Abstract
In the maize endosperm, the Opaque2 (O2) basic leucine zipper transcriptional activator regulates the expression of a subset of the zein seed storage protein gene family. Immunodetection of wild-type or mutant O2 polypeptides fractionated by SDS-PAGE resolved a closely spaced doublet migrating in the 68- to 72-kD range, whereas by using isoelectric focusing, seven to nine isoforms were detected for each allele. Phosphatase treatment simplified the protein patterns to a single band corresponding to the nonphosphorylated component. In vivo and in vitro labeling confirmed that O2 can be phosphorylated. In protein gel blots probed with DNA, only the nonphosphorylated and hypophosphorylated O2 polypeptides were able to bind an oligonucleotide containing the O2 binding sequence. Upon in situ dephosphorylation of the focused isoforms by phosphatase treatment of the isoelectric focusing filter, the hyperphosphorylated forms acquired DNA binding activity. The ratio among the various isoforms remained constant throughout the developmental stages of endosperm growth but changed from daytime to nighttime, with a significant increase of the hyperphosphorylated forms during the night period. These results indicate that O2 exists in vivo as a pool of differently phosphorylated polypeptides and demonstrate that O2 DNA binding activity is modulated by a phosphorylation/dephosphorylation mechanism that appears to be influenced by environmental conditions.
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- Alderson A., Sabelli P. A., Dickinson J. R., Cole D., Richardson M., Kreis M., Shewry P. R., Halford N. G. Complementation of snf1, a mutation affecting global regulation of carbon metabolism in yeast, by a plant protein kinase cDNA. Proc Natl Acad Sci U S A. 1991 Oct 1;88(19):8602–8605. doi: 10.1073/pnas.88.19.8602. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bairoch A. PROSITE: a dictionary of sites and patterns in proteins. Nucleic Acids Res. 1991 Apr 25;19 (Suppl):2241–2245. doi: 10.1093/nar/19.suppl.2241. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bernard L., Ciceri P., Viotti A. Molecular analysis of wild-type and mutant alleles at the Opaque-2 regulatory locus of maize reveals different mutations and types of O2 products. Plant Mol Biol. 1994 Mar;24(6):949–959. doi: 10.1007/BF00014448. [DOI] [PubMed] [Google Scholar]
- Bjellqvist B., Hughes G. J., Pasquali C., Paquet N., Ravier F., Sanchez J. C., Frutiger S., Hochstrasser D. The focusing positions of polypeptides in immobilized pH gradients can be predicted from their amino acid sequences. Electrophoresis. 1993 Oct;14(10):1023–1031. doi: 10.1002/elps.11501401163. [DOI] [PubMed] [Google Scholar]
- Datta N., Cashmore A. R. Binding of a pea nuclear protein to promoters of certain photoregulated genes is modulated by phosphorylation. Plant Cell. 1989 Nov;1(11):1069–1077. doi: 10.1105/tpc.1.11.1069. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Despres C., Subramaniam R., Matton D. P., Brisson N. The Activation of the Potato PR-10a Gene Requires the Phosphorylation of the Nuclear Factor PBF-1. Plant Cell. 1995 May;7(5):589–598. doi: 10.1105/tpc.7.5.589. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Dietrich A., Mayer J. E., Hahlbrock K. Fungal elicitor triggers rapid, transient, and specific protein phosphorylation in parsley cell suspension cultures. J Biol Chem. 1990 Apr 15;265(11):6360–6368. [PubMed] [Google Scholar]
- Driouich A., Faye L., Staehelin L. A. The plant Golgi apparatus: a factory for complex polysaccharides and glycoproteins. Trends Biochem Sci. 1993 Jun;18(6):210–214. doi: 10.1016/0968-0004(93)90191-o. [DOI] [PubMed] [Google Scholar]
