Recombinant Promoter (MUASCsV8CP) Driven Totiviral Killer Protein 4 (KP4) Imparts Resistance Against Fungal Pathogens in Transgenic Tobacco

Abstract

Development of disease-resistant plant varieties achieved by engineering anti-microbial transgenes under the control of strong promoters can suffice the inhibition of pathogen growth and simultaneously ensure enhanced crop production. For evaluating the prospect of such strong promoters, we comprehensively characterized the full-length transcript promoter of Cassava Vein Mosaic Virus (CsVMV; -565 to +166) and identified CsVMV8 (-215 to +166) as the highest expressing fragment in both transient and transgenic assays. Further, we designed a new chimeric promoter ‘MUASCsV8CP’ through inter-molecular hybridization among the upstream activation sequence (UAS) of Mirabilis Mosaic Virus (MMV; -297 to -38) and CsVMV8, as the core promoter (CP). The MUASCsV8CP was found to be ∼2.2 and ∼2.4 times stronger than the CsVMV8 and CaMV35S promoters, respectively, while its activity was found to be equivalent to that of the CaMV35S2 promoter. Furthermore, we generated transgenic tobacco plants expressing the totiviral ‘Killer protein KP4’ (KP4) under the control of the MUASCsV8CP promoter. Recombinant KP4 was found to accumulate both in the cytoplasm and apoplast of plant cells. The agar-based killing zone assays revealed enhanced resistance of plant-derived KP4 against two deuteromycetous foliar pathogenic fungi viz. Alternaria alternata and Phoma exigua var. exigua. Also, transgenic plants expressing KP4 inhibited the growth progression of these fungi and conferred significant fungal resistance in detached-leaf and whole plant assays. Taken together, we establish the potential of engineering “in-built” fungal stress-tolerance in plants by expressing KP4 under a novel chimeric caulimoviral promoter in a transgenic approach.

Unraveling the expression of differentially expressed proteins and enzymatic activity in response to Phytophthora nicotianae across different flue-cured tobacco cultivars

“…Soil environmental conditions such as rhizospheric microbial diversity, availability of nutrients and organic matter, soil texture, and soil pH are the main factors responsible for soilborne diseases [ 10 ]. Over the past few decades, important integrated disease management methods such as resistant cultivars [ 11 ], chemical pesticides (synthetic fungicides) [ 12 ], crop rotation, and soil replacement [ 13 ] have been adopted to control this disease. However, these methods have many disadvantages, such as the long-term application of synthetic fungicides may lead to resistance in the pathogen, affect soil microbial diversity, and are environmentally unfriendly [ 1 , 14 ].…”

“…Cultivation and selection of disease-resistance cultivars play an important role to control tobacco black shank disease [ 11 ]. Many studies have proven that the incidence of black shank disease was higher in susceptible cultivars than resistance cultivars [ 17 ].…”

Tinkering Cis Motifs Jigsaw Puzzle Led to Root-Specific Drought-Inducible Novel Synthetic Promoters

“…Most recombinant promoters aim to achieve maximal transcription by optimizing the copy number and spacing of the key cis elements in their sequences [17,33]. Several techniques have been employed to design and test these promoters [17,28,29,33,34]. Approximately 200 such promoters have been successfully developed by engineering different abiotic [35,36] and biotic [17,37] stress-responsive cis elements or domains using these techniques.…”

Effects of Cultured Root and Soil Microbial Communities on the Disease of Nicotiana tabacum Caused by Phytophthora nicotianae

“…Soil environments, including soil type, nutrients, and populations of bacteria and fungi, are dominant factors for black shank occurrence and development (Kyselková et al, 2009). The traditional approaches to control N. tabacum diseases generally include synthetic fungicides (You et al, 2016), soil removal and replacement, crop rotation (Niu et al, 2017), and/or disease-resistant varieties (Deb et al, 2018). However, many disadvantages have been found during the use of those traditional approaches, such as the emergence of fungicide-resistant populations, disruption of local ecosystems, large labor requirements, and threats to human health.…”