PLANT PHYSIOLOGY , Vol 102, Issue 3 843-850, Copyright © 1993 by American Society of Plant Biologists
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MOLECULAR BIOLOGY AND GENE REGULATION |
Characteristics of an Hsp70 Homolog Localized in Higher Plant Chloroplasts That Is Similar to DnaK, the Hsp70 of Prokaryotes
H. Wang, M. Goffreda and T. Leustek
Center for Agricultural Molecular Biology, Rutgers University New Brunswick New Jersey 08903
Members of the 70-kD heat-shock protein (Hsp70) family are important
cellular factors that are thought to mediate protein folding and assembly.
A chloroplast-localized Hsp70 homolog (Chsp70) was recently identified
based on its similarity to DnaK, the Hsp70 homolog of Escherichia coli (D.
Amir-Shapira, T. Leustek, B. Dalie, H. Weissbach, N. Brot [1990] Proc Natl
Acad Sci USA 87: 1749-1752). To learn more about the function of Chsp70, we
purified the protein from Spinacia oleracea chloroplasts by ATP-agarose
affinity chromatography. A single, 75,000-D protein was isolated which
becomes phosphorylated on a threonine residue when incubated with
[[gamma]-32P]ATP and 10 mM Ca2+, a property similar to DnaK. Chloroplast
fractionation and immunoblot analysis showed that Chsp70 is a soluble
stromal protein. Chsp70-specific antiserum was used to clone a partial cDNA
that shows greater homology with Hsp70 from prokaryotes than with
cytoplasmic Hsp70 from eukaryotes. The antiserum and cDNA were used to
study Chsp70 expression. Following heat shock of spinach seedlings at
37[deg] C, Chsp70 synthesis increases 12-fold, the level of Chsp70 mRNA
increases 5-fold, and the level of Chsp70 protein increases less than
2-fold. Chsp70 is constitutively expressed in all spinach tissues,
indicating that it is likely to be localized in all plastid types. The
highest levels occur in seeds, leaves, florets, and seedlings grown in the
light. Lower levels occur in roots, stems, and etiolated seedlings.
