AACA Element Negatively Regulates Expression of Protein Phosphatase 2C (PP2C) like Promoter in Arabidopsis thaliana

Purva Bhalothia, Anshu Alok, Sandhya Mehrotra, Rajesh Mehrotra

Department of Biological Sciences, Birla Institute of Technology & Sciences (BITS), Pilani, India.
Department of Biological Sciences, Birla Institute of Technology & Sciences (BITS), Pilani, India..
National Agri-Food Biotechnology Institute (NABI), Department of Biotechnology, Government of India, Mohali, India.

DOI: 10.4236/ajps.2013.43071                

Abstract

Genome wide analysis of Arabidopsis thaliana reveals a unique genetic arrangement of ACGT motif in the protein phosphatase 2C (PP2C) like promoter (accession number AT5G59220). In the present study, full length, 900 base pair, 500 base pair, 400 base pair and NRM deletion variants of PP2C like promoter were constructed to investigate the activity of PP2C like promoter in the presence of inducers like abscisic acid, jasmonic acid and salicylic acid. The PP2C promoter has three ACGT elements in close vicinity. They are so positioned that they are separated by a spacer of 30 base pair and 5 base pair respectively from each other. The study has shown that ACGT_N30ACGT genetic architecture is essential for the promoter to be induced in response to abscisic acid. The synergistic and antagonistic effects of cis elements were observed. AACA is a positive regulatory element in endosperm and is known to act as negative regulatory element in other tissues. In this study AACA, have been found to negatively regulate the expression of reporter gene EGFP in both induced and under uninduced conditions.

Keywords

Abscisic Acid; ACGT; AACA; Abiotic Stress

Conflicts of Interest

The authors declare no conflicts of interest.

