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Molecular cloning and characterization of RGA1 encoding a G protein α subunit from rice (Oryza sativa L. IR-36)

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Abstract

A cDNA clone, RGA1, was isolated by using a GPA1 cDNA clone of Arabidopsis thaliana G protein α subunit as a probe from a rice (Oryza sativa L. IR-36) seedling cDNA library prepared from roots and leaves. Sequence analysis of genomic clone reveals that the RGA1 gene has 14 exons and 13 introns, and encodes a polypeptide of 380 amino acid residues with a calculated molecular weight of 44.5 kDa. The encoded protein exhibits a considerable degree of amino acid sequence similarity to all the other known G protein α subunits. A putative TATA sequence (ATATGA), a potential CAAT box sequence (AGCAATAC), and a cis-acting element, CCACGTGG (ABRE), known to be involved in ABA induction are found in the promoter region. The RGA1 protein contains all the consensus regions of G protein α subunits except the cysteine residue near the C-terminus for ADP-ribosylation by pertussis toxin. The RGA1 polypeptide expressed in Escherichia coli was, however, ADP-ribosylated by 10 μM [adenylate-32P] NAD and activated cholera toxin. Southern analysis indicates that there are no other genes similar to the RGA1 gene in the rice genome. Northern analysis reveals that the RGA1 mRNA is 1.85 kb long and expressed in vegetative tissues, including leaves and roots, and that its expression is regulated by light.

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References

  1. Anuntalabhochai S, Terryn N, VanMontagu M, Inze D: Molecular characterization of an Arabidopsis thaliana cDNA encoding a small GTP-binding protein, Rhal. Plant J 1: 167–174 (1991).

    Google Scholar 

  2. Balch WE: Small GTP-binding proteins in vesicular transport. Trends Biochem Sci 15: 473–477 (1990).

    Article  PubMed  Google Scholar 

  3. Dallmann G, Sticher H, Marshallsay C, Nagy F: Molecular characterization of tobacco cDNAs encoding two small GTP-binding proteins. Plant Mol Biol 19: 847–857 (1992).

    PubMed  Google Scholar 

  4. Ebert PR, Ha SB, An G: Identification of an essential upstream element in the nopaline synthetase promoter by stable and transient assays. Proc Natl Acad Sci USA 84: 5745–5749 (1987).

    Google Scholar 

  5. Fong HKW, Yoshimoto KK, Eversole-Cire P, Simon MI: Identification of a GTP-binding protein α subunit that lacks an apparant ADP-ribosylation site for pertussis toxin. Proc Natl Acad Sci USA 85: 3066–3070 (1988).

    PubMed  Google Scholar 

  6. Freissmuth M, Casey PJ, Gilman AG: G proteins control diverse pathways of transmembrane signalling. FASEB J 3: 2125–2131 (1989).

    PubMed  Google Scholar 

  7. George WF, Michael BY, Himan S: α-tubulin influences nucleotide binding to β-tubulin: an assay using picomoles of unpurified protein. Proc Natl Acad Sci USA 87: 5041–5045 (1990).

    PubMed  Google Scholar 

  8. Gilman AG: G proteins: transducers of receptor-generated signals. Annu Rev Biochem 56: 615–649 (1987).

    PubMed  Google Scholar 

  9. Grand RJA, Owen D: The biochemistry of ras p21. Biochem J 279: 609–631 (1991).

    PubMed  Google Scholar 

  10. Graziano MP, Casey PJ, Gilman AG: Expression of cDNA for G protein in E. coli. J Biol Chem 262: 11375–11381 (1987).

    PubMed  Google Scholar 

  11. Gunther N, Chri B, Chua NH: Calcium/calmodulin-dependent and-independent phytochrome signal transduction pathways. Cell 73: 937–952 (1993).

    PubMed  Google Scholar 

  12. Hall A: The cellular functions of small GTP-binding proteins. Science 249: 635–640 (1990).

    PubMed  Google Scholar 

  13. Itoh H, Kozasa T, Nagata S, Nakamura S, Katada T, Ui M, Iwai S, Ohtsuka E, Kawasaki K, Suzuki J, Kaziro Y: Molecular cloning and sequence determination of cDNAs for α subunits of the guanine nucleotide-binding proteins, Gs, Gi, and Go from rat brain. Proc Natl Acad Sci USA 83: 3376–3780 (1986).

