PUB16 gene expression under abiotic stress and their putative role as an ARM repeat protein in Arabidopsis thaliana self-pollination pathway

María Gabriela Acosta, Miguel Ángel Ahumada, Sergio Luis Lassaga, Víctor Hugo Casco

Agricultural Sciences School; National University of Entre Rios; Oro Verde, Argentina.
Microscopy Laboratory Applied to Cell and Molecular Studies, Engineering School; National University of Entre Rios; Oro Verde, Argentina.

DOI: 10.4236/abb.2012.35079                

Abstract

The armadillo repeat super-family proteins (ARM repeat super-family proteins) possess tandem armadillo repeats and have been postulated to play different roles in plant development, morphogenesis, defense, cell death, and signal transduction through hormone signalling. In The Arabidopsis Information Resource (TAIR), we found 113 loci closely related to ARM repeat family proteins. This extensive group of proteins was studied in flowers tissues by western blot using antibodies directed against the most conserved region of the ARM repeat family proteins. The amino acid residues sequences from TAIR were aligned and the resulting phylogenetic tree allows us to inferring their evolutionary relationships. The main finding was the high similarity between the gene product of PUB16 (At5g01830, A. thaliana) and ARC1 (Brassica napus). In order to search a possible role for PUB16 we carried out stress bioassays using hormonal and saline approaches. Gene expression using RT-PCR showed that some of the ARM repeat super-family proteins are expressed both under salt or hormonal stress conditions. Particularly these studies allowed to detect and semi-quantify PUB16 gene expression in normal or stress growth conditions. In this approach it was revealed that, only in presence of GA, the expression of mRNA-PUB16 became evident. To morphologically verify the increasing number of germinated pollen grain in gibberellins treated flowers, we used epi-fluorescence microscopy assay. These results suggest that PUB16 may participate in GA signaling pathway favoring self-pollination.

Keywords

Self-Pollination; ARM Repeat; Gibberellins

Conflicts of Interest

The authors declare no conflicts of interest.