- Edery I., Zwiebel L. J., Dembinska M. E., Rosbash M. Temporal phosphorylation of the Drosophila period protein. Proc Natl Acad Sci U S A. 1994 Mar 15;91(6):2260–2264. doi: 10.1073/pnas.91.6.2260. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gallusci P., Salamini F., Thompson R. D. Differences in cell type-specific expression of the gene Opaque 2 in maize and transgenic tobacco. Mol Gen Genet. 1994 Aug 15;244(4):391–400. doi: 10.1007/BF00286691. [DOI] [PubMed] [Google Scholar]
- Grasser K. D., Maier U. G., Feix G. A nuclear casein type II kinase from maize endosperm phosphorylating HMG proteins. Biochem Biophys Res Commun. 1989 Jul 14;162(1):456–463. doi: 10.1016/0006-291x(89)92019-6. [DOI] [PubMed] [Google Scholar]
- Harrison M. J., Lawton M. A., Lamb C. J., Dixon R. A. Characterization of a nuclear protein that binds to three elements within the silencer region of a bean chalcone synthase gene promoter. Proc Natl Acad Sci U S A. 1991 Mar 15;88(6):2515–2519. doi: 10.1073/pnas.88.6.2515. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Harter K., Kircher S., Frohnmeyer H., Krenz M., Nagy F., Schäfer E. Light-regulated modification and nuclear translocation of cytosolic G-box binding factors in parsley. Plant Cell. 1994 Apr;6(4):545–559. doi: 10.1105/tpc.6.4.545. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hunter T., Karin M. The regulation of transcription by phosphorylation. Cell. 1992 Aug 7;70(3):375–387. doi: 10.1016/0092-8674(92)90162-6. [DOI] [PubMed] [Google Scholar]
- Karin M., Hunter T. Transcriptional control by protein phosphorylation: signal transmission from the cell surface to the nucleus. Curr Biol. 1995 Jul 1;5(7):747–757. doi: 10.1016/s0960-9822(95)00151-5. [DOI] [PubMed] [Google Scholar]
- Klimczak L. J., Collinge M. A., Farini D., Giuliano G., Walker J. C., Cashmore A. R. Reconstitution of Arabidopsis casein kinase II from recombinant subunits and phosphorylation of transcription factor GBF1. Plant Cell. 1995 Jan;7(1):105–115. doi: 10.1105/tpc.7.1.105. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Klimczak L. J., Schindler U., Cashmore A. R. DNA binding activity of the Arabidopsis G-box binding factor GBF1 is stimulated by phosphorylation by casein kinase II from broccoli. Plant Cell. 1992 Jan;4(1):87–98. doi: 10.1105/tpc.4.1.87. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kodrzycki R., Boston R. S., Larkins B. A. The opaque-2 mutation of maize differentially reduces zein gene transcription. Plant Cell. 1989 Jan;1(1):105–114. doi: 10.1105/tpc.1.1.105. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Krebs E. G., Eisenman R. N., Kuenzel E. A., Litchfield D. W., Lozeman F. J., Lüscher B., Sommercorn J. Casein kinase II as a potentially important enzyme concerned with signal transduction. Cold Spring Harb Symp Quant Biol. 1988;53(Pt 1):77–84. doi: 10.1101/sqb.1988.053.01.012. [DOI] [PubMed] [Google Scholar]
- Ma H. Protein phosphorylation in plants: enzymes, substrates and regulators. Trends Genet. 1993 Jul;9(7):228–230. doi: 10.1016/0168-9525(93)90075-s. [DOI] [PubMed] [Google Scholar]
- Maddaloni M., Di Fonzo N., Hartings H., Lazzaroni N., Salamini F., Thompson R., Motto M. The sequence of the zein regulatory gene opaque-2 (O2) of Zea Mays. Nucleic Acids Res. 1989 Sep 25;17(18):7532–7532. doi: 10.1093/nar/17.18.7532. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Pulverer B. J., Kyriakis J. M., Avruch J., Nikolakaki E., Woodgett J. R. Phosphorylation of c-jun mediated by MAP kinases. Nature. 1991 Oct 17;353(6345):670–674. doi: 10.1038/353670a0. [DOI] [PubMed] [Google Scholar]
