Molecular characterization and systematic analysis of NtAP2/ERF in tobacco and functional determination of NtRAV-4 under drought stress

The APETALA2/ethylene response factor (AP2/ERF) transcription factor (TF) superfamily play crucial roles in plant growth and development as well as biotic and abiotic stresses response. Here, we systematically characterized 375 AP2/ERF TFs in the Nicotiana tabacum genome. Phylogenetic tree topology and conserved domain number allowed TF classifications into three families of 29 AP2, 341 ERF, and 5 RAV genes, which were unevenly distributed throughout 24 tobacco chromosomes. Gene family expansions were retained from whole genome or segmental duplications followed by tandem duplication. Gene structure and motif analysis revealed intra-group conservation. MicroRNA target site prediction identified nine miR172 family members targeting six NtAP2-family genes; 41 NtAP2/ERFs participated in protein co-regulatory networks. NtAP2/ERF gene global expression profiles ascertained by RNA-seq displayed diverse expression patterns across tissues and under different abiotic and biotic stresses (including drought, cold, and Phytopthora parasitica inoculation). As determined by qRT-PCR, the expression of NtAP2/ERF were induced by five hormone and four abiotic stress. RNA interference of NtRAV-4 in tobacco accelerates seed germination, enhance root development and leaf photosynthetic ability. Suppression of NtRAV-4 increases drought tolerance by improving antioxidant defense ability and reduced relative electrolyte leakage under drought stress. These results enhance understanding of NtAP2/ERF gene function and will facilitate genetic improvement of tobacco stress tolerance.