References

[1]	P. K. Busk and M. Pages, “Regulation of Abscisic Acid-Induced Transcription,” Plant Molecular Biology, Vol. 37, No. 3, 1998, pp. 425-435. doi:10.1023/A:1006058700720
[2]	Q. Shen and T. H. Ho, “Functional Dissection of an Abscisic Acid (ABA)-Inducible Gene Reveals Two Independent ABA Responsive Complexes Each Containing a G-Box and a Novel cis Acting Element,” Plant Cell, Vol. 7, No. 3, 1995, pp. 295-307.
[3]	M. J. Guiltinan, W. R. Marcotte and R. S. Quatrano, “A Plant Leucine Zipper Protein that Recognizes an Abscisic Acid Response Element,” Science, Vol. 250, No. 4978, 1990, pp. 267-271. doi:10.1126/science.2145628
[4]	R. G. K. Donald and A. R. Cashmore, “Mutation of Either G Box or I Box Sequences Profoundly Affects Expression from the Arabidopsis rbcS-1A Promoter,” The EMBO Journal, Vol. 9, No. 6, 1990, pp. 1717-1726.
[5]	I. Jupin and N. H. Chua, “Activation of the CaMV as-1 cis Element by Salicylic Acid: Differential DNA-Binding of a Factor Related to TGA1a,” The EMBO Journal, Vol. 15, No. 20, 1996, pp. 5679-5689.
[6]	T. Umezawa, K. Nakashima, T. Miyakawa, T. Kuromori, M. Tanokura, K. Shinozaki and K. Yamaguchi-Shinozaki, “Molecular Basis of the Core Regulatory Network in ABA Responses: Sensing, Signaling and Transport,” Plant and Cell Physiology, Vol. 51, No. 11, 2010, pp. 1821- 1839. doi:10.1093/pcp/pcq156
[7]	R. Mehrotra, K. Kiran, C. P. Chaturvedi, S. A. Ansari, N. Lodhi, S. Sawant and R. Tuli, “Effect of Copy Number and Spacing of the ACGT and GT cis Elements on Transient Expression of Minimal Promoter in Plants,” Journal of Genetics, Vol. 84, No. 2, 2005, pp. 183-187. doi:10.1007/BF02715844
[8]	T. Hobo, M. Asada, Y. Kowyama and T. Hattori, “ACGT Containing Abscisic Acid Response Element (ABRE) and Coupling Element 3 (CE3) Are Functionally Equivalent,” The Plant Journal, Vol. 19, No. 6, 1999, pp. 679-689. doi:10.1046/j.1365-313x.1999.00565.x
[9]	R. Mehrotra and J. Panwar, “Dimerization of GT Element Interferes Negatively with Gene Activation,” Journal of Genetics, Vol. 88, No. 2, 2009, pp. 257-260. doi:10.1007/s12041-009-0037-7
[10]	R. Mehrotra and S. Mehrotra, “Promoter Activation by ACGT in Response to Salicylic and Abscisic Acids Is Differentially Regulated by the Spacing between Two Copies of the Motif,” Journal of Plant Physiology, Vol. 167, No. 14, 2010, pp. 1214-1218. doi:10.1007/s12041-009-0037-7
[11]	R. Mehrotra, A. Yadav, P. Bhalothia, R. Karan and S. Mehrotra, “Evidence for Directed Evolution of Larger Size Motif in Arabidopsis thaliana Genome,” The Scientific World Journal, Vol. 2012, 2012, Article ID: 983528 doi:10.1100/2012/983528
[12]	T. Izawa, R. Foster and N. H. Chua, “Plant bZIP Protein DNA Binding Specificity,” Journal of Molecular Biology, Vol. 20, No. 4, 1993, pp. 1131-1144. doi:10.1006/jmbi.1993.1230
[13]	R. Foster, T. Izawa and N. H. Chua, “Plant Basic Leucine Zipper Proteins Gather at ACGT Elements,” The EMBO Journal, Vol. 8, No. 2, 1994, pp. 192-200.
[14]	M. Suzuki, M. G. Ketterling and D. R. McCarty, “Quantitative Statistical Analysis of cis-Regulatory Sequences in ABA/VP1- and CBF/DREB1-Regulated Genes of Arabidopsis,” Plant Physiology, Vol. 139, No. 1, 2005, pp. 437- 447. doi:10.1104/pp.104.058412
[15]	R. Mehrotra, G. Gupta, R. Sethi, P. Bhalothia, N. Kumar and S. Mehrotra, “Designer Promoter: An Artwork of cis Engineering,” Plant Molecular Biology, Vol. 75, No. 6, 2011, pp. 527-536. doi:10.1007/s11103-011-9755-3
[16]	M. G. Murray and W. F. Thompson, “Thompson Rapid Isolation of High Molecular Weight Plant DNA,” Nucleic Acids Research, Vol. 8, No. 19, 1980, pp. 4321-4326. doi:10.1093/nar/8.19.4321
[17]	S. V. Sawant, P. K. Singh and R. Tuli, “Pretreatment of Micropojectiles to Improve the Delivery of DNA in Plant Transformation,” Biotechniques, Vol. 29, No. 2, 2000, pp. 246-248.
[18]	T. Murashige and F. Skoog, “A Revised Medium for Rapid Growth and Bioassays with Tobacco Tissue Cultures,” Plant Physiology, Vol. 15, No. 43, 1962, pp. 473- 497.
[19]	C. Reichel, J. Mathur, P. Eckes, K. Langenkemper, C. Koncz, J. Schell, B. Reiss and C. Maas, “Enhanced Green Fluorescence by the Expression of an Aequorea victoria Green Fluorescent Protein Mutant in Mono- and Dicotyledonous Plant Cells,” Proceedings of the National Academy of Sciences of the United States of America, Vol. 93, No. 12, 1996, pp. 5888-5893. doi:10.1073/pnas.93.12.5888
[20]	J. J. Wu, Y. W. Liu and M. X. Sun, “Improved and High throughput Quanti-tative Measurements of Weak GFP Expression in Transgenic Plant Materials,” Plant Cell Report, Vol. 30, No. 7, 2011, pp. 1253-1260. doi:10.1046/j.1365-313x.2000.00797.x
[21]	D. S. Prestridge, “SIGNAL SCAN: A Computer Program That Scans DNA Sequences for Eukaryotic Transcriptional Elements,” Computer Applications in the Biosciences, Vol. 7, No. 2, 1991, pp. 203-206.
[22]	K. Higo, Y. Ugawa, M. Iwamoto and T. Korenaga, “Plant cis-Acting Regulatory DNA Elements (PLACE) Database,” Nucleic Acids Research, Vol. 27, No. 1, 1999, pp. 297-300. doi:10.1093/nar/27.1.297
[23]	C. Wu, H. Washida, Y. Onodera, K. Harada and F. Takaiwa, “Quantitative Nature of the Prolamin-Box, ACGT and AACA Motifs in a Rice Glutelin Gene Promoter: Minimal cis-Element Requirements for Endosperm-Specific Gene Expression,” The Plant Journal, Vol. 23, No. 3, 2000, pp. 415-421. doi:10.1046/j.1365-313x.2000.00797.x
[24]	T. Yoshihara, H. Washida and F. Takaiwa, “A 45-bp Proximal Region Containing AACA and GCN4 Motif Is Sufficient to Confer Endosperm-Specific Expression of the Rice Storage Protein Glutelin Gene, GluA-3,” FEBS Letters, Vol. 383, No. 3, 1996, pp. 2133-2138. doi:10.1016/0014-5793(96)00233-5
[25]	S. R. Kim, Y. Kim and G. An, “Identification of Methyl Jasmonate and Salicylic Acid Response Elements from the Nopaline Synthase (nos) Promoter,” Plant Physiology, Vol. 103, No. 1, 1993, pp. 97-103. doi:10.1104/pp.103.1.97
[26]	S. Sawant, K. Kiran, R. Mehrotra, C. P. Chaturvedi, S. A. Ansari, P. Singh, N. Lodhi and R. Tuli, “A Variety of Synergistic and Antagonistic Interactions Mediated by cis-Acting DNA Motifs Regulate Gene Expression in Plant Cells and Modulate Stability of the Transcription Complex Formed on a Basal Promoter,” Journal of Experimental Botany, Vol. 56, No. 419, pp. 2345-2353. doi:10.1093/jxb/eri227