- Pysh L. D., Aukerman M. J., Schmidt R. J. OHP1: a maize basic domain/leucine zipper protein that interacts with opaque2. Plant Cell. 1993 Feb;5(2):227–236. doi: 10.1105/tpc.5.2.227. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rihs H. P., Jans D. A., Fan H., Peters R. The rate of nuclear cytoplasmic protein transport is determined by the casein kinase II site flanking the nuclear localization sequence of the SV40 T-antigen. EMBO J. 1991 Mar;10(3):633–639. doi: 10.1002/j.1460-2075.1991.tb07991.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Salamini F. Controlling elements at the opaque-2 locus of maize: their involvement in the origin of spontaneous mutation. Cold Spring Harb Symp Quant Biol. 1981;45(Pt 2):467–476. doi: 10.1101/sqb.1981.045.01.062. [DOI] [PubMed] [Google Scholar]
- Sarokin L. P., Chua N. H. Binding sites for two novel phosphoproteins, 3AF5 and 3AF3, are required for rbcS-3A expression. Plant Cell. 1992 Apr;4(4):473–483. doi: 10.1105/tpc.4.4.473. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Schmidt R. J., Ketudat M., Aukerman M. J., Hoschek G. Opaque-2 is a transcriptional activator that recognizes a specific target site in 22-kD zein genes. Plant Cell. 1992 Jun;4(6):689–700. doi: 10.1105/tpc.4.6.689. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sorger P. K., Pelham H. R. Yeast heat shock factor is an essential DNA-binding protein that exhibits temperature-dependent phosphorylation. Cell. 1988 Sep 9;54(6):855–864. doi: 10.1016/s0092-8674(88)91219-6. [DOI] [PubMed] [Google Scholar]
- Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W. Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 1990;185:60–89. doi: 10.1016/0076-6879(90)85008-c. [DOI] [PubMed] [Google Scholar]
- Sun L., Doxsee R. A., Harel E., Tobin E. M. CA-1, a novel phosphoprotein, interacts with the promoter of the cab140 gene in Arabidopsis and is undetectable in det1 mutant seedlings. Plant Cell. 1993 Jan;5(1):109–121. doi: 10.1105/tpc.5.1.109. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Tjaden G., Coruzzi G. M. A novel AT-rich DNA binding protein that combines an HMG I-like DNA binding domain with a putative transcription domain. Plant Cell. 1994 Jan;6(1):107–118. doi: 10.1105/tpc.6.1.107. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Unger E., Parsons R. L., Schmidt R. J., Bowen B., Roth B. A. Dominant Negative Mutants of Opaque2 Suppress Transactivation of a 22-kD Zein Promoter by Opaque2 in Maize Endosperm Cells. Plant Cell. 1993 Aug;5(8):831–841. doi: 10.1105/tpc.5.8.831. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Varagona M. J., Raikhel N. V. The basic domain in the bZIP regulatory protein Opaque2 serves two independent functions: DNA binding and nuclear localization. Plant J. 1994 Feb;5(2):207–214. doi: 10.1046/j.1365-313x.1994.05020207.x. [DOI] [PubMed] [Google Scholar]
- Varagona M. J., Schmidt R. J., Raikhel N. V. Monocot regulatory protein Opaque-2 is localized in the nucleus of maize endosperm and transformed tobacco plants. Plant Cell. 1991 Feb;3(2):105–113. doi: 10.1105/tpc.3.2.105. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Varagona M. J., Schmidt R. J., Raikhel N. V. Nuclear localization signal(s) required for nuclear targeting of the maize regulatory protein Opaque-2. Plant Cell. 1992 Oct;4(10):1213–1227. doi: 10.1105/tpc.4.10.1213. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Woods A., Munday M. R., Scott J., Yang X., Carlson M., Carling D. Yeast SNF1 is functionally related to mammalian AMP-activated protein kinase and regulates acetyl-CoA carboxylase in vivo. J Biol Chem. 1994 Jul 29;269(30):19509–19515. [PubMed] [Google Scholar]
- Yu L. M., Lamb C. J., Dixon R. A. Purification and biochemical characterization of proteins which bind to the H-box cis-element implicated in transcriptional activation of plant defense genes. Plant J. 1993 Jun;3(6):805–816. doi: 10.1111/j.1365-313x.1993.00805.x. [DOI] [PubMed] [Google Scholar]
- Zhang S., Jin C. D., Roux S. J. Casein Kinase II-Type Protein Kinase from Pea Cytoplasm and Its Inactivation by Alkaline Phosphatase in Vitro. Plant Physiol. 1993 Nov;103(3):955–962. doi: 10.1104/pp.103.3.955. [DOI] [PMC free article] [PubMed] [Google Scholar]