BIOLOGIA PLANTARUM XX (X): XXX-XXX, 20XX 1 Gene expression of the oil palm transcription factor EgAP2-1 during fruit ripening and in response to ethylene and ABA treatments V. OMIDVAR 1,2 , S.N.A. ABDULLAH 1,2 *, M. EBRAHIMI 1 , C.L. HO 2,3 , and M. MAHMOOD 3 Department of Agriculture Technology, Faculty of Agriculture 1 , Laboratory of Plantation Crops, Institute of Tropical Agriculture 2 , and Department of Cell and Molecular Biology, Faculty of Biotechnology and Biomolecular Sciences 3 , University Putra Malaysia, 43400 UPM, Selangor, Malaysia Abstract A cDNA encoding an ERE-binding protein (EgAP2-1) was isolated from the oil palm fruit mesocarp treated with ethylene using yeast one-hybrid assay. EgAP2-1 belongs to the AP2 subfamily of the APETALA2/ethylene-responsive factor (AP2/ERF) proteins and contains two highly conserved AP2/EREBP DNA-binding domains (DNA-BD). Sequence comparison of EgAP2-1 with other AP2 proteins revealed high conservation of the two AP2/EREBP domains and linker region among these proteins. Its protein was localized to the nucleus of onion epidermis cells and showed ERE-specific binding, transcriptional activation, and transactivation properties in yeast and in vitro. Its mRNA was highly expressed in oil palm mesocarp with elevated levels in ripening fruits but not in leaves and roots. EgAP2-1 was induced in mesocarp in response to ethylene and abscisic acid but not other hormonal stimuli, including methyl jasmonate and salicylic acid, and abiotic stresses including drought, cold, and high-salinity. Our results demonstrated a link between the regulation of EgAP2-1 expression and ethylene- and/or ABA-coordinated control of the fruit ripening and suggest a regulatory role for EgAP2-1 during fruit ripening and development in oil palm. Additional key words: abiotic stress, AP2/ERF, ethylene-responsive element, gene expression, yeast one-hybrid assay. Introduction The APETALA2/ethylene-responsive factor (AP2/ERF) superfamily of transcription factors play important regulatory roles in plant responses to various biotic and abiotic stresses as well as in plant growth and development (Yamaguchi and Shinozaki 2006). Completion of genome projects has led to the identification of 147, 167, 132, and 200 AP2/ERF members in Arabidopsis, rice, grapevine, and poplar genomes, respectively (Sakuma et al. 2002, Nakano et al. 2006, Riano-Pachon et al. 2007, Guo et al. 2008, Zhuang et al. 2008). The AP2/ERFs have been categorized into five subfamilies based on the number of AP2/EREBP DNA-BDs including ethylene-responsive factor (ERF), dehydration-responsive element binding protein/c-repeat binding factor (DREB/CBF), APETALA2 (AP2), ABI3/VP-related (RAV), and others (Nakano et al. 2006, Sharoni et al. 2011). The ERF and DREB/CBF subfamilies contain only one AP2/EREBP domain whereas the AP2 has two such domains. The RAV subfamily contains an additional B3 domain along with the AP2/EREBP domain. The AP2/EREBP domain consists of 60 - 70 amino acids and was firstly identified in the Arabidopsis AP2 protein (Jofuku et al. 1994) and later in ethylene responsive element binding proteins (EREBPs) from Nicotiana tabaccum (Ohme-Takagi and Shinshi 1995). The ERFs bind to the ethylene response element (ERE), also called a GCC box, found in the promoter of many pathogenesis-related and other ethylene-regulated genes, such as 1-aminocyclopropane-1-carboxylic acid (ACC) synthase and ACC oxidase, and modulate the plant response to various biotic stresses and developmental processes (Tournier et al. 2003, Qin et al. 2006, El-Sharkawy et al. 2008). Some ERFs, such as the ⎯⎯⎯⎯ Received 8 June 2012, accepted 16 January 2013. Abbreviations: ABA - abscisic acid; AD - activation domain; AP2/ERF - APETALA2/ethylene-responsive factor; BD - binding domain; CRT - c-repeat; CBF - CRT-binding factor; DRE - dehydration-responsive element; DREB - DRE-binding protein; EMSA - electrophoretic mobility shift assay; ERE - ethylene-responsive element; EREBP - ERE-binding protein; ERF - ethylene- responsive factor; GFP - green fluorescent protein; MJ - methyl jasmonate; SA - salicylic acid. Acknowledgments: We thank the Ministry of Science, Technology, and Innovation of Malaysia for providing the research grant through Agriculture Biotechnology Institute. * Corresponding author: fax: (+603) 8938 1612, e-mail: sakmar@agri.upm.edu.my