References

[1]	Coates, J.C. (2007) Plant Cell Monographs: Plant Growth Signalling Ed. Bogre and Beemster, Springer, 299-314.
[2]	Samuel, M., Salt, J., Shiu, S. and Goring, D. (2006) Multifunctional arm repeat domains in plants. International Review of Cytology, 253, 1-26.
[3]	Coates, J.C., Laplaze, L. and Haseloff, J. (2006) Armadillo-related proteins promote lateral root development in Arabidopsis. Proceedings of the National Academy of Sciences USA, 103, 1621–1626.
[4]	González-Lamothe, R., Tsitsigiannis, D.I., Ludwig, A.A., Panicot, M., Shirasu, K. and Jones, J.D.G. (2006) The U-box protein CMPG1 is required for efficient activation of defense mechanisms triggered by multiple resistance genes in tobacco and tomato. The Plant Cell, 18, 1067–1083.
[5]	Liu, P., Sherman-Broyles, S. and Nasrallah, M.E. (2007) A cryptic modifier causing transient self-incompatibility in Arabidopsis thaliana. Current Biology, 17, 734–740.
[6]	Yan, J., Wang, J., Li, Q., Hwang, J.R., Patterson, C. and Zhang, H. (2003) AtCHIP, a U-box-containing E3 ubiquitin ligase, plays a critical role in temperature stress tolerance in Arabidopsis. Plant Physiology, 132, 861–869.
[7]	Dong, J., Kim, S.T. and Lord, E.M. (2005) Plantacyanin plays a role in reproduction in Arabidopsis. Plant Physiology, 138,778-789.
[8]	Wheeler, M.J., Franklin-Tong, V.E. and Franklin, F.C.H. (2001) The molecular and genetic basis of pollen–pistil interactions. New Phytologist, 151, 565–584.
[9]	Nasrallah, M.E., Liu, P., Sherman-Broyles, S., Boggs, N.A. and Nasrallah, J.B. (2004) Natural variation in expression of self-Incompatibility in Arabidopsis thaliana: implications for the evolution of selfing. Proceedings of the National Academy of Sciences USA, 101, 16070–16074.
[10]	Sanchez, A.M., Bosch, M., Bots, M., Nieuwland, J., Feron, R. and Mariani, C. (2004) Pistil Factors Controlling Pollination. The Plant Cell, 16, S98-S106.
[11]	Huang, J., Zhao, L., Yang, Q. and Xue ,Y. (2006) AhSSK1, a novel SKP1-like protein that interacts with the Slocus F-box protein SLF. The Plant Journal, 46, 780–793.
[12]	Kusaba, M., Dwyer, K., Hendershot, J., Vrebalov, J., Nasrallah, J.B. and Nasrallah, M.E. (2001) Self-incompatibility in the genus Arabidopsis: characterization of the S locus in the outcrossing A. lyrata and its autogamous relative A. thaliana. The Plant Cell, 13, 627-643.
[13]	Heslop-Harrison, Y. and Shivanna, K.R. (1977) The receptive surface of the angiosperm stigma. Annals of Botany, 41, 1233-1258.
[14]	Dickinson, H. (1995) Dry stigmas, water and self-incompatibility in Brassica. Sexual Plant Reproduction, 8, 1–10.
[15]	Hulskamp, M., Kopczak, S.D., Horejsi, T.F., Kihl, B.K. and Pruitt, R.E. (1995) Identification of genes required for pollen–stigma recognition in Arabidopsis thaliana. The Plant Journal, 8, 703–714.
[16]	Sampson, D.R. (1962) Intergeneric pollen–stigma incompatibility in Cruciferae. Canadian Journal of Genetics and Cytology, 4, 38–49.
[17]	Hiscock, S.J. and Dickinson, H.G. (1993) Unilateral incompatibility within the Brassicaceae: further evidence for the involvement of the self incompatibility (S)-locus. Theoretical and Applied Genetics, 86, 744–753.
[18]	Lelivelt, C.L.C. (1993) Studies of pollen grain germination, pollen-tube growth, micropylar penetration and seed set in intraspeci?c and intergeneric crosses within three Cruciferae species. Euphytica, 67, 185–197.
[19]	Nasrallah, J.B. (2000) Cell–cell signaling in the self-incompatibility response. Current Opinion of Plant Biology, 3, 368–373.
[20]	Hill, J.P. and Lord, E.M. (1987) Dynamics of pollen tube growth in the wild radish, Raphanus raphanistrum (Brassicaceae) 2.Morphology, cytochemistry, and ultrastructure of transmitting tissues, and path of pollen tube growth. American Journal of Botany, 74, 988–997.