    PubMed  Google Scholar 

  14. Kaziro Y, Itoh H, Kozasa T, Nakafuka M, Satoh T: Structure and function of signal-transducing GTP-binding proteins. Annu Rev Biochem 60: 349–400 (1991).

    Article  PubMed  Google Scholar 

  15. Lochrie MA, Simon MI: G protein multiplicity in eukaryotic signal transduction system. Biochemistry 27: 4957–4965 (1988).

    PubMed  Google Scholar 

  16. Lowy DR, Willumsen BM: New clue RAS lipid glue. Nature 341: 384–385 (1989).

    Article  PubMed  Google Scholar 

  17. Luetcke HA, Chow KC, Michel FS, Moss KA, Kern HF, Scheele GA: Selection of AUG initiation codons differs in plant and animals. EMBO J 6: 43–48 (1987).

    PubMed  Google Scholar 

  18. Ma H, Yanofsky MF, Huang H: Isolation and sequence analysis of TAG1 cDNAs encoding a tomato G protein α subunit. Gene 107: 189–195 (1991).

    Article  PubMed  Google Scholar 

  19. Ma H, Yanofsky MF, Meyerowitz EM: Molecular cloning and characterization of GPA1, a G protein α subunit gene from Arabidopsis thaliana. Proc Natl Acad Sci USA 87: 3821–3825 (1990).

    PubMed  Google Scholar 

  20. McKenzie FR: Basic techniques to study G-protein function. In: Milligan G (ed) Signal Transduction: A practical Approach p. 42. IRL Press, Oxford (1992).

    Google Scholar 

  21. McKenzie FR: Basic techniques to study G-protein function. In: Milligan G (ed) Signal Transduction: A Practical Approach, pp. 46–47. IRL press, Oxford (1992).

    Google Scholar 

  22. Medynski DC, Sullivan K, Smith D, VanDop C, Chang FH, Fung BKK, Seeburg PH, Bourne HR: Amino acid sequence of the α subunit of transducin deduced from the cDNA sequence. Proc Natl Acad Sci USA 82: 4311–4315 (1985).

    PubMed  Google Scholar 

  23. Murray MG, Thompson WF: Rapid isolation of high molecular weight plant DNA. Nucl Acids Res 8: 4321–4325 (1980).

    PubMed  Google Scholar 

  24. Nakafuka M, Itoh H, Nakamura S, Kaziro Y: Occurrence in Saccharomyces cerevisiae of a gene homologous to the cDNA coding for the α subunit of mammalian G proteins. Proc Natl Acad Sci USA 84: 2140–2144 (1987).

    PubMed  Google Scholar 

  25. Neer EJ, Clapham DE: Roles of G protein subunits in transmembrane signalling. Nature 333: 129–134 (1988).

    Article  PubMed  Google Scholar 

  26. Nukata T, Tanabe T, Takahashi H, Noda M, Haga K, Haga T, Ichiyama A, Kangawa K, Hiranaga M, Matsuo H, Numa S: Primary structure of the α-subunit of bovine adenylate-cyclase-inhibiting G-protein deduced from the cDNA sequence. FEBS Lett 197: 305–310 (1986).

    Article  PubMed  Google Scholar 

  27. Palme K, Diefenthal T, Vingron M, Sander C, Schell J: Molecular cloning and structural analysis of genes from Zea mays (L.) coding for members of the ras-related ypt family. Proc Natl Acad Sci USA 89: 787–791 (1992).

    PubMed  Google Scholar 

  28. Philip SH: An efficient method to purify active eukaryotic proteins from the inclusion bodies in Escherichia coli. Biotechniques 11 No 6 (1991).

    Google Scholar 

  29. Provost NM, Somers DE, Hurley JB: A Drosophila melanogaster G protein α subunit gene is expressed primarily in embryo and pupae. J Biol Chem 263: 12070–12076 (1988).

