Root-preferential expression of Newcastle virus glycoproteins driven by NtREL1 promoter in tobacco hairy roots and evaluation of oral delivery in mice

Abstract

Newcastle disease virus (NDV) is a lethal virus in avian species with a disastrous effect on the poultry industry. NDV is enveloped by a host-derived membrane with two glycosylated haemagglutinin-neuraminidase (HN) and Fusion (F) proteins. NDV infection usually leads to death within 2-6 days, so the preexisting antibodies provide the most critical protection for this infection. The HN and F glycoproteins are considered the main targets of the immune system. In the present study, two constructs harboring the HN or F epitopes are sub-cloned separately under the control of a root-specific promoter NtREL1 or CaMV35S (35S Cauliflower Mosaic Virus promoter) as a constitutive promoter. The recombinant vectors were transformed into the Agrobacterium tumefaciens strain LBA4404 and then introduced to tobacco (Nicotiana tabacum L.) leaf disk explants. PCR with specific primers was performed to confirm the presence of the hn and f genes in the genome of the regenerated plants. Then, the positive lines were transformed via non-recombinant A. rhizogenes (strain ATCC15834) to develop hairy roots.HN and F were expressed at 0.37% and 0.33% of TSP using the CaMV35S promoter and at 0.75% and 0.54% of TSP using the NtREL1 promoter, respectively. Furthermore, the mice fed transgenic hairy roots showed a high level of antibody responses (IgG and IgA) against rHN and rF proteins.

Keywords: Hairy roots; Newcastle disease virus; Oral delivery; Recombinant HN and F proteins; Tissue-specific promoter.