[21]	Elleman, C.J., Dickinson, H.G. (1990) The role of the exine coating in pollen–stigma interactions in Brassica oleracea L. New Phytologist, 114, 511–518.
[22]	Elleman, C.J., Franklin-Tong, V. and Dickinson, H.G. (1992) Pollination in species with dry stigmas:the nature of the early stigmatic response and the pathway taken by pollen tubes. New Phytologist, 121, 413–424.
[23]	Kandasamy, M.K., Nasrallah, J.B. and Nasrallah, M.E. (1994) Pollen–pistil Interactions and developmental regulation of pollen tube growth in Arabidopsis. Development, 120, 3405–3418.
[24]	Hulskamp, M., Schneitz, K. and Pruitt, R.E. (1995b) Genetic evidence for along-range activity that directs pollen tube guidance in Arabidopsis. The Plant Cell, 7, 57–64.
[25]	Lennon, K.A., Roy, S., Hepler, P.K. and Lord, E.M. (1998) The structure of the transmitting tissue of Arabidopsis thaliana (L.) and the path of pollen tube growth. Sexual Plant Reproduction, 11, 49–59.
[26]	Tsuchimatsu, T., Suwabe, K., Shimizu-Inatsugi, R., Isokawa, S., Pavlidis, P., Stadler, T,, Suzuki, G., Takayama, S., Watanabe, M. and Shimizu, K. (2010) Evolution of self-compatibility in Arabidopsis by a mutation in the male speci?city gene. Nature, 464, 1342–1346.
[27]	Bateman, A.J. (1955) Self-incompatibility systems in angiosperms. III. Cruciferae. Heredity, 9, 52–68.
[28]	Thompson, K.F. and Taylor, J.P. (1966) Non-linear dominance relationships between Salleles. Heredity, 21, 345–362.
[29]	Tarutani, Y., Shiba, H., Iwano, M., Kakizaki, T., Suzuki, G., Watanabe, M., Isogai, A., and Takayama, S. (2010) Transacting small RNA determines dominance relationships in Brassica self-incompatibility. Nature, 466, 983–986.
[30]	Suzuki, G., Kai, N., Hirose, T., Fukui, K., Nishio, T., Takayama, S., Isogai, A., Watanabe, M. and Hinata, K. (1999) Genomic organization of the S locus: identification and characterization of genes in SLG/SRK region of S9 haplotype of Brassica campestris (syn. rapa). Genetics, 153, 391–400.
[31]	Schopfer, C.R., Nasrallah, M.E. and Nasrallah, J.B. (1999) The male determinant of self-incompatibility in Brassica. Science, 286, 1697–1700.
[32]	Takayama, S., Shiba, H., Iwano, M., Shimosato, H., Che, F.S., Kai, N., Watanabe, M., Suzuki, G., Hinata, K. and Isogai, A. (2000) The pollen determinant of self-incompatibility in Brassica campestris. Proceedings of the National Academy of Sciences USA, 97, 1920–1925.
[33]	Takayama, S., Shimosato, H., Shiba, H., Funato, M., Che, F.S., Watanabe, M., Iwano, M. and Isogai, A. (2001) Direct ligand–receptor complex interaction controls Brassica self-incompatibility. Nature, 413, 534–538.
[34]	Shiba, H., Takayama, S., Iwano, M., Shimosato, H., Funato, M., Nakagawa, T., Che, F., Suzuki, G., Watanabe, M., Hinata, K. and Isogai, A. (2001) A pollen coat protein, SP11/SCR, determines the pollen S-speci?city in the self-incompatibility of Brassica species. Plant Physiology, 125, 2095–2103.
[35]	Stone, S.L., Anderson, E.M., Mullen, R.T. and Goring, D.R. (2003) ARC1 Is an E3 ubiquitin ligase and promotes the ubiquitination of proteins during the rejection of self-incompatible Brassica pollen. The Plant Cell, 15, 885–898.
[36]	Gu, T.S., Mazzurco, M., Sulaman, W., Matias, D.D. and Goring, D.R. (1998) Binding of an arm repeat protein to the kinase domain of the S-locus receptor kinase. Proceedings of the National Academy of Sciences USA, 95, 382–387.
[37]	Samuel, M.A., Mudgil, Y., Salt, J.N., Delmas, F., Ramachandran, S., Chilelli, A. and Goring, D.R. (2008) Interactions between the S-domain receptor kinases and AtPUB-ARM E3 ubiquitin ligase ssuggest a conserved signaling pathway in Arabidopsis. Plant Physiology, 147, 2084–2095.
[38]	Murase, K., Shiba, H., Iwano, M., Che, F.S., Watanabe, M., Isogai, A. and Takayama, S. (2004) A membrane-anchored protein kinase involved in Brassica self-incompatibility signaling. Science, 303,1516–1519.
