First published online February 24, 2002; 10.1104/pp.010770
Plant Physiol, March 2002, Vol. 128, pp. 1008-1021
An Arabidopsis Calcium-Dependent Protein Kinase Is Associated
with the Endoplasmic Reticulum1
Sheen X.
Lu and
Estelle M.
Hrabak*
Department of Plant Biology, University of New Hampshire, 46 College Road, Durham, New Hampshire 03824
Arabidopsis contains 34 genes that are predicted to encode
calcium-dependent protein kinases (CDPKs). CDPK enzymatic activity previously has been detected in many locations in plant cells, including the cytosol, the cytoskeleton, and the membrane fraction. However, little is known about the subcellular locations of individual CDPKs or the mechanisms involved in targeting them to those locations. We investigated the subcellular location of one Arabidopsis CDPK, AtCPK2, in detail. Membrane-associated AtCPK2 did not partition with
the plasma membrane in a two-phase system. Sucrose gradient fractionation of microsomes demonstrated that AtCPK2 was associated with the endoplasmic reticulum (ER). AtCPK2 does not contain
transmembrane domains or known ER-targeting signals, but does have
predicted amino-terminal acylation sites. AtCPK2 was myristoylated in a cell-free extract and myristoylation was prevented by converting the glycine at the proposed site of myristate attachment to alanine (G2A). In plants, the G2A mutation decreased AtCPK2 membrane
association by approximately 50%. A recombinant protein, consisting of
the first 10 amino acids of AtCPK2 fused to the amino-terminus of  -glucuronidase, was also targeted to the ER, indicating that the
amino terminus of AtCPK2 can specify ER localization of a soluble
protein. These results indicate that AtCPK2 is localized to the ER,
that myristoylation is likely to be involved in the membrane
association of AtCPK2, and that the amino terminal region of AtCPK2 is
sufficient for correct membrane targeting.
1
This work was supported by the U.S. Department
of Agriculture-National Research Initiative (grant no. 9801263 to
E.M.H.). This is paper no. 2,073 of the New Hampshire Agriculture
Experiment Station.
*
Corresponding author; e-mail emhrabak{at}cisunix.unh.edu; fax
603-862-3784.
© 2002 American Society of Plant Physiologists
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