Molecular cloning, expression profile and promoter analysis of a novel ethylene responsive transcription factor gene GhERF4 from cotton (Gossypium hirstum)

Ethylene-responsive element binding factors (ERFs) are plant-specific transcription factors, many of which have been linked to stress responses. A novel ERF gene, designated GhERF4, was isolated by RACE-PCR from Gossypium hirstum. The GhERF4 cDNA has a total length of 1061 bp with an open reading frame of 669 bp, encoding a protein of 222 amino acids with a molecular weight of 23.5 kDa and a calculated pI of 9.03. Sequence alignment shows that GhERF4 contains a 58 amino acid long AP2/ERF domain and a RKRP nuclear localization signal, and belongs to a group II protein in the ERF subfamily as typified by the C-terminal ERF-associated Amphiphilic Repression (EAR) motif. Southern blot analysis indicates that GhERF4 is a single copy gene in cotton genome. Using green fluorescent protein fusion, we demonstrate that GhERF4 accumulates specifically in the nucleus of onion epidermis cells. Semi-quantitative RT-PCR reveals that GhERF4 is constitutively expressed in true leaves, roots, seeds and stems. The transcripts of GhERF4 accumulate highly and rapidly when plants are treated with exogenous ethylene, salt, cold, drought stresses and exogenous abscisic acid (ABA) treatment, suggesting that GhERF4 is regulated by certain components of the stress signaling pathway. Promoter analysis indicates that the 5′ upstream region of GhERF4 possesses some elements induced by physiological and environmental factors. These results indicate that GhERF4 may play an important role in response to ethylene, ABA and environmental stresses.