Biotic and abiotic stresses are main factors limiting crop plants yields and adaptability. Antimicrobial peptides (AMPs) play a pivotal role in plant immune responses to diverse stresses, and hence, becoming novel and essential molecules for studying plant responses to environmental harsh conditions. In this study, for the rst time, overexpression of recombinant defensin NaD1gene under control of 3x 35S promoter was incorporated into tobacco plants resulting in generating NaD1 transgenic lines. Stable expression of NaD1 in transgenic tobacco lines was con rmed by RT-PCR, and next, presence of NaD1 recombinant peptide was veri ed by ELISA and western blot analysis in transgenic lines. In Silico bioinformatic analysis revealed that the most abundant components in Cisregulatory elements in eleven NaD1 homologs in Nicotiana attenuate (NaDEF genes) are MYB, MYC and ABRE elements suggesting that NaD1 promoter is involved in regulation of abiotic stresses. Overexpression of the NaD1 in transgenic plants led to a signi cant (P ≤ 0.01) increase in the content of chlorophyll a, b, and total chlorophyll under drought stress. Correspondingly, the index of chlorophyll stability signi cantly increased in 3 transgenic lines.Moreover, activities of Catalase (CAT), Peroxidase (POD), Ascorbate peroxidase (APX) and S uperoxide dismutase (SOD) were signi cantly enhanced in response to drought stress in transgenic lines. Among three transgenic lines, line 1 showed the highest chlorophyll level and chlorophyll stability index and a high level of POD, CAT, and SOD enzyme activity under drought stress. The data together suggest that increased antioxidant activity of the enzymes might presumably lead to eliminate ROS levels and maintain the chlorophyll content and stability in response to drought stress. Therefore, the antimicrobial peptide defensin NaD1 can be considered an essential factor in regulation of plant responses to drought stress and could be used in developing transgenic lines resistant to abiotic stresses.

Biotic and abiotic stresses are main factors limiting crop plants yields and adaptability. Antimicrobial peptides (AMPs) play a pivotal role in plant immune responses to diverse stresses, and hence, becoming novel and essential molecules for studying plant responses to environmental harsh conditions. In this study, for the first time, overexpression of recombinant defensin NaD1gene under control of 3x 35S promoter was incorporated into tobacco plants resulting in generating NaD1 transgenic lines. Stable expression of NaD1 in transgenic tobacco lines was confirmed by RT-PCR, and next, presence of NaD1 recombinant peptide was verified by ELISA and western blot analysis in transgenic lines. In Silico bioinformatic analysis revealed that the most abundant components in Cis-regulatory elements in eleven NaD1 homologs in Nicotiana attenuate (NaDEF genes) are MYB, MYC and ABRE elements suggesting that NaD1 promoter is involved in regulation of abiotic stresses. Overexpression of the NaD1 in transgenic plants led to a significant (P ≤ 0.01) increase in the content of chlorophyll a, b, and total chlorophyll under drought stress. Correspondingly, the index of chlorophyll stability significantly increased in 3 transgenic lines. Moreover, activities of Catalase (CAT), Peroxidase (POD), Ascorbate peroxidase (APX) and S uperoxide dismutase (SOD) were significantly enhanced in response to drought stress in transgenic lines. Among three transgenic lines, line 1 showed the highest chlorophyll level and chlorophyll stability index and a high level of POD, CAT, and SOD enzyme activity under drought stress. The data together suggest that increased antioxidant activity of the enzymes might presumably lead to eliminate ROS levels and maintain the chlorophyll content and stability in response to drought stress. Therefore, the antimicrobial peptide defensin NaD1 can be considered an essential factor in regulation of plant responses to drought stress and could be used in developing transgenic lines resistant to abiotic stresses.