[39]	Kakita, M., Murase, K., Iwano, M., Matsumoto, T., Watanabe, M., Shiba, H., Isogai, A. and Takayama, S. (2007) Two distinct forms of Mlocus protein kinase localize to the plasma membrane and interact directly with S-locus receptor kinase to transducer self-incompatibility signaling in Brassica rapa. The Plant Cell, 19, 3961–3973.
[40]	Kakita, M., Shimosato, H., Murase, K., Isogai, A., Takayama, S. (2007) Direct interaction between the S-locus receptor kinase and M locus protein kinase involved in Brassica self-incompatibility signaling. Plant Biotechnology, 24, 185–190.
[41]	Riggleman, B., Wieschaus, E. and Schedl, P. (1989) Molecular analysis of the armadillo locus: uniformly distributed transcripts and a protein with novel internal repeats are associated with a Drosophila segment polarity gene. Genes Development, 3, 96-113.
[42]	Mudgil, Y., Shiu, S.H., Stone, S.L., Salt, J.N. and Goring, D.R. (2004) A large complement of the predicted Arabidopsis ARM repeat proteins are members of the U-box E3 ubiquitin ligase family. Plant Physiology, 134, 59–66.
[43]	Acosta, M.G., Langhi, D., Lassaga, S.L. and Casco, V.H. (2010a) Bioinformatics and morphological studies of pollination mechanism as a process of cell-cell adhesion in Arabidopsis thaliana. Agricultural Science Magazine, 14, 3-15.
[44]	Epstein, E. (1972) Mineral nutrition of plants: Principles and perspectives. J. Wiley and Sons, Inc., New York, 68-82.
[45]	Thompson, J.D., Higgins, D.G. and Gibson, T.J. (1994) CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Research, 22, 4673-4680.
[46]	Perriere, G. and Gouy, M. (1996) www-query: An on-line retrieval system for biological sequence banks. Biochimie, 78, 364-369.
[47]	Smyth, D.R., Bowman, J.L. and Meyerowitz, E.M. (1990) Early flower development in Arabidopsis. The Plant Cell, 2, 755-767.
[48]	Bowman, J. (1994) Arabidopsis: An atlas of morphology and development. Springer-Verlag, New York, 258-259.
[49]	Eschrich, W. and Currier, H.B. (1964) Identification of callose by it diachrome and fluorchrome reactions. Stain Technology, 39, 308-309.
[50]	Banzai, T., Hershkovits, G., Katcoff, D.J., Hanagata, N., Dubinsky, Z., and Karube, I. (2002) Identification and characterization of mRNA transcripts differentially expressed in response to high salinity by means of differential display in the mangrove, Bruguiera gymnorrhiza. Plant Science, 162, 499–505.
[51]	Sambrook, D.W. and Russell, J. (2001) Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory Press, 102-114.
[52]	Weigel, D. (2002) Arabidopsis: a laboratory manual. Cold spring harbour Laboratory Press. New York, 102-145.
[53]	Bradford, M.M. (1976) Rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72, 248–254.
[54]	Laemmli, U.K. (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature, 227, 680–685.
[55]	Azevedo, C., Santos-Rosa, M.J. and Shirasu, K. (2001) The U-box protein family in plants. Trends in Plant Science, 6, 354–358.
[56]	Andersen, P., Kragelund, B.B., Olsen, A.N., Larsen, F.H., Chua, N., Poulsen, F.M. and Skriver, K. (2004) Structure and biochemical function of aprototypical Arabidopsis U-box domain. Journal of Biological Chemistry, 279, 40053–4006.
[57]	Wiborg, J., O’Shea, C. and Skriver, K. (2008) Biochemical function of typical and variant Arabidopsis thaliana U-box E3 ubiquitin-protein ligases. Biochem J 413: 447–457
[58]	Thomas SG, Franklin-Tong VE (2004) Self-incompatibility triggers programmed cell death in Papaver pollen. Nature, 429, 305–309.
[59]	Yang, C.W., González-Lamothe, R., Ewan, R.A., Rowland, O., Yoshioka, H., Shenton, M., Ye, H., O’Donnell, E., Jones, J.D.G. and Sadanandom, A. (2006) The E3 ubiquitin ligase activity of Arabidopsis PLANTU-BOX17 and its functional tobacco homolog ACRE276 are required for cell death and defense. The Plant Cell, 18, 1084–1098.
