science, 95 (2). pp. 219-230. ISSN 0021-9533 http://resolver.caltech.edu/CaltechAUTHORS:20120424-133701024
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Permeabilized, MgATP-reactivated cells of Paramecium (models) respond to cyclic AMP and cyclic GMP by increasing forward swimming speed. In association with the motile response, cyclic AMP and 8-bromo-cyclic GMP (8-Br-cyclic GMP) stimulated protein phosphorylation. Cyclic AMP addition to permeabilized cells reproducibly stimulated the phosphorylation of 10 proteins, ranging in molecular weight from 15 to 110K (K = 10^3 M_r). 8-Br-cyclic GMP, which selectively activates the cyclic GMP-dependent protein kinase of Paramecium, stimulated the phosphorylation of a subset of the proteins phosphorylated by cyclic AMP. Ca^(2+) addition caused backward swimming and stimulated the phosphorylation of four substrates, including a 25K target that may also be phosphorylated in response to cyclic nucleotide addition. Ba^(2+) and Sr^(2+) also induced backward swimming, but did not cause detectable phosphorylation. To identify ciliary targets of cyclic nucleotide-dependent protein kinase activity, permeabilized cells were deciliated following reactivation of motility with Mg-[y-^(32)P]ATP in the presence or absence of cyclic nucleotide. Soluble proteins of the deciliation supernatant were enriched in 15 cyclic AMP-stimulated phosphoproteins, ranging in molecular weight from 15 to 95K. Most of the ciliary substrates were axonemal and could be released by high salt solution. A 29K protein that copurified in sucrose gradients with the 22S dynein, and a high molecular weight protein (greater than 300K) in the 19 S region were phosphorylated when cyclic AMP was added to permeabilized, motile cells. These data suggest that regulation of ciliary motility by cyclic AMP may include phosphorylation of dynein-associated proteins.
|Additional Information:||© 1990 by Company of Biologists. Received 5 December 1988. Accepted, in revised form, 18 October 1989. We thank C. Norris, T. C. Evans, P. A. Mason, M. Hochstrasser, L. A. P. Miglietta and J. B. Peterson for critique of the manuscript; B. C. Soltvedt for excellent editorial assistance; Dr P. Ludden for the use of his scanning densitometer; and M. Hochstrasser and R. E. Gundersen for antibodies against the cyclic AMP- and Ca^(2+)-dependent protein kinases, respectively. This research was supported by NIH grants GM34906, GM32514 and by a grant from the Graduate School of the University of Wisconsin-Madison.|
|Subject Keywords:||Paramecium, cyclic AMP, cyclic GMP, Ca2+, dynein, cilia, Triton-permeabilized cells, swimming speed, ciliary reversal, protein phosphorylation.|
|Official Citation:||N.M. Bonini and D.L. Nelson Phosphoproteins associated with cyclic nucleotide stimulation of ciliary motility in Paramecium J Cell Sci 1990 95:219-230|
|Usage Policy:||No commercial reproduction, distribution, display or performance rights in this work are provided.|
|Deposited By:||Ruth Sustaita|
|Deposited On:||24 Apr 2012 21:22|
|Last Modified:||26 Dec 2012 15:07|
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