Vacuole Transport; H± ATPase Regulation
Since 1987, Dr. Johnson's research interests have centered on vacuolar membrane proteins in plants. We have characterized two relatively small (27- and 30 kD), abundant integral membrane proteins. The 27-kD protein, known as TIP (tonoplast intrinsic protein), was the first plant protein to be identified as an aquaporin (water channel). It is widespread among higher plants, exists in several isoforms in each species, and the seed-specific form is subject to control by phosphorylation.
TIP is phosphorylated by a Ca2+ dependent, tonoplast-bound protein kinase in seeds. We have localized the phosphorylation site to a single serine residue near the N-terminus. TIP expressed in transgenic tobacco leaf cells is also phosphorylated in vivo and in isolated vacuoles, and site-specific mutagenesis has shown that the same serine residue is utilized in this transgenic context. We are taking advantage of the transgenic vacuole system to probe the regulatory aspects of vacuolar phosphorylation events.
In a joint project with D. Rayle, we are looking into the idea that auxin, the major plant growth hormone, regulates cell enlargement, at least partly, through elevating the level of the plasma membrane proton pump. We are in the process of isolating isoforms of this ATPase expressed in auxin-treated corn coleoptiles, then using unique sequences within each isoform to test auxin-induced up- or down-regulation of the various isoform messages.
Johnson, K.D. and M.J. Chrispeels, Substrate specificities of N-acetylglucosaminyl-, fucosyl-, and xylosyltransferases that modify glycoproteins in the Golgi apparatus of bean cotyledons. Plant Physiol. 84, 1301-1308 (1987).
Faye L., K.D. Johnson, A Sturm, and M.J. Chrispeels, Structure, biosynthesis, and function of asparagine-linked glycans of plant glycoproteins. Physiol. Plant. 75, 309-314 (1989).
Johnson, K.D., E.M. Herman and M.J. Chrispeels, An abundant, highly-conserved tonoplast protein in seeds. Plant Physiol. 91, 1006-1013 (1989).
Lauriere M., C. Lauriere, K.D. Johnson, M.J. Chrispeels, and A. Sturm, Characterization of a xylose-specific antiserum that reacts with the complex asparagine-linked glycans of extracellular and vacuolar glycoproteins. Plant Physiol. 90, 1182-1188 (1989).
Johnson, K.D., H. Hofte and M.J. Chrispeels. An intrinsic tonoplast protein of protein storage vacuoles in seeds is structurally related to a bacterial solute transporter (Glpf). Plant Cell 2, 525-532 (1990).
Pao, G.M., L-F Wu, K.D. Johnson, H. Höfte, M.J. Chrispeels, G. Sweet, N.N. Sandal, and M.H. Saier, Jr., Evolution of the MIP family of integral membrane transport proteins. Molec. Microbiol. 5, 33-37 (1990).
Johnson, K.D. and M.J. Chrispeels, Tonoplast-bound protein kinase phosphorylated tonoplast intriusic protein. Plant Physiol. 100, 1787-1795 (1992).
Maurel, C, H. Höfte, K.D. Johnson and M.J. Chrispeels, Aquaporins: Water channel proteins in the tonoplast of plant cells. Plant Molecular Biology. Molecular-Genetic Analysis of Plant Development and Metabolism. NATO ASI Series: Cell Biology 81, 481-490 (1993).
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