[60]	Itoh, H., Ueguchi-Tanaka, M., Sato, Y., Ashikari, M. and Matsuoka, M. (2002) The gibberellin signaling pathway is regulated by the appearance and disappearance of SLENDER RICE1 in nuclei. The Plant Cell, 14, 57–70.
[61]	Spartz, A.K., Lee, S.H., Wenger, J.P., Gonzalez, N., Itoh, H., Inzé, D., Peer, W.A., Murphy, A.S., Overvoorde, P.J. and Gray, W.M. (2012) The SAUR19 subfamily of SMALL AUXIN UP RNA genes promote cell expansion. The Plant Journal doi: 10.1111/j.1365-313X.2012.04946.x
[62]	Komeda, Y. (2004) Genetic regulation of time to flowering Arabidopsis thaliana Annual Review of Plant Biology, 55, 521–35.
[63]	Yu, H., Ito, T., Wellmer, F. and Meyerowitz, E.M. (2004) Floral homeotic genes are targets of gibberellins signaling in flower Proceedings of the National Academy of Sciences USA, 101, 7827–7832.
[64]	Weiss, D. and Ori, N. (2007) Mechanisms of cross talk between gibberellin and other hormones. Plant Physiology, 144, 1240-1246.
[65]	Acosta, M.G., Ahumada, M.A., Lassaga, S.L. and Casco, V.H. (2010b) Abiotic stress effect on gene expression of ARM repeats proteins in A. thaliana. Book of Abstracts - XXIII Meeting Argentina Plant Physiology, 1, 205.
[66]	Cabrillac, D., Cock, J.M., Dumas, C. and Gaude, T. (2001) The S locus receptor kinase is inhibited by thioredoxins and activated by pollen coat proteins. Nature, 410, 220–223.
[67]	Griffiths, J., Murase, K., Rieu, I., Zentella, R., Zhang, Z.L., Powers, S.J., Gong, F., Phillips, A.L., Hedden, P. and Sun, T.P. (2007) Genetic characterization and functional analysis of the GID1 gibberellin receptors in Arabidopsis. The Plant Cell, 18, 3399-3414.
[68]	Blazquez, M.A., Green, R., Nilsson, O., Sussman, M.R. and Weigel, D. (1998) Gibberellins promote flowering of Arabidopsis by activating the LEAFY promoter. The Plant Cell, 10, 791–800.
[69]	Sun, T.P. and Gubler, F. (2004) Molecular mechanism of gibberellin signaling in plants. Annual Review of Plant Biology, 55, 197–223.
[70]	Ueguchi-Tanaka, M., Ashikari, M., Nakajima, M., Itoh, H., Katoh, E., Kobayashi, M., Chow, T.Y., Hsing, Y.I., Kitano, H. and Yamaguchi, I. (2005) GIBBERELLIN INSENSITIVE DWARF1 encodes a soluble receptor for gibberellin. Nature, 437, 693–698.
[71]	Hartweck, L.M. and Olszewski, N.E. (2006) GIBBERELLIN INSENSITIVE DWARF1 is a gibberellin receptor that illuminates and raises questions about GA signaling. The Plant Cell, 18, 278–282.
[72]	Yee, D. and Goring, D.R. (2009) The diversity of PUB E3 ubiquitin ligases: from upstream activators to downstream target substrates. Journal of Experimental Botany, 60, 1109-1121.
[73]	Cho, S.K., Chung, H.S., Ryu, M.Y., Park, M.J., Lee, M.M., Bahk, Y.Y., Kim, J., Pai, H.S. and Kim, W.T. (2006) Heterologous expression and molecular and cellular characterization of CaPUB1 encoding a hot pepper U-BoxE3 ubiquitin ligase homolog. Plant Physiology, 142, 1664–1682.
[74]	Cho, S.K., Ryu, M.Y., Song, C., Kwak, J.M. and Kim, W.T. (2008) Arabidopsis PUB22 and PUB23 are homologous U-box E3 ubiquitin ligases that play combinatory roles in response to drought stress. The Plant Cell, 20, 1899–1914.
[75]	Shen, G., Yan, J., Pasapula, V., Luo, J., He, C., Clarke, A.K. and Zhang, H. (2007) The chloroplast protease subunit ClpP4 is a substrate of the E3 ligase AtCHIP and plays an important role in chloroplast function. The Plant Journal, 49, 228–237.
[76]	Alonso-Ramírez, A., Rodríguez, D., Reyes, D., Jiménez, J.A., Nicolás, G., López-Climent, M., Gómez-Cadenas, A. and Nicolás, C. (2009) Cross-talk between gibberellins and salicylic acid in early stress responses in Arabidopsis thaliana seeds. Plant Signal Behavior, 4, 750–751.
[77]	Letunic, I. and Bork, P. (2007) Interactive Tree Of Life (iTOL): an on line tool for phylogenetic tree display and annotation. Bioinformatics, 23, 127–128.