    PubMed  Google Scholar 

  30. Pugh BF, Tjian R: Transcription from a TATA-less promoter requires a multisubunit TFIID complex. Genes Devel 5: 1935–1945 (1991).

    PubMed  Google Scholar 

  31. Pupillo M, Kumagai A, Pitt GS, Firtel RA, Devreotes PN: Multiple α subunits of guanine nucleotide-binding proteins in Dictyostelium. Proc Natl Acad Sci USA 86: 4892–4896 (1989).

    PubMed  Google Scholar 

  32. Sano H, Youssefian S: A novel ras-related rgpl gene encoding a GTP-binding protein has reduced expression in 5′-azacytidine-induced dwarf rice. Mol Gen Genet 228: 227–237 (1991).

    Article  PubMed  Google Scholar 

  33. Schleifer LS, Kahn RA, Hanski E, Northup JK, Sternweis PC, Gilman AG: Requirements for cholera toxin-dependent ADP-ribosylation of the purified regulatory component of adenylate cyclase. J Biol Chem 257: 20–23 (1982).

    PubMed  Google Scholar 

  34. Shirley BW, Ham DP, Senecoff JF, Berry-Lowe SL, Zurfluh LL, Shah DM, Meagher RB. Comparison of the expression of two highly homologous members of soybean ribulose-1,5-bisphosphate carboxylase small subunit gene family. Plant Mol Biol 14: 909–925 (1990).

    PubMed  Google Scholar 

  35. Terryn N, Anuntalabhochai S, VanMontagu M, Inze D: Analysis of a Nicotiana plumbaginifolia cDNA encoding a novel small GTP-binding protein. FEBS Lett 299: 287–290 (1992).

    Article  PubMed  Google Scholar 

  36. Towler DA, Adams SP, Eubanks SR, Towery DS, Towery DS, Jackson-Machelski E, Glaser L, Gordon JI: Myristoyl CoA: Protein N-myristoyltransferase activities from rat liver and yeast posses overlapping yet distinct peptide substrate specificities. J Biol Chem 263: 1784–1790 (1988).

    PubMed  Google Scholar 

  37. Valencia A, Chardin P, Wittighofer A, Sander C: The ras protein family: evolutionary tree and role of conserved amino acids. Biochemistry 30: 4367–4648 (1991).

    PubMed  Google Scholar 

  38. VanDop C, Yamanaka G, Steinberg F, Sekura RD, Manclark CR, Stryer L, Bourne HR: ADP-ribosylation of transducin by pertussis toxin blocks the light-stimulated hydrolysis of GTP and cGMP in retinal photoreceptors. J Biol Chem 259: 23–26 (1984).

    PubMed  Google Scholar 

  39. Verwoerd TC, Dekker BMM, Hoekema A: A small scale procedure for the rapid isolation of plant RNAs. Nucl Acids Res 17: 2362 (1989).

    PubMed  Google Scholar 

  40. Zhou Q, Lieberman PM, Boyer TG, Berk AJ: Holo-TFIID supports transcriptional stimulation by diverse activators and from a TATA-less promoter. Genes Devel 6: 1964–1974 (1992).

    PubMed  Google Scholar 

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Authors and Affiliations

  1. Plant Molecular Biology and Biotechnology Research Center, Gyeongsang National University, 660-701, Chinju, Korea

    Hak-Soo Seo, Ho-Yeon Kim, Jin-Yong Jeong, Sang-Yeol Lee, Moo-Je Cho & Jeong-Dong Bahk

  2. Department of Biochemistry, Gyeongsang National University, 660-701, Chinju, Korea

    Ho-Yeon Kim, Jin-Yong Jeong, Sang-Yeol Lee, Moo-Je Cho & Jeong-Dong Bahk

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  1. Hak-Soo Seo
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Seo, HS., Kim, HY., Jeong, JY. et al. Molecular cloning and characterization of RGA1 encoding a G protein α subunit from rice (Oryza sativa L. IR-36). Plant Mol Biol 27, 1119–1131 (1995). https://doi.org/10.1007/BF00020885

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  • DOI: https://doi.org/10.1007/